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Sample records for germline flcn gene

  1. Genetic screening of the FLCN gene identify six novel variants and a Danish founder mutation

    DEFF Research Database (Denmark)

    Rossing, Maria; Albrechtsen, Anders; Skytte, Anne-Bine

    2016-01-01

    Pathogenic germline mutations in the folliculin (FLCN) tumor suppressor gene predispose to Birt-Hogg-Dubé (BHD) syndrome, a rare disease characterized by the development of cutaneous hamartomas (fibrofolliculomas), multiple lung cysts, spontaneous pneumothoraces and renal cell cancer. In this stu...... understanding of BHD syndrome and management of BHD patients.Journal of Human Genetics advance online publication, 13 October 2016; doi:10.1038/jhg.2016.118....

  2. Novel folliculin (FLCN) mutation and familial spontaneous pneumothorax.

    Science.gov (United States)

    Zhu, J-F; Shen, X-Q; Zhu, F; Tian, L

    2017-01-01

    Familial spontaneous pneumothorax is one of the characteristics of Birt-Hogg-Dubé syndrome (BHDS), which is an autosomal dominant disease caused by the mutation of folliculin (FLCN). To investigate the mutation of FLCN gene in a familial spontaneous pneumothorax. Prospective case study. Clinical and genetic data of a Chinese family with four patients who presented spontaneous pneumothorax in the absence of skin lesions or renal tumors were collected. CT scan of patient's lung was applied for observation of pneumothorax. DNA sequencing of the coding exons (4-14 exons) of FLCN was performed for all 11 members of the family and 100 unrelated healthy controls. CT scan of patient's lung showed spontaneous pneumothorax. A mutation (c. 510C > G) that leads to a premature stop codon (p. Y170X) was found in the proband using DNA sequencing of coding exons (4-14 exons) of FLCN. This mutation was also observed in the other affected members of the family. A nonsense mutation of FLCN was found in a spontaneous pneumothorax family. Our results expand the mutational spectrum of FLCN in patients with BHDS. © The Author 2016. Published by Oxford University Press on behalf of the Association of Physicians. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  3. Birt-Hogg-Dubé syndrome: novel FLCN frameshift deletion in daughter and father with renal cell carcinomas.

    Science.gov (United States)

    Näf, Ernst; Laubscher, Dominik; Hopfer, Helmut; Streit, Markus; Matyas, Gabor

    2016-01-01

    Germline mutation of the FLCN gene causes Birt-Hogg-Dubé syndrome (BHD), a rare autosomal dominant condition characterized by skin fibrofolliculomas, lung cysts, spontaneous pneumothorax and renal tumours. We identified a hitherto unreported pathogenic FLCN frameshift deletion c.563delT (p.Phe188Serfs*35) in a family of a 46-year-old woman presented with macrohematuria due to bilateral chromophobe renal carcinomas. A heritable renal cancer was suspected due to the bilaterality of the tumour and as the father of this woman had suffered from renal cancer. Initially, however, BHD was overlooked by the medical team despite the highly suggestive clinical presentation. We assume that BHD is underdiagnosed, at least partially, due to low awareness of this variable condition and to insufficient use of appropriate genetic testing. Our study indicates that BHD and FLCN testing should be routinely considered in patients with positive family or personal history of renal tumours. In addition, we demonstrate how patients and their families can play a driving role in initiating genetic diagnosis, presymptomatic testing of at-risk relatives, targeted disease management, and genetic counselling of rare diseases such as BHD.

  4. Human germline gene editing: Recommendations of ESHG and ESHRE

    NARCIS (Netherlands)

    de Wert, Guido; Pennings, Guido; Clarke, Angus; Eichenlaub-Ritter, Ursula; van El, Carla G.; Forzano, Francesca; Goddijn, Mariëtte; Heindryckx, Björn; Howard, Heidi C.; Radojkovic, Dragica; Rial-Sebbag, Emmanuelle; Tarlatzis, Basil C.; Cornel, Martina C.

    2018-01-01

    Technological developments in gene editing raise high expectations for clinical applications, first of all for somatic gene editing but in theory also for germline gene editing (GLGE). GLGE is currently not allowed in many countries. This makes clinical applications in these countries impossible

  5. Germ-line gene therapy and the medical imperative.

    Science.gov (United States)

    Munson, Ronald; Davis, Lawrence H

    1992-06-01

    Somatic cell gene therapy has yielded promising results. If germ cell gene therapy can be developed, the promise is even greater: hundreds of genetic diseases might be virtually eliminated. But some claim the procedure is morally unacceptable. We thoroughly and sympathetically examine several possible reasons for this claim but find them inadequate. There is no moral reason, then, not to develop and employ germ-line gene therapy. Taking the offensive, we argue next that medicine has a prima facie moral obligation to do so.

  6. Renal cancer and pneumothorax risk in Birt-Hogg-Dubé syndrome; an analysis of 115 FLCN mutation carriers from 35 BHD families

    NARCIS (Netherlands)

    Houweling, A. C.; Gijezen, L. M.; Jonker, M. A.; van Doorn, M. B. A.; Oldenburg, R. A.; van Spaendonck-Zwarts, K. Y.; Leter, E. M.; van Os, T. A.; van Grieken, N. C. T.; Jaspars, E. H.; de Jong, M. M.; Bongers, E. M. H. F.; Johannesma, P. C.; Postmus, P. E.; van Moorselaar, R. J. A.; van Waesberghe, J-H T. M.; Starink, T. M.; van Steensel, M. A. M.; Gille, J. J. P.; Menko, F. H.

    2011-01-01

    Birt-Hogg-Dubé (BHD) syndrome is an autosomal dominant condition caused by germline FLCN mutations, and characterised by fibrofolliculomas, pneumothorax and renal cancer. The renal cancer risk, cancer phenotype and pneumothorax risk of BHD have not yet been fully clarified. The main focus of this

  7. Renal cancer and pneumothorax risk in Birt-Hogg-Dube syndrome; an analysis of 115 FLCN mutation carriers from 35 BHD families

    NARCIS (Netherlands)

    Houweling, A. C.; Gijezen, L. M.; Jonker, M. A.; van Doorn, M. B. A.; Oldenburg, R. A.; van Spaendonck-Zwarts, K. Y.; Leter, E. M.; van Os, T. A.; van Grieken, N. C. T.; Jaspars, E. H.; de Jong, M. M.; Johannesma, P. C.; Postmus, P. E.; van Moorselaar, R. J. A.; van Waesberghe, J-H T. M.; Starink, T. M.; van Steensel, M. A. M.; Gille, J. J. P.; Menko, F. H.; Bongers, Ernie M. H. F.

    2011-01-01

    BACKGROUND: Birt-Hogg-Dube (BHD) syndrome is an autosomal dominant condition caused by germline FLCN mutations, and characterised by fibrofolliculomas, pneumothorax and renal cancer. The renal cancer risk, cancer phenotype and pneumothorax risk of BHD have not yet been fully clarified. The main

  8. Responsible innovation in human germline gene editing: Background document to the recommendations of ESHG and ESHRE

    NARCIS (Netherlands)

    de Wert, Guido; Heindryckx, Björn; Pennings, Guido; Clarke, Angus; Eichenlaub-Ritter, Ursula; van El, Carla G.; Forzano, Francesca; Goddijn, Mariëtte; Howard, Heidi C.; Radojkovic, Dragica; Rial-Sebbag, Emmanuelle; Dondorp, Wybo; Tarlatzis, Basil C.; Cornel, Martina C.

    2018-01-01

    Technological developments in gene editing raise high expectations for clinical applications, including editing of the germline. The European Society of Human Reproduction and Embryology (ESHRE) and the European Society of Human Genetics (ESHG) together developed a Background document and

  9. Germline and somatic mutations in the MTOR gene in focal cortical dysplasia and epilepsy

    DEFF Research Database (Denmark)

    Møller, Rikke S; Weckhuysen, Sarah; Chipaux, Mathilde

    2016-01-01

    OBJECTIVE: To assess the prevalence of somatic MTOR mutations in focal cortical dysplasia (FCD) and of germline MTOR mutations in a broad range of epilepsies. METHODS: We collected 20 blood-brain paired samples from patients with FCD and searched for somatic variants using deep-targeted gene panel...... sequencing. Germline mutations in MTOR were assessed in a French research cohort of 93 probands with focal epilepsies and in a diagnostic Danish cohort of 245 patients with a broad range of epilepsies. Data sharing among collaborators allowed us to ascertain additional germline variants in MTOR. RESULTS: We...... detected recurrent somatic variants (p.Ser2215Phe, p.Ser2215Tyr, and p.Leu1460Pro) in the MTOR gene in 37% of participants with FCD II and showed histologic evidence for activation of the mTORC1 signaling cascade in brain tissue. We further identified 5 novel de novo germline missense MTOR variants in 6...

  10. Constitutional FLCN mutations in patients with suspected Birt-Hogg-Dubé syndrome ascertained for non-cutaneous manifestations.

    Science.gov (United States)

    Maffé, A; Toschi, B; Circo, G; Giachino, D; Giglio, S; Rizzo, A; Carloni, A; Poletti, V; Tomassetti, S; Ginardi, C; Ungari, S; Genuardi, M

    2011-04-01

    Birt-Hogg-Dubé syndrome (BHDS) is characterized by a clinical triad including cutaneous hamartomas originating from hair follicles, lung cysts/pneumothorax, and kidney tumors. Inactivating mutations of the tumor suppressor gene FLCN are identified in most families with BHDS. Usually, patients are referred for genetic examination by dermatologists because of the presence of typical multiple skin tumors with or without additional symptoms. However, because of phenotypic variability and incomplete penetrance, the clinical presentation of BHDS is not yet fully defined. Criteria for genetic testing and diagnosis that take into account variable manifestations have recently been proposed by the European BHD Consortium. We sequenced the FLCN gene coding region in a series of 19 patients selected for kidney and/or lung manifestations. Overall, FLCN mutations were found in 9 of 19 (47%) families and were detected only in probands who had either >2 components of the clinical triad or a single component (renal or pulmonary) along with a family history of another main BHDS manifestation. Typical cutaneous lesions were present only in 8 of 21 FLCN mutation carriers aged >20 years identified in the mutation-positive families. In addition, we provide clinical and molecular evidence that parotid oncocytoma, so far reported in six BHDS cases, is associated with this condition, based on the observation of a patient with bilateral parotid involvement and marked reduction of the wild-type FLCN allele signal in tumor DNA. Overall, the results obtained in this study contribute to the definition of the phenotypic characteristics that should be considered for BHDS diagnosis and FLCN mutation testing. © 2010 John Wiley & Sons A/S.

  11. Responsible innovation in human germline gene editing: Background document to the recommendations of ESHG and ESHRE.

    Science.gov (United States)

    De Wert, Guido; Heindryckx, Björn; Pennings, Guido; Clarke, Angus; Eichenlaub-Ritter, Ursula; van El, Carla G; Forzano, Francesca; Goddijn, Mariëtte; Howard, Heidi C; Radojkovic, Dragica; Rial-Sebbag, Emmanuelle; Dondorp, Wybo; Tarlatzis, Basil C; Cornel, Martina C

    2018-04-01

    Technological developments in gene editing raise high expectations for clinical applications, including editing of the germline. The European Society of Human Reproduction and Embryology (ESHRE) and the European Society of Human Genetics (ESHG) together developed a Background document and Recommendations to inform and stimulate ongoing societal debates. This document provides the background to the Recommendations. Germline gene editing is currently not allowed in many countries. This makes clinical applications in these countries impossible now, even if germline gene editing would become safe and effective. What were the arguments behind this legislation, and are they still convincing? If a technique could help to avoid serious genetic disorders, in a safe and effective way, would this be a reason to reconsider earlier standpoints? This Background document summarizes the scientific developments and expectations regarding germline gene editing, legal regulations at the European level, and ethics for three different settings (basic research, preclinical research and clinical applications). In ethical terms, we argue that the deontological objections (e.g., gene editing goes against nature) do not seem convincing while consequentialist objections (e.g., safety for the children thus conceived and following generations) require research, not all of which is allowed in the current legal situation in European countries. Development of this Background document and Recommendations reflects the responsibility to help society understand and debate the full range of possible implications of the new technologies, and to contribute to regulations that are adapted to the dynamics of the field while taking account of ethical considerations and societal concerns.

  12. Antibody Heavy Chain Variable Domains of Different Germline Gene Origins Diversify through Different Paths

    Directory of Open Access Journals (Sweden)

    Ufuk Kirik

    2017-11-01

    Full Text Available B cells produce antibodies, key effector molecules in health and disease. They mature their properties, including their affinity for antigen, through hypermutation events; processes that involve, e.g., base substitution, codon insertion and deletion, often in association with an isotype switch. Investigations of antibody evolution define modes whereby particular antibody responses are able to form, and such studies provide insight important for instance for development of efficient vaccines. Antibody evolution is also used in vitro for the design of antibodies with improved properties. To better understand the basic concepts of antibody evolution, we analyzed the mutational paths, both in terms of amino acid substitution and insertions and deletions, taken by antibodies of the IgG isotype. The analysis focused on the evolution of the heavy chain variable domain of sets of antibodies, each with an origin in 1 of 11 different germline genes representing six human heavy chain germline gene subgroups. Investigated genes were isolated from cells of human bone marrow, a major site of antibody production, and characterized by next-generation sequencing and an in-house bioinformatics pipeline. Apart from substitutions within the complementarity determining regions, multiple framework residues including those in protein cores were targets of extensive diversification. Diversity, both in terms of substitutions, and insertions and deletions, in antibodies is focused to different positions in the sequence in a germline gene-unique manner. Altogether, our findings create a framework for understanding patterns of evolution of antibodies from defined germline genes.

  13. First report of a de novo germline mutation in the MLH1 gene

    NARCIS (Netherlands)

    Stulp, Rein P; Vos, Yvonne J; Mol, Bart; Karrenbeld, Arend; de Raad, Monique; van der Mijle, Huub J C; Sijmons, Rolf H

    2006-01-01

    Hereditary non-polyposis colorectal carcinoma (HNPCC) is an autosomal dominant disorder associated with colorectal and endometrial cancer and a range of other tumor types. Germline mutations in the DNA mismatch repair (MMR) genes, particularly MLH1, MSH2, and MSH6, underlie this disorder. The vast

  14. Germline mutations in 40 cancer susceptibility genes among Chinese patients with high hereditary risk breast cancer.

    Science.gov (United States)

    Li, Junyan; Jing, Ruilin; Wei, Hongyi; Wang, Minghao; Qi, Xiaowei; Liu, Haoxi; Liu, Jian; Ou, Jianghua; Jiang, Weihua; Tian, Fuguo; Sheng, Yuan; Li, Hengyu; Xu, Hong; Zhang, Ruishan; Guan, Aihua; Liu, Ke; Jiang, Hongchuan; Ren, Yu; He, Jianjun; Huang, Weiwei; Liao, Ning; Cai, Xiangjun; Ming, Jia; Ling, Rui; Xu, Yan; Hu, Chunyan; Zhang, Jianguo; Guo, Baoliang; Ouyang, Lizhi; Shuai, Ping; Liu, Zhenzhen; Zhong, Ling; Zeng, Zhen; Zhang, Ting; Xuan, Zhaoling; Tan, Xuanni; Liang, Junbin; Pan, Qinwen; Chen, Li; Zhang, Fan; Fan, Linjun; Zhang, Yi; Yang, Xinhua; Li, Jingbo; Chen, Chongjian; Jiang, Jun

    2018-05-12

    Multigene panel testing of breast cancer predisposition genes have been extensively conducted in Europe and America, which is relatively rare in Asia however. In this study, we assessed the frequency of germline mutations in 40 cancer predisposition genes, including BRCA1 and BRCA2, among a large cohort of Chinese patients with high hereditary risk of BC. From 2015 to 2016, consecutive BC patients from 26 centers of China with high hereditary risk were recruited (n=937). Clinical information was collected and next-generation sequencing (NGS) was performed using blood samples of participants to identify germline mutations. In total, we acquired 223 patients with putative germline mutations, including 159 in BRCA1/2, 61 in 15 other BC susceptibility genes and 3 in both BRCA1/2 and non-BRCA1/2 gene. Major mutant non-BRCA1/2 genes were TP53 (n=18), PALB2 (n=11), CHEK2 (n=6), ATM (n=6), and BARD1 (n=5). No factors predicted pathologic mutations in non-BRCA1/2 genes when treated as a whole. TP53 mutations were associated with HER-2 positive BC and younger age at diagnosis; and CHEK2 and PALB2 mutations were enriched in patients with luminal BC. Among high hereditary risk Chinese BC patients, 23.8% contained germline mutations, including 6.8% in non-BRCA1/2 genes. TP53 and PALB2 had a relatively high mutation rates (1.9% and 1.2%). Although no factors predicted for detrimental mutations in non-BRCA1/2 genes, some clinical features were associated with mutations of several particular genes. This article is protected by copyright. All rights reserved. © 2018 UICC.

  15. Protection of germline gene expression by the C. elegans Argonaute CSR-1.

    Science.gov (United States)

    Wedeles, Christopher J; Wu, Monica Z; Claycomb, Julie M

    2013-12-23

    In Caenorhabditis elegans, the Piwi-interacting small RNA (piRNA)-mediated germline surveillance system encodes more than 30,000 unique 21-nucleotide piRNAs, which silence a variety of foreign nucleic acids. What mechanisms allow endogenous germline-expressed transcripts to evade silencing by the piRNA pathway? One likely candidate in a protective mechanism is the Argonaute CSR-1, which interacts with 22G-small RNAs that are antisense to nearly all germline-expressed genes. Here, we use an in vivo RNA tethering assay to demonstrate that the recruitment of CSR-1 to a transcript licenses expression of the transcript, protecting it from piRNA-mediated silencing. Licensing occurs mainly at the level of transcription, as we observe changes in pre-mRNA levels consistent with transcriptional activation when CSR-1 is tethered. Furthermore, the recruitment of CSR-1 to a previously silenced locus transcriptionally activates its expression. Together, these results demonstrate a rare positive role for an endogenous Argonaute pathway in heritably licensing and protecting germline transcripts.

  16. Developmental expression of "germline"- and "sex determination"-related genes in the ctenophore Mnemiopsis leidyi.

    Science.gov (United States)

    Reitzel, Adam M; Pang, Kevin; Martindale, Mark Q

    2016-01-01

    An essential developmental pathway in sexually reproducing animals is the specification of germ cells and the differentiation of mature gametes, sperm and oocytes. The "germline" genes vasa, nanos and piwi are commonly identified in primordial germ cells, suggesting a molecular signature for the germline throughout animals. However, these genes are also expressed in a diverse set of somatic stem cells throughout the animal kingdom leaving open significant questions for whether they are required for germline specification. Similarly, members of the Dmrt gene family are essential components regulating sex determination and differentiation in bilaterian animals, but the functions of these transcription factors, including potential roles in sex determination, in early diverging animals remain unknown. The phylogenetic position of ctenophores and the genome sequence of the lobate Mnemiopsis leidyi motivated us to determine the compliment of these gene families in this species and determine expression patterns during development. Our phylogenetic analyses of the vasa, piwi and nanos gene families show that Mnemiopsis has multiple genes in each family with multiple lineage-specific paralogs. Expression domains of Mnemiopsis nanos, vasa and piwi, during embryogenesis from fertilization to the cydippid stage, were diverse, with little overlapping expression and no or little expression in what we think are the germ cells or gametogenic regions. piwi paralogs in Mnemiopsis had distinct expression domains in the ectoderm during development. We observed overlapping expression domains in the apical organ and tentacle apparatus of the cydippid for a subset of "germline genes," which are areas of high cell proliferation, suggesting that these genes are involved with "stem cell" specification and maintenance. Similarly, the five Dmrt genes show diverse non-overlapping expression domains, with no clear evidence for expression in future gametogenic regions of the adult. We also

  17. Germline Mutations in Cancer Predisposition Genes are Frequent in Sporadic Sarcomas

    OpenAIRE

    Chan, Sock Hoai; Lim, Weng Khong; Ishak, Nur Diana Binte; Li, Shao-Tzu; Goh, Wei Lin; Tan, Gek San; Lim, Kiat Hon; Teo, Melissa; Young, Cedric Ng Chuan; Malik, Simeen; Tan, Mann Hong; Teh, Jonathan Yi Hui; Chin, Francis Kuok Choon; Kesavan, Sittampalam; Selvarajan, Sathiyamoorthy

    2017-01-01

    Associations of sarcoma with inherited cancer syndromes implicate genetic predisposition in sarcoma development. However, due to the apparently sporadic nature of sarcomas, little attention has been paid to the role genetic susceptibility in sporadic sarcoma. To address this, we performed targeted-genomic sequencing to investigate the prevalence of germline mutations in known cancer-associated genes within an Asian cohort of sporadic sarcoma patients younger than 50 years old. We observed 13....

  18. Oyster vasa-like gene as a marker of the germline cell development in Crassostrea gigas

    International Nuclear Information System (INIS)

    Fabioux, C.; Huvet, A.; Lelong, C.; Robert, R.; Pouvreau, S.; Daniel, J.Y.; Minguant, C.; Le Pennec, M.

    2004-01-01

    The oyster vasa-like gene was previously demonstrated to be specifically expressed in germline cells of adult oysters Crassostrea gigas. In the present study, this gene was used as a molecular marker to establish the developmental pattern of germline cells during oyster ontogenesis, using whole-mount in situ hybridization and real-time PCR. The Oyvlg transcripts appeared to be localized to the vegetal pole of unfertilized oocytes and maternally transmitted to embryos. At early development, these maternal transcripts were observed to segregate into a single blastomere, from the CD macromere of 2-cell stage to the 4d mesentoblast of blastula. From late blastula stage, the mesentoblast divided into two cell clumps that migrated to both sides of the larvae body and that would correspond to primordial germ cells (PGCs). Based on these results, we postulate that the germline of C. gigas is specified at early development by maternal cytoplasmic determinants including Oyvlg mRNAs, in putative PGCs that would differentiate into germinal stem cells in juvenile oysters

  19. Germline Transgenic Methods for Tracking Cells and Testing Gene Function during Regeneration in the Axolotl

    Science.gov (United States)

    Khattak, Shahryar; Schuez, Maritta; Richter, Tobias; Knapp, Dunja; Haigo, Saori L.; Sandoval-Guzmán, Tatiana; Hradlikova, Kristyna; Duemmler, Annett; Kerney, Ryan; Tanaka, Elly M.

    2013-01-01

    The salamander is the only tetrapod that regenerates complex body structures throughout life. Deciphering the underlying molecular processes of regeneration is fundamental for regenerative medicine and developmental biology, but the model organism had limited tools for molecular analysis. We describe a comprehensive set of germline transgenic strains in the laboratory-bred salamander Ambystoma mexicanum (axolotl) that open up the cellular and molecular genetic dissection of regeneration. We demonstrate tissue-dependent control of gene expression in nerve, Schwann cells, oligodendrocytes, muscle, epidermis, and cartilage. Furthermore, we demonstrate the use of tamoxifen-induced Cre/loxP-mediated recombination to indelibly mark different cell types. Finally, we inducibly overexpress the cell-cycle inhibitor p16INK4a, which negatively regulates spinal cord regeneration. These tissue-specific germline axolotl lines and tightly inducible Cre drivers and LoxP reporter lines render this classical regeneration model molecularly accessible. PMID:24052945

  20. Detection of mismatch repair gene germline mutation carrier among Chinese population with colorectal cancer

    International Nuclear Information System (INIS)

    Jin, Hei-Ying; Zhao, Ronghua; Liu, Xiufang; Li, Vicky Ka Ming; Ding, Yijiang; Yang, Bolin; Geng, Jianxiang; Lai, Rensheng; Ding, Shuqing; Ni, Min

    2008-01-01

    Hereditary nonpolyposis colorectal cancer (HNPCC) is an autosomal dominant syndrome. The National Cancer Institute (NCI) has recommended the Revised Bethesda guidelines for screening HNPCC. There has been a great deal of research on the value of these tests in other countries. However, literature about the Chinese population is scarce. Our objective is to detect and study microsatellite instability (MSI) and mismatch repair (MMR) gene germline mutation carriers among a Chinese population with colorectal cancer. In 146 prospectively recruited consecutive patients with clinically proven colorectal cancer, MSI carriers were identified by analysis of tumor tissue using multiplex fluorescence polymerase chain reaction (PCR) using the NCI recommended panel and classified into microsatellite instability-low (MSI-L), microsatellite instability-high (MSI-H) and microsatellite stable (MSS) groups. Immunohistochemical staining for MSH2, MSH6 and MLH1 on tissue microarrays (TMAs) was performed, and methylation of the MLH1 promoter was analyzed by quantitative methylation specific PCR (MSP). Germline mutation analysis of blood samples was performed for MSH2, MSH6 and MLH1 genes. Thirty-four out of the 146 colorectal cancers (CRCs, 23.2%) were MSI, including 19 MSI-H CRCs and 15 MSI-L CRCS. Negative staining for MSH2 was found in 8 CRCs, negative staining for MSH6 was found in 6 CRCs. One MSI-H CRC was negative for both MSH6 and MSH2. Seventeen CRCs stained negatively for MLH1. MLH1 promoter methylation was determined in 34 MSI CRCs. Hypermethylation of the MLH1 promoter occurred in 14 (73.7%) out of 19 MSI-H CRCs and 5 (33.3%) out of 15 MSI-L CRCs. Among the 34 MSI carriers and one MSS CRC with MLH1 negative staining, 8 had a MMR gene germline mutation, which accounted for 23.5% of all MSI colorectal cancers and 5.5% of all the colorectal cancers. Five patients harbored MSH2 germline mutations, and three patients harbored MSH6 germline mutations. None of the patients had an MLH

  1. The role of germline promoters and I exons in cytokine-induced gene-specific class switch recombination.

    Science.gov (United States)

    Dunnick, Wesley A; Shi, Jian; Holden, Victoria; Fontaine, Clinton; Collins, John T

    2011-01-01

    Germline transcription precedes class switch recombination (CSR). The promoter regions and I exons of these germline transcripts include binding sites for activation- and cytokine-induced transcription factors, and the promoter regions/I exons are essential for CSR. Therefore, it is a strong hypothesis that the promoter/I exons regions are responsible for much of cytokine-regulated, gene-specific CSR. We tested this hypothesis by swapping the germline promoter and I exons for the murine γ1 and γ2a H chain genes in a transgene of the entire H chain C-region locus. We found that the promoter/I exon for γ1 germline transcripts can direct robust IL-4-induced recombination to the γ2a gene. In contrast, the promoter/I exon for the γ2a germline transcripts works poorly in the context of the γ1 H chain gene, resulting in expression of γ1 H chains that is level. Nevertheless, the small amount of recombination to the chimeric γ1 gene is induced by IFN-γ. These results suggest that cytokine regulation of CSR, but not the magnitude of CSR, is regulated by the promoter/I exons.

  2. Activation of germline-specific genes is required for limb regeneration in the Mexican axolotl

    Science.gov (United States)

    Zhu, Wei; Pao, Gerald M; Satoh, Akira; Cummings, Gillian; Monaghan, James R; Harkins, Timothy T; Bryant, Susan V; Voss, S Randal; Gardiner, David M; Hunter, Tony

    2013-01-01

    The capacity for tissue and organ regeneration in humans is dwarfed by comparison to that of salamanders. Emerging evidence suggests that mechanisms learned from the early phase of salamander limb regeneration – wound healing, cellular dedifferentiation and blastemal formation – will reveal therapeutic approaches for tissue regeneration in humans. Here we describe a unique transcriptional fingerprint of regenerating limb tissue in the Mexican axolotl (Ambystoma mexicanum) that is indicative of cellular reprogramming of differentiated cells to a germline-like state. Two genes that are required for self-renewal of germ cells in mice and flies, Piwi-like 1 (PL1) and Piwi-like 2 (PL2), are expressed in limb blastemal cells, the basal layer keratinocytes and the thickened apical epithelial cap in the wound epidermis in the regenerating limb. Depletion of PL1 and PL2 by morpholino oligonucleotides decreased cell proliferation and increased cell death in the blastema leading to a significant retardation of regeneration. Examination of key molecules that are known to be required for limb development or regeneration further revealed that FGF8 is transcriptionally downregulated in the presence of the morpholino oligos, indicating PL1 and PL2 might participate in FGF signaling during limb regeneration. Given the requirement for FGF signaling in limb development and regeneration, the results suggest that PL1 and PL2 function to establish a unique germline-like state that is associated with successful regeneration. PMID:22841627

  3. Preferential Allele Expression Analysis Identifies Shared Germline and Somatic Driver Genes in Advanced Ovarian Cancer

    Science.gov (United States)

    Halabi, Najeeb M.; Martinez, Alejandra; Al-Farsi, Halema; Mery, Eliane; Puydenus, Laurence; Pujol, Pascal; Khalak, Hanif G.; McLurcan, Cameron; Ferron, Gwenael; Querleu, Denis; Al-Azwani, Iman; Al-Dous, Eman; Mohamoud, Yasmin A.; Malek, Joel A.; Rafii, Arash

    2016-01-01

    Identifying genes where a variant allele is preferentially expressed in tumors could lead to a better understanding of cancer biology and optimization of targeted therapy. However, tumor sample heterogeneity complicates standard approaches for detecting preferential allele expression. We therefore developed a novel approach combining genome and transcriptome sequencing data from the same sample that corrects for sample heterogeneity and identifies significant preferentially expressed alleles. We applied this analysis to epithelial ovarian cancer samples consisting of matched primary ovary and peritoneum and lymph node metastasis. We find that preferentially expressed variant alleles include germline and somatic variants, are shared at a relatively high frequency between patients, and are in gene networks known to be involved in cancer processes. Analysis at a patient level identifies patient-specific preferentially expressed alleles in genes that are targets for known drugs. Analysis at a site level identifies patterns of site specific preferential allele expression with similar pathways being impacted in the primary and metastasis sites. We conclude that genes with preferentially expressed variant alleles can act as cancer drivers and that targeting those genes could lead to new therapeutic strategies. PMID:26735499

  4. Meta-analysis of expression of l(3)mbt tumor-associated germline genes supports the model that a soma-to-germline transition is a hallmark of human cancers.

    Science.gov (United States)

    Feichtinger, Julia; Larcombe, Lee; McFarlane, Ramsay J

    2014-05-15

    Evidence is starting to emerge indicating that tumorigenesis in metazoans involves a soma-to-germline transition, which may contribute to the acquisition of neoplastic characteristics. Here, we have meta-analyzed gene expression profiles of the human orthologs of Drosophila melanogaster germline genes that are ectopically expressed in l(3)mbt brain tumors using gene expression datasets derived from a large cohort of human tumors. We find these germline genes, some of which drive oncogenesis in D. melanogaster, are similarly ectopically activated in a wide range of human cancers. Some of these genes normally have expression restricted to the germline, making them of particular clinical interest. Importantly, these analyses provide additional support to the emerging model that proposes a soma-to-germline transition is a general hallmark of a wide range of human tumors. This has implications for our understanding of human oncogenesis and the development of new therapeutic and biomarker targets with clinical potential. © 2013 The Authors. Published by Wiley Periodicals, Inc. on behalf of UICC.

  5. Allele-specific expression in the germline of patients with familial pancreatic cancer: An unbiased approach to cancer gene discovery

    OpenAIRE

    Tan, Aik Choon; Fan, Jian-Bing; Karikari, Collins; Bibikova, Marina; Garcia, Eliza Wickham; Zhou, Lixin; Barker, David; Serre, David; Feldmann, Georg; Hruban, Ralph H.; Klein, Alison P.; Goggins, Michael; Couch, Fergus J.; Hudson, Thomas J.; Winslow, Raimond L.

    2007-01-01

    Physiologic allele-specific expression (ASE) in germline tissues occurs during random X-chromosome inactivation1 and in genomic imprinting,2 wherein the two alleles of a gene in a heterozygous individual are not expressed equally. Recent studies have confirmed the existence of ASE in apparently non-imprinted autosomal genes;3–14 however, the extent of ASE in the human genome is unknown. We explored ASE in lymphoblastoid cell lines of 145 individuals using an oligonucleotide array based assay....

  6. CNVs affecting cancer predisposing genes (CPGs) detected as incidental findings in routine germline diagnostic chromosomal microarray (CMA) testing.

    Science.gov (United States)

    Innes, Josie; Reali, Lisa; Clayton-Smith, Jill; Hall, Georgina; Lim, Derek Hk; Burghel, George J; French, Kim; Khan, Unzela; Walker, Daniel; Lalloo, Fiona; Evans, D Gareth R; McMullan, Dominic; Maher, Eamonn R; Woodward, Emma R

    2018-02-01

    Identification of CNVs through chromosomal microarray (CMA) testing is the first-line investigation in individuals with learning difficulties/congenital abnormalities. Although recognised that CMA testing may identify CNVs encompassing a cancer predisposition gene (CPG), limited information is available on the frequency and nature of such results. We investigated CNV gains and losses affecting 39 CPGs in 3366 pilot index case individuals undergoing CMA testing, and then studied an extended cohort (n=10 454) for CNV losses at 105 CPGs and CNV gains at 9 proto-oncogenes implicated in inherited cancer susceptibility. In the pilot cohort, 31/3366 (0.92%) individuals had a CNV involving one or more of 16/39 CPGs. 30/31 CNVs involved a tumour suppressor gene (TSG), and 1/30 a proto-oncogene (gain of MET ). BMPR1A , TSC2 and TMEM127 were affected in multiple cases. In the second stage analysis, 49/10 454 (0.47%) individuals in the extended cohort had 50 CNVs involving 24/105 CPGs. 43/50 CNVs involved a TSG and 7/50 a proto-oncogene (4 gains, 3 deletions). The most frequently involved genes, FLCN (n=10) and SDHA (n=7), map to the Smith-Magenis and cri-du-chat regions, respectively. Incidental identification of a CNV involving a CPG is not rare and poses challenges for future cancer risk estimation. Prospective data collection from CPG-CNV cohorts ascertained incidentally and through syndromic presentations is required to determine the risks posed by specific CNVs. In particular, ascertainment and investigation of adults with CPG-CNVs and adults with learning disability and cancer, could provide important information to guide clinical management and surveillance. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2018. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  7. A novel germline mutation in the aryl hydrocarbon receptor-interacting protein (AIP) gene in an Italian family with gigantism.

    Science.gov (United States)

    Urbani, C; Russo, D; Raggi, F; Lombardi, M; Sardella, C; Scattina, I; Lupi, I; Manetti, L; Tomisti, L; Marcocci, C; Martino, E; Bogazzi, F

    2014-10-01

    Acromegaly usually occurs as a sporadic disease, but it may be a part of familial pituitary tumor syndromes in rare cases. Germline mutations in the aryl hydrocarbon receptor-interacting protein (AIP) gene have been associated with a predisposition to familial isolated pituitary adenoma. The aim of the present study was to evaluate the AIP gene in a patient with gigantism and in her relatives. Direct sequencing of AIP gene was performed in fourteen members of the family, spanning among three generations. The index case was an 18-year-old woman with gigantism due to an invasive GH-secreting pituitary adenoma and a concomitant tall-cell variant of papillary thyroid carcinoma. A novel germline mutation in the AIP gene (c.685C>T, p.Q229X) was identified in the proband and in two members of her family, who did not present clinical features of acromegaly or other pituitary disorders. Eleven subjects had no mutation in the AIP gene. Two members of the family with clinical features of acromegaly refused either the genetic or the biochemical evaluation. The Q229X mutation was predicted to generate a truncated AIP protein, lacking the last two tetratricopeptide repeat domains and the final C-terminal α-7 helix. We identified a new AIP germline mutation predicted to produce a truncated AIP protein, lacking its biological properties due to the disruption of the C-terminus binding sites for both the chaperones and the client proteins of AIP.

  8. The C. elegans CSR-1 argonaute pathway counteracts epigenetic silencing to promote germline gene expression.

    Science.gov (United States)

    Seth, Meetu; Shirayama, Masaki; Gu, Weifeng; Ishidate, Takao; Conte, Darryl; Mello, Craig C

    2013-12-23

    Organisms can develop adaptive sequence-specific immunity by reexpressing pathogen-specific small RNAs that guide gene silencing. For example, the C. elegans PIWI-Argonaute/piwi-interacting RNA (piRNA) pathway recruits RNA-dependent RNA polymerase (RdRP) to foreign sequences to amplify a transgenerational small-RNA-induced epigenetic silencing signal (termed RNAe). Here, we provide evidence that, in addition to an adaptive memory of silenced sequences, C. elegans can also develop an opposing adaptive memory of expressed/self-mRNAs. We refer to this mechanism, which can prevent or reverse RNAe, as RNA-induced epigenetic gene activation (RNAa). We show that CSR-1, which engages RdRP-amplified small RNAs complementary to germline-expressed mRNAs, is required for RNAa. We show that a transgene with RNAa activity also exhibits accumulation of cognate CSR-1 small RNAs. Our findings suggest that C. elegans adaptively acquires and maintains a transgenerational CSR-1 memory that recognizes and protects self-mRNAs, allowing piRNAs to recognize foreign sequences innately, without the need for prior exposure

  9. Functioning Mediastinal Paraganglioma Associated with a Germline Mutation of von Hippel-Lindau Gene

    Directory of Open Access Journals (Sweden)

    Thibault Bahougne

    2018-05-01

    Full Text Available We report the case of a 21-year old woman presenting with high blood pressure and raised normetanephrine levels. Indium-111-pentetreotide single photon-emission computed tomography with computed tomography (SPECT/CT and 2-deoxy-2-[fluorine-18]fluoro-d-glucose (FDG positron emission tomography/computed tomography (PET/CT imaging showing isolated tracer-uptake by a 2 cm tumor close to the costovertebral angle of the third thoracic vertebra. Thoracic surgery led to normalization of normetanephrine levels. Histological findings were consistent with the presence of a paraganglioma. Mutations in SDHA, SDHB, SDHC, SDHD, RET, SDHAF2, TMEM127, MAX, NF1, FH, MDH2, and EPAS1 were absent, but a heterozygous missense mutation, c.311G > T, was found in exon 1 of the von Hippel-Lindau gene, VHL, resulting in a glycine to valine substitution in the VHL protein at position 104, p.Gly104Val. This same mutation was found in both the mother and the 17-year old sister in whom a small retinal hemangioblastoma was also found. We diagnose an unusual functional mediastinal paraganglioma in this young patient with a germline VHL gene mutation, a mutation previously described as inducing polycythemia and/or pheochromocytoma but not paraganglioma or retinal hemangioblastoma.

  10. Germ-line mutations of the p53 tumor suppressor gene in patients with high risk for cancer inactivate the p53 protein.

    Science.gov (United States)

    Frebourg, T; Kassel, J; Lam, K T; Gryka, M A; Barbier, N; Andersen, T I; Børresen, A L; Friend, S H

    1992-07-15

    Germ-line mutations in the p53 tumor suppressor gene have been observed in patients with Li-Fraumeni syndrome, brain tumors, second malignancies, and breast cancers. It is unclear whether all of these mutations have inactivated p53 and thereby provide an increased risk for cancer. Therefore, it is necessary to establish the biological significance of these germ-line mutations by the functional and structural analysis of the resulting mutant p53 proteins. We analyzed the ability of seven germ-line mutant proteins observed in patients with Li-Fraumeni syndrome, second primary neoplasms, or familial breast cancer to block the growth of malignant cells and compared the structural properties of the mutant proteins to that of the wild-type protein. Six of seven missense mutations disrupted the growth inhibitory properties and structure of the wild-type protein. One germ-line mutation retained the features of the wild-type p53. Genetic analysis of the breast cancer family in which this mutation was observed indicated that this germ-line mutation was not associated with the development of cancer. These results demonstrate that germ-line p53 mutations observed in patients with Li-Fraumeni syndrome and with second malignancies have inactivated the p53 tumor suppressor gene. The inability of the germ-line p53 mutants to block the growth of malignant cells can explain why patients with these germ-line mutations have an increased risk for cancer. The observation of a functionally silent germ-line mutation indicates that, before associating a germ-line tumor suppressor gene mutation with cancer risk, it is prudent to consider its functional significance.

  11. Germ-line mutations of the p53 tumor suppressor gene in patients with high risk for cancer inactivate the p53 protein.

    Science.gov (United States)

    Frebourg, T; Kassel, J; Lam, K T; Gryka, M A; Barbier, N; Andersen, T I; Børresen, A L; Friend, S H

    1992-01-01

    Germ-line mutations in the p53 tumor suppressor gene have been observed in patients with Li-Fraumeni syndrome, brain tumors, second malignancies, and breast cancers. It is unclear whether all of these mutations have inactivated p53 and thereby provide an increased risk for cancer. Therefore, it is necessary to establish the biological significance of these germ-line mutations by the functional and structural analysis of the resulting mutant p53 proteins. We analyzed the ability of seven germ-line mutant proteins observed in patients with Li-Fraumeni syndrome, second primary neoplasms, or familial breast cancer to block the growth of malignant cells and compared the structural properties of the mutant proteins to that of the wild-type protein. Six of seven missense mutations disrupted the growth inhibitory properties and structure of the wild-type protein. One germ-line mutation retained the features of the wild-type p53. Genetic analysis of the breast cancer family in which this mutation was observed indicated that this germ-line mutation was not associated with the development of cancer. These results demonstrate that germ-line p53 mutations observed in patients with Li-Fraumeni syndrome and with second malignancies have inactivated the p53 tumor suppressor gene. The inability of the germ-line p53 mutants to block the growth of malignant cells can explain why patients with these germ-line mutations have an increased risk for cancer. The observation of a functionally silent germ-line mutation indicates that, before associating a germ-line tumor suppressor gene mutation with cancer risk, it is prudent to consider its functional significance. Images PMID:1631137

  12. Specific Tandem 3'UTR Patterns and Gene Expression Profiles in Mouse Thy1+ Germline Stem Cells.

    Directory of Open Access Journals (Sweden)

    Yan Huang

    Full Text Available A recently developed strategy of sequencing alternative polyadenylation (APA sites (SAPAS with second-generation sequencing technology can be used to explore complete genome-wide patterns of tandem APA sites and global gene expression profiles. spermatogonial stem cells (SSCs maintain long-term reproductive abilities in male mammals. The detailed mechanisms by which SSCs self-renew and generate mature spermatozoa are not clear. To understand the specific alternative polyadenylation pattern and global gene expression profile of male germline stem cells (GSCs, mainly referred to SSCs here, we isolated and purified mouse Thy1+ cells from testis by magnetic-activated cell sorting (MACS and then used the SAPAS method for analysis, using pluripotent embryonic stem cells (ESCs and differentiated mouse embryonic fibroblast cells (MEFs as controls. As a result, we obtained 99,944 poly(A sites, approximately 40% of which were newly detected in our experiments. These poly(A sites originated from three mouse cell types and covered 17,499 genes, including 831 long non-coding RNA (lncRNA genes. We observed that GSCs tend to have shorter 3'UTR lengths while MEFs tend towards longer 3'UTR lengths. We also identified 1337 genes that were highly expressed in GSCs, and these genes were highly consistent with the functional characteristics of GSCs. Our detailed bioinformatics analysis identified APA site-switching events at 3'UTRs and many new specifically expressed genes in GSCs, which we experimentally confirmed. Furthermore, qRT-PCR was performed to validate several events of the 334 genes with distal-to-proximal poly(A switch in GSCs. Consistently APA reporter assay confirmed the total 3'UTR shortening in GSCs compared to MEFs. We also analyzed the cis elements around the proximal poly(A site preferentially used in GSCs and found C-rich elements may contribute to this regulation. Overall, our results identified the expression level and polyadenylation site

  13. Specific Tandem 3'UTR Patterns and Gene Expression Profiles in Mouse Thy1+ Germline Stem Cells

    Science.gov (United States)

    Lin, Zhuoheng; Feng, Xuyang; Jiang, Xue; Songyang, Zhou; Huang, Junjiu

    2015-01-01

    A recently developed strategy of sequencing alternative polyadenylation (APA) sites (SAPAS) with second-generation sequencing technology can be used to explore complete genome-wide patterns of tandem APA sites and global gene expression profiles. spermatogonial stem cells (SSCs) maintain long-term reproductive abilities in male mammals. The detailed mechanisms by which SSCs self-renew and generate mature spermatozoa are not clear. To understand the specific alternative polyadenylation pattern and global gene expression profile of male germline stem cells (GSCs, mainly referred to SSCs here), we isolated and purified mouse Thy1+ cells from testis by magnetic-activated cell sorting (MACS) and then used the SAPAS method for analysis, using pluripotent embryonic stem cells (ESCs) and differentiated mouse embryonic fibroblast cells (MEFs) as controls. As a result, we obtained 99,944 poly(A) sites, approximately 40% of which were newly detected in our experiments. These poly(A) sites originated from three mouse cell types and covered 17,499 genes, including 831 long non-coding RNA (lncRNA) genes. We observed that GSCs tend to have shorter 3'UTR lengths while MEFs tend towards longer 3'UTR lengths. We also identified 1337 genes that were highly expressed in GSCs, and these genes were highly consistent with the functional characteristics of GSCs. Our detailed bioinformatics analysis identified APA site-switching events at 3'UTRs and many new specifically expressed genes in GSCs, which we experimentally confirmed. Furthermore, qRT-PCR was performed to validate several events of the 334 genes with distal-to-proximal poly(A) switch in GSCs. Consistently APA reporter assay confirmed the total 3'UTR shortening in GSCs compared to MEFs. We also analyzed the cis elements around the proximal poly(A) site preferentially used in GSCs and found C-rich elements may contribute to this regulation. Overall, our results identified the expression level and polyadenylation site profiles and

  14. Hyperthyroidism caused by a germline activating mutation of the thyrotropin receptor gene: difficulties in diagnosis and therapy.

    Science.gov (United States)

    Bertalan, Rita; Sallai, Agnes; Sólyom, János; Lotz, Gábor; Szabó, István; Kovács, Balázs; Szabó, Eva; Patócs, Attila; Rácz, Károly

    2010-03-01

    Germline activating mutations of the thyrotropin receptor (TSHR) gene have been considered as the only known cause of sporadic nonautoimmune hyperthyroidism in the pediatric population. Here we describe the long-term follow-up and evaluation of a patient with sporadic nonautoimmune primary hyperthyroidism who was found to have a de novo germline activating mutation of the TSHR gene. The patient was an infant who presented at the age of 10 months in an unconscious state with exsiccation, wet skin, fever, and tachycardia. Nonautoimmune primary hyperthyroidism was diagnosed, and brain magnetic resonance imaging and computed tomography showed also Arnold-Chiari malformation type I. Continuous propylthiouracil treatment resulted in a prolonged clinical cure lasting for 10 years. At the age of 11 years and 5 months the patient underwent subtotal thyroidectomy because of symptoms of trachea compression caused by a progressive multinodular goiter. However, 2 months after surgery, hormonal evaluation indicated recurrent hyperthyroidism and the patient was treated with propylthiouracil during the next 4 years. At the age of 15 years the patient again developed symptoms of trachea compression. Radioiodine treatment resulted in a regression of the recurrent goiter and a permanent cure of hyperthyroidism without relapse during the last 3 years of his follow-up. Sequencing of exon 10 of the TSHR gene showed a de novo heterozygous germline I630L mutation, which has been previously described as activating mutation at somatic level in toxic thyroid nodules. The I630L mutation of the TSHR gene occurs not only at somatic level in toxic thyroid nodules, but also its presence in germline is associated with nonautoimmune primary hyperthyroidism. Our case report demonstrates that in this disorder a continuous growth of the thyroid occurs without any evidence of elevated TSH due to antithyroid drug overdosing. This may justify previous recommendations for early treatment of affected

  15. Germline stem cell gene PIWIL2 mediates DNA repair through relaxation of chromatin.

    Directory of Open Access Journals (Sweden)

    De-Tao Yin

    Full Text Available DNA damage response (DDR is an intrinsic barrier of cell to tumorigenesis initiated by genotoxic agents. However, the mechanisms underlying the DDR are not completely understood despite of extensive investigation. Recently, we have reported that ectopic expression of germline stem cell gene PIWIL2 is associated with tumor stem cell development, although the underlying mechanisms are largely unknown. Here we show that PIWIL2 is required for the repair of DNA-damage induced by various types of genotoxic agents. Upon ultraviolet (UV irradiation, silenced PIWIL2 gene in normal human fibroblasts was transiently activated after treatment with UV light. This activation was associated with DNA repair, because Piwil2-deficienct mouse embryonic fibroblasts (mili(-/- MEFs were defective in cyclobutane pyrimidine dimers (CPD repair after UV treatment. As a result, the UV-treated mili(-/- MEFs were more susceptible to apoptosis, as characterized by increased levels of DNA damage-associated apoptotic proteins, such as active caspase-3, cleaved Poly (ADP-ribose polymerase (PARP and Bik. The impaired DNA repair in the mili(-/- MEFs was associated with the reductions of histone H3 acetylation and chromatin relaxation, although the DDR pathway downstream chromatin relaxation appeared not to be directly affected by Piwil2. Moreover, guanine-guanine (Pt-[GG] and double strand break (DSB repair were also defective in the mili(-/- MEFs treated by genotoxic chemicals Cisplatin and ionizing radiation (IR, respectively. The results indicate that Piwil2 can mediate DNA repair through an axis of Piwil2 → histone acetylation → chromatin relaxation upstream DDR pathways. The findings reveal a new role for Piwil2 in DNA repair and suggest that Piwil2 may act as a gatekeeper against DNA damage-mediated tumorigenesis.

  16. Exome sequencing reveals frequent deleterious germline variants in cancer susceptibility genes in women with invasive breast cancer undergoing neoadjuvant chemotherapy.

    Science.gov (United States)

    Ellingson, Marissa S; Hart, Steven N; Kalari, Krishna R; Suman, Vera; Schahl, Kimberly A; Dockter, Travis J; Felten, Sara J; Sinnwell, Jason P; Thompson, Kevin J; Tang, Xiaojia; Vedell, Peter T; Barman, Poulami; Sicotte, Hugues; Eckel-Passow, Jeanette E; Northfelt, Donald W; Gray, Richard J; McLaughlin, Sarah A; Moreno-Aspitia, Alvaro; Ingle, James N; Moyer, Ann M; Visscher, Daniel W; Jones, Katie; Conners, Amy; McDonough, Michelle; Wieben, Eric D; Wang, Liewei; Weinshilboum, Richard; Boughey, Judy C; Goetz, Matthew P

    2015-09-01

    When sequencing blood and tumor samples to identify targetable somatic variants for cancer therapy, clinically relevant germline variants may be uncovered. We evaluated the prevalence of deleterious germline variants in cancer susceptibility genes in women with breast cancer referred for neoadjuvant chemotherapy and returned clinically actionable results to patients. Exome sequencing was performed on blood samples from women with invasive breast cancer referred for neoadjuvant chemotherapy. Germline variants within 142 hereditary cancer susceptibility genes were filtered and reviewed for pathogenicity. Return of results was offered to patients with deleterious variants in actionable genes if they were not aware of their result through clinical testing. 124 patients were enrolled (median age 51) with the following subtypes: triple negative (n = 43, 34.7%), HER2+ (n = 37, 29.8%), luminal B (n = 31, 25%), and luminal A (n = 13, 10.5%). Twenty-eight deleterious variants were identified in 26/124 (21.0%) patients in the following genes: ATM (n = 3), BLM (n = 1), BRCA1 (n = 4), BRCA2 (n = 8), CHEK2 (n = 2), FANCA (n = 1), FANCI (n = 1), FANCL (n = 1), FANCM (n = 1), FH (n = 1), MLH3 (n = 1), MUTYH (n = 2), PALB2 (n = 1), and WRN (n = 1). 121/124 (97.6%) patients consented to return of research results. Thirteen (10.5%) had actionable variants, including four that were returned to patients and led to changes in medical management. Deleterious variants in cancer susceptibility genes are highly prevalent in patients with invasive breast cancer referred for neoadjuvant chemotherapy undergoing exome sequencing. Detection of these variants impacts medical management.

  17. Germ-line mutations of the p53 tumor suppressor gene in patients with high risk for cancer inactivate the p53 protein.

    OpenAIRE

    Frebourg, T; Kassel, J; Lam, K T; Gryka, M A; Barbier, N; Andersen, T I; Børresen, A L; Friend, S H

    1992-01-01

    Germ-line mutations in the p53 tumor suppressor gene have been observed in patients with Li-Fraumeni syndrome, brain tumors, second malignancies, and breast cancers. It is unclear whether all of these mutations have inactivated p53 and thereby provide an increased risk for cancer. Therefore, it is necessary to establish the biological significance of these germ-line mutations by the functional and structural analysis of the resulting mutant p53 proteins. We analyzed the ability of seven germ-...

  18. FLCN and AMPK Confer Resistance to Hyperosmotic Stress via Remodeling of Glycogen Stores.

    Directory of Open Access Journals (Sweden)

    Elite Possik

    2015-10-01

    Full Text Available Mechanisms of adaptation to environmental changes in osmolarity are fundamental for cellular and organismal survival. Here we identify a novel osmotic stress resistance pathway in Caenorhabditis elegans (C. elegans, which is dependent on the metabolic master regulator 5'-AMP-activated protein kinase (AMPK and its negative regulator Folliculin (FLCN. FLCN-1 is the nematode ortholog of the tumor suppressor FLCN, responsible for the Birt-Hogg-Dubé (BHD tumor syndrome. We show that flcn-1 mutants exhibit increased resistance to hyperosmotic stress via constitutive AMPK-dependent accumulation of glycogen reserves. Upon hyperosmotic stress exposure, glycogen stores are rapidly degraded, leading to a significant accumulation of the organic osmolyte glycerol through transcriptional upregulation of glycerol-3-phosphate dehydrogenase enzymes (gpdh-1 and gpdh-2. Importantly, the hyperosmotic stress resistance in flcn-1 mutant and wild-type animals is strongly suppressed by loss of AMPK, glycogen synthase, glycogen phosphorylase, or simultaneous loss of gpdh-1 and gpdh-2 enzymes. Our studies show for the first time that animals normally exhibit AMPK-dependent glycogen stores, which can be utilized for rapid adaptation to either energy stress or hyperosmotic stress. Importantly, we show that glycogen accumulates in kidneys from mice lacking FLCN and in renal tumors from a BHD patient. Our findings suggest a dual role for glycogen, acting as a reservoir for energy supply and osmolyte production, and both processes might be supporting tumorigenesis.

  19. Germline variants in MRE11/RAD50/NBN complex genes in childhood leukemia

    International Nuclear Information System (INIS)

    Mosor, Maria; Ziółkowska-Suchanek, Iwona; Nowicka, Karina; Dzikiewicz-Krawczyk, Agnieszka; Januszkiewicz–Lewandowska, Danuta; Nowak, Jerzy

    2013-01-01

    The MRE11, RAD50, and NBN genes encode proteins of the MRE11-RAD50-NBN (MRN) complex involved in cellular response to DNA damage and the maintenance of genome stability. In our previous study we showed that the germline p.I171V mutation in NBN may be considered as a risk factor in the development of childhood acute lymphoblastic leukemia (ALL) and some specific haplotypes of that gene may be associated with childhood leukemia. These findings raise important questions about the role of mutations in others genes of the MRN complex in childhood leukemia. The aim of this study was to answer the question whether MRE11 and RAD50 alterations may be associated with childhood ALL or AML. We estimated the frequency of constitutional mutations and polymorphisms in selected regions of MRE11, RAD50, and NBN in the group of 220 children diagnosed with childhood leukemias and controls (n=504/2200). The analysis was performed by specific amplification of region of interest by PCR and followed by multi-temperature single-strand conformation polymorphism (PCR-MSSCP) technique. We performed two molecular tests to examine any potential function of the detected the c.551+19G>A SNP in RAD50 gene. To our knowledge, this is the first analysis of the MRE11, RAD50 and NBN genes in childhood leukemia. The frequency of either the AA genotype or A allele of RAD50-rs17166050 were significantly different in controls compared to leukemia group (ALL+AML) (p<0.0019 and p<0.0019, respectively). The cDNA analysis of AA or GA genotypes carriers has not revealed evidence of splicing abnormality of RAD50 pre-mRNA. We measured the allelic-specific expression of G and A alleles at c.551+19G>A and the statistically significant overexpression of the G allele has been observed. Additionally we confirmed the higher incidence of the p.I171V mutation in the leukemia group (7/220) than among controls (12/2400) (p<0.0001). The formerly reported sequence variants in the RAD50 and MRE11 gene may not constitute a

  20. Maternal Germline-Specific Genes in the Asian Malaria Mosquito Anopheles stephensi: Characterization and Application for Disease Control

    Science.gov (United States)

    Biedler, James K.; Qi, Yumin; Pledger, David; Macias, Vanessa M.; James, Anthony A.; Tu, Zhijian

    2014-01-01

    Anopheles stephensi is a principal vector of urban malaria on the Indian subcontinent and an emerging model for molecular and genetic studies of mosquito biology. To enhance our understanding of female mosquito reproduction, and to develop new tools for basic research and for genetic strategies to control mosquito-borne infectious diseases, we identified 79 genes that displayed previtellogenic germline-specific expression based on RNA-Seq data generated from 11 life stage–specific and sex-specific samples. Analysis of this gene set provided insights into the biology and evolution of female reproduction. Promoters from two of these candidates, vitellogenin receptor and nanos, were used in independent transgenic cassettes for the expression of artificial microRNAs against suspected mosquito maternal-effect genes, discontinuous actin hexagon and myd88. We show these promoters have early germline-specific expression and demonstrate 73% and 42% knockdown of myd88 and discontinuous actin hexagon mRNA in ovaries 48 hr after blood meal, respectively. Additionally, we demonstrate maternal-specific delivery of mRNA and protein to progeny embryos. We discuss the application of this system of maternal delivery of mRNA/miRNA/protein in research on mosquito reproduction and embryonic development, and for the development of a gene drive system based on maternal-effect dominant embryonic arrest. PMID:25480960

  1. A novel heterozygous germline deletion in MSH2 gene in a five generation Chinese family with Lynch syndrome

    OpenAIRE

    Wu, Bin; Ji, Wuyang; Liang, Shengran; Ling, Chao; You, Yan; Xu, Lai; Zhong, Min-Er; Xiao, Yi; Qiu, Hui-Zhong; Lu, Jun-Yang; Banerjee, Santasree

    2017-01-01

    Lynch syndrome (LS) is one of the most common familial forms of colorectal cancer predisposing syndrome with an autosomal dominant mode of inheritance. LS is caused by the germline mutations in DNA mismatch repair (MMR) genes including MSH2, MLH1, MSH6 and PMS2. Clinically, LS is characterized by high incidence of early-onset colorectal cancer as well as endometrial, small intestinal and urinary tract cancers, usually occur in the third to fourth decade of the life. Here we describe a five ge...

  2. In Genes We Trust: Germline Engineering, Eugenics, and the Future of the Human Genome.

    Science.gov (United States)

    Powell, Russell

    2015-12-01

    Liberal proponents of genetic engineering maintain that developing human germline modification technologies is morally desirable because it will result in a net improvement in human health and well-being. Skeptics of germline modification, in contrast, fear evolutionary harms that could flow from intervening in the human germline, and worry that such programs, even if well intentioned, could lead to a recapitulation of the scientifically and morally discredited projects of the old eugenics. Some bioconservatives have appealed as well to the value of retaining our "given" human biological nature as a reason for restraining the development and use of human genetic modification technologies even where they would tend to increase well-being. In this article, I argue that germline intervention will be necessary merely to sustain the levels of genetic health that we presently enjoy for future generations-a goal that should appeal to bioliberals and bioconservatives alike. This is due to the population-genetic consequences of relaxed selection pressures in human populations caused by the increasing efficacy and availability of conventional medicine. This heterodox conclusion, which I present as a problem of intergenerational justice, has been overlooked in medicine and bioethics due to certain misconceptions about human evolution, which I attempt to rectify, as well as the sordid history of Darwinian approaches to medicine and social policy, which I distinguish from the present argument. © The Author 2015. Published by Oxford University Press, on behalf of the Journal of Medicine and Philosophy Inc. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  3. Heterozygous Germline Mutations in the CBL Tumor-Suppressor Gene Cause a Noonan Syndrome-like Phenotype

    Science.gov (United States)

    Martinelli, Simone; De Luca, Alessandro; Stellacci, Emilia; Rossi, Cesare; Checquolo, Saula; Lepri, Francesca; Caputo, Viviana; Silvano, Marianna; Buscherini, Francesco; Consoli, Federica; Ferrara, Grazia; Digilio, Maria C.; Cavaliere, Maria L.; van Hagen, Johanna M.; Zampino, Giuseppe; van der Burgt, Ineke; Ferrero, Giovanni B.; Mazzanti, Laura; Screpanti, Isabella; Yntema, Helger G.; Nillesen, Willy M.; Savarirayan, Ravi; Zenker, Martin; Dallapiccola, Bruno; Gelb, Bruce D.; Tartaglia, Marco

    2010-01-01

    RAS signaling plays a key role in controlling appropriate cell responses to extracellular stimuli and participates in early and late developmental processes. Although enhanced flow through this pathway has been established as a major contributor to oncogenesis, recent discoveries have revealed that aberrant RAS activation causes a group of clinically related developmental disorders characterized by facial dysmorphism, a wide spectrum of cardiac disease, reduced growth, variable cognitive deficits, ectodermal and musculoskeletal anomalies, and increased risk for certain malignancies. Here, we report that heterozygous germline mutations in CBL, a tumor-suppressor gene that is mutated in myeloid malignancies and encodes a multivalent adaptor protein with E3 ubiquitin ligase activity, can underlie a phenotype with clinical features fitting or partially overlapping Noonan syndrome (NS), the most common condition of this disease family. Independent CBL mutations were identified in two sporadic cases and two families from among 365 unrelated subjects who had NS or suggestive features and were negative for mutations in previously identified disease genes. Phenotypic heterogeneity and variable expressivity were documented. Mutations were missense changes altering evolutionarily conserved residues located in the RING finger domain or the linker connecting this domain to the N-terminal tyrosine kinase binding domain, a known mutational hot spot in myeloid malignancies. Mutations were shown to affect CBL-mediated receptor ubiquitylation and dysregulate signal flow through RAS. These findings document that germline mutations in CBL alter development to cause a clinically variable condition that resembles NS and that possibly predisposes to malignancies. PMID:20619386

  4. Assessing the spectrum of germline variation in Fanconi anemia genes among patients with head and neck carcinoma before age 50.

    Science.gov (United States)

    Chandrasekharappa, Settara C; Chinn, Steven B; Donovan, Frank X; Chowdhury, Naweed I; Kamat, Aparna; Adeyemo, Adebowale A; Thomas, James W; Vemulapalli, Meghana; Hussey, Caroline S; Reid, Holly H; Mullikin, James C; Wei, Qingyi; Sturgis, Erich M

    2017-10-15

    Patients with Fanconi anemia (FA) have an increased risk for head and neck squamous cell carcinoma (HNSCC). The authors sought to determine the prevalence of undiagnosed FA and FA carriers among patients with HNSCC as well as an age cutoff for FA genetic screening. Germline DNA samples from 417 patients with HNSCC aged <50 years were screened for sequence variants by targeted next-generation sequencing of the entire length of 16 FA genes. The sequence revealed 194 FA gene variants in 185 patients (44%). The variant spectrum was comprised of 183 nonsynonymous point mutations, 9 indels, 1 large deletion, and 1 synonymous variant that was predicted to effect splicing. One hundred eight patients (26%) had at least 1 rare variant that was predicted to be damaging, and 57 (14%) had at least 1 rare variant that was predicted to be damaging and had been previously reported. Fifteen patients carried 2 rare variants or an X-linked variant in an FA gene. Overall, an age cutoff for FA screening was not identified among young patients with HNSCC, because there were no significant differences in mutation rates when patients were stratified by age, tumor site, ethnicity, smoking status, or human papillomavirus status. However, an increased burden, or mutation load, of FA gene variants was observed in carriers of the genes FA complementation group D2 (FANCD2), FANCE, and FANCL in the HNSCC patient cohort relative to the 1000 Genomes population. FA germline functional variants offer a novel area of study in HNSCC tumorigenesis. FANCE and FANCL, which are components of the core complex, are known to be responsible for the recruitment and ubiquitination, respectively, of FANCD2, a critical step in the FA DNA repair pathway. In the current cohort, the increased mutation load of FANCD2, FANCE, and FANCL variants among younger patients with HNSCC indicates the importance of the FA pathway in HNSCC. Cancer 2017;123:3943-54. © 2017 American Cancer Society. © 2017 American Cancer Society.

  5. Discovery of the First Germline-Restricted Gene by Subtractive Transcriptomic Analysis in the Zebra Finch, Taeniopygia guttata.

    Science.gov (United States)

    Biederman, Michelle K; Nelson, Megan M; Asalone, Kathryn C; Pedersen, Alyssa L; Saldanha, Colin J; Bracht, John R

    2018-05-21

    Developmentally programmed genome rearrangements are rare in vertebrates, but have been reported in scattered lineages including the bandicoot, hagfish, lamprey, and zebra finch (Taeniopygia guttata) [1]. In the finch, a well-studied animal model for neuroendocrinology and vocal learning [2], one such programmed genome rearrangement involves a germline-restricted chromosome, or GRC, which is found in germlines of both sexes but eliminated from mature sperm [3, 4]. Transmitted only through the oocyte, it displays uniparental female-driven inheritance, and early in embryonic development is apparently eliminated from all somatic tissue in both sexes [3, 4]. The GRC comprises the longest finch chromosome at over 120 million base pairs [3], and previously the only known GRC-derived sequence was repetitive and non-coding [5]. Because the zebra finch genome project was sourced from male muscle (somatic) tissue [6], the remaining genomic sequence and protein-coding content of the GRC remain unknown. Here we report the first protein-coding gene from the GRC: a member of the α-soluble N-ethylmaleimide sensitive fusion protein (NSF) attachment protein (α-SNAP) family hitherto missing from zebra finch gene annotations. In addition to the GRC-encoded α-SNAP, we find an additional paralogous α-SNAP residing in the somatic genome (a somatolog)-making the zebra finch the first example in which α-SNAP is not a single-copy gene. We show divergent, sex-biased expression for the paralogs and also that positive selection is detectable across the bird α-SNAP lineage, including the GRC-encoded α-SNAP. This study presents the identification and evolutionary characterization of the first protein-coding GRC gene in any organism. Copyright © 2018 Elsevier Ltd. All rights reserved.

  6. The Budapest Meeting 2005. Intensified networking on ethics of science : The case of reproductive cloning, germline gene therapy and human dignity

    NARCIS (Netherlands)

    van Steendam, Guido; Dinnyes, Andras; Mallet, Jacques; Roosendaal, Hans E.

    2006-01-01

    This paper reports on the meeting of the Sounding Board of the EU Reprogenetics Project that was held in Budapest, Hungary, 6–9 November 2005. The Reprogenetics Project runs from 2004 until 2007 and has a brief to study the ethical aspects of human reproductive cloning and germline gene therapy.

  7. Almost 2% of Spanish breast cancer families are associated to germline pathogenic mutations in the ATM gene.

    Science.gov (United States)

    Tavera-Tapia, A; Pérez-Cabornero, L; Macías, J A; Ceballos, M I; Roncador, G; de la Hoya, M; Barroso, A; Felipe-Ponce, V; Serrano-Blanch, R; Hinojo, C; Miramar-Gallart, M D; Urioste, M; Caldés, T; Santillan-Garzón, S; Benitez, J; Osorio, A

    2017-02-01

    There is still a considerable percentage of hereditary breast and ovarian cancer (HBOC) cases not explained by BRCA1 and BRCA2 genes. In this report, next-generation sequencing (NGS) techniques were applied to identify novel variants and/or genes involved in HBOC susceptibility. Using whole exome sequencing, we identified a novel germline mutation in the moderate-risk gene ATM (c.5441delT; p.Leu1814Trpfs*14) in a family negative for mutations in BRCA1/2 (BRCAX). A case-control association study was performed to establish its prevalence in Spanish population, in a series of 1477 BRCAX families and 589 controls further screened, and NGS panels were used for ATM mutational screening in a cohort of 392 HBOC Spanish BRCAX families and 350 patients affected with diseases not related to breast cancer. Although the interrogated mutation was not prevalent in case-control association study, a comprehensive mutational analysis of the ATM gene revealed 1.78% prevalence of mutations in the ATM gene in HBOC and 1.94% in breast cancer-only BRCAX families in Spanish population, where data about ATM mutations were very limited. ATM mutation prevalence in Spanish population highlights the importance of considering ATM pathogenic variants linked to breast cancer susceptibility.

  8. Targeted next generation sequencing identifies functionally deleterious germline mutations in novel genes in early-onset/familial prostate cancer.

    Directory of Open Access Journals (Sweden)

    Paula Paulo

    2018-04-01

    Full Text Available Considering that mutations in known prostate cancer (PrCa predisposition genes, including those responsible for hereditary breast/ovarian cancer and Lynch syndromes, explain less than 5% of early-onset/familial PrCa, we have sequenced 94 genes associated with cancer predisposition using next generation sequencing (NGS in a series of 121 PrCa patients. We found monoallelic truncating/functionally deleterious mutations in seven genes, including ATM and CHEK2, which have previously been associated with PrCa predisposition, and five new candidate PrCa associated genes involved in cancer predisposing recessive disorders, namely RAD51C, FANCD2, FANCI, CEP57 and RECQL4. Furthermore, using in silico pathogenicity prediction of missense variants among 18 genes associated with breast/ovarian cancer and/or Lynch syndrome, followed by KASP genotyping in 710 healthy controls, we identified "likely pathogenic" missense variants in ATM, BRIP1, CHEK2 and TP53. In conclusion, this study has identified putative PrCa predisposing germline mutations in 14.9% of early-onset/familial PrCa patients. Further data will be necessary to confirm the genetic heterogeneity of inherited PrCa predisposition hinted in this study.

  9. C. elegans FOG-3/Tob can either promote or inhibit germline proliferation, depending on gene dosage and genetic context.

    Science.gov (United States)

    Snow, J J; Lee, M-H; Verheyden, J; Kroll-Conner, P L; Kimble, J

    2013-05-23

    Vertebrate Tob/BTG proteins inhibit cell proliferation when overexpressed in tissue-culture cells, and they can function as tumor suppressors in mice. The single Caenorhabditis elegans Tob/BTG ortholog, FOG-3, by contrast, was identified from its loss-of-function phenotype as a regulator of sperm fate specification. Here we report that FOG-3 also regulates proliferation in the germline tissue. We first demonstrate that FOG-3 is a positive regulator of germline proliferation. Thus, fog-3 null mutants possess fewer germ cells than normal, a modest but reproducible decrease observed for each of two distinct fog-3 null alleles. A similar decrease also occurred in fog-3/+ heterozygotes, again for both fog-3 alleles, revealing a haplo-insufficient effect on proliferation. Therefore, FOG-3 normally promotes proliferation, and two copies of the fog-3 gene are required for this function. We next overexpressed FOG-3 by removal of FBF, the collective term for FBF-1 and FBF-2, two nearly identical PUF RNA-binding proteins. We find that overexpressed FOG-3 blocks proliferation in fbf-1 fbf-2 mutants; whereas germ cells stop dividing and instead differentiate in fbf-1 fbf-2 double mutants, they continue to proliferate in fog-3; fbf-1 fbf-2 triple mutants. Therefore, like its vertebrate Tob/BTG cousins, overexpressed FOG-3 is 'antiproliferative'. Indeed, some fog-3; fbf-1 fbf-2 mutants possess small tumors, suggesting that FOG-3 can act as a tumor suppressor. Finally, we show that FOG-3 and FBF work together to promote tumor formation in animals carrying oncogenic Notch mutations. A similar effect was not observed when germline tumors were induced by manipulation of other regulators; therefore, this FOG-3 tumor-promoting effect is context dependent. We conclude that FOG-3 can either promote or inhibit proliferation in a manner that is sensitive to both genetic context and gene dosage. The discovery of these FOG-3 effects on proliferation has implications for our understanding of

  10. Novel germline mutation (Leu512Met) in the thyrotropin receptor gene (TSHR) leading to sporadic non-autoimmune hyperthyroidism.

    Science.gov (United States)

    Roberts, Stephanie A; Moon, Jennifer E; Dauber, Andrew; Smith, Jessica R

    2017-03-01

    Primary nonautoimmune hyperthyroidism is a rare cause of neonatal hyperthyroidism. This results from an activating mutation in the thyrotropin-receptor (TSHR). It can be inherited in an autosomal dominant manner or occur sporadically as a de novo mutation. Affected individuals display a wide phenotype from severe neonatal to mild subclinical hyperthyroidism. We describe a 6-month-old boy with a de novo mutation in the TSHR gene who presented with accelerated growth, enlarging head circumference, tremor and thyrotoxicosis. Genomic DNA from the patient's and parents' peripheral blood leukocytes was extracted. Exons 9 and 10 of the TSHR gene were amplified by PCR and sequenced. Sequencing exon 10 of the TSHR gene revealed a novel heterozygous missense mutation substituting cytosine to adenine at nucleotide position 1534 in the patient's peripheral blood leukocytes. This leads to a substitution of leucine to methionine at amino acid position 512. The mutation was absent in the parents. In silico modeling by PolyPhen-2 and SIFT predicted the mutation to be deleterious. The p.Leu512Met mutation (c.1534C>A) of the TSHR gene has not been previously described in germline or somatic mutations. This case presentation highlights the possibility of mild thyrotoxicosis in affected individuals and contributes to the understanding of sporadic non-autoimmune primary hyperthyroidism.

  11. Novel germline mutation (Leu512Met) in the thyrotropin receptor gene (TSHR) leading to sporadic non-autoimmune hyperthyroidism

    Science.gov (United States)

    Roberts, Stephanie A.; Moon, Jennifer E.; Dauber, Andrew; Smith, Jessica R.

    2018-01-01

    Background Primary nonautoimmune hyperthyroidism is a rare cause of neonatal hyperthyroidism. This results from an activating mutation in the thyrotropin-receptor (TSHR). It can be inherited in an autosomal dominant manner or occur sporadically as a de novo mutation. Affected individuals display a wide phenotype from severe neonatal to mild subclinical hyperthyroidism. We describe a 6-month-old boy with a de novo mutation in the TSHR gene who presented with accelerated growth, enlarging head circumference, tremor and thyrotoxicosis. Methods Genomic DNA from the patient’s and parents’ peripheral blood leukocytes was extracted. Exons 9 and 10 of the TSHR gene were amplified by PCR and sequenced. Results Sequencing exon 10 of the TSHR gene revealed a novel heterozygous missense mutation substituting cytosine to adenine at nucleotide position 1534 in the patient’s peripheral blood leukocytes. This leads to a substitution of leucine to methionine at amino acid position 512. The mutation was absent in the parents. In silico modeling by PolyPhen-2 and SIFT predicted the mutation to be deleterious. Conclusions The p.Leu512Met mutation (c.l534C>A) of the TSHR gene has not been previously described in germline or somatic mutations. This case presentation highlights the possibility of mild thyrotoxicosis in affected individuals and contributes to the understanding of sporadic non-autoimmune primary hyperthyroidism. PMID:28195550

  12. DNA methylation patterns of candidate genes regulated by thymine DNA glycosylase in patients with TP53 germline mutations

    Energy Technology Data Exchange (ETDEWEB)

    Fortes, F.P. [CIPE, Laboratrio de Oncogentica Molecular, A.C. Camargo Cancer Center, São Paulo, SP (Brazil); Kuasne, H. [CIPE, Laboratrio NeoGene, A.C. Camargo Cancer Center, São Paulo, SP (Brazil); Departamento de Urologia, Faculdade de Medicina, Universidade Estadual Paulista, Botucatu, SP (Brazil); Marchi, F.A. [CIPE, Laboratrio NeoGene, A.C. Camargo Cancer Center, São Paulo, SP (Brazil); Programa Inter-Institucional em Bioinformtica, Instituto de Matemtica e Estatstica, Universidade So Paulo, So Paulo, SP (Brazil); Miranda, P.M. [CIPE, Laboratrio NeoGene, A.C. Camargo Cancer Center, São Paulo, SP (Brazil); Rogatto, S.R. [CIPE, Laboratrio NeoGene, A.C. Camargo Cancer Center, São Paulo, SP (Brazil); Departamento de Urologia, Faculdade de Medicina, Universidade Estadual Paulista, Botucatu, SP (Brazil); Achatz, M.I. [CIPE, Laboratrio de Oncogentica Molecular, A.C. Camargo Cancer Center, São Paulo, SP (Brazil); Departamento de Oncogentica, A.C. Camargo Cancer Center, So Paulo, SP (Brazil)

    2015-04-28

    Li-Fraumeni syndrome (LFS) is a rare, autosomal dominant, hereditary cancer predisposition disorder. In Brazil, the p.R337H TP53 founder mutation causes the variant form of LFS, Li-Fraumeni-like syndrome. The occurrence of cancer and age of disease onset are known to vary, even in patients carrying the same mutation, and several mechanisms such as genetic and epigenetic alterations may be involved in this variability. However, the extent of involvement of such events has not been clarified. It is well established that p53 regulates several pathways, including the thymine DNA glycosylase (TDG) pathway, which regulates the DNA methylation of several genes. This study aimed to identify the DNA methylation pattern of genes potentially related to the TDG pathway (CDKN2A, FOXA1, HOXD8, OCT4, SOX2, and SOX17) in 30 patients with germline TP53mutations, 10 patients with wild-type TP53, and 10 healthy individuals. We also evaluated TDG expression in patients with adrenocortical tumors (ADR) with and without the p.R337H TP53 mutation. Gene methylation patterns of peripheral blood DNA samples assessed by pyrosequencing revealed no significant differences between the three groups. However, increased TDG expression was observed by quantitative reverse transcription PCR in p.R337H carriers with ADR. Considering the rarity of this phenotype and the relevance of these findings, further studies using a larger sample set are necessary to confirm our results.

  13. Isolation of oogenesis-specific genes transcribed in the germ-line of Calliphora erythrocephala and Drosophila melanogaster

    International Nuclear Information System (INIS)

    Tucker, M.A.

    1988-01-01

    Poly(A) + RNA from early or mid-stage ovarian follicles of C. erythrocephala was used to generate radiolabelled oogenesis-specific cDNA probes for screening the phage libraries. A cDNA probe made from mid-stage embryo poly(A) + RNA was used as the differential screening probe. Thus plaques hybridizing to the two oogenesis-specific probes but not the mid-stage embryo probe were selected as potentially containing oogenesis-specific genes. Two further rounds of screening were used to eliminate false positives and, after plaque purification, restriction digests of the remaining clones were screened by Southern blot hybridization to identify DNA fragments transcribed in an oogenesis-specific manner. In situ hybridization to sections of ovarian follicles has been used to determine the cell types within the follicles in which the various genes are expressed. Radiolabelled RNA probes for four of the C. erythrocephala oogenesis-specific clones and the two D. melanogaster clones have been hybridized to ovarian follicles. Further studies have been concentrated on the two germ-line transcribed, oogenesis-specific clones isolated from the D. melanogaster clone library. Detailed genetic mapping of the DA clone and of these mutations was performed to determine which mutations might represent the DA gene. cDNA clones have been isolated for the transcribed region of clone DA and have been used to further define the transcription unit from this region of the D. melanogaster genome

  14. Mutation Detection in Patients With Advanced Cancer by Universal Sequencing of Cancer-Related Genes in Tumor and Normal DNA vs Guideline-Based Germline Testing.

    Science.gov (United States)

    Mandelker, Diana; Zhang, Liying; Kemel, Yelena; Stadler, Zsofia K; Joseph, Vijai; Zehir, Ahmet; Pradhan, Nisha; Arnold, Angela; Walsh, Michael F; Li, Yirong; Balakrishnan, Anoop R; Syed, Aijazuddin; Prasad, Meera; Nafa, Khedoudja; Carlo, Maria I; Cadoo, Karen A; Sheehan, Meg; Fleischut, Megan H; Salo-Mullen, Erin; Trottier, Magan; Lipkin, Steven M; Lincoln, Anne; Mukherjee, Semanti; Ravichandran, Vignesh; Cambria, Roy; Galle, Jesse; Abida, Wassim; Arcila, Marcia E; Benayed, Ryma; Shah, Ronak; Yu, Kenneth; Bajorin, Dean F; Coleman, Jonathan A; Leach, Steven D; Lowery, Maeve A; Garcia-Aguilar, Julio; Kantoff, Philip W; Sawyers, Charles L; Dickler, Maura N; Saltz, Leonard; Motzer, Robert J; O'Reilly, Eileen M; Scher, Howard I; Baselga, Jose; Klimstra, David S; Solit, David B; Hyman, David M; Berger, Michael F; Ladanyi, Marc; Robson, Mark E; Offit, Kenneth

    2017-09-05

    Guidelines for cancer genetic testing based on family history may miss clinically actionable genetic changes with established implications for cancer screening or prevention. To determine the proportion and potential clinical implications of inherited variants detected using simultaneous sequencing of the tumor and normal tissue ("tumor-normal sequencing") compared with genetic test results based on current guidelines. From January 2014 until May 2016 at Memorial Sloan Kettering Cancer Center, 10 336 patients consented to tumor DNA sequencing. Since May 2015, 1040 of these patients with advanced cancer were referred by their oncologists for germline analysis of 76 cancer predisposition genes. Patients with clinically actionable inherited mutations whose genetic test results would not have been predicted by published decision rules were identified. Follow-up for potential clinical implications of mutation detection was through May 2017. Tumor and germline sequencing compared with the predicted yield of targeted germline sequencing based on clinical guidelines. Proportion of clinically actionable germline mutations detected by universal tumor-normal sequencing that would not have been detected by guideline-directed testing. Of 1040 patients, the median age was 58 years (interquartile range, 50.5-66 years), 65.3% were male, and 81.3% had stage IV disease at the time of genomic analysis, with prostate, renal, pancreatic, breast, and colon cancer as the most common diagnoses. Of the 1040 patients, 182 (17.5%; 95% CI, 15.3%-19.9%) had clinically actionable mutations conferring cancer susceptibility, including 149 with moderate- to high-penetrance mutations; 101 patients tested (9.7%; 95% CI, 8.1%-11.7%) would not have had these mutations detected using clinical guidelines, including 65 with moderate- to high-penetrance mutations. Frequency of inherited mutations was related to case mix, stage, and founder mutations. Germline findings led to discussion or initiation of

  15. Germline mutations in candidate predisposition genes in individuals with cutaneous melanoma and at least two independent additional primary cancers.

    Science.gov (United States)

    Pritchard, Antonia L; Johansson, Peter A; Nathan, Vaishnavi; Howlie, Madeleine; Symmons, Judith; Palmer, Jane M; Hayward, Nicholas K

    2018-01-01

    higher overall burden of mutations in all cancer genes. We identified several pathogenic variants that likely predispose to at least one of the tumours in patients with multiple cancers. We additionally present evidence that there may be a higher burden of variants of unknown significance in 'cancer genes' in patients with multiple cancer types. Further screens of this nature need to be carried out to build evidence to show if the cancers observed in these patients form part of a cancer spectrum associated with single germline variants in these genes, whether multiple layers of susceptibility exist (oligogenic or polygenic), or if the occurrence of multiple different cancers is due to random chance.

  16. Germline heterozygous variants in genes associated with familial hemophagocytic lymphohistiocytosis as a cause of increased bleeding

    DEFF Research Database (Denmark)

    Fager Ferrari, Marcus; Leinoe, Eva; Rossing, Maria

    2018-01-01

    Familial hemophagocytic lymphohistiocytosis (FHL) is caused by biallelic variants in genes regulating granule secretion in cytotoxic lymphocytes. In FHL3-5, the affected genes UNC13D, STX11 and STXBP2 have further been shown to regulate the secretion of platelet granules, giving rise to compromised...

  17. Analysis of functional germline variants in APOBEC3 and driver genes on breast cancer risk in Moroccan study population

    International Nuclear Information System (INIS)

    Marouf, Chaymaa; Göhler, Stella; Filho, Miguel Inacio Da Silva; Hajji, Omar; Hemminki, Kari; Nadifi, Sellama; Försti, Asta

    2016-01-01

    Breast cancer (BC) is the most prevalent cancer in women and a major public health problem in Morocco. Several Moroccan studies have focused on studying this disease, but more are needed, especially at the genetic and molecular levels. Therefore, we investigated the potential association of several functional germline variants in the genes commonly mutated in sporadic breast cancer. In this case–control study, we examined 36 single nucleotide polymorphisms (SNPs) in 13 genes (APOBEC3A, APOBEC3B, ARID1B, ATR, MAP3K1, MLL2, MLL3, NCOR1, RUNX1, SF3B1, SMAD4, TBX3, TTN), which were located in the core promoter, 5’-and 3’UTR or which were nonsynonymous SNPs to assess their potential association with inherited predisposition to breast cancer development. Additionally, we identified a ~29.5-kb deletion polymorphism between APOBEC3A and APOBEC3B and explored possible associations with BC. A total of 226 Moroccan breast cancer cases and 200 matched healthy controls were included in this study. The analysis showed that12 SNPs in 8 driver genes, 4 SNPs in APOBEC3B gene and 1 SNP in APOBEC3A gene were associated with BC risk and/or clinical outcome at P ≤ 0.05 level. RUNX1-rs8130963 (odds ratio (OR) = 2.25; 95 % CI 1.42-3.56; P = 0.0005; dominant model), TBX3-rs8853 (OR = 2.04; 95 % CI 1.38-3.01; P = 0.0003; dominant model), TBX3-rs1061651 (OR = 2.14; 95 % CI1.43-3.18; P = 0.0002; dominant model), TTN-rs12465459 (OR = 2.02; 95 % confidence interval 1.33-3.07; P = 0.0009; dominant model), were the most significantly associated SNPs with BC risk. A strong association with clinical outcome were detected for the genes SMAD4 -rs3819122 with tumor size (OR = 0.45; 95 % CI 0.25-0.82; P = 0.009) and TTN-rs2244492 with estrogen receptor (OR = 0.45; 95 % CI 0.25-0.82; P = 0.009). Our results suggest that genetic variations in driver and APOBEC3 genes were associated with the risk of BC and may have impact on clinical outcome. However, the reported association between the

  18. Tumour risks and genotype-phenotype correlations associated with germline variants in succinate dehydrogenase subunit genes SDHB, SDHC and SDHD.

    Science.gov (United States)

    Andrews, Katrina A; Ascher, David B; Pires, Douglas Eduardo Valente; Barnes, Daniel R; Vialard, Lindsey; Casey, Ruth T; Bradshaw, Nicola; Adlard, Julian; Aylwin, Simon; Brennan, Paul; Brewer, Carole; Cole, Trevor; Cook, Jackie A; Davidson, Rosemarie; Donaldson, Alan; Fryer, Alan; Greenhalgh, Lynn; Hodgson, Shirley V; Irving, Richard; Lalloo, Fiona; McConachie, Michelle; McConnell, Vivienne P M; Morrison, Patrick J; Murday, Victoria; Park, Soo-Mi; Simpson, Helen L; Snape, Katie; Stewart, Susan; Tomkins, Susan E; Wallis, Yvonne; Izatt, Louise; Goudie, David; Lindsay, Robert S; Perry, Colin G; Woodward, Emma R; Antoniou, Antonis C; Maher, Eamonn R

    2018-06-01

    Germline pathogenic variants in SDHB/SDHC / SDHD are the most frequent causes of inherited phaeochromocytomas/paragangliomas. Insufficient information regarding penetrance and phenotypic variability hinders optimum management of mutation carriers. We estimate penetrance for symptomatic tumours and elucidate genotype-phenotype correlations in a large cohort of SDHB/SDHC / SDHD mutation carriers. A retrospective survey of 1832 individuals referred for genetic testing due to a personal or family history of phaeochromocytoma/paraganglioma. 876 patients (401 previously reported) had a germline mutation in SDHB/SDHC / SDHD (n=673/43/160). Tumour risks were correlated with in silico structural prediction analyses. Tumour risks analysis provided novel penetrance estimates and genotype-phenotype correlations. In addition to tumour type susceptibility differences for individual genes, we confirmed that the SDHD: p.Pro81Leu mutation has a distinct phenotype and identified increased age-related tumour risks with highly destabilising SDHB missense mutations. By Kaplan-Meier analysis, the penetrance (cumulative risk of clinically apparent tumours) in SDHB and (paternally inherited) SDHD mutation-positive non-probands (n=371/67 with detailed clinical information) by age 60 years was 21.8% (95% CI 15.2% to 27.9%) and 43.2% (95% CI 25.4% to 56.7%), respectively. Risk of malignant disease at age 60 years in non-proband SDHB mutation carriers was 4.2%(95% CI 1.1% to 7.2%). With retrospective cohort analysis to adjust for ascertainment, cumulative tumour risks for SDHB mutation carriers at ages 60 years and 80 years were 23.9% (95% CI 20.9% to 27.4%) and 30.6% (95% CI 26.8% to 34.7%). Overall risks of clinically apparent tumours for SDHB mutation carriers are substantially lower than initially estimated and will improve counselling of affected families. Specific genotype-tumour risk associations provides a basis for novel investigative strategies into succinate dehydrogenase

  19. Epigenetic modulation of cancer-germline antigen gene expression in tumorigenic human mesenchymal stem cells: implications for cancer therapy

    DEFF Research Database (Denmark)

    Gjerstorff, Morten; Burns, Jorge S; Nielsen, Ole

    2009-01-01

    Cancer-germline antigens are promising targets for cancer immunotherapy, but whether such therapies will also eliminate the primary tumor stem cell population remains undetermined. We previously showed that long-term cultures of telomerized adult human bone marrow mesenchymal stem cells can...... spontaneously evolve into tumor-initiating, mesenchymal stem cells (hMSC-TERT20), which have characteristics of clinical sarcoma cells. In this study, we used the hMSC-TERT20 tumor stem cell model to investigate the potential of cancer-germline antigens to serve as tumor stem cell targets. We found...... of cancer-germline antigens in hMSC-TERT20 cells, while their expression levels in primary human mesenchymal stem cells remained unaffected. The expression pattern of cancer-germline antigens in tumorigenic mesenchymal stem cells and sarcomas, plus their susceptibility to enhancement by epigenetic...

  20. Germline Missense Changes in the APC Gene and Their Relationship to Disease.

    Science.gov (United States)

    Scott, Rodney J; Crooks, Renee; Rose, Lindy; Attia, John; Thakkinstian, Ammarin; Thomas, Lesley; Spigelman, Allan D; Meldrum, Cliff J

    2004-05-15

    Familial adenomatous polyposis (FAP) is characterized by the presence of hundreds to thousands of adenomas that carpet the entire colon and rectum. Nonsense and frameshift mutations in the adenomatous polyposis coli (APC) gene account for the majority of mutations identified to date and predispose primarily to the typical disease phenotype. Some APC mutations are associated with a milder form of the disease known as attenuated FAP. Virtually all mutations that have been described in the APC gene result in the formation of a premature stop codon and very little is known about missense mutations apart from a common Ashkenazi Jewish mutation (1307 K) and a British E1317Q missense change. The incidence of missense mutations in the APC gene has been underreported since the APC gene lends itself to analysis using an artificial transcription and translation assay known as the Protein Truncation Test (PTT) or the In Vitro Synthetic Protein assay (IVSP).In this report we have used denaturing high performance liquid chromatography to analyse the entire coding sequence of the APC gene to determine if a cohort of patients adhering to the diagnostic criteria of FAP to assess the frequency of missense mutations in the APC gene. Altogether 112 patients were studied and 22 missense mutations were identified. From the total of 22 missense changes, 13 were silent changes and the remaining 9 resulted in amino acid substitutions. One or more of these changes were identified multiple times in 62.5% of the population under study.The results reveal that missense mutations in the APC gene appear not to radically alter protein function but may be associated with more subtle processing of RNA transcripts which in turn could result in the expression of differentially spliced forms of the APC gene which may interfere with the functional activity of the APC protein.

  1. Germline Missense Changes in the APC Gene and Their Relationship to Disease

    Directory of Open Access Journals (Sweden)

    Scott Rodney J

    2004-05-01

    Full Text Available Abstract Familial adenomatous polyposis (FAP is characterized by the presence of hundreds to thousands of adenomas that carpet the entire colon and rectum. Nonsense and frameshift mutations in the adenomatous polyposis coli (APC gene account for the majority of mutations identified to date and predispose primarily to the typical disease phenotype. Some APC mutations are associated with a milder form of the disease known as attenuated FAP. Virtually all mutations that have been described in the APC gene result in the formation of a premature stop codon and very little is known about missense mutations apart from a common Ashkenazi Jewish mutation (1307 K and a British E1317Q missense change. The incidence of missense mutations in the APC gene has been underreported since the APC gene lends itself to analysis using an artificial transcription and translation assay known as the Protein Truncation Test (PTT or the In Vitro Synthetic Protein assay (IVSP. In this report we have used denaturing high performance liquid chromatography to analyse the entire coding sequence of the APC gene to determine if a cohort of patients adhering to the diagnostic criteria of FAP to assess the frequency of missense mutations in the APC gene. Altogether 112 patients were studied and 22 missense mutations were identified. From the total of 22 missense changes, 13 were silent changes and the remaining 9 resulted in amino acid substitutions. One or more of these changes were identified multiple times in 62.5% of the population under study. The results reveal that missense mutations in the APC gene appear not to radically alter protein function but may be associated with more subtle processing of RNA transcripts which in turn could result in the expression of differentially spliced forms of the APC gene which may interfere with the functional activity of the APC protein.

  2. Germline variants in the ATM gene and breast cancer susceptibility in Moroccan women: A meta-analysis

    Directory of Open Access Journals (Sweden)

    Chaymaa Marouf

    2017-10-01

    Full Text Available Background: The ATM gene encoding a large protein kinase is mutated in ataxia-telangiectasia (AT, an autosomale recessive disease characterized by neurological and immunological symptoms, and cancer predisposition. Previous studies suggest that heterozygous carriers of ATM mutations have an increased risk of breast cancer compared with non carriers, but the contribution of specific variants has been difficult to estimate. However, two functional ATM variants, c.7271T > G and c.1066–6T > G (IVS10–6T > G, are associated with increased risk for the development of breast cancer. Methods: To investigate the role of ATM in breast cancer susceptibility, we genotyped 163 case patients with breast cancer and 150 healthy control individuals for the c.7271T > G and c.1066–6T > G (IVS10–6T > G ATM variants using polymerase chain reaction (PCR-restriction fragment length polymorphism (RFLP analysis. Results: We did not detect the ATM c.7271T > G and c.1066–6T > G (IVS10–6T > G mutations in any of 150 healthy control individuals and 163 breast cancer patients, including 59 women diagnosed with breast cancer at an early age ( G (IVS10–6T > G mutation and the rare c.7271T > G variant are not a risk factor for developing breast cancer in the Moroccan population. Larger and/or combined association studies are needed to clarify this issue. Keywords: Breast cancers, ATM gene, Germline mutation, Genetic susceptibility, Moroccan population

  3. Human Germline Genome Editing.

    Science.gov (United States)

    Ormond, Kelly E; Mortlock, Douglas P; Scholes, Derek T; Bombard, Yvonne; Brody, Lawrence C; Faucett, W Andrew; Garrison, Nanibaa' A; Hercher, Laura; Isasi, Rosario; Middleton, Anna; Musunuru, Kiran; Shriner, Daniel; Virani, Alice; Young, Caroline E

    2017-08-03

    With CRISPR/Cas9 and other genome-editing technologies, successful somatic and germline genome editing are becoming feasible. To respond, an American Society of Human Genetics (ASHG) workgroup developed this position statement, which was approved by the ASHG Board in March 2017. The workgroup included representatives from the UK Association of Genetic Nurses and Counsellors, Canadian Association of Genetic Counsellors, International Genetic Epidemiology Society, and US National Society of Genetic Counselors. These groups, as well as the American Society for Reproductive Medicine, Asia Pacific Society of Human Genetics, British Society for Genetic Medicine, Human Genetics Society of Australasia, Professional Society of Genetic Counselors in Asia, and Southern African Society for Human Genetics, endorsed the final statement. The statement includes the following positions. (1) At this time, given the nature and number of unanswered scientific, ethical, and policy questions, it is inappropriate to perform germline gene editing that culminates in human pregnancy. (2) Currently, there is no reason to prohibit in vitro germline genome editing on human embryos and gametes, with appropriate oversight and consent from donors, to facilitate research on the possible future clinical applications of gene editing. There should be no prohibition on making public funds available to support this research. (3) Future clinical application of human germline genome editing should not proceed unless, at a minimum, there is (a) a compelling medical rationale, (b) an evidence base that supports its clinical use, (c) an ethical justification, and (d) a transparent public process to solicit and incorporate stakeholder input. Copyright © 2017 American Society of Human Genetics. All rights reserved.

  4. The Number of Overlapping AID Hotspots in Germline IGHV Genes Is Inversely Correlated with Mutation Frequency in Chronic Lymphocytic Leukemia.

    Science.gov (United States)

    Yuan, Chaohui; Chu, Charles C; Yan, Xiao-Jie; Bagnara, Davide; Chiorazzi, Nicholas; MacCarthy, Thomas

    2017-01-01

    The targeting of mutations by Activation-Induced Deaminase (AID) is a key step in generating antibody diversity at the Immunoglobulin (Ig) loci but is also implicated in B-cell malignancies such as chronic lymphocytic leukemia (CLL). AID has previously been shown to preferentially deaminate WRC (W = A/T, R = A/G) hotspots. WGCW sites, which contain an overlapping WRC hotspot on both DNA strands, mutate at much higher frequency than single hotspots. Human Ig heavy chain (IGHV) genes differ in terms of WGCW numbers, ranging from 4 for IGHV3-48*03 to as many as 12 in IGHV1-69*01. An absence of V-region mutations in CLL patients ("IGHV unmutated", or U-CLL) is associated with a poorer prognosis compared to "IGHV mutated" (M-CLL) patients. The reasons for this difference are still unclear, but it has been noted that particular IGHV genes associate with U-CLL vs M-CLL. For example, patients with IGHV1-69 clones tend to be U-CLL with a poor prognosis, whereas patients with IGHV3-30 tend to be M-CLL and have a better prognosis. Another distinctive feature of CLL is that ~30% of (mostly poor prognosis) patients can be classified into "stereotyped" subsets, each defined by HCDR3 similarity, suggesting selection, possibly for a self-antigen. We analyzed >1000 IGHV genes from CLL patients and found a highly significant statistical relationship between the number of WGCW hotspots in the germline V-region and the observed mutation frequency in patients. However, paradoxically, this correlation was inverse, with V-regions with more WGCW hotspots being less likely to be mutated, i.e., more likely to be U-CLL. The number of WGCW hotspots in particular, are more strongly correlated with mutation frequency than either non-overlapping (WRC) hotspots or more general models of mutability derived from somatic hypermutation data. Furthermore, this correlation is not observed in sequences from the B cell repertoires of normal individuals and those with autoimmune diseases.

  5. Four novel germline mutations in the MLH1 and PMS2 mismatch repair genes in patients with hereditary nonpolyposis colorectal cancer.

    Science.gov (United States)

    Montazer Haghighi, Mahdi; Radpour, Ramin; Aghajani, Katayoun; Zali, Narges; Molaei, Mahsa; Zali, Mohammad Reza

    2009-08-01

    Hereditary nonpolyposis colorectal cancer (HNPCC) is the most common cause of early onset hereditary colorectal cancer. In the majority of HNPCC families, microsatellite instability (MSI) and germline mutation in one of the DNA mismatch repair (MMR) genes are found. The entire coding sequence of MMR genes (MLH1, MLH2, MLH6, and PMS2) was analyzed using direct sequencing. Also, tumor tests were done as MSI and immunohistochemistry testing. We were able to find three novel MLH1 and one novel PMS2 germline mutations in three Iranian HNPCC patients. The first was a transversion mutation c.346A>C (T116P) and happened in the highly conserved HATPase-c region of MLH1 protein. The second was a transversion mutation c.736A>T (I246L), which caused an amino acid change of isoleucine to leucine. The third mutation (c.2145,6 delTG) was frameshift and resulted in an immature stop codon in five codons downstream. All of these three mutations were detected in the MLH1 gene. The other mutation was a transition mutation, c.676G>A (G207E), which has been found in exon six of the PMS2 gene and caused an amino acid change of glycine to glutamic acid. MSI assay revealed high instability in microsatellite for two patients and microsatellite stable for one patient. In all patients, an abnormal expression of the MMR proteins in HNPCC was related to the above novel mutations.

  6. Association of germline mutation in the PTEN tumour suppressor gene and Proteus and Proteus-like syndromes

    NARCIS (Netherlands)

    Zhou, X.; Hampel, H.; Thiele, H.; Gorlin, R. J.; Hennekam, R. C.; Parisi, M.; Winter, R. M.; Eng, C.

    2001-01-01

    The molecular aetiology of Proteus syndrome (PS) remains elusive. Germline mutations in PTEN cause Cowden syndrome and Bannayan-Riley-Ruvalcaba syndrome, which are hereditary hamartoma syndromes. Some features-eg, macrocephaly, lipomatosis, and vascular malformations-can be seen in all three

  7. An Abundant Class of Non-coding DNA Can Prevent Stochastic Gene Silencing in the C. elegans Germline

    DEFF Research Database (Denmark)

    Frøkjær-Jensen, Christian; Jain, Nimit; Hansen, Loren

    2016-01-01

    /or structure. Here, we demonstrate that a pervasive non-coding DNA feature in Caenorhabditis elegans, characterized by 10-base pair periodic An/Tn-clusters (PATCs), can license transgenes for germline expression within repressive chromatin domains. Transgenes containing natural or synthetic PATCs are resistant...

  8. The Budapest Meeting 2005 intensified networking on ethics of science: the case of reproductive cloning, germline gene therapy and human dignity.

    Science.gov (United States)

    Van Steendam, Guido; Dinnyés, András; Mallet, Jacques; Meloni, Rolando; Casabona, Carlos Romeo; González, Jorge Guerra; Kure, Josef; Szathmáry, Eörs; Vorstenbosch, Jan; Molnár, Péter; Edbrooke, David; Sándor, Judit; Oberfrank, Ferenc; Cole-Turner, Ron; Hargittai, István; Littig, Beate; Ladikas, Miltos; Mordini, Emilio; Roosendaal, Hans E; Salvi, Maurizio; Gulyás, Balázs; Malpede, Diana

    2006-10-01

    This paper reports on the meeting of the Sounding Board of the EU Reprogenetics Project that was held in Budapest, Hungary, 6-9 November 2005. The Reprogenetics Project runs from 2004 until 2007 and has a brief to study the ethical aspects of human reproductive cloning and germline gene therapy. Discussions during The Budapest Meeting are reported in depth in this paper as well as the initiatives to involve the participating groups and others in ongoing collaborations with the goal of forming an integrated network of European resources in the fields of ethics of science.

  9. Screening for germline mutations of MLH1, MSH2, MSH6 and PMS2 genes in Slovenian colorectal cancer patients: implications for a population specific detection strategy of Lynch syndrome.

    Science.gov (United States)

    Berginc, Gasper; Bracko, Matej; Ravnik-Glavac, Metka; Glavac, Damjan

    2009-01-01

    Microsatellite instability (MSI) is present in more than 90% of colorectal cancers of patients with Lynch syndrome, and is therefore a feasible marker for the disease. Mutations in MLH1, MSH2, MSH6 and PMS2, which are one of the main causes of deficient mismatch repair and subsequent MSI, have been linked to the disease. In order to establish the role of each of the 4 genes in Slovenian Lynch syndrome patients, we performed MSI analysis on 593 unselected CRC patients and subsequently searched for the presence of point mutations, larger genomic rearrangements and MLH1 promoter hypermethylation in patients with MSI-high tumours. We detected 43 (7.3%) patients with MSI-H tumours, of which 7 patients (1.3%) harboured germline defects: 2 in MLH1, 4 in MSH2, 1 in PMS2 and none in MSH6. Twenty-nine germline sequence variations of unknown significance and 17 deleterious somatic mutations were found. MLH1 promoter methylation was detected in 56% of patients without detected germline defects and in 1 (14%) suspected Lynch syndrome. Due to the minor role of germline MSH6 mutations, we adapted the Lynch syndrome detection strategy for the Slovenian population of CRC patients, whereby germline alterations should be first sought in MLH1 and MSH2 followed by a search for larger genomic rearrangements in these two genes. When no germline mutations are found tumors should be further tested for the presence of germline defects in PMS2 and MSH6. The choice about which gene should be tested first can be guided more accurately by the immunohistochemical analysis. Our study demonstrates that the incidence of MMR mutations in a population should be known prior to the application of one of several suggested strategies for detection of Lynch syndrome.

  10. Germline variants in Hamartomatous Polyposis Syndrome-associated genes from patients with one or few hamartomatous polyps

    DEFF Research Database (Denmark)

    Jelsig, Anne Marie; Brusgaard, Klaus; Hansen, Tine Plato

    2016-01-01

    OBJECTIVE: A subgroup of patients with hamartomatous polyps in the GI tract has a hereditary Hamartomatous Polyposis Syndrome with an increased risk of cancer. The distinction between patients with one or few polyps and patients with a syndrome can be difficult. A pathogenic germline mutation can...... significance of genetic variants can be difficult to interpret. A family history of polyps, cancer, or extraintestinal findings or a minimum of 3-5 polyps seems to be relevant information to include before genetic testing....

  11. Biallelic germline and somatic mutations in malignant mesothelioma: multiple mutations in transcription regulators including mSWI/SNF genes.

    Science.gov (United States)

    Yoshikawa, Yoshie; Sato, Ayuko; Tsujimura, Tohru; Otsuki, Taiichiro; Fukuoka, Kazuya; Hasegawa, Seiki; Nakano, Takashi; Hashimoto-Tamaoki, Tomoko

    2015-02-01

    We detected low levels of acetylation for histone H3 tail lysines in malignant mesothelioma (MM) cell lines resistant to histone deacetylase inhibitors. To identify the possible genetic causes related to the low histone acetylation levels, whole-exome sequencing was conducted with MM cell lines established from eight patients. A mono-allelic variant of BRD1 was common to two MM cell lines with very low acetylation levels. We identified 318 homozygous protein-damaging variants/mutations (18-78 variants/mutations per patient); annotation analysis showed enrichment of the molecules associated with mammalian SWI/SNF (mSWI/SNF) chromatin remodeling complexes and co-activators that facilitate initiation of transcription. In seven of the patients, we detected a combination of variants in histone modifiers or transcription factors/co-factors, in addition to variants in mSWI/SNF. Direct sequencing showed that homozygous mutations in SMARCA4, PBRM1 and ARID2 were somatic. In one patient, homozygous germline variants were observed for SMARCC1 and SETD2 in chr3p22.1-3p14.2. These exhibited extended germline homozygosity and were in regions containing somatic mutations, leading to a loss of BAP1 and PBRM1 expression in MM cell line. Most protein-damaging variants were heterozygous in normal tissues. Heterozygous germline variants were often converted into hemizygous variants by mono-allelic deletion, and were rarely homozygous because of acquired uniparental disomy. Our findings imply that MM might develop through the somatic inactivation of mSWI/SNF complex subunits and/or histone modifiers, including BAP1, in subjects that have rare germline variants of these transcription regulators and/or transcription factors/co-factors, and in regions prone to mono-allelic deletion during oncogenesis. © 2014 UICC.

  12. Endometrial tumour BRAF mutations and MLH1 promoter methylation as predictors of germline mismatch repair gene mutation status: a literature review.

    Science.gov (United States)

    Metcalf, Alexander M; Spurdle, Amanda B

    2014-03-01

    Colorectal cancer (CRC) that displays high microsatellite instability (MSI-H) can be caused by either germline mutations in mismatch repair (MMR) genes, or non-inherited transcriptional silencing of the MLH1 promoter. A correlation between MLH1 promoter methylation, specifically the 'C' region, and BRAF V600E status has been reported in CRC studies. Germline MMR mutations also greatly increase risk of endometrial cancer (EC), but no systematic review has been undertaken to determine if these tumour markers may be useful predictors of MMR mutation status in EC patients. Endometrial cancer cohorts meeting review inclusion criteria encompassed 2675 tumours from 20 studies for BRAF V600E, and 447 tumours from 11 studies for MLH1 methylation testing. BRAF V600E mutations were reported in 4/2675 (0.1%) endometrial tumours of unknown MMR mutation status, and there were 7/823 (0.9%) total sequence variants in exon 11 and 27/1012 (2.7%) in exon 15. Promoter MLH1 methylation was not observed in tumours from 32 MLH1 mutation carriers, or for 13 MSH2 or MSH6 mutation carriers. MMR mutation-negative individuals with tumour MLH1 and PMS2 IHC loss displayed MLH1 methylation in 48/51 (94%) of tumours. We have also detailed specific examples that show the importance of MLH1 promoter region, assay design, and quantification of methylation. This review shows that BRAF mutations occurs so infrequently in endometrial tumours they can be discounted as a useful marker for predicting MMR-negative mutation status, and further studies of endometrial cohorts with known MMR mutation status are necessary to quantify the utility of tumour MLH1 promoter methylation as a marker of negative germline MMR mutation status in EC patients.

  13. Frequency and phenotypic spectrum of germline mutations in POLE and seven other polymerase genes in 266 patients with colorectal adenomas and carcinomas.

    Science.gov (United States)

    Spier, Isabel; Holzapfel, Stefanie; Altmüller, Janine; Zhao, Bixiao; Horpaopan, Sukanya; Vogt, Stefanie; Chen, Sophia; Morak, Monika; Raeder, Susanne; Kayser, Katrin; Stienen, Dietlinde; Adam, Ronja; Nürnberg, Peter; Plotz, Guido; Holinski-Feder, Elke; Lifton, Richard P; Thiele, Holger; Hoffmann, Per; Steinke, Verena; Aretz, Stefan

    2015-07-15

    In a number of families with colorectal adenomatous polyposis or suspected Lynch syndrome/HNPCC, no germline alteration in the APC, MUTYH, or mismatch repair (MMR) genes are found. Missense mutations in the polymerase genes POLE and POLD1 have recently been identified as rare cause of multiple colorectal adenomas and carcinomas, a condition termed polymerase proofreading-associated polyposis (PPAP). The aim of the present study was to evaluate the clinical relevance and phenotypic spectrum of polymerase germline mutations. Therefore, targeted sequencing of the polymerase genes POLD1, POLD2, POLD3, POLD4, POLE, POLE2, POLE3 and POLE4 was performed in 266 unrelated patients with polyposis or fulfilled Amsterdam criteria. The POLE mutation c.1270C>G;p.Leu424Val was detected in four unrelated patients. The mutation was present in 1.5% (4/266) of all patients, 4% (3/77) of all familial cases and 7% (2/30) of familial polyposis cases. The colorectal phenotype in 14 affected individuals ranged from typical adenomatous polyposis to a HNPCC phenotype, with high intrafamilial variability. Multiple colorectal carcinomas and duodenal adenomas were common, and one case of duodenal carcinoma was reported. Additionally, various extraintestinal lesions were evident. Nine further putative pathogenic variants were identified. The most promising was c.1306C>T;p.Pro436Ser in POLE. In conclusion, a PPAP was identified in a substantial number of polyposis and familial colorectal cancer patients. Screening for polymerase proofreading mutations should therefore be considered, particularly in unexplained familial cases. The present study broadens the phenotypic spectrum of PPAP to duodenal adenomas and carcinomas, and identified novel, potentially pathogenic variants in four polymerase genes. © 2014 UICC.

  14. Prevalence of Germline Mutations in Genes Engaged in DNA Damage Repair by Homologous Recombination in Patients with Triple-Negative and Hereditary Non-Triple-Negative Breast Cancers.

    Directory of Open Access Journals (Sweden)

    Pawel Domagala

    Full Text Available This study sought to assess the prevalence of common germline mutations in several genes engaged in the repair of DNA double-strand break by homologous recombination in patients with triple-negative breast cancers and hereditary non-triple-negative breast cancers. Tumors deficient in this type of DNA damage repair are known to be especially sensitive to DNA cross-linking agents (e.g., platinum drugs and to poly(ADP-ribose polymerase (PARP inhibitors.Genetic testing was performed for 36 common germline mutations in genes engaged in the repair of DNA by homologous recombination, i.e., BRCA1, BRCA2, CHEK2, NBN, ATM, PALB2, BARD1, and RAD51D, in 202 consecutive patients with triple-negative breast cancers and hereditary non-triple-negative breast cancers.Thirty five (22.2% of 158 patients in the triple-negative group carried mutations in genes involved in DNA repair by homologous recombination, while 10 (22.7% of the 44 patients in the hereditary non-triple-negative group carried such mutations. Mutations in BRCA1 were most frequent in patients with triple-negative breast cancer (18.4%, and mutations in CHEK2 were most frequent in patients with hereditary non-triple-negative breast cancers (15.9%. In addition, in the triple-negative group, mutations in CHEK2, NBN, and ATM (3.8% combined were found, while mutations in BRCA1, NBN, and PALB2 (6.8% combined were identified in the hereditary non-triple-negative group.Identifying mutations in genes engaged in DNA damage repair by homologous recombination other than BRCA1/2 can substantially increase the proportion of patients with triple-negative breast cancer and hereditary non-triple-negative breast cancer who may be eligible for therapy using PARP inhibitors and platinum drugs.

  15. Tumor mismatch repair immunohistochemistry and DNA MLH1 methylation testing of patients with endometrial cancer diagnosed at age younger than 60 years optimizes triage for population-level germline mismatch repair gene mutation testing.

    Science.gov (United States)

    Buchanan, Daniel D; Tan, Yen Y; Walsh, Michael D; Clendenning, Mark; Metcalf, Alexander M; Ferguson, Kaltin; Arnold, Sven T; Thompson, Bryony A; Lose, Felicity A; Parsons, Michael T; Walters, Rhiannon J; Pearson, Sally-Ann; Cummings, Margaret; Oehler, Martin K; Blomfield, Penelope B; Quinn, Michael A; Kirk, Judy A; Stewart, Colin J; Obermair, Andreas; Young, Joanne P; Webb, Penelope M; Spurdle, Amanda B

    2014-01-10

    Clinicopathologic data from a population-based endometrial cancer cohort, unselected for age or family history, were analyzed to determine the optimal scheme for identification of patients with germline mismatch repair (MMR) gene mutations. Endometrial cancers from 702 patients recruited into the Australian National Endometrial Cancer Study (ANECS) were tested for MMR protein expression using immunohistochemistry (IHC) and for MLH1 gene promoter methylation in MLH1-deficient cases. MMR mutation testing was performed on germline DNA of patients with MMR-protein deficient tumors. Prediction of germline mutation status was compared for combinations of tumor characteristics, age at diagnosis, and various clinical criteria (Amsterdam, Bethesda, Society of Gynecologic Oncology, ANECS). Tumor MMR-protein deficiency was detected in 170 (24%) of 702 cases. Germline testing of 158 MMR-deficient cases identified 22 truncating mutations (3% of all cases) and four unclassified variants. Tumor MLH1 methylation was detected in 99 (89%) of 111 cases demonstrating MLH1/PMS2 IHC loss; all were germline MLH1 mutation negative. A combination of MMR IHC plus MLH1 methylation testing in women younger than 60 years of age at diagnosis provided the highest positive predictive value for the identification of mutation carriers at 46% versus ≤ 41% for any other criteria considered. Population-level identification of patients with MMR mutation-positive endometrial cancer is optimized by stepwise testing for tumor MMR IHC loss in patients younger than 60 years, tumor MLH1 methylation in individuals with MLH1 IHC loss, and germline mutations in patients exhibiting loss of MSH6, MSH2, or PMS2 or loss of MLH1/PMS2 with absence of MLH1 methylation.

  16. Congenital Neonatal Hyperthyroidism Caused by Germline Mutations in the TSH Receptor Gene: Case Report and Review of the Literature

    Science.gov (United States)

    Chester, Jeremy; Rotenstein, Deborah; Ringkananont, Usanee; Steuer, Guy; Carlin, Beatrice; Stewart, Lindsay; Grasberger, Helmut; Refetoff, Samuel

    2018-01-01

    Neonatal hyperthyroidism, a rare and serious disorder occurs in two forms. An autoimmune form associated with maternal Graves’ disease, resulting from transplacental passage of maternal thyroid-stimulating antibodies, and a nonautoimmune form, resulting from mutations in the stimulatory G protein or the thyrotropin receptor (TSHR) causing constitutive activation of intracellular signaling cascades. To date, 29 separate cases of thyrotoxicosis caused by germline mutations of the TSHR have been documented. These cases have expressed themselves in a range of clinical consequences. This report describes a new case of a newborn with nonautoimmune hyperthyroidism secondary to a constitutively active TSHR mutation (S281N) whose clinical course was complicated by severe respiratory compromise. Typical clinical findings in this disorder are discussed by a review of all previously published cases. PMID:18655531

  17. Evolutionary pattern of mutation in the factor IX genes of great apes: How does it compare to the pattern of recent germline mutation in patients with hemophilia B?

    Energy Technology Data Exchange (ETDEWEB)

    Grouse, L.H.; Ketterling, R.P.; Sommer, S.S. [Mayo Clinic/Foundation, Rochester, MN (United States)

    1994-09-01

    Most mutations causing hemophilia B have arisen within the past 150 years. By correcting for multiple biases, the underlying rates of spontaneous germline mutation have been estimated in the factor IX gene. From these rates, an underlying pattern of mutation has emerged. To determine if this pattern compares to a underlying pattern found in the great apes, sequence changes were determined in intronic regions of the factor IX gene. The following species were studied: Gorilla gorilla, Pan troglodytes (chimpanzee), Pongo pygmacus (orangutan) and Homo sapiens. Intronic sequences at least 200 bp from a splice junction were randomly chosen, amplified by cross-species PCR, and sequenced. These regions are expected to be subject to little if any selective pressure. Early diverged species of Old World monkeys were also studied to help determine the direction of mutational changes. A total of 62 sequence changes were observed. Initial data suggest that the average pattern since evolution of the great apes has a paucity of transitions at CpG dinucleotides and an excess of microinsertions to microdeletions when compared to the pattern observed in humans during the past 150 years (p<.05). A larger study is in progress to confirm these results.

  18. TumorNext-Lynch-MMR: a comprehensive next generation sequencing assay for the detection of germline and somatic mutations in genes associated with mismatch repair deficiency and Lynch syndrome.

    Science.gov (United States)

    Gray, Phillip N; Tsai, Pei; Chen, Daniel; Wu, Sitao; Hoo, Jayne; Mu, Wenbo; Li, Bing; Vuong, Huy; Lu, Hsiao-Mei; Batth, Navanjot; Willett, Sara; Uyeda, Lisa; Shah, Swati; Gau, Chia-Ling; Umali, Monalyn; Espenschied, Carin; Janicek, Mike; Brown, Sandra; Margileth, David; Dobrea, Lavinia; Wagman, Lawrence; Rana, Huma; Hall, Michael J; Ross, Theodora; Terdiman, Jonathan; Cullinane, Carey; Ries, Savita; Totten, Ellen; Elliott, Aaron M

    2018-04-17

    The current algorithm for Lynch syndrome diagnosis is highly complex with multiple steps which can result in an extended time to diagnosis while depleting precious tumor specimens. Here we describe the analytical validation of a custom probe-based NGS tumor panel, TumorNext-Lynch-MMR, which generates a comprehensive genetic profile of both germline and somatic mutations that can accelerate and streamline the time to diagnosis and preserve specimen. TumorNext-Lynch-MMR can detect single nucleotide variants, small insertions and deletions in 39 genes that are frequently mutated in Lynch syndrome and colorectal cancer. Moreover, the panel provides microsatellite instability status and detects loss of heterozygosity in the five Lynch genes; MSH2 , MSH6 , MLH1 , PMS2 and EPCAM . Clinical cases are described that highlight the assays ability to differentiate between somatic and germline mutations, precisely classify variants and resolve discordant cases.

  19. No germline mutations in the histone acetyltransferase gene EP300 in BRCA1 and BRCA2 negative families with breast cancer and gastric, pancreatic, or colorectal cancer

    International Nuclear Information System (INIS)

    Campbell, Ian G; Choong, David; Chenevix-Trench, Georgia

    2004-01-01

    Mutations in BRCA1, BRCA2, ATM, TP53, CHK2 and PTEN account for many, but not all, multiple-case breast and ovarian cancer families. The histone acetyltransferase gene EP300 may function as a tumour suppressor gene because it is sometimes somatically mutated in breast, colorectal, gastric and pancreatic cancers, and is located on a region of chromosome 22 that frequently undergoes loss of heterozygosity in many cancer types. We hypothesized that germline mutations in EP300 may account for some breast cancer families that include cases of gastric, pancreatic and/or colorectal cancer. We screened the entire coding region of EP300 for mutations in the youngest affected members of 23 non-BRCA1/BRCA2 breast cancer families with at least one confirmed case of gastric, pancreatic and/or colorectal cancer. These families were ascertained in Australia through the Kathleen Cuningham Foundation Consortium for Research into Familial Breast Cancer. Denaturing HPLC analysis identified a heterozygous alteration at codon 211, specifically a GGC to AGC (glycine to serine) alteration, in two individuals. This conservative amino acid change was not within any known functional domains of EP300. The frequency of the Ser211 variant did not differ significanlty between a series of 352 breast cancer patients (4.0%) and 254 control individuals (2.8%; P = 0.5). The present study does not support a major role for EP300 mutations in breast and ovarian cancer families with a history of gastric, pancreatic and/or colorectal cancer

  20. Familial isolated primary hyperparathyroidism/hyperparathyroidism-jaw tumour syndrome caused by germline gross deletion or point mutations of CDC73 gene in Chinese.

    Science.gov (United States)

    Kong, Jing; Wang, Ou; Nie, Min; Shi, Jie; Hu, Yingying; Jiang, Yan; Li, Mei; Xia, Weibo; Meng, Xunwu; Xing, Xiaoping

    2014-08-01

    Hyperparathyroidism-jaw tumour syndrome (HPT-JT) and familial isolated primary hyperparathyroidism (FIHP) are two subtypes of familial primary hyperparathyroidism, which are rarely reported in Chinese population. Here, we reported three FIHP families and one HPT-JT family with long-term follow-up and genetic analysis. A total of 22 patients, from four FIHP/HPT-JT families of Chinese descent, were recruited and genomic DNA was extracted from their peripheral blood lymphocytes. Direct sequencing for MEN1, CDC73, CASR gene was conducted. Reverse transcription PCR (RT-PCR) and quantitative real-time PCR (qRT-PCR) were used to study the effect of splice site mutations and gross deletion mutations. Immunohistochemistry was performed to analyse parafibromin expression in parathyroid tumours. Genotype-phenotype correlations were assessed through clinical characteristics and long-term follow-up data. Genetic analysis revealed four CDC73 germline mutations that were responsible for the four kindreds, including two novel point mutation (c.157 G>T and IVS3+1 G>A), one recurrent point mutation (c.664 C>T) and one deletion mutation (c.307+?_513-?del exons 4, 5, 6). RT-PCR confirmed that IVS3+1 G>A generated an aberrant transcript with exon3 deletion. Immunohistochemical analysis demonstrated reduced nuclear parafibromin expression in tumours supporting the pathogenic effects of these mutations. This study supplies information on mutations and phenotypes of HPT-JT/FIHP syndrome in Chinese. Screening for gross deletion and point mutations of the CDC73 gene is necessary in susceptible subjects. © 2014 John Wiley & Sons Ltd.

  1. Generation of germline ablated male pigs by CRISPR/Cas9 editing of the NANOS2 gene

    Science.gov (United States)

    Genome editing tools have revolutionized the generation of genetically modified animals including livestock. In particular, the domestic pig is a proven model of human physiology and an agriculturally important species. In this study, we utilized the CRISPR/Cas9 system to edit the NANOS2 gene in p...

  2. Rare germline alterations in cancer-related genes associated with the risk of multiple primary tumor development

    DEFF Research Database (Denmark)

    Villacis, Rolando A. R.; Basso, Tatiane R; Canto, Luisa M

    2017-01-01

    Multiple primary tumors (MPT) have been described in carriers of inherited cancer predisposition genes. However, the genetic etiology of a large proportion of MPT cases remains unclear. We reviewed 267 patients with hereditary cancer predisposition syndromes (HCPS) that underwent genetic counseli...

  3. Higher occurrence of childhood cancer in families with germline mutations in BRCA2, MMR and CDKN2A genes

    DEFF Research Database (Denmark)

    Magnusson, S.; Borg, A.; Kristoffersson, U.

    2008-01-01

    The contribution of hereditary factors for development of childhood tumors is limited to some few known syndromes associated with predominance of tumors in childhood. Occurrence of childhood tumors in hereditary cancer syndromes such as BRCA1/2 associated breast and ovarian cancer, DNA-mismatch r......-mismatch repair (MMR) genes associated hereditary non polyposis colorectal cancer and CDKN2A associated familial malignant melanoma are very little studied. Herein we report the prevalence of childhood tumors (diagnosed...

  4. Prevalence of deleterious germline variants in risk genes including BRCA1/2 in consecutive ovarian cancer patients (AGO-TR-1.

    Directory of Open Access Journals (Sweden)

    Philipp Harter

    Full Text Available Identification of families at risk for ovarian cancer offers the opportunity to consider prophylactic surgery thus reducing ovarian cancer mortality. So far, identification of potentially affected families in Germany was solely performed via family history and numbers of affected family members with breast or ovarian cancer. However, neither the prevalence of deleterious variants in BRCA1/2 in ovarian cancer in Germany nor the reliability of family history as trigger for genetic counselling has ever been evaluated.Prospective counseling and germline testing of consecutive patients with primary diagnosis or with platinum-sensitive relapse of an invasive epithelial ovarian cancer. Testing included 25 candidate and established risk genes. Among these 25 genes, 16 genes (ATM, BRCA1, BRCA2, CDH1, CHEK2, MLH1, MSH2, MSH6, NBN, PMS2, PTEN, PALB2, RAD51C, RAD51D, STK11, TP53 were defined as established cancer risk genes. A positive family history was defined as at least one relative with breast cancer or ovarian cancer or breast cancer in personal history.In total, we analyzed 523 patients: 281 patients with primary diagnosis of ovarian cancer and 242 patients with relapsed disease. Median age at primary diagnosis was 58 years (range 16-93 and 406 patients (77.6% had a high-grade serous ovarian cancer. In total, 27.9% of the patients showed at least one deleterious variant in all 25 investigated genes and 26.4% in the defined 16 risk genes. Deleterious variants were most prevalent in the BRCA1 (15.5%, BRCA2 (5.5%, RAD51C (2.5% and PALB2 (1.1% genes. The prevalence of deleterious variants did not differ significantly between patients at primary diagnosis and relapse. The prevalence of deleterious variants in BRCA1/2 (and in all 16 risk genes in patients <60 years was 30.2% (33.2% versus 10.6% (18.9% in patients ≥60 years. Family history was positive in 43% of all patients. Patients with a positive family history had a prevalence of deleterious variants

  5. Identification of germline transcriptional regulatory elements in Aedes aegypti

    Science.gov (United States)

    Akbari, Omar S.; Papathanos, Philippos A.; Sandler, Jeremy E.; Kennedy, Katie; Hay, Bruce A.

    2014-02-01

    The mosquito Aedes aegypti is the principal vector for the yellow fever and dengue viruses, and is also responsible for recent outbreaks of the alphavirus chikungunya. Vector control strategies utilizing engineered gene drive systems are being developed as a means of replacing wild, pathogen transmitting mosquitoes with individuals refractory to disease transmission, or bringing about population suppression. Several of these systems, including Medea, UDMEL, and site-specific nucleases, which can be used to drive genes into populations or bring about population suppression, utilize transcriptional regulatory elements that drive germline-specific expression. Here we report the identification of multiple regulatory elements able to drive gene expression specifically in the female germline, or in the male and female germline, in the mosquito Aedes aegypti. These elements can also be used as tools with which to probe the roles of specific genes in germline function and in the early embryo, through overexpression or RNA interference.

  6. Genotype and phenotype spectrum of NRAS germline variants

    NARCIS (Netherlands)

    Altmuller, F.; Lissewski, C.; Bertola, D.; Flex, E.; Stark, Z.; Spranger, S.; Baynam, G.; Buscarilli, M.; Dyack, S.; Gillis, J.; Yntema, H.G.; Pantaleoni, F.; Loon, R.L. van; MacKay, S.; Mina, K.; Schanze, I.; Tan, T.Y.; Walsh, M.; White, S.M.; Niewisch, M.R.; Garcia-Minaur, S.; Plaza, D.; Ahmadian, M.R.; Cave, H.; Tartaglia, M.; Zenker, M.

    2017-01-01

    RASopathies comprise a group of disorders clinically characterized by short stature, heart defects, facial dysmorphism, and varying degrees of intellectual disability and cancer predisposition. They are caused by germline variants in genes encoding key components or modulators of the highly

  7. Functional characteristics of three new germline mutations of the thyrotropin receptor gene causing autosomal dominant toxic thyroid hyperplasia

    Energy Technology Data Exchange (ETDEWEB)

    Tonacchera, M.; Van Sande, J.; Cetani, F. [Universite Libre de Bruxelles, Brussels (Belgium)] [and others

    1996-02-01

    We report three unrelated families in which hyperthyroidism associated with thyroid hyperplasia was transmitted in an autosomal dominant fashion, in the absence of signs of autoimmunity. Exon 10 of the TSH receptor gene was directly sequenced after PCR amplification from DNA of peripheral leukocytes. In one family, a C to A transversion resulted in an S505R substitution in the third transmembrane segment; in the second, an A to T transversion caused an N650Y substitution in the sixth transmembrane segment; and in the third family, an A to G transition resulted in an N670S substitution in the seventh transmembrane segment. When expressed by transfection in COS-7 cells, each mutated receptor displayed an increase in constitutive stimulation of cAMP production; no effect on basal accumulation of inositol phosphates (IP) could be detected. In binding studies, cells transfected with wild-type of mutated receptors showed similar levels of expression, with the mutated receptors displaying similar or slightly increased affinity for bovine TSH (bTSH) binding. Cells transfected with S505R and N650Y mutants showed a similar cAMP maximal TSH-stimulated accumulation over the cells transfected with the wild type, whereas N670S transfectants showed a blunted response with an increase in EC{sub 50}. A higher IP response to 100 mU/mL bTSH over that obtained with the wild-type receptor was obtained in cells transfected with N650Y; in contrast, cells transfected with S505R showed a blunted IP production (50% less), and the N670S mutant completely lost the ability to stimulate IP accumulation in response to bTSH. The differential effects of individual mutations on stimulation by bTSH of cAMP or IP accumulation suggest that individual mutant receptors may achieve different active conformations with selective abilities to couple to G{sub s}{alpha} and to G{sub q}{alpha}. 17 refs., 8 figs.

  8. Germline APC mutations in hepatoblastoma.

    Science.gov (United States)

    Yang, Adeline; Sisson, Rebecca; Gupta, Anita; Tiao, Greg; Geller, James I

    2018-04-01

    Conflicting reports on the frequency of germline adenomatous polyposis coli (APC) gene mutations in patients with hepatoblastoma (HB) have called into question the clinical value of APC mutation testing on apparently sporadic HB. An Institutional Review Board approved retrospective review of clinical data collected from patients with HB who received APC testing at our institution was conducted. All HB patients seen at Cincinnati Children's Hospital Medical Center were eligible for testing. Potential genotype/phenotype correlations were assessed. As of July 2015, 29 patients with HB had received constitutional APC testing. Four (14%) were found to have APC pathogenic truncations of the APC protein and in addition two (7%) had APC missense variants of unknown clinical significance. Two patients (7%) had family histories indicative of familial adenomatous polyposis (FAP). Response to chemotherapy tracked differently in APC pathogenic cases, with a slower imaging response despite an equivalent or slightly faster α-fetoprotein (AFP) response. The prevalence of pathogenic APC variants in apparently sporadic HB may be higher than previously detected. Differences in time to imaging response, despite similar AFP response, may impact surgical planning. All patients with HB warrant germline APC mutation testing for underlying FAP. © 2017 Wiley Periodicals, Inc.

  9. Characterization of a rabbit germ-line VH gene that is a candidate donor for VH gene conversion in mutant Alicia rabbits.

    Science.gov (United States)

    Chen, H T; Alexander, C B; Mage, R G

    1995-06-15

    Normal rabbits preferentially rearrange the 3'-most VH gene, VH1, to encode Igs with VHa allotypes, which constitute the majority of rabbit serum Igs. A gene conversion-like mechanism is employed to diversify the primary Ab repertoire. In mutant Alicia rabbits that derived from a rabbit with VHa2 allotype, the VH1 gene was deleted. Our previous studies showed that the first functional gene (VH4) or VH4-like genes were rearranged in 2- to 8-wk-old homozygous Alicia. The VH1a2-like sequences that were found in splenic mRNA from 6-wk and older Alicia rabbits still had some residues that were typical of VH4. The appearances of sequences resembling that of VH1a2 may have been caused by gene conversions that altered the sequences of the rearranged VH or there may have been rearrangement of upstream VH1a2-like genes later in development. To investigate this further, we constructed a cosmid library and isolated a VH1a2-like gene, VH12-1-6, with a sequence almost identical to VH1a2. This gene had a deleted base in the heptamer of its recombination signal sequence. However, even if this defect diminished or eliminated its ability to rearrange, the a2-like gene could have acted as a donor for gene-conversion-like alteration of rearranged VH genes. Sequence comparisons suggested that this gene or a gene like it could have acted as a donor for gene conversion in mutant Alicia and in normal rabbits.

  10. Loss of Folliculin Disrupts Hematopoietic Stem Cell Quiescence and Homeostasis Resulting in Bone Marrow Failure.

    Science.gov (United States)

    Baba, Masaya; Toyama, Hirofumi; Sun, Lei; Takubo, Keiyo; Suh, Hyung-Chan; Hasumi, Hisashi; Nakamura-Ishizu, Ayako; Hasumi, Yukiko; Klarmann, Kimberly D; Nakagata, Naomi; Schmidt, Laura S; Linehan, W Marston; Suda, Toshio; Keller, Jonathan R

    2016-04-01

    Folliculin (FLCN) is an autosomal dominant tumor suppressor gene that modulates diverse signaling pathways required for growth, proliferation, metabolism, survival, motility, and adhesion. FLCN is an essential protein required for murine embryonic development, embryonic stem cell (ESC) commitment, and Drosophila germline stem cell maintenance, suggesting that Flcn may be required for adult stem cell homeostasis. Conditional inactivation of Flcn in adult hematopoietic stem/progenitor cells (HSPCs) drives hematopoietic stem cells (HSC) into proliferative exhaustion resulting in the rapid depletion of HSPC, loss of all hematopoietic cell lineages, acute bone marrow (BM) failure, and mortality after 40 days. HSC that lack Flcn fail to reconstitute the hematopoietic compartment in recipient mice, demonstrating a cell-autonomous requirement for Flcn in HSC maintenance. BM cells showed increased phosphorylation of Akt and mTorc1, and extramedullary hematopoiesis was significantly reduced by treating mice with rapamycin in vivo, suggesting that the mTorc1 pathway was activated by loss of Flcn expression in hematopoietic cells in vivo. Tfe3 was activated and preferentially localized to the nucleus of Flcn knockout (KO) HSPCs. Tfe3 overexpression in HSPCs impaired long-term hematopoietic reconstitution in vivo, recapitulating the Flcn KO phenotype, and supporting the notion that abnormal activation of Tfe3 contributes to the Flcn KO phenotype. Flcn KO mice develop an acute histiocytic hyperplasia in multiple organs, suggesting a novel function for Flcn in macrophage development. Thus, Flcn is intrinsically required to maintain adult HSC quiescence and homeostasis, and Flcn loss leads to BM failure and mortality in mice. Published 2016. This article is a U.S. Government work and is in the public domain in the USA.

  11. Human Germline Genome Editing

    OpenAIRE

    Ormond, Kelly E.; Mortlock, Douglas P.; Scholes, Derek T.; Bombard, Yvonne; Brody, Lawrence C.; Faucett, W. Andrew; Garrison, Nanibaa’ A.; Hercher, Laura; Isasi, Rosario; Middleton, Anna; Musunuru, Kiran; Shriner, Daniel; Virani, Alice; Young, Caroline E.

    2017-01-01

    With CRISPR/Cas9 and other genome-editing technologies, successful somatic and germline genome editing are becoming feasible. To respond, an American Society of Human Genetics (ASHG) workgroup developed this position statement, which was approved by the ASHG Board in March 2017. The workgroup included representatives from the UK Association of Genetic Nurses and Counsellors, Canadian Association of Genetic Counsellors, International Genetic Epidemiology Society, and US National Society of Gen...

  12. Aggressive pituitary adenomas occurring in young patients in a large Polynesian kindred with a germline R271W mutation in the AIP gene.

    OpenAIRE

    Jennings, J. E.; Georgitsi, M.; Holdaway, I.; Daly, Adrian; Tichomirowa, M.; Beckers, Albert; Aaltonen, Lauri A; Karhu, A.; Cameron, F. J.

    2009-01-01

    OBJECTIVE: Mutations in the aryl hydrocarbon receptor-interacting protein (AIP) were recently shown to confer a pituitary adenoma predisposition in patients with familial isolated pituitary adenomas (FIPA). We report a large Samoan FIPA kindred from Australia/New Zealand with an R271W mutation that was associated with aggressive pituitary tumors. DESIGN AND METHODS: Case series with germline screening of AIP and haplotype analyses among R271W families. RESULTS: This previously unreported kind...

  13. A novel c. 204 Ile68Met germline variant in exon 2 of the mutL homolog 1 gene in a colorectal cancer patient

    Czech Academy of Sciences Publication Activity Database

    Vodička, Pavel; Caja, F.; Vymetálková, Veronika; Procházka, Pavel; Vodičková, Ludmila; Schwarzová, L.; Slyšková, Jana; Kumar, R.; Schneiderová, M.

    2015-01-01

    Roč. 9, č. 1 (2015), s. 183-186 ISSN 1792-1074 R&D Projects: GA ČR GPP304/11/P715; GA ČR(CZ) GAP304/12/1585 Institutional support: RVO:68378041 Keywords : mutL homolog 1 * germline mutation * colorectal cancer Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 1.482, year: 2015

  14. Germline CDKN1B/p27Kip1 mutation in multiple endocrine neoplasia

    NARCIS (Netherlands)

    Georgitsi, Marianthi; Raitila, Anniina; Karhu, Auli; van der Luijt, Rob B.; Aalfs, Cora M.; Sane, Timo; Vierimaa, Outi; Mäkinen, Markus J.; Tuppurainen, Karoliina; Paschke, Ralph; Gimm, Oliver; Koch, Christian A.; Gündogdu, Sadi; Lucassen, Anneke; Tischkowitz, Marc; Izatt, Louise; Aylwin, Simon; Bano, Gul; Hodgson, Shirley; de Menis, Ernesto; Launonen, Virpi; Vahteristo, Pia; Aaltonen, Lauri A.

    2007-01-01

    Germline mutations in the MEN1 gene predispose to multiple endocrine neoplasia type 1 (MEN1) syndrome, but in up to 20-25% of clinical MEN1 cases, no MEN1 mutations can be found. Recently, a germline mutation in the CDKN1B gene, encoding p27(Kip1), was reported in one suspected MEN1 family with two

  15. Integrated tumor and germline whole-exome sequencing identifies mutations in MAPK and PI3K pathway genes in an adolescent with rosette-forming glioneuronal tumor of the fourth ventricle

    Science.gov (United States)

    Lin, Frank Y.; Bergstrom, Katie; Person, Richard; Bavle, Abhishek; Ballester, Leomar Y.; Scollon, Sarah; Raesz-Martinez, Robin; Jea, Andrew; Birchansky, Sherri; Wheeler, David A.; Berg, Stacey L.; Chintagumpala, Murali M.; Adesina, Adekunle M.; Eng, Christine; Roy, Angshumoy; Plon, Sharon E.; Parsons, D. Williams

    2016-01-01

    The integration of genome-scale studies such as whole-exome sequencing (WES) into the clinical care of children with cancer has the potential to provide insight into the genetic basis of an individual's cancer with implications for clinical management. This report describes the results of clinical tumor and germline WES for a patient with a rare tumor diagnosis, rosette-forming glioneuronal tumor of the fourth ventricle (RGNT). Three pathogenic gene alterations with implications for clinical care were identified: somatic activating hotspot mutations in FGFR1 (p.N546K) and PIK3CA (p.H1047R) and a germline pathogenic variant in PTPN11 (p.N308S) diagnostic for Noonan syndrome. The molecular landscape of RGNT is not well-described, but these data are consistent with prior observations regarding the importance of the interconnected MAPK and PI3K/AKT/mTOR signaling pathways in this rare tumor. The co-occurrence of FGFR1, PIK3CA, and PTPN11 alterations provides further evidence for consideration of RGNT as a distinct molecular entity from pediatric low-grade gliomas and suggests potential therapeutic strategies for this patient in the event of tumor recurrence as novel agents targeting these pathways enter pediatric clinical trials. Although RGNT has not been definitively linked with cancer predisposition syndromes, two prior cases have been reported in patients with RASopathies (Noonan syndrome and neurofibromatosis type 1 [NF1]), providing an additional link between these tumors and the mitogen-activated protein kinase (MAPK) signaling pathway. In summary, this case provides an example of the potential for genome-scale sequencing technologies to provide insight into the biology of rare tumors and yield both tumor and germline results of potential relevance to patient care. PMID:27626068

  16. Germline pathogenic variants in PALB2 and other cancer-predisposing genes in families with hereditary diffuse gastric cancer without CDH1 mutation: a whole-exome sequencing study.

    Science.gov (United States)

    Fewings, Eleanor; Larionov, Alexey; Redman, James; Goldgraben, Mae A; Scarth, James; Richardson, Susan; Brewer, Carole; Davidson, Rosemarie; Ellis, Ian; Evans, D Gareth; Halliday, Dorothy; Izatt, Louise; Marks, Peter; McConnell, Vivienne; Verbist, Louis; Mayes, Rebecca; Clark, Graeme R; Hadfield, James; Chin, Suet-Feung; Teixeira, Manuel R; Giger, Olivier T; Hardwick, Richard; di Pietro, Massimiliano; O'Donovan, Maria; Pharoah, Paul; Caldas, Carlos; Fitzgerald, Rebecca C; Tischkowitz, Marc

    2018-04-26

    Germline pathogenic variants in the E-cadherin gene (CDH1) are strongly associated with the development of hereditary diffuse gastric cancer. There is a paucity of data to guide risk assessment and management of families with hereditary diffuse gastric cancer that do not carry a CDH1 pathogenic variant, making it difficult to make informed decisions about surveillance and risk-reducing surgery. We aimed to identify new candidate genes associated with predisposition to hereditary diffuse gastric cancer in affected families without pathogenic CDH1 variants. We did whole-exome sequencing on DNA extracted from the blood of 39 individuals (28 individuals diagnosed with hereditary diffuse gastric cancer and 11 unaffected first-degree relatives) in 22 families without pathogenic CDH1 variants. Genes with loss-of-function variants were prioritised using gene-interaction analysis to identify clusters of genes that could be involved in predisposition to hereditary diffuse gastric cancer. Protein-affecting germline variants were identified in probands from six families with hereditary diffuse gastric cancer; variants were found in genes known to predispose to cancer and in lesser-studied DNA repair genes. A frameshift deletion in PALB2 was found in one member of a family with a history of gastric and breast cancer. Two different MSH2 variants were identified in two unrelated affected individuals, including one frameshift insertion and one previously described start-codon loss. One family had a unique combination of variants in the DNA repair genes ATR and NBN. Two variants in the DNA repair gene RECQL5 were identified in two unrelated families: one missense variant and a splice-acceptor variant. The results of this study suggest a role for the known cancer predisposition gene PALB2 in families with hereditary diffuse gastric cancer and no detected pathogenic CDH1 variants. We also identified new candidate genes associated with disease risk in these families. UK Medical

  17. Lack of association between autonomously functioning thyroid nodules and germline polymorphisms of the thyrotropin receptor and Gαs genes in a mild to moderate iodine-deficient Caucasian population.

    Science.gov (United States)

    Vicchio, Teresa Manuela; Giovinazzo, Salvatore; Certo, Rosaria; Cucinotta, Mariapaola; Micali, Carmelo; Baldari, Sergio; Benvenga, Salvatore; Trimarchi, Francesco; Campennì, Alfredo; Ruggeri, Rosaria Maddalena

    2014-07-01

    Mutations of the thyrotropin receptor (TSHR) and/or Gαs gene have been found in a number of, but not all, autonomously functioning thyroid nodules (AFTNs). Recently, in a 15-year-old girl with a hyperfunctioning papillary thyroid carcinoma, we found two somatic and germline single nucleotide polymorphisms (SNPs): a SNP of the TSHR gene (exon 7, codon 187) and a SNP of Gαs gene (exon 8, codon 185). The same silent SNP of the TSHR gene had been reported in patients with AFTN or familial non-autoimmune hyperthyroidism. No further data about the prevalence of the two SNPs in AFTNs as well as in the general population are available in the literature. To clarify the possible role of these SNPs in predisposing to AFTN. Germline DNA was extracted from blood leukocytes of 115 patients with AFTNs (43 males and 72 females, aged 31-85 years, mean ± SD = 64 ± 13) and 100 sex-matched healthy individuals from the same geographic area, which is marginally iodine deficient. The genotype distribution of the two SNPs was investigated by restriction fragment length polymorphism-polymerase chain reaction. The prevalence of the two SNPs in our study population was low and not different to that found in healthy individuals: 8 % of patients vs. 9 % of controls were heterozygous for the TSHR SNP and 4 % patients vs. 6 % controls were heterozygous for the Gαs SNP. One patient harbored both SNPs. These results suggest that these two SNPs do not confer susceptibility for the development of AFTN.

  18. Germline Variants in Targeted Tumor Sequencing Using Matched Normal DNA.

    Science.gov (United States)

    Schrader, Kasmintan A; Cheng, Donavan T; Joseph, Vijai; Prasad, Meera; Walsh, Michael; Zehir, Ahmet; Ni, Ai; Thomas, Tinu; Benayed, Ryma; Ashraf, Asad; Lincoln, Annie; Arcila, Maria; Stadler, Zsofia; Solit, David; Hyman, David M; Hyman, David; Zhang, Liying; Klimstra, David; Ladanyi, Marc; Offit, Kenneth; Berger, Michael; Robson, Mark

    2016-01-01

    Tumor genetic sequencing identifies potentially targetable genetic alterations with therapeutic implications. Analysis has concentrated on detecting tumor-specific variants, but recognition of germline variants may prove valuable as well. To estimate the burden of germline variants identified through routine clinical tumor sequencing. Patients with advanced cancer diagnoses eligible for studies of targeted agents at Memorial Sloan Kettering Cancer Center are offered tumor-normal sequencing with MSK-IMPACT, a 341-gene panel. We surveyed the germline variants seen in 187 overlapping genes with Mendelian disease associations in 1566 patients who had undergone tumor profiling between March and October 2014. The number of presumed pathogenic germline variants (PPGVs) and variants of uncertain significance per person in 187 genes associated with single-gene disorders and the proportions of individuals with PPGVs in clinically relevant gene subsets, in genes consistent with known tumor phenotypes, and in genes with evidence of second somatic hits in their tumors. The mean age of the 1566 patients was 58 years, and 54% were women. Presumed pathogenic germline variants in known Mendelian disease-associated genes were identified in 246 of 1566 patients (15.7%; 95% CI, 14.0%-17.6%), including 198 individuals with mutations in genes associated with cancer susceptibility. Germline findings in cancer susceptibility genes were concordant with the individual's cancer type in only 81 of 198 cases (40.9%; 95% CI, 34.3%-47.9%). In individuals with PPGVs retained in the tumor, somatic alteration of the other allele was seen in 39 of 182 cases (21.4%; 95% CI, 16.1%-28.0%), of which 13 cases did not show a known correlation of the germline mutation and a known syndrome. Mutations in non-cancer-related Mendelian disease genes were seen in 55 of 1566 cases (3.5%; 95% CI, 27.1%-45.4%). Almost every individual had more than 1 variant of uncertain significance (1565 of 1566 patients; 99

  19. Somatic and germline mosaicism for a mutation of the PHEX gene can lead to genetic transmission of X-linked hypophosphatemic rickets that mimics an autosomal dominant trait.

    Science.gov (United States)

    Goji, Katsumi; Ozaki, Kayo; Sadewa, Ahmad H; Nishio, Hisahide; Matsuo, Masafumi

    2006-02-01

    Familial hypophosphatemic rickets is usually transmitted as an X-linked dominant disorder (XLH), although autosomal dominant forms have also been observed. Genetic studies of these disorders have identified mutations in PHEX and FGF23 as the causes of X-linked dominant disorder and autosomal dominant forms, respectively. The objective of the study was to describe the molecular genetic findings in a family affected by hypophosphatemic rickets with presumed autosomal dominant inheritance. We studied a family in which the father and the elder of his two daughters, but not the second daughter, were affected by hypophosphatemic rickets. The pedigree interpretation of the family suggested that genetic transmission of the disorder occurred as an autosomal dominant trait. Direct nucleotide sequencing of FGF23 and PHEX revealed that the elder daughter was heterozygous for an R567X mutation in PHEX, rather than FGF23, suggesting that the genetic transmission occurred as an X-linked dominant trait. Unexpectedly, the father was heterozygous for this mutation. Single-nucleotide primer extension and denaturing HPLC analysis of the father using DNA from single hair roots revealed that he was a somatic mosaic for the mutation. Haplotype analysis confirmed that the father transmitted the genotypes for 18 markers on the X chromosome equally to his two daughters. The fact that the father transmitted the mutation to only one of his two daughters indicated that he was a germline mosaic for the mutation. Somatic and germline mosaicism for an X-linked dominant mutation in PHEX may mimic autosomal dominant inheritance.

  20. Clerics urge ban on altering germline cells.

    Science.gov (United States)

    Norman, C

    1983-06-24

    A resolution calling for a ban on genetic engineering of human reproductive cells has been signed by leaders of almost every major church group in the United States. Some of the religious leaders, while not certain that a total moratorium should be placed on altering germline cells, signed the statement in order to stimulate public debate on the issue. Legislation has recently been introduced in Congress to set up a committee to monitor genetic engineering and its human applications, but author Jeremy Rifkin, the impetus behind the church leaders' resolution, argues that such tampering threatens the gene pool and should be banned altogether.

  1. Germline fumarate hydratase mutations in patients with ovarian mucinous cystadenoma

    DEFF Research Database (Denmark)

    Ylisaukko-oja, Sanna K.; Cybulski, Cezary; Lehtonen, Rainer

    2006-01-01

    Germline mutations in the fumarate hydratase (FH) gene were recently shown to predispose to the dominantly inherited syndrome, hereditary leiomyomatosis and renal cell cancer (HLRCC). HLRCC is characterized by benign leiomyomas of the skin and the uterus, renal cell carcinoma, and uterine...... leiomyosarcoma. The aim of this study was to identify new families with FH mutations, and to further examine the tumor spectrum associated with FH mutations. FH germline mutations were screened from 89 patients with RCC, skin leiomyomas or ovarian tumors. Subsequently, 13 ovarian and 48 bladder carcinomas were...

  2. Rapid detection of most frequent Slovenian germ-line mutations in BRCA1 gene using real-time PCR and melting curve analysis

    International Nuclear Information System (INIS)

    Novakovic, S.; Stegel, V.

    2005-01-01

    Background. Detection of inherited mutations in cancer susceptibility genes is of great importance in some types of cancers including the colorectal cancer (mutations of APC gene in familial adenomatous polyposis - FAP, mutations in mismatch repair genes in hereditary nonpolyposis colorectal cancer - HNPCC), malignant melanoma (mutations in CDKN2A and CDK4 genes) and breast cancer (mutations in BRCA1 and BRCA2 genes). Methods. This article presents the technical data for the detection of five mutations in BRCA1 gene in breast cancer patients and their relatives. The mutations - 1806C>T, 300T>G, 300T>A, 310G>A, 5382insC - were determined by the real-time PCR and the melting curve analysis. Results and conclusion. In comparison to direct sequencing, this method proved to be sensitive and rapid enough for the routine daily determination of mutations in DNA isolated from the peripheral blood. (author)

  3. Selfish genetic elements favor the evolution of a distinction between soma and germline.

    Science.gov (United States)

    Johnson, Louise J

    2008-08-01

    Many multicellular organisms have evolved a dedicated germline. This can benefit the whole organism, but its advantages to genetic parasites have not been explored. Here I model the evolutionary success of a selfish element, such as a transposable element or endosymbiont, which is capable of creating or strengthening a germline-soma distinction in a primitively multicellular host, and find that it will always benefit the element to do so. Genes causing germline sequestration can therefore spread in a population even if germline sequestration is maladaptive for the host organism. Costly selfish elements are expected to survive only in sexual populations, so sexual species may experience an additional push toward germline-soma distinction, and hence toward cell differentiation and multicellularity.

  4. Birt-Hogg-Dube syndrome: diagnosis and management

    DEFF Research Database (Denmark)

    Menko, F.H.; Steensel, M.A. van; Giraud, S.

    2009-01-01

    Birt-Hogg-Dube syndrome (BHD) is an autosomal dominant condition characterised clinically by skin fibrofolliculomas, pulmonary cysts, spontaneous pneumothorax, and renal cancer. The condition is caused by germline mutations in the FLCN gene, which encodes folliculin; the function of this protein...... is largely unknown, although FLCN has been linked to the mTOR pathway. The availability of DNA-based diagnosis has allowed insight into the great variation in expression of FLCN, both within and between families. Patients can present with skin signs and also with pneumothorax or renal cancer. Preventive...

  5. Rapidly progressive adenomatous polyposis in a patient with germline mutations in both the APC and MLH1 genes: the worst of two worlds.

    NARCIS (Netherlands)

    Scheenstra, R; Rijcken, FE; Koornstra, JJ; Hollema, H; Fodde, R; Menko, F.H.; Sijmons, RH; Bijleveld, CM; Kleibeuker, J.H.

    2003-01-01

    The two most common inherited forms of colorectal cancer are familial adenomatous polyposis and hereditary non-polyposis colorectal cancer. Simultaneous inheritance of both an APC gene mutation and a mismatch repair gene (for example, MLH1) mutation has never been described. In the present case

  6. An unusual case of an ACTH-secreting macroadenoma with a germline variant in the aryl hydrocarbon receptor-interacting protein (AIP) gene

    DEFF Research Database (Denmark)

    Dinesen, Pia T; Dal, Jakob; Gabrovska, Plamena

    2015-01-01

    growth rather than symptoms of hypersecretion. The particular AIP gene variant identified in our patient is shared by four other reported cases of CD. Future studies are needed to assess whether the reported AIP gene variant is more than just coincidental. LEARNING POINTS: CD is occasionally dominated...

  7. Lessons for Inductive Germline Determination

    Science.gov (United States)

    Seervai, Riyad N.H.; Wessel, Gary M.

    2015-01-01

    SUMMARY Formation of the germline in an embryo marks a fresh round of reproductive potential, yet the developmental stage and location within the embryo where the primordial germ cells (PGCs) form differs wildly among species. In most animals, the germline is formed either by an inherited mechanism, in which maternal provisions within the oocyte drive localized germ-cell fate once acquired in the embryo, or an inductive mechanism that involves signaling between cells that directs germ-cell fate. The inherited mechanism has been widely studied in model organisms such as Drosophila melanogaster, Caenorhabditis elegans, Xenopus laevis, and Danio rerio. Given the rapid generation time and the effective adaptation for laboratory research of these organisms, it is not coincidental that research on these organisms has led the field in elucidating mechanisms for germline specification. The inductive mechanism, however, is less well understood and is studied primarily in the mouse (Mus musculus). In this review, we compare and contrast these two fundamental mechanisms for germline determination, beginning with the key molecular determinants that play a role in the formation of germ cells across all animal taxa. We next explore the current understanding of the inductive mechanism of germ-cell determination in mice, and evaluate the hypotheses for selective pressures on these contrasting mechanisms. We then discuss the hypothesis that the transition between these determination mechanisms, which has happened many times in phylogeny, is more of a continuum than a binary change. Finally, we propose an analogy between germline determination and sex determination in vertebrates—two of the milestones of reproduction and development—in which animals use contrasting strategies to activate similar pathways. PMID:23450642

  8. Germline TERT promoter mutations are rare in familial melanoma

    DEFF Research Database (Denmark)

    Harland, Mark; Petljak, Mia; Robles-Espinoza, Carla Daniela

    2016-01-01

    Germline CDKN2A mutations occur in 40 % of 3-or-more case melanoma families while mutations of CDK4, BAP1, and genes involved in telomere function (ACD, TERF2IP, POT1), have also been implicated in melanomagenesis. Mutation of the promoter of the telomerase reverse transcriptase (TERT) gene (c.-57...... T>G variant) has been reported in one family. We tested for the TERT promoter variant in 675 multicase families wild-type for the known high penetrance familial melanoma genes, 1863 UK population-based melanoma cases and 529 controls. Germline lymphocyte telomere length was estimated in carriers....... The c.-57 T>G TERT promoter variant was identified in one 7-case family with multiple primaries and early age of onset (earliest, 15 years) but not among population cases or controls. One family member had multiple primary melanomas, basal cell carcinomas and a bladder tumour. The blood leukocyte...

  9. Exploration of the Germline Genome of the Ciliate Chilodonella uncinata through Single-Cell Omics (Transcriptomics and Genomics

    Directory of Open Access Journals (Sweden)

    Xyrus X. Maurer-Alcalá

    2018-01-01

    Full Text Available Separate germline and somatic genomes are found in numerous lineages across the eukaryotic tree of life, often separated into distinct tissues (e.g., in plants, animals, and fungi or distinct nuclei sharing a common cytoplasm (e.g., in ciliates and some foraminifera. In ciliates, germline-limited (i.e., micronuclear-specific DNA is eliminated during the development of a new somatic (i.e., macronuclear genome in a process that is tightly linked to large-scale genome rearrangements, such as deletions and reordering of protein-coding sequences. Most studies of germline genome architecture in ciliates have focused on the model ciliates Oxytricha trifallax, Paramecium tetraurelia, and Tetrahymena thermophila, for which the complete germline genome sequences are known. Outside of these model taxa, only a few dozen germline loci have been characterized from a limited number of cultivable species, which is likely due to difficulties in obtaining sufficient quantities of “purified” germline DNA in these taxa. Combining single-cell transcriptomics and genomics, we have overcome these limitations and provide the first insights into the structure of the germline genome of the ciliate Chilodonella uncinata, a member of the understudied class Phyllopharyngea. Our analyses reveal the following: (i large gene families contain a disproportionate number of genes from scrambled germline loci; (ii germline-soma boundaries in the germline genome are demarcated by substantial shifts in GC content; (iii single-cell omics techniques provide large-scale quality germline genome data with limited effort, at least for ciliates with extensively fragmented somatic genomes. Our approach provides an efficient means to understand better the evolution of genome rearrangements between germline and soma in ciliates.

  10. Evidence for a hierarchical transcriptional circuit in Drosophila male germline involving testis-specific TAF and two gene-specific transcription factors, Mod and Acj6.

    Science.gov (United States)

    Jiang, Mei; Gao, Zhengliang; Wang, Jian; Nurminsky, Dmitry I

    2018-01-01

    To analyze transcription factors involved in gene regulation by testis-specific TAF (tTAF), tTAF-dependent promoters were mapped and analyzed in silico. Core promoters show decreased AT content, paucity of classical promoter motifs, and enrichment with translation control element CAAAATTY. Scanning of putative regulatory regions for known position frequency matrices identified 19 transcription regulators possibly contributing to tTAF-driven gene expression. Decreased male fertility associated with mutation in one of the regulators, Acj6, indicates its involvement in male reproduction. Transcriptome study of testes from male mutants for tTAF, Acj6, and previously characterized tTAF-interacting factor Modulo implies the existence of a regulatory hierarchy of tTAF, Modulo and Acj6, in which Modulo and/or Acj6 regulate one-third of tTAF-dependent genes. © 2017 Federation of European Biochemical Societies.

  11. The spectrum of HNF1A gene mutations in Greek patients with MODY3: relative frequency and identification of seven novel germline mutations.

    Science.gov (United States)

    Tatsi, Christina; Kanaka-Gantenbein, Christina; Vazeou-Gerassimidi, Adriani; Chrysis, Dionysios; Delis, Dimitrios; Tentolouris, Nikolaos; Dacou-Voutetakis, Catherine; Chrousos, George P; Sertedaki, Amalia

    2013-11-01

    Maturity-Onset Diabetes of the Young (MODY) is the most common type of monogenic diabetes accounting for 1-2% of the population with diabetes. The relative incidence of HNF1A-MODY (MODY3) is high in European countries; however, data are not available for the Greek population. The aims of this study were to determine the relative frequency of MODY3 in Greece, the type of the mutations observed, and their relation to the phenotype of the patients. Three hundred ninety-five patients were referred to our center because of suspected MODY during a period of 15 yr. The use of Denaturing Gradient Gel Electrophoresis of polymerase chain reaction amplified DNA revealed 72 patients carrying Glucokinase gene mutations (MODY2) and 8 patients carrying HNF1A gene mutations (MODY3). After using strict criteria, 54 patients were selected to be further evaluated by direct sequencing or by multiplex ligation probe amplification (MLPA) for the presence of HNF1A gene mutations. In 16 unrelated patients and 13 of their relatives, 15 mutations were identified in the HNF1A gene. Eight of these mutations were previously reported, whereas seven were novel. Clinical features, such as age of diabetes at diagnosis or severity of hyperglycemia, were not related to the mutation type or location. In our cohort of patients fulfilling strict clinical criteria for MODY, 12% carried an HNF1A gene mutation, suggesting that defects of this gene are responsible for a significant proportion of monogenic diabetes in the Greek population. No clear phenotype-genotype correlations were identified. © 2013 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  12. Homozygous germ-line mutation of the PMS2 mismatch repair gene: a unique case report of constitutional mismatch repair deficiency (CMMRD)

    OpenAIRE

    Ramchander, N. C.; Ryan, N. A. J.; Crosbie, E. J.; Evans, D. G.

    2017-01-01

    BackgroundConstitutional mismatch repair deficiency syndrome results from bi-allelic inheritance of mutations affecting the key DNA mismatch repair genes: MLH1, MSH2, MSH6 or PMS2. Individuals with bi-allelic mutations have a dysfunctional mismatch repair system from birth; as a result, constitutional mismatch repair deficiency syndrome is characterised by early onset malignancies. Fewer than 150 cases have been reported in the literature over the past 20 years. This is the first report of th...

  13. Germ-line variants identified by next generation sequencing in a panel of estrogen and cancer associated genes correlate with poor clinical outcome in Lynch syndrome patients.

    Science.gov (United States)

    Jóri, Balazs; Kamps, Rick; Xanthoulea, Sofia; Delvoux, Bert; Blok, Marinus J; Van de Vijver, Koen K; de Koning, Bart; Oei, Felicia Trups; Tops, Carli M; Speel, Ernst Jm; Kruitwagen, Roy F; Gomez-Garcia, Encarna B; Romano, Andrea

    2015-12-01

    The risk to develop colorectal and endometrial cancers among subjects testing positive for a pathogenic Lynch syndrome mutation varies, making the risk prediction difficult. Genetic risk modifiers alter the risk conferred by inherited Lynch syndrome mutations, and their identification can improve genetic counseling. We aimed at identifying rare genetic modifiers of the risk of Lynch syndrome endometrial cancer. A family based approach was used to assess the presence of genetic risk modifiers among 35 Lynch syndrome mutation carriers having either a poor clinical phenotype (early age of endometrial cancer diagnosis or multiple cancers) or a neutral clinical phenotype. Putative genetic risk modifiers were identified by Next Generation Sequencing among a panel of 154 genes involved in endometrial physiology and carcinogenesis. A simple pipeline, based on an allele frequency lower than 0.001 and on predicted non-conservative amino-acid substitutions returned 54 variants that were considered putative risk modifiers. The presence of two or more risk modifying variants in women carrying a pathogenic Lynch syndrome mutation was associated with a poor clinical phenotype. A gene-panel is proposed that comprehends genes that can carry variants with putative modifying effects on the risk of Lynch syndrome endometrial cancer. Validation in further studies is warranted before considering the possible use of this tool in genetic counseling.

  14. A germline FANCA alteration that is associated with increased sensitivity to DNA damaging agents.

    OpenAIRE

    Wilkes, David C; Sailer, Verena; Xue, Hui; Cheng, Hongwei; Collins, Colin C; Gleave, Martin; Wang, Yuzhuo; Demichelis, Francesca; Beltran, Himisha; Rubin, Mark Andrew; Rickman, David S

    2017-01-01

    Defects in genes involved in DNA damage repair (DDR) pathway are emerging as novel biomarkers and targets for new prostate cancer drug therapies. A previous report revealed an association between an exceptional response to cisplatin treatment and a somatic loss of heterozygosity (LOH) of FANCA in a patient with metastatic prostate cancer who also harbored a germline FANCA variant (S1088F). Although germline FANCA mutations are the most frequent alterations in patients with Fanconi anemia, ger...

  15. Prevalence of deleterious ATM germline mutations in gastric cancer patients.

    Science.gov (United States)

    Huang, Dong-Sheng; Tao, Hou-Quan; He, Xu-Jun; Long, Ming; Yu, Sheng; Xia, Ying-Jie; Wei, Zhang; Xiong, Zikai; Jones, Sian; He, Yiping; Yan, Hai; Wang, Xiaoyue

    2015-12-01

    Besides CDH1, few hereditary gastric cancer predisposition genes have been previously reported. In this study, we discovered two germline ATM mutations (p.Y1203fs and p.N1223S) in a Chinese family with a history of gastric cancer by screening 83 cancer susceptibility genes. Using a published exome sequencing dataset, we found deleterious germline mutations of ATM in 2.7% of 335 gastric cancer patients of different ethnic origins. The frequency of deleterious ATM mutations in gastric cancer patients is significantly higher than that in general population (p=0.0000435), suggesting an association of ATM mutations with gastric cancer predisposition. We also observed biallelic inactivation of ATM in tumors of two gastric cancer patients. Further evaluation of ATM mutations in hereditary gastric cancer will facilitate genetic testing and risk assessment.

  16. Germline Variants of Prostate Cancer in Japanese Families.

    Directory of Open Access Journals (Sweden)

    Takahide Hayano

    Full Text Available Prostate cancer (PC is the second most common cancer in men. Family history is the major risk factor for PC. Only two susceptibility genes were identified in PC, BRCA2 and HOXB13. A comprehensive search of germline variants for patients with PC has not been reported in Japanese families. In this study, we conducted exome sequencing followed by Sanger sequencing to explore responsible germline variants in 140 Japanese patients with PC from 66 families. In addition to known susceptibility genes, BRCA2 and HOXB13, we identified TRRAP variants in a mutually exclusive manner in seven large PC families (three or four patients per family. We also found shared variants of BRCA2, HOXB13, and TRRAP from 59 additional small PC families (two patients per family. We identified two deleterious HOXB13 variants (F127C and G132E. Further exploration of the shared variants in rest of the families revealed deleterious variants of the so-called cancer genes (ATP1A1, BRIP1, FANCA, FGFR3, FLT3, HOXD11, MUTYH, PDGFRA, SMARCA4, and TCF3. The germline variant profile provides a new insight to clarify the genetic etiology and heterogeneity of PC among Japanese men.

  17. A novel germline mutation (c.A527G) in STK11 gene causes Peutz-Jeghers syndrome in a Chinese girl: A case report.

    Science.gov (United States)

    Zhao, Zi-Ye; Jiang, Yu-Liang; Li, Bai-Rong; Yang, Fu; Li, Jing; Jin, Xiao-Wei; Sun, Shu-Han; Ning, Shou-Bin

    2017-12-01

    Peutz-Jeghers syndrome (PJS) is a Mendelian autosomal dominant disease caused by mutations in the tumor suppressor gene, serine/threonine kinase 11 (STK11). The features of this syndrome include gastrointestinal (GI) hamartomas, melanin spots on the lips and the extremities, and an increased risk of developing cancer. Early onset of disease is often characterized by mucocutaneous pigmentation and intussusception due to GI polyps in childhood. A girl with a positive family history grew oral pigmentation at 1 and got intussusception by small bowel hamartomas at 5. She was diagnosed with PJS based on oral pigmentation and a positive family history of PJS. Enteroscopy was employed to treat the GI polyps. Sanger sequencing was used to investigate STK11 mutation in this family. A large jejunal polyp together with other smaller ones was resected, and the girl recovered uneventfully. We discovered a heterozygous substitution in STK11, c.A527G in exon 4, in the girl and her father who was also a PJS patient, and the amine acid change was an aspartic acid-glycine substitution in codon 176. This mutation was not found in other healthy family members and 50 unrelated non-PJS controls, and it is not recorded in databases, which prove it a novel mutation. Evolutionary conservation analysis of amino acid residues showed this aspartic acid is a conserved one between species, and protein structure prediction by SWISS-MODEL indicated an obvious change in local structure. In addition, PolyPhen-2 score for this mutation is 1, which indicates it probably damaging. PJS can cause severe complication like intussusception in young children, and early screening for small bowel may be beneficial for these patients. The mutation of STK11 found in this girl is a novel one, which enlarges the spectrum of STK11. Our analysis supported it a causative one in PJS.

  18. Homozygous germ-line mutation of the PMS2 mismatch repair gene: a unique case report of constitutional mismatch repair deficiency (CMMRD).

    Science.gov (United States)

    Ramchander, N C; Ryan, N A J; Crosbie, E J; Evans, D G

    2017-04-05

    Constitutional mismatch repair deficiency syndrome results from bi-allelic inheritance of mutations affecting the key DNA mismatch repair genes: MLH1, MSH2, MSH6 or PMS2. Individuals with bi-allelic mutations have a dysfunctional mismatch repair system from birth; as a result, constitutional mismatch repair deficiency syndrome is characterised by early onset malignancies. Fewer than 150 cases have been reported in the literature over the past 20 years. This is the first report of the founder PMS2 mutation - NM_000535.5:c.1500del (p.Val501TrpfsTer94) in exon 11 and its associated cancers in this family. The proband is 30 years old and is alive today. She is of Pakistani ethnic origin and a product of consanguinity. She initially presented aged 24 with painless bleeding per-rectum from colorectal polyps and was referred to clinical genetics. Clinical examination revealed two café-au-lait lesions, lichen planus, and a dermoid cyst. Her sister had been diagnosed in childhood with an aggressive brain tumour followed by colorectal cancer. During follow up, the proband developed 37 colorectal adenomatous polyps, synchronous ovarian and endometrial adenocarcinomas, and ultimately a metachronous gastric adenocarcinoma. DNA sequencing of peripheral lymphocytes revealed a bi-allelic inheritance of the PMS2 mutation NM_000535.5:c.1500del (p.Val501TrpfsTer94) in exon 11. Ovarian tumour tissue demonstrated low microsatellite instability. To date, she has had a total abdominal hysterectomy, bilateral salpingo-oophorectomy, and a total gastrectomy. Aspirin and oestrogen-only hormone replacement therapy provide some chemoprophylaxis and manage postmenopausal symptoms, respectively. An 18-monthly colonoscopy surveillance programme has led to the excision of three high-grade dysplastic colorectal tubular adenomatous polyps. The proband's family pedigree displays multiple relatives with cancers including a likely case of 'true' Turcot syndrome. Constitutional mismatch repair

  19. Molecular analysis on germline mutation caused by low-dose irradiation

    International Nuclear Information System (INIS)

    Uchiyama, R.; Fujikawa, K.; Nishimura, M.; Adzuma, H.; Shimada, Y.; Yamauchi, M.

    2003-01-01

    Full text: Genetic heterogeneity and a low frequency of germline mutation at single-copy gene loci have limited the direct measurement of germline mutation in human populations. Two conflicting results have been reported for the effect of ionizing radiation on germline mutation in human populations. A study conducted on the first-generation progeny of the survivors of the atomic bombs at Hiroshima and Nagasaki found no significant increase in germline mutations. On the other hand, a significant increase in germline mutation was reported among the human population in the Belarus area after the Chernobyl accident in 1986. We investigated the germline mutation at the molecular level using experimental mouse strains with different genetic backgrounds to assess the risk of ionizing radiation on human populations. The C3H male parents were exposed to X ray (0, 0.3, 1, and 3Gy) and mated with unexposed C57BL females after two weeks interval, so as to detect the germline mutation occurred at the spermatid stage. Genomic DNA samples were prepared from the both parents and F1s, and the genomic DNA sequences were compared between parents and offspring at the specific genomic gene loci, such as adenine phosphoribosyl transferase (aprt) gene and cytidine triphosphate synthetase (ctps) gene, using the automated DNA sequencer. Also hypervariable Pc-1 (Ms6-hm) minisatellite repeat locus was analyzed by using Southern blot hybridization technique. Our preliminary results indicated that the changes of the restriction DNA fragment length in offspring did not reflect the occurrence of the mutation, such as point mutation, insertion, and deletion, in the genomic gene loci including the intervening sequence (intron)

  20. Flow cytometry sorting of nuclei enables the first global characterization of Paramecium germline DNA and transposable elements.

    Science.gov (United States)

    Guérin, Frédéric; Arnaiz, Olivier; Boggetto, Nicole; Denby Wilkes, Cyril; Meyer, Eric; Sperling, Linda; Duharcourt, Sandra

    2017-04-26

    DNA elimination is developmentally programmed in a wide variety of eukaryotes, including unicellular ciliates, and leads to the generation of distinct germline and somatic genomes. The ciliate Paramecium tetraurelia harbors two types of nuclei with different functions and genome structures. The transcriptionally inactive micronucleus contains the complete germline genome, while the somatic macronucleus contains a reduced genome streamlined for gene expression. During development of the somatic macronucleus, the germline genome undergoes massive and reproducible DNA elimination events. Availability of both the somatic and germline genomes is essential to examine the genome changes that occur during programmed DNA elimination and ultimately decipher the mechanisms underlying the specific removal of germline-limited sequences. We developed a novel experimental approach that uses flow cell imaging and flow cytometry to sort subpopulations of nuclei to high purity. We sorted vegetative micronuclei and macronuclei during development of P. tetraurelia. We validated the method by flow cell imaging and by high throughput DNA sequencing. Our work establishes the proof of principle that developing somatic macronuclei can be sorted from a complex biological sample to high purity based on their size, shape and DNA content. This method enabled us to sequence, for the first time, the germline DNA from pure micronuclei and to identify novel transposable elements. Sequencing the germline DNA confirms that the Pgm domesticated transposase is required for the excision of all ~45,000 Internal Eliminated Sequences. Comparison of the germline DNA and unrearranged DNA obtained from PGM-silenced cells reveals that the latter does not provide a faithful representation of the germline genome. We developed a flow cytometry-based method to purify P. tetraurelia nuclei to high purity and provided quality control with flow cell imaging and high throughput DNA sequencing. We identified 61

  1. Germline V repertoires: Origin, maintenance, diversification.

    Science.gov (United States)

    Steele, E J; Lindley, R A

    2018-06-01

    In our view, Melvin Cohn (Scand J Immunol. 2018;87:e12640) has set out the logical guidelines towards a resolution of the very real enigma of the selectability of vertebrate germline Ig V repertoires under the current evolutionary paradigm…" A somatically derived repertoire scrambles this (germline VL + VH) substrate so that its specificities are lost, making it un-selectable in the germline. Consequently, evolution faced an incompatibility." It is argued here in Reply that a reverse transcriptase-based soma-to-germline process (S->G) targeting germline V segment arrays goes some considerable way to resolving fundamental contradictions on the origin, maintenance and then real-time adaptive diversification of these limited sets of V segments encoded within various V repertoire arrays. © 2018 The Foundation for the Scandinavian Journal of Immunology.

  2. Germline progenitors escape the widespread phenomenon of homolog pairing during Drosophila development.

    Directory of Open Access Journals (Sweden)

    Eric F Joyce

    Full Text Available Homolog pairing, which plays a critical role in meiosis, poses a potential risk if it occurs in inappropriate tissues or between nonallelic sites, as it can lead to changes in gene expression, chromosome entanglements, and loss-of-heterozygosity due to mitotic recombination. This is particularly true in Drosophila, which supports organismal-wide pairing throughout development. Discovered over a century ago, such extensive pairing has led to the perception that germline pairing in the adult gonad is an extension of the pairing established during embryogenesis and, therefore, differs from the mechanism utilized in most species to initiate pairing specifically in the germline. Here, we show that, contrary to long-standing assumptions, Drosophila meiotic pairing in the gonad is not an extension of pairing established during embryogenesis. Instead, we find that homologous chromosomes are unpaired in primordial germ cells from the moment the germline can be distinguished from the soma in the embryo and remain unpaired even in the germline stem cells of the adult gonad. We further establish that pairing originates immediately after the stem cell stage. This pairing occurs well before the initiation of meiosis and, strikingly, continues through the several mitotic divisions preceding meiosis. These discoveries indicate that the spatial organization of the Drosophila genome differs between the germline and the soma from the earliest moments of development and thus argue that homolog pairing in the germline is an active process as versus a passive continuation of pairing established during embryogenesis.

  3. Germline Manipulation and Our Future Worlds.

    Science.gov (United States)

    Harris, John

    2015-01-01

    Two genetic technologies capable of making heritable changes to the human genome have revived interest in, and in some quarters a very familiar panic concerning, so-called germline interventions. These technologies are: most recently the use of CRISPR/Cas9 to edit genes in non-viable IVF zygotes and Mitochondrial Replacement Therapy (MRT) the use of which was approved in principle in a landmark vote earlier this year by the United Kingdom Parliament. The possibility of using either of these techniques in humans has encountered the most violent hostility and suspicion. However it is important to be aware that much of this hostility dates back to the fears associated with In Vitro Fertilization (IVF) and other reproductive technologies and by cloning; fears which were baseless at the time concerning both IVF and cloning the use of both of which have proved to be highly beneficial to humanity and which have been effectively regulated and controlled. This paper argues that CRISPR should by pursued through researh until it is safe enough for use in humans but there is no reason to suppose at this stage that such use will be unsafe or unethical (Collins 2015).

  4. A 44 bp intestine-specific hermaphrodite-specific enhancer from the C. elegans vit-2 vitellogenin gene is directly regulated by ELT-2, MAB-3, FKH-9 and DAF-16 and indirectly regulated by the germline, by daf-2/insulin signaling and by the TGF-β/Sma/Mab pathway.

    Science.gov (United States)

    Goszczynski, Barbara; Captan, Vasile V; Danielson, Alicia M; Lancaster, Brett R; McGhee, James D

    2016-05-01

    The Caenorhabditis elegans vitellogenin genes are transcribed in the intestine of adult hermaphrodites but not of males. A 44-bp region from the vit-2 gene promoter is able largely to reconstitute this tissue-, stage- and sex-specific-expression. This "enhancer" contains a binding site for the DM-domain factor MAB-3, the male-specific repressor of vitellogenesis, as well as an activator site that we show is the direct target of the intestinal GATA factor ELT-2. We further show that the enhancer is directly activated by the winged-helix/forkhead-factor FKH-9, (whose gene has been shown by others to be a direct target of DAF-16), by an unknown activator binding to the MAB-3 site, and by the full C. elegans TGF-β/Sma/Mab pathway acting within the intestine. The vit-2 gene has been shown by others to be repressed by the daf-2/daf-16 insulin signaling pathway, which so strongly influences aging and longevity in C. elegans. We show that the activity of the 44 bp vit-2 enhancer is abolished by loss of daf-2 but is restored by simultaneous loss of daf-16. DAF-2 acts from outside of the intestine but DAF-16 acts both from outside of the intestine and from within the intestine where it binds directly to the same non-canonical target site that interacts with FKH-9. Activity of the 44 bp vit-2 enhancer is also inhibited by loss of the germline, in a manner that is only weakly influenced by DAF-16 but that is strongly influenced by KRI-1, a key downstream effector in the pathway by which germline loss increases C. elegans lifespan. The complex behavior of this enhancer presumably allows vitellogenin gene transcription to adjust to demands of body size, germline proliferation and nutritional state but we suggest that the apparent involvement of this enhancer in aging and longevity "pathways" could be incidental. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Germline RAD51B truncating mutation in a family with cutaneous melanoma

    DEFF Research Database (Denmark)

    Wadt, Karin A W; Aoude, Lauren G; Golmard, Lisa

    2015-01-01

    Known melanoma predisposition genes only account for around 40% of high-density melanoma families. Other rare mutations are likely to play a role in melanoma predisposition. RAD51B plays an important role in DNA repair through homologous recombination, and inactivation of RAD51B has been implicated...... in tumorigenesis. Thus RAD51B is a good candidate melanoma susceptibility gene, and previously, a germline splicing mutation in RAD51B has been identified in a family with early-onset breast cancer. In order to find genetic variants associated with melanoma predisposition, whole-exome sequencing was carried out...... on blood samples from a three-case cutaneous melanoma family. We identified a novel germline RAD51B nonsense mutation, and we demonstrate reduced expression of RAD51B in melanoma cells indicating inactivation of RAD51B. This is only the second report of a germline truncating RAD51B mutation. While...

  6. Xeroderma Pigmentosum: Low Prevalence of Germline XPA Mutations in a Brazilian XP Population

    Directory of Open Access Journals (Sweden)

    Karina Miranda Santiago

    2015-04-01

    Full Text Available Xeroderma pigmentosum (XP is a rare autosomal recessive disorder characterized by DNA repair defects that cause photophobia, sunlight-induced cancers, and neurodegeneration. Prevalence of germline mutations in the nucleotide excision repair gene XPA vary significantly in different populations. No Brazilian patients have been reported to carry a germline mutation in this gene. In this study, the germline mutational status of XPA was determined in Brazilian patients exhibiting major clinical features of XP syndrome. The study was conducted on 27 unrelated patients from select Brazilian families. A biallelic inactivating transition mutation c.619C>T (p.Arg207Ter was identified in only one patient with a history of neurological impairment and mild skin abnormalities. These findings suggest that XP syndrome is rarely associated with inherited disease-causing XPA mutations in the Brazilian population. Additionally, this report demonstrates the effectiveness of genotype-phenotype correlation as a valuable tool to guide direct genetic screening.

  7. Xeroderma pigmentosum: low prevalence of germline XPA mutations in a Brazilian XP population.

    Science.gov (United States)

    Santiago, Karina Miranda; França de Nóbrega, Amanda; Rocha, Rafael Malagoli; Rogatto, Silvia Regina; Achatz, Maria Isabel

    2015-04-22

    Xeroderma pigmentosum (XP) is a rare autosomal recessive disorder characterized by DNA repair defects that cause photophobia, sunlight-induced cancers, and neurodegeneration. Prevalence of germline mutations in the nucleotide excision repair gene XPA vary significantly in different populations. No Brazilian patients have been reported to carry a germline mutation in this gene. In this study, the germline mutational status of XPA was determined in Brazilian patients exhibiting major clinical features of XP syndrome. The study was conducted on 27 unrelated patients from select Brazilian families. A biallelic inactivating transition mutation c.619C>T (p.Arg207Ter) was identified in only one patient with a history of neurological impairment and mild skin abnormalities. These findings suggest that XP syndrome is rarely associated with inherited disease-causing XPA mutations in the Brazilian population. Additionally, this report demonstrates the effectiveness of genotype-phenotype correlation as a valuable tool to guide direct genetic screening.

  8. A germline FANCA alteration that is associated with increased sensitivity to DNA damaging agents.

    Science.gov (United States)

    Wilkes, David C; Sailer, Verena; Xue, Hui; Cheng, Hongwei; Collins, Colin C; Gleave, Martin; Wang, Yuzhuo; Demichelis, Francesca; Beltran, Himisha; Rubin, Mark A; Rickman, David S

    2017-09-01

    Defects in genes involved in DNA damage repair (DDR) pathway are emerging as novel biomarkers and targets for new prostate cancer drug therapies. A previous report revealed an association between an exceptional response to cisplatin treatment and a somatic loss of heterozygosity (LOH) of FANCA in a patient with metastatic prostate cancer who also harbored a germline FANCA variant (S1088F). Although germline FANCA mutations are the most frequent alterations in patients with Fanconi anemia, germline alterations are less common in prostate cancer. We hypothesized that the germline S1088F FANCA variant in combination with FANCA LOH was deleterious for FANCA function and contributed to the patient's exceptional response to cisplatin. We show that although it properly localizes to the nucleus, the S1088F FANCA mutant protein disrupts the FANC protein complex resulting in increased sensitivity to DNA damaging agents. Because molecular stratification is emerging as a strategy for treating men with metastatic, castrate-resistant prostate cancer harboring specific DDR gene defects, our findings suggest that more biomarker studies are needed to better define clinically relevant germline and somatic alterations. © 2017 Wilkes et al.; Published by Cold Spring Harbor Laboratory Press.

  9. Foxn1[Cre] Expression in the Male Germline.

    Science.gov (United States)

    Shi, Jianjun; Getun, Irina; Torres, Bivian; Petrie, Howard T

    2016-01-01

    Foxn1 (forkhead box N1), also known as the nude gene or winged-helix nude (Whn), is a forkhead transcription factor thought to be restricted to keratinocytes in the skin and thymus. Consistent with this tissue distribution, spontaneous or targeted mutation of Foxn1 results in the absence of both hair and a thymus. Genetic manipulation of the Foxn1 locus thus represents a powerful tool for tissue specific gene control in the skin and thymus, and tools such as Cre recombinase under control of the Foxn1 locus are widely used for this purpose. Unexpectedly, we show that Foxn1[Cre] exhibits unexpected activity in male germ cells, resulting in ubiquitous targeting of loxP-flanked alleles in all tissues in offspring from Foxn1[Cre] expressing male mice. Inheritance of recombined loxP alleles occurs independently of Cre inheritance (i.e., offspring lacking Cre nonetheless exhibit recombined alleles), suggesting that Foxn1[Cre] induced recombination in male germ cells must occur prior to meiosis in diploid germ cells. Together with previously published data, our results show that Foxn1, and alleles under its control, are expressed in the pre-meiotic male germline, revealing a new tool for germline targeting of genes, and raising important concerns for gender selection when using Foxn1 regulatory elements.

  10. Lynch syndrome caused by germline PMS2 mutations: delineating the cancer risk

    NARCIS (Netherlands)

    Broeke, S.W. ten; Brohet, R.M.; Tops, C.M.; Klift, H.M. van der; Velthuizen, M.E.; Bernstein, I.; Capella Munar, G.; Garcia, E.; Hoogerbrugge, N.; Letteboer, T.G.; Menko, F.H.; Lindblom, A.; Mensenkamp, A.R.; Moller, P.; Os, T.A. van; Rahner, N.; Redeker, B.J.; Sijmons, R.H.; Spruijt, L.; Suerink, M.; Vos, Y.J.; Wagner, A.; Hes, F.J.; Vasen, H.F.A.; Nielsen, M.; Wijnen, J.T.

    2015-01-01

    PURPOSE: The clinical consequences of PMS2 germline mutations are poorly understood compared with other Lynch-associated mismatch repair gene (MMR) mutations. The aim of this European cohort study was to define the cancer risk faced by PMS2 mutation carriers. METHODS: Data were collected from 98

  11. Lynch Syndrome Caused by Germline PMS2 Mutations: Delineating the Cancer Risk

    NARCIS (Netherlands)

    ten Broeke, Sanne W.; Brohet, Richard M.; Tops, Carli M.; van der Klift, Heleen M.; Velthuizen, Mary E.; Bernstein, Inge; Capellá Munar, Gabriel; Gomez Garcia, Encarna; Hoogerbrugge, Nicoline; Letteboer, Tom G. W.; Menko, Fred H.; Lindblom, Annika; Mensenkamp, Arjen R.; Moller, Pal; van Os, Theo A.; Rahner, Nils; Redeker, Bert J. W.; Sijmons, Rolf H.; Spruijt, Liesbeth; Suerink, Manon; Vos, Yvonne J.; Wagner, Anja; Hes, Frederik J.; Vasen, Hans F.; Nielsen, Maartje; Wijnen, Juul T.

    2015-01-01

    Purpose The clinical consequences of PMS2 germline mutations are poorly understood compared with other Lynch-associated mismatch repair gene (MMR) mutations. The aim of this European cohort study was to define the cancer risk faced by PMS2 mutation carriers. Methods Data were collected from 98 PMS2

  12. Lynch Syndrome Caused by Germline PMS2 Mutations : Delineating the Cancer Risk

    NARCIS (Netherlands)

    ten Broeke, Sanne W.; Brohet, Richard M.; Tops, Carli M.; van der Klift, Heleen M.; Velthuizen, Mary E.; Bernstein, Inge; Capella Munar, Gabriel; Garcia, Encarna Gomez; Hoogerbrugge, Nicoline; Letteboer, Tom G. W.; Menko, Fred H.; Lindblom, Annika; Mensenkamp, Arjen R.; Moller, Pal; Van Os, Theo A.; Rahner, Nils; Redeker, Bert J. W.; Sijmons, Rolf H.; Spruijt, Liesbeth; Suerink, Manon; Vos, Yvonne J.; Wagner, Anja; Hes, Frederik J.; Vasen, Hans F.; Nielsen, Maartje; Wijnen, Juul T.

    2015-01-01

    Purpose The clinical consequences of PMS2 germline mutations are poorly understood compared with other Lynch-associated mismatch repair gene (MMR) mutations. The aim of this European cohort study was to define the cancer risk faced by PMS2 mutation carriers. Methods Data were collected from 98 PMS2

  13. Elucidating the impact of neurofibromatosis-1 germline mutations on neurofibromin function and dopamine-based learning.

    Science.gov (United States)

    Anastasaki, Corina; Woo, Albert S; Messiaen, Ludwine M; Gutmann, David H

    2015-06-15

    Neurofibromatosis type 1 (NF1) is a common autosomal dominant neurologic condition characterized by significant clinical heterogeneity, ranging from malignant cancers to cognitive deficits. Recent studies have begun to reveal rare genotype-phenotype correlations, suggesting that the specific germline NF1 gene mutation may be one factor underlying disease heterogeneity. The purpose of this study was to define the impact of the germline NF1 gene mutation on brain neurofibromin function relevant to learning. Herein, we employ human NF1-patient primary skin fibroblasts, induced pluripotent stem cells and derivative neural progenitor cells (NPCs) to demonstrate that NF1 germline mutations have dramatic effects on neurofibromin expression. Moreover, while all NF1-patient NPCs exhibit increased RAS activation and reduced cyclic AMP generation, there was a neurofibromin dose-dependent reduction in dopamine (DA) levels. Additionally, we leveraged two complementary Nf1 genetically-engineered mouse strains in which hippocampal-based learning and memory is DA-dependent to establish that neuronal DA levels and signaling as well as mouse spatial learning are controlled in an Nf1 gene dose-dependent manner. Collectively, this is the first demonstration that different germline NF1 gene mutations differentially dictate neurofibromin function in the brain. © The Author 2015. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  14. Sex chromosomes and germline transcriptomics explored by single-cell sequencing and RNA-tomography

    NARCIS (Netherlands)

    Vértesy, Ábel

    2018-01-01

    In our study of germ cell differentiation, we applied two recently developed technologies on the germline of various model organisms: single-cell mRNA sequencing and RNA-tomography. For the first time we could look at gene expression with such a high resolution, and this led us to discover the

  15. A recurrent germline PAX5 mutation confers susceptibility to pre-B cell acute lymphoblastic leukemia

    NARCIS (Netherlands)

    Shah, S.; Schrader, K.A.; Waanders, E.; Timms, A.E.; Vijai, J.; Miething, C.; Wechsler, J.; Yang, J.; Hayes, J.; Klein, R.J.; Zhang, J.; Wei, L.; Wu, G.; Rusch, M.; Nagahawatte, P.; Ma, J; Chen, S.C.; Song, G.; Cheng, J.; Meyers, P.; Bhojwani, D.; Jhanwar, S.; Maslak, P.; Fleisher, M.; Littman, J.; Offit, L.; Rau-Murthy, R.; Fleischut, M.H.; Corines, M.; Murali, R.; Gao, X.; Manschreck, C.; Kitzing, T.; Murty, V.V.; Raimondi, S.C.; Kuiper, R.P.; Simons, A.; Schiffman, J.D.; Onel, K.; Plon, S.E.; Wheeler, D.A.; Ritter, D.; Ziegler, D.S.; Tucker, K.; Sutton, R.; Chenevix-Trench, G.; Li, J.; Huntsman, D.G.; Hansford, S.; Senz, J.; Walsh, T.; Lee (Helen Dowling Instituut), M. van der; Hahn, C.N.; Roberts, K.G.; King, M.C.; Lo, S.M.; Levine, R.L.; Viale, A.; Socci, N.D.; Nathanson, K.L.; Scott, H.S.; Daly, M.; Lipkin, S.M.; Lowe, S.W.; Downing, J.R.; Altshuler, D.; Sandlund, J.T.; Horwitz, M.S.; Mullighan, C.G.; Offit, K.

    2013-01-01

    Somatic alterations of the lymphoid transcription factor gene PAX5 (also known as BSAP) are a hallmark of B cell precursor acute lymphoblastic leukemia (B-ALL), but inherited mutations of PAX5 have not previously been described. Here we report a new heterozygous germline variant, c.547G>A

  16. Germline competence of mouse ES and iPS cell lines: Chimera technologies and genetic background.

    Science.gov (United States)

    Carstea, Ana Claudia; Pirity, Melinda K; Dinnyes, Andras

    2009-12-31

    In mice, gene targeting by homologous recombination continues to play an essential role in the understanding of functional genomics. This strategy allows precise location of the site of transgene integration and is most commonly used to ablate gene expression ("knock-out"), or to introduce mutant or modified alleles at the locus of interest ("knock-in"). The efficacy of producing live, transgenic mice challenges our understanding of this complex process, and of the factors which influence germline competence of embryonic stem cell lines. Increasingly, evidence indicates that culture conditions and in vitro manipulation can affect the germline-competence of Embryonic Stem cell (ES cell) lines by accumulation of chromosome abnormalities and/or epigenetic alterations of the ES cell genome. The effectiveness of ES cell derivation is greatly strain-dependent and it may also influence the germline transmission capability. Recent technical improvements in the production of germline chimeras have been focused on means of generating ES cells lines with a higher germline potential. There are a number of options for generating chimeras from ES cells (ES chimera mice); however, each method has its advantages and disadvantages. Recent developments in induced pluripotent stem (iPS) cell technology have opened new avenues for generation of animals from genetically modified somatic cells by means of chimera technologies. The aim of this review is to give a brief account of how the factors mentioned above are influencing the germline transmission capacity and the developmental potential of mouse pluripotent stem cell lines. The most recent methods for generating specifically ES and iPS chimera mice, including the advantages and disadvantages of each method are also discussed.

  17. Molecular and clinical characteristics of MSH6 variants : An analysis of 25 index carriers of a germline variant

    NARCIS (Netherlands)

    Olderode - Berends, Maria; Wu, Ying; Sijmons, RH; Mensink, RGJ; van der Sluis, T; Hordijk-Hos, JM; de Vries, EGE; Hollema, H; Karrenbeld, Arend; Buys, CHCM; van der Zee, AGJ; Hofstra, RMW; Kleibeuker, JH

    The MSH6 gene is one of the mismatch-repair genes involved in hereditary nonpolyposis colorectal cancer (HNPCC). Three hundred sixteen individuals who were known or suspected to have HNPCC were analyzed for MSH6 germline mutations. For 25 index patients and 8 relatives with MSH6 variants, molecular

  18. The protein kinase MBK-1 contributes to lifespan extension in daf-2 mutant and germline-deficient Caenorhabditis elegans.

    Science.gov (United States)

    Mack, Hildegard I D; Zhang, Peichuan; Fonslow, Bryan R; Yates, John R

    2017-05-25

    In Caenorhabditis elegans , reduction of insulin/IGF-1 like signaling and loss of germline stem cells both increase lifespan by activating the conserved transcription factor DAF-16 (FOXO). While the mechanisms that regulate DAF-16 nuclear localization in response to insulin/IGF-1 like signaling are well characterized, the molecular pathways that act in parallel to regulate DAF-16 transcriptional activity, and the pathways that couple DAF-16 activity to germline status, are not fully understood at present. Here, we report that inactivation of MBK-1, the C. elegans ortholog of the human FOXO1-kinase DYRK1A substantially shortens the prolonged lifespan of daf-2 and glp-1 mutant animals while decreasing wild-type lifespan to a lesser extent. On the other hand, lifespan-reduction by mutation of the MBK-1-related kinase HPK-1 was not preferential for long-lived mutants. Interestingly, mbk-1 loss still allowed for DAF-16 nuclear accumulation but reduced expression of certain DAF-16 target genes in germline-less, but not in daf-2 mutant animals. These findings indicate that mbk-1 and daf-16 functionally interact in the germline- but not in the daf-2 pathway. Together, our data suggest mbk-1 as a novel regulator of C. elegans longevity upon both, germline ablation and DAF-2 inhibition, and provide evidence for mbk-1 regulating DAF-16 activity in germline-deficient animals.

  19. Somatic and Germline Diversification of a Putative Immunoreceptor within One Phylum: Dscam in Arthropods.

    Science.gov (United States)

    Brites, Daniela; Du Pasquier, Louis

    2015-01-01

    Arthropod Dscam, the homologue of the human Down Syndrome cell adhesion molecule, is a receptor used by the nervous and immune systems. Unlike in vertebrates, evolutionary pressure has selected and maintained a vast Dscam diversity of isoforms, known to specifying neuronal identity during the nervous system differentiation. This chapter examines the different modes of Dscam diversification in the context of arthropods' evolution and that of their immune system, where its role is controversial. In the single Dscam gene of insects and crustaceans, mutually exclusive alternative splicing affects three clusters of duplicated exons encoding the variable parts of the receptor. The Dscam gene produces over 10,000 isoforms. In the more basal arthropods such as centipedes, Dscam diversity results from a combination of many germline genes (over 80) with, in about half of those, the possibility of alternative splicing affecting only one exon cluster. In the even more basal arthropods, such as chelicerates, no splicing possibility is detected, but there exist dozens of germline Dscam genes. Compared to controlling the expression of multiple germline genes, the somatic mutually alternative splicing within a single gene may offer a simplified way of expressing a large Dscam repertoire. Expressed by hemocytes, Dscam is considered a phagocytic receptor but is also encountered in solution. More information is necessary about its binding to pathogens, its role in phagocytosis, its possible role in specifying hemocyte identity, its kinetics of expression, and the regulation of its RNA splicing to understand how its diversity is linked to immunity.

  20. C. elegans DAF-16/FOXO interacts with TGF-ß/BMP signaling to induce germline tumor formation via mTORC1 activation.

    Directory of Open Access Journals (Sweden)

    Wenjing Qi

    2017-05-01

    Full Text Available Activation of the FOXO transcription factor DAF-16 by reduced insulin/IGF signaling (IIS is considered to be beneficial in C. elegans due to its ability to extend lifespan and to enhance stress resistance. In the germline, cell-autonomous DAF-16 activity prevents stem cell proliferation, thus acting tumor-suppressive. In contrast, hypodermal DAF-16 causes a tumorous germline phenotype characterized by hyperproliferation of the germline stem cells and rupture of the adjacent basement membrane. Here we show that cross-talk between DAF-16 and the transforming growth factor ß (TGFß/bone morphogenic protein (BMP signaling pathway causes germline hyperplasia and results in disruption of the basement membrane. In addition to activating MADM/NRBP/hpo-11 gene alone, DAF-16 also directly interacts with both R-SMAD proteins SMA-2 and SMA-3 in the nucleus to regulate the expression of mTORC1 pathway. Knocking-down of BMP genes or each of the four target genes in the hypodermis was sufficient to inhibit germline proliferation, indicating a cell-non-autonomously controlled regulation of stem cell proliferation by somatic tissues. We propose the existence of two antagonistic DAF-16/FOXO functions, a cell-proliferative somatic and an anti-proliferative germline activity. Whereas germline hyperplasia under reduced IIS is inhibited by DAF-16 cell-autonomously, activation of somatic DAF-16 in the presence of active IIS promotes germline proliferation and eventually induces tumor-like germline growth. In summary, our results suggest a novel pathway crosstalk of DAF-16 and TGF-ß/BMP that can modulate mTORC1 at the transcriptional level to cause stem-cell hyperproliferation. Such cell-type specific differences may help explaining why human FOXO activity is considered to be tumor-suppressive in most contexts, but may become oncogenic, e.g. in chronic and acute myeloid leukemia.

  1. C. elegans DAF-16/FOXO interacts with TGF-ß/BMP signaling to induce germline tumor formation via mTORC1 activation.

    Science.gov (United States)

    Qi, Wenjing; Yan, Yijian; Pfeifer, Dietmar; Donner V Gromoff, Erika; Wang, Yimin; Maier, Wolfgang; Baumeister, Ralf

    2017-05-01

    Activation of the FOXO transcription factor DAF-16 by reduced insulin/IGF signaling (IIS) is considered to be beneficial in C. elegans due to its ability to extend lifespan and to enhance stress resistance. In the germline, cell-autonomous DAF-16 activity prevents stem cell proliferation, thus acting tumor-suppressive. In contrast, hypodermal DAF-16 causes a tumorous germline phenotype characterized by hyperproliferation of the germline stem cells and rupture of the adjacent basement membrane. Here we show that cross-talk between DAF-16 and the transforming growth factor ß (TGFß)/bone morphogenic protein (BMP) signaling pathway causes germline hyperplasia and results in disruption of the basement membrane. In addition to activating MADM/NRBP/hpo-11 gene alone, DAF-16 also directly interacts with both R-SMAD proteins SMA-2 and SMA-3 in the nucleus to regulate the expression of mTORC1 pathway. Knocking-down of BMP genes or each of the four target genes in the hypodermis was sufficient to inhibit germline proliferation, indicating a cell-non-autonomously controlled regulation of stem cell proliferation by somatic tissues. We propose the existence of two antagonistic DAF-16/FOXO functions, a cell-proliferative somatic and an anti-proliferative germline activity. Whereas germline hyperplasia under reduced IIS is inhibited by DAF-16 cell-autonomously, activation of somatic DAF-16 in the presence of active IIS promotes germline proliferation and eventually induces tumor-like germline growth. In summary, our results suggest a novel pathway crosstalk of DAF-16 and TGF-ß/BMP that can modulate mTORC1 at the transcriptional level to cause stem-cell hyperproliferation. Such cell-type specific differences may help explaining why human FOXO activity is considered to be tumor-suppressive in most contexts, but may become oncogenic, e.g. in chronic and acute myeloid leukemia.

  2. Germline mutations of BRCA1 gene exon 11 are not associated with platinum response neither with survival advantage in patients with primary ovarian cancer: understanding the clinical importance of one of the biggest human exons. A study of the Tumor Bank Ovarian Cancer (TOC) Consortium.

    Science.gov (United States)

    Dimitrova, Desislava; Ruscito, Ilary; Olek, Sven; Richter, Rolf; Hellwag, Alexander; Türbachova, Ivana; Woopen, Hannah; Baron, Udo; Braicu, Elena Ioana; Sehouli, Jalid

    2016-09-01

    Germline mutations in BRCA1 gene have been reported in up to 20 % of epithelial ovarian cancer (EOC) patients. Distinct clinical characteristics have been attributed to this special EOC population. We hypothesized that mutations in different BRCA1 gene exons may differently affect the clinical course of the disease. The aim of this study was to analyze, in a large cohort of primary EOCs, the clinical impact of mutations in BRCA1 gene exon 11, the largest exon of the gene sequence encoding the 60 % of BRCA1 protein. Two hundred sixty-three primary EOC patients, treated between 2000 and 2008 at Charité University Hospital of Berlin, were included. Patients' blood samples were obtained from the Tumor Ovarian Cancer (TOC) Network ( www.toc-network.de ). Direct sequencing of BRCA1 gene exon 11 was performed for each patient to detect mutations. Based on their BRCA1 exon 11 mutational status, patients were compared regarding clinico-pathological variables and survival. Mutations in BRCA1 exon 11 were found in 18 out of 263 patients (6.8 %). Further 10/263 (3.8 %) cases showed variants of uncertain significance (VUS). All exon 11 BRCA1-positive tumors (100 %) were Type 2 ovarian carcinomas (p = 0.05). Age at diagnosis was significantly younger in Type 2 exon 11 mutated patients (p = 0.01). On multivariate analysis, BRCA1 exon 11 mutational status was not found to be an independent predictive factor for optimal cytoreduction, platinum response, or survival. Mutations in BRCA1 gene exon 11 seem to predispose women to exclusively develop a Type 2 ovarian cancer at younger age. Exon 11 BRCA1-mutated EOC patients showed distinct clinico-pathological features but similar clinical outcome with respect to sporadic EOC patients.

  3. DAF-16 and TCER-1 Facilitate Adaptation to Germline Loss by Restoring Lipid Homeostasis and Repressing Reproductive Physiology in C. elegans

    Science.gov (United States)

    Amrit, Francis Raj Gandhi; Steenkiste, Elizabeth Marie; Ratnappan, Ramesh; Chen, Shaw-Wen; McClendon, T. Brooke; Kostka, Dennis; Yanowitz, Judith; Olsen, Carissa Perez; Ghazi, Arjumand

    2016-01-01

    Elimination of the proliferating germline extends lifespan in C. elegans. This phenomenon provides a unique platform to understand how complex metazoans retain metabolic homeostasis when challenged with major physiological perturbations. Here, we demonstrate that two conserved transcription regulators essential for the longevity of germline-less adults, DAF-16/FOXO3A and TCER-1/TCERG1, concurrently enhance the expression of multiple genes involved in lipid synthesis and breakdown, and that both gene classes promote longevity. Lipidomic analyses revealed that key lipogenic processes, including de novo fatty acid synthesis, triglyceride production, desaturation and elongation, are augmented upon germline removal. Our data suggest that lipid anabolic and catabolic pathways are coordinately augmented in response to germline loss, and this metabolic shift helps preserve lipid homeostasis. DAF-16 and TCER-1 also perform essential inhibitory functions in germline-ablated animals. TCER-1 inhibits the somatic gene-expression program that facilitates reproduction and represses anti-longevity genes, whereas DAF-16 impedes ribosome biogenesis. Additionally, we discovered that TCER-1 is critical for optimal fertility in normal adults, suggesting that the protein acts as a switch supporting reproductive fitness or longevity depending on the presence or absence of the germline. Collectively, our data offer insights into how organisms adapt to changes in reproductive status, by utilizing the activating and repressive functions of transcription factors and coordinating fat production and degradation. PMID:26862916

  4. The oogenic germline starvation response in C. elegans.

    Directory of Open Access Journals (Sweden)

    Hannah S Seidel

    Full Text Available Many animals alter their reproductive strategies in response to environmental stress. Here we have investigated how L4 hermaphrodites of Caenorhabditis elegans respond to starvation. To induce starvation, we removed food at 2 h intervals from very early- to very late-stage L4 animals. The starved L4s molted into adulthood, initiated oogenesis, and began producing embryos; however, all three processes were severely delayed, and embryo viability was reduced. Most animals died via 'bagging,' because egg-laying was inhibited, and embryos hatched in utero, consuming their parent hermaphrodites from within. Some animals, however, avoided bagging and survived long term. Long-term survival did not rely on embryonic arrest but instead upon the failure of some animals to produce viable progeny during starvation. Regardless of the bagging fate, starved animals showed two major changes in germline morphology: All oogenic germlines were dramatically reduced in size, and these germlines formed only a single oocyte at a time, separated from the remainder of the germline by a tight constriction. Both changes in germline morphology were reversible: Upon re-feeding, the shrunken germlines regenerated, and multiple oocytes formed concurrently. The capacity for germline regeneration upon re-feeding was not limited to the small subset of animals that normally survive starvation: When bagging was prevented ectopically by par-2 RNAi, virtually all germlines still regenerated. In addition, germline shrinkage strongly correlated with oogenesis, suggesting that during starvation, germline shrinkage may provide material for oocyte production. Finally, germline shrinkage and regeneration did not depend upon crowding. Our study confirms previous findings that starvation uncouples germ cell proliferation from germline stem cell maintenance. Our study also suggests that when nutrients are limited, hermaphrodites scavenge material from their germlines to reproduce. We discuss

  5. CHEK2 1100DELC germline mutation: a frequency study in hereditary breast and colon cancer Brazilian families Mutação germinativa 1100delC no gene CHEK2: estudo da frequência em famílias brasileiras com câncer de mama e cólon hereditários

    Directory of Open Access Journals (Sweden)

    Jamile Abud

    2012-12-01

    Full Text Available CONTEXT: CHEK2 encodes a cell cycle checkpoint kinase that plays an important role in the DNA damage repair pathway, activated mainly by ATM (Ataxia Telangiectasia Mutated in response to double-stranded DNA breaks. A germline mutation in CHEK2, 1100delC, has been described as a low penetrance allele in a significant number of families with breast and colorectal cancer in certain countries and is also associated with increased risk of contralateral breast cancer in women previously affected by the disease. About 5%-10% of all breast and colorectal cancers are associated with hereditary predisposition and its recognition is of great importance for genetic counseling and cancer risk management. OBJECTIVES: Here, we have assessed the frequency of the CHEK2 1100delC mutation in the germline of 59 unrelated Brazilian individuals with clinical criteria for the hereditary breast and colorectal cancer syndrome. METHODS: A long-range PCR strategy followed by gene sequencing was used. RESULTS: The 1100delC mutation was encountered in the germline of one (1.7% individual in this high risk cohort. This indicates that the CHEK2 1100delC is not commonly encountered in Brazilian families with multiple diagnoses of breast and colorectal cancer. CONCLUSION: These results should be confirmed in a larger series of families and further testing should be undertaken to investigate the molecular mechanisms underlying the hereditary breast and colorectal cancer phenotype.INTRODUÇÃO: CHEK2 codifica uma proteína quinase envolvida em um ponto de checagem do ciclo celular que desempenha um papel importante na via de reparação do DNA, danos ativados principalmente por ATM (Ataxia Telangiectasia Mutado em resposta a danos na dupla hélice do DNA. A mutação germinativa 1100delC no gene CHEK2 tem sido descrita como um alelo de baixa penetrância em um número significativo de famílias com câncer de mama e cólon em certos países e também está associada com risco

  6. Human germline hedgehog pathway mutations predispose to fatty liver.

    Science.gov (United States)

    Guillen-Sacoto, Maria J; Martinez, Ariel F; Abe, Yu; Kruszka, Paul; Weiss, Karin; Everson, Joshua L; Bataller, Ramon; Kleiner, David E; Ward, Jerrold M; Sulik, Kathleen K; Lipinski, Robert J; Solomon, Benjamin D; Muenke, Maximilian

    2017-10-01

    Non-alcoholic fatty liver disease (NAFLD) is the most common form of liver disease. Activation of hedgehog (Hh) signaling has been implicated in the progression of NAFLD and proposed as a therapeutic target; however, the effects of Hh signaling inhibition have not been studied in humans with germline mutations that affect this pathway. Patients with holoprosencephaly (HPE), a disorder associated with germline mutations disrupting Sonic hedgehog (SHH) signaling, were clinically evaluated for NAFLD. A combined mouse model of Hh signaling attenuation (Gli2 heterozygous null: Gli2 +/- ) and diet-induced NAFLD was used to examine aspects of NAFLD and hepatic gene expression profiles, including molecular markers of hepatic fibrosis and inflammation. Patients with HPE had a higher prevalence of liver steatosis compared to the general population, independent of obesity. Exposure of Gli2 +/- mice to fatty liver-inducing diets resulted in increased liver steatosis compared to wild-type mice. Similar to humans, this effect was independent of obesity in the mutant mice and was associated with decreased expression of pro-fibrotic and pro-inflammatory genes, and increased expression of PPARγ, a potent anti-fibrogenic and anti-inflammatory regulator. Interestingly, tumor suppressors p53 and p16INK4 were found to be downregulated in the Gli2 +/- mice exposed to a high-fat diet. Our results indicate that germline mutations disrupting Hh signaling promotes liver steatosis, independent of obesity, with reduced fibrosis. While Hh signaling inhibition has been associated with a better NAFLD prognosis, further studies are required to evaluate the long-term effects of mutations affecting this pathway. Lay summary: Non-alcoholic fatty liver disease (NAFLD) is characterized by excess fat deposition in the liver predominantly due to high calorie intake and a sedentary lifestyle. NAFLD progression is usually accompanied by activation of the Sonic hedgehog (SHH) pathway leading to fibrous

  7. Pediatric MDS: GATA screen the germline.

    Science.gov (United States)

    Stieglitz, Elliot; Loh, Mignon L

    2016-03-17

    In this issue of Blood, Wlodarski and colleagues demonstrate that as many as 72% of adolescents diagnosed with myelodysplastic syndrome (MDS) and monosomy 7 harbor germline mutations in GATA2. Although pediatric MDS is a very rare diagnosis, occurring in 0.8 to 4 cases per million, Wlodarski et al screened >600 cases of primary or secondary MDS in children and adolescents who were enrolled in the European Working Group on MDS consortium over a period of 15 years. The overall frequency of germline GATA2 mutations in children with primary MDS was 7%, and 15% in those presenting with advanced disease. Notably, mutations in GATA2 were absent in patients with therapy-related MDS or acquired aplastic anemia.

  8. Lynch Syndrome Caused by Germline PMS2 Mutations

    DEFF Research Database (Denmark)

    Ten Broeke, Sanne W; Brohet, Richard M; Tops, Carli M

    2015-01-01

    PURPOSE: The clinical consequences of PMS2 germline mutations are poorly understood compared with other Lynch-associated mismatch repair gene (MMR) mutations. The aim of this European cohort study was to define the cancer risk faced by PMS2 mutation carriers. METHODS: Data were collected from 98...... PMS2 families ascertained from family cancer clinics that included a total of 2,548 family members and 377 proven mutation carriers. To adjust for potential ascertainment bias, a modified segregation analysis model was used to calculate colorectal cancer (CRC) and endometrial cancer (EC) risks....... Standardized incidence ratios (SIRs) were calculated to estimate risks for other Lynch syndrome-associated cancers. RESULTS: The cumulative risk (CR) of CRC for male mutation carriers by age 70 years was 19%. The CR among female carriers was 11% for CRC and 12% for EC. The mean age of CRC development was 52...

  9. Genomic hypomethylation in the human germline associates with selective structural mutability in the human genome.

    Directory of Open Access Journals (Sweden)

    Jian Li

    Full Text Available The hotspots of structural polymorphisms and structural mutability in the human genome remain to be explained mechanistically. We examine associations of structural mutability with germline DNA methylation and with non-allelic homologous recombination (NAHR mediated by low-copy repeats (LCRs. Combined evidence from four human sperm methylome maps, human genome evolution, structural polymorphisms in the human population, and previous genomic and disease studies consistently points to a strong association of germline hypomethylation and genomic instability. Specifically, methylation deserts, the ~1% fraction of the human genome with the lowest methylation in the germline, show a tenfold enrichment for structural rearrangements that occurred in the human genome since the branching of chimpanzee and are highly enriched for fast-evolving loci that regulate tissue-specific gene expression. Analysis of copy number variants (CNVs from 400 human samples identified using a custom-designed array comparative genomic hybridization (aCGH chip, combined with publicly available structural variation data, indicates that association of structural mutability with germline hypomethylation is comparable in magnitude to the association of structural mutability with LCR-mediated NAHR. Moreover, rare CNVs occurring in the genomes of individuals diagnosed with schizophrenia, bipolar disorder, and developmental delay and de novo CNVs occurring in those diagnosed with autism are significantly more concentrated within hypomethylated regions. These findings suggest a new connection between the epigenome, selective mutability, evolution, and human disease.

  10. The Fanconi anemia DNA damage repair pathway in the spotlight for germline predisposition to colorectal cancer.

    Science.gov (United States)

    Esteban-Jurado, Clara; Franch-Expósito, Sebastià; Muñoz, Jenifer; Ocaña, Teresa; Carballal, Sabela; López-Cerón, Maria; Cuatrecasas, Miriam; Vila-Casadesús, Maria; Lozano, Juan José; Serra, Enric; Beltran, Sergi; Brea-Fernández, Alejandro; Ruiz-Ponte, Clara; Castells, Antoni; Bujanda, Luis; Garre, Pilar; Caldés, Trinidad; Cubiella, Joaquín; Balaguer, Francesc; Castellví-Bel, Sergi

    2016-10-01

    Colorectal cancer (CRC) is one of the most common neoplasms in the world. Fanconi anemia (FA) is a very rare genetic disease causing bone marrow failure, congenital growth abnormalities and cancer predisposition. The comprehensive FA DNA damage repair pathway requires the collaboration of 53 proteins and it is necessary to restore genome integrity by efficiently repairing damaged DNA. A link between FA genes in breast and ovarian cancer germline predisposition has been previously suggested. We selected 74 CRC patients from 40 unrelated Spanish families with strong CRC aggregation compatible with an autosomal dominant pattern of inheritance and without mutations in known hereditary CRC genes and performed germline DNA whole-exome sequencing with the aim of finding new candidate germline predisposition variants. After sequencing and data analysis, variant prioritization selected only those very rare alterations, producing a putative loss of function and located in genes with a role compatible with cancer. We detected an enrichment for variants in FA DNA damage repair pathway genes in our familial CRC cohort as 6 families carried heterozygous, rare, potentially pathogenic variants located in BRCA2/FANCD1, BRIP1/FANCJ, FANCC, FANCE and REV3L/POLZ. In conclusion, the FA DNA damage repair pathway may play an important role in the inherited predisposition to CRC.

  11. Germline transformation of the Mediterranean fruit fly, Ceratitis capitata

    International Nuclear Information System (INIS)

    McCombs, Susan D.

    2000-01-01

    Gene transfer methodology for insects was first developed in Drosophila melanogaster Meigen using a transposon-mediated system based on the P element (Spradling and Rubin 1982, Rubin and Spradling 1982). In addition to the P element, three unrelated transposons have been used successfully in genetic transformation of D. melanogaster: hobo (Blackman et al. 1989), Minos (Loukeris et al. 1992), and mariner (Lidholm et al. 1993). Routine gene transfer in Drosophila created a great deal of optimism amongst researchers who sought to employ transgenic techniques in other arthropods. However, what followed were years of consistently disappointing results in other insect species. For example, the P element system was tried unsuccessfully in several species, but was eventually shown to be non-functional outside the genus Drosophila (O'Brochta and Handler 1988). Ensuing research in non-drosophilids emphasised testing of other Drosophila systems and development of transposons isolated from other species. After nearly 15 years of intensive effort, the first successes have only recently been reported. Three Drosophila-derived transposon-based systems: hobo from D. melanogaster, mariner from Drosophila mauritiana Tsacas and David and Minos from Drosophila hydei Sturtevant have produced germline transformation in Drosophila virilis Sturtevant (Gomez and Handler 1997, Lozovskaya et al. 1996), Aedes aegypti L. (Coates et al. 1998), and Ceratitis capitata (Wied.) (Loukeris et al. 1995), respectively. Germline transformation was accomplished with two transposon-based systems from non-drosophilids, Hermes from Musca domestica L. and piggyBac from Trichoplusia ni Huebner in A. aegypti and C. capitata, respectively

  12. Repression of germline RNAi pathways in somatic cells by retinoblastoma pathway chromatin complexes.

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    Xiaoyun Wu

    Full Text Available The retinoblastoma (Rb tumor suppressor acts with a number of chromatin cofactors in a wide range of species to suppress cell proliferation. The Caenorhabditis elegans retinoblastoma gene and many of these cofactors, called synMuv B genes, were identified in genetic screens for cell lineage defects caused by growth factor misexpression. Mutations in many synMuv B genes, including lin-35/Rb, also cause somatic misexpression of the germline RNA processing P granules and enhanced RNAi. We show here that multiple small RNA components, including a set of germline-specific Argonaute genes, are misexpressed in the soma of many synMuv B mutant animals, revealing one node for enhanced RNAi. Distinct classes of synMuv B mutants differ in the subcellular architecture of their misexpressed P granules, their profile of misexpressed small RNA and P granule genes, as well as their enhancement of RNAi and the related silencing of transgenes. These differences define three classes of synMuv B genes, representing three chromatin complexes: a LIN-35/Rb-containing DRM core complex, a SUMO-recruited Mec complex, and a synMuv B heterochromatin complex, suggesting that intersecting chromatin pathways regulate the repression of small RNA and P granule genes in the soma and the potency of RNAi. Consistent with this, the DRM complex and the synMuv B heterochromatin complex were genetically additive and displayed distinct antagonistic interactions with the MES-4 histone methyltransferase and the MRG-1 chromodomain protein, two germline chromatin regulators required for the synMuv phenotype and the somatic misexpression of P granule components. Thus intersecting synMuv B chromatin pathways conspire with synMuv B suppressor chromatin factors to regulate the expression of small RNA pathway genes, which enables heightened RNAi response. Regulation of small RNA pathway genes by human retinoblastoma may also underlie its role as a tumor suppressor gene.

  13. A regulatory network of Drosophila germline stem cell self-renewal

    OpenAIRE

    Yan, Dong; Neumüller, Ralph A.; Buckner, Michael; Ayers, Kathleen; Li, Hua; Hu, Yanhui; Yang-Zhou, Donghui; Pan, Lei; Wang, Xiaoxi; Kelley, Colleen; Vinayagam, Arunachalam; Binari, Richard; Randklev, Sakara; Perkins, Lizabeth A.; Xie, Ting

    2014-01-01

    Stem cells possess the capacity to generate two cells of distinct fate upon division; one cell retaining stem cell identity and the other cell destined to differentiate. These cell fates are established by cell-type-specific genetic networks. To comprehensively identify components of these networks, we performed a large-scale RNAi screen in Drosophila female germline stem cells (GSCs) covering ~25% of the genome. The screen identified 366 genes that affect GSC maintenance, differentiation or ...

  14. Effect of BRCA germline mutations on breast cancer prognosis

    Science.gov (United States)

    Baretta, Zora; Mocellin, Simone; Goldin, Elena; Olopade, Olufunmilayo I.; Huo, Dezheng

    2016-01-01

    Abstract Background: The contribution of BRCA germline mutational status to breast cancer patients’ prognosis is unclear. We aimed to systematically review and perform meta-analysis of the available evidence of effects of BRCA germline mutations on multiple survival outcomes of breast cancer patients as a whole and in specific subgroups of interest, including those with triple negative breast cancer, those with Ashkenazi Jewish ancestry, and patients with stage I–III disease. Methods: Sixty studies met all inclusion criteria and were considered for this meta-analysis. These studies involved 105,220 breast cancer patients, whose 3588 (3.4%) were BRCA mutations carriers. The associations between BRCA genes mutational status and overall survival (OS), breast cancer-specific survival (BCSS), recurrence-free survival (RFS), and distant metastasis-free survival (DMFS) were evaluated using random-effect models. Results: BRCA1 mutation carriers have worse OS than BRCA-negative/sporadic cases (hazard ratio, HR 1.30, 95% CI: 1.11–1.52) and worse BCSS than sporadic/BRCA-negative cases among patients with stage I–III breast cancer (HR 1.45, 95% CI: 1.01–2.07). BRCA2 mutation carriers have worse BCSS than sporadic/BRCA-negative cases (HR 1.29, 95% CI: 1.03–1.62), although they have similar OS. Among triple negative breast cancer, BRCA1/2 mutations carriers had better OS than BRCA-negative counterpart (HR 0.49, 95% CI: 0.26–0.92). Among Ashkenazi Jewish women, BRCA1/2 mutations carriers presented higher risk of death from breast cancer (HR 1.44, 95% CI: 1.05–1.97) and of distant metastases (HR 1.82, 95% CI: 1.05–3.16) than sporadic/BRCA-negative patients. Conclusion: Our results support the evaluation of BRCA mutational status in patients with high risk of harboring BRCA germline mutations to better define the prognosis of breast cancer in these patients. PMID:27749552

  15. The molecular anatomy of spontaneous germline mutations in human testes.

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    Jian Qin

    2007-09-01

    Full Text Available The frequency of the most common sporadic Apert syndrome mutation (C755G in the human fibroblast growth factor receptor 2 gene (FGFR2 is 100-1,000 times higher than expected from average nucleotide substitution rates based on evolutionary studies and the incidence of human genetic diseases. To determine if this increased frequency was due to the nucleotide site having the properties of a mutation hot spot, or some other explanation, we developed a new experimental approach. We examined the spatial distribution of the frequency of the C755G mutation in the germline by dividing four testes from two normal individuals each into several hundred pieces, and, using a highly sensitive PCR assay, we measured the mutation frequency of each piece. We discovered that each testis was characterized by rare foci with mutation frequencies 10(3 to >10(4 times higher than the rest of the testis regions. Using a model based on what is known about human germline development forced us to reject (p < 10(-6 the idea that the C755G mutation arises more frequently because this nucleotide simply has a higher than average mutation rate (hot spot model. This is true regardless of whether mutation is dependent or independent of cell division. An alternate model was examined where positive selection acts on adult self-renewing Ap spermatogonial cells (SrAp carrying this mutation such that, instead of only replacing themselves, they occasionally produce two SrAp cells. This model could not be rejected given our observed data. Unlike the disease site, similar analysis of C-to-G mutations at a control nucleotide site in one testis pair failed to find any foci with high mutation frequencies. The rejection of the hot spot model and lack of rejection of a selection model for the C755G mutation, along with other data, provides strong support for the proposal that positive selection in the testis can act to increase the frequency of premeiotic germ cells carrying a mutation

  16. Comprehensive analysis of BRCA1, BRCA2 and TP53 germline mutation and tumor characterization: a portrait of early-onset breast cancer in Brazil.

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    Dirce Maria Carraro

    Full Text Available Germline mutations in BRCA1, BRCA2 and TP53 genes have been identified as one of the most important disease-causing issues in young breast cancer patients worldwide. The specific defective biological processes that trigger germline mutation-associated and -negative tumors remain unclear. To delineate an initial portrait of Brazilian early-onset breast cancer, we performed an investigation combining both germline and tumor analysis. Germline screening of the BRCA1, BRCA2, CHEK2 (c.1100delC and TP53 genes was performed in 54 unrelated patients <35 y; their tumors were investigated with respect to transcriptional and genomic profiles as well as hormonal receptors and HER2 expression/amplification. Germline mutations were detected in 12 out of 54 patients (22% [7 in BRCA1 (13%, 4 in BRCA2 (7% and one in TP53 (2% gene]. A cancer familial history was present in 31.4% of the unrelated patients, from them 43.7% were carriers for germline mutation (37.5% in BRCA1 and in 6.2% in the BRCA2 genes. Fifty percent of the unrelated patients with hormone receptor-negative tumors carried BRCA1 mutations, percentage increasing to 83% in cases with familial history of cancer. Over-representation of DNA damage-, cellular and cell cycle-related processes was detected in the up-regulated genes of BRCA1/2-associated tumors, whereas cell and embryo development-related processes were over-represented in the up-regulated genes of BRCA1/2-negative tumors, suggesting distinct mechanisms driving the tumorigenesis. An initial portrait of the early-onset breast cancer patients in Brazil was generated pointing out that hormone receptor-negative tumors and positive familial history are two major risk factors for detection of a BRCA1 germline mutation. Additionally, the data revealed molecular factors that potentially trigger the tumor development in young patients.

  17. Long-term culture of chicken primordial germ cells isolated from embryonic blood and production of germline chimaeric chickens.

    Science.gov (United States)

    Naito, Mitsuru; Harumi, Takashi; Kuwana, Takashi

    2015-02-01

    Production of germline chimaeric chickens by the transfer of cultured primordial germ cells (PGC) is a useful system for germline manipulation. A novel culture system was developed for chicken PGC isolated from embryonic blood. The isolated PGC were cultured on feeder cells derived from chicken embryonic fibroblast. The cultured PGC formed colonies and they proliferated about 300-times during the first 30 days. The cultured PGC retained the ability to migrate to recipient gonads and were also chicken VASA homologue (CVH)-positive. Female PGC were present in the mixed-sex PGC populations cultured for more than 90 days and gave rise to viable offspring efficiently via germline chimaeric chickens. Male cultured PGC were transferred to recipient embryos and produced putative chimaeric chickens. The DNA derived from the cultured PGC was detected in the sperm samples of male putative chimaeric chickens, but no donor derived offspring were obtained. Donor-derived offspring were also obtained from germline chimaeric chickens by the transfer of frozen-thawed cultured PGC. The culture method for PGC developed in the present study is useful for manipulation of the germline in chickens, such as preservation of genetic resources and gene transfer. Copyright © 2014 Elsevier B.V. All rights reserved.

  18. Asymmetric distribution of pl10 and bruno2, new members of a conserved core of early germline determinants in cephalochordates

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    Simon eDailey

    2016-01-01

    Full Text Available Molecular fingerprinting of conserved germline and somatic ¨stemness¨ markers in different taxa have been key in defining the mechanism of germline specification (preformation or epigenesis, as well as expression domains of somatic progenitors. The distribution of molecular markers for primordial germ cells (PGCs, including vasa, nanos and piwil1, as well as Vasa antibody staining, support a determinative mechanism of germline specification in the cephalochordate Branchiostoma lanceolatum, similarly to other amphioxus species. pl10 and bruno2, but not bruno4/6, are also expressed in a pattern consistent with these other germline genes, adding to our repertoire of PGC markers in lancelets. Expression of nanos, vasa and the remaining markers (musashi, pufA, pufB, pumilio and piwil2 may define populations of putative somatic progenitors in the tailbud, the amphioxus posterior growth zone, or zones of proliferative activity. Finally, we also identify a novel expression domain for musashi, a classic neural stem cell marker, during notochord development in amphioxus. These results are discussed in the context of germline determination in other taxa, stem cell regulation and regenerative capacity in adult amphioxus.

  19. Germline truncating-mutations in BRCA1 and MSH6 in a patient with early onset endometrial cancer

    International Nuclear Information System (INIS)

    Kast, Karin; Schackert, Hans K; Neuhann, Teresa M; Görgens, Heike; Becker, Kerstin; Keller, Katja; Klink, Barbara; Aust, Daniela; Distler, Wolfgang; Schröck, Evelin

    2012-01-01

    Hereditary Breast and Ovarian Cancer Syndrome (HBOCS) and Hereditary Non-Polyposis Colorectal Cancer Syndrome (HNPCC, Lynch Syndrome) are two tumor predisposition syndromes responsible for the majority of hereditary breast and colorectal cancers. Carriers of both germline mutations in breast cancer genes BRCA1 or BRCA2 and in mismatch repair (MMR) genes MLH1, MSH2, MSH6 or PMS2 are very rare. We identified germline mutations in BRCA1 and in MSH6 in a patient with increased risk for HBOC diagnosed with endometrial cancer at the age of 46 years. Although carriers of mutations in both MMR and BRCA genes are rare in Caucasian populations and anamnestical and histopathological findings may guide clinicians to identify these families, both syndromes can only be diagnosed through a complete gene analysis of the respective genes

  20. Germline truncating-mutations in BRCA1 and MSH6 in a patient with early onset endometrial cancer

    Energy Technology Data Exchange (ETDEWEB)

    Kast, Karin [Department of Gynecology and Obstetrics, University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden (Germany); Schackert, Hans K [Department of Surgical Research, University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden (Germany); Neuhann, Teresa M [Institute for Clinical Genetics, Technische Universität Dresden, Dresden (Germany); Medical Genetic Center, Munich (Germany); Görgens, Heike [Department of Surgical Research, University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden (Germany); Becker, Kerstin [Institute for Clinical Genetics, Technische Universität Dresden, Dresden (Germany); Keller, Katja [Department of Gynecology and Obstetrics, University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden (Germany); Klink, Barbara [Institute for Clinical Genetics, Technische Universität Dresden, Dresden (Germany); Aust, Daniela [Institute of Pathology, University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden (Germany); Distler, Wolfgang [Department of Gynecology and Obstetrics, University Hospital Carl Gustav Carus, Technische Universität Dresden, Dresden (Germany); Schröck, Evelin [Institute for Clinical Genetics, Technische Universität Dresden, Dresden (Germany)

    2012-11-20

    Hereditary Breast and Ovarian Cancer Syndrome (HBOCS) and Hereditary Non-Polyposis Colorectal Cancer Syndrome (HNPCC, Lynch Syndrome) are two tumor predisposition syndromes responsible for the majority of hereditary breast and colorectal cancers. Carriers of both germline mutations in breast cancer genes BRCA1 or BRCA2 and in mismatch repair (MMR) genes MLH1, MSH2, MSH6 or PMS2 are very rare. We identified germline mutations in BRCA1 and in MSH6 in a patient with increased risk for HBOC diagnosed with endometrial cancer at the age of 46 years. Although carriers of mutations in both MMR and BRCA genes are rare in Caucasian populations and anamnestical and histopathological findings may guide clinicians to identify these families, both syndromes can only be diagnosed through a complete gene analysis of the respective genes.

  1. Germline truncating-mutations in BRCA1 and MSH6 in a patient with early onset endometrial cancer

    Directory of Open Access Journals (Sweden)

    Kast Karin

    2012-11-01

    Full Text Available Abstract Background Hereditary Breast and Ovarian Cancer Syndrome (HBOCS and Hereditary Non-Polyposis Colorectal Cancer Syndrome (HNPCC, Lynch Syndrome are two tumor predisposition syndromes responsible for the majority of hereditary breast and colorectal cancers. Carriers of both germline mutations in breast cancer genes BRCA1 or BRCA2 and in mismatch repair (MMR genes MLH1, MSH2, MSH6 or PMS2 are very rare. Case presentation We identified germline mutations in BRCA1 and in MSH6 in a patient with increased risk for HBOC diagnosed with endometrial cancer at the age of 46 years. Conclusions Although carriers of mutations in both MMR and BRCA genes are rare in Caucasian populations and anamnestical and histopathological findings may guide clinicians to identify these families, both syndromes can only be diagnosed through a complete gene analysis of the respective genes.

  2. Germline truncating-mutations in BRCA1 and MSH6 in a patient with early onset endometrial cancer.

    Science.gov (United States)

    Kast, Karin; Neuhann, Teresa M; Görgens, Heike; Becker, Kerstin; Keller, Katja; Klink, Barbara; Aust, Daniela; Distler, Wolfgang; Schröck, Evelin; Schackert, Hans K

    2012-11-20

    Hereditary Breast and Ovarian Cancer Syndrome (HBOCS) and Hereditary Non-Polyposis Colorectal Cancer Syndrome (HNPCC, Lynch Syndrome) are two tumor predisposition syndromes responsible for the majority of hereditary breast and colorectal cancers. Carriers of both germline mutations in breast cancer genes BRCA1 or BRCA2 and in mismatch repair (MMR) genes MLH1, MSH2, MSH6 or PMS2 are very rare. We identified germline mutations in BRCA1 and in MSH6 in a patient with increased risk for HBOC diagnosed with endometrial cancer at the age of 46 years. Although carriers of mutations in both MMR and BRCA genes are rare in Caucasian populations and anamnestical and histopathological findings may guide clinicians to identify these families, both syndromes can only be diagnosed through a complete gene analysis of the respective genes.

  3. Germline PMS2 mutation screened by mismatch repair protein immunohistochemistry of colorectal cancer in Japan.

    Science.gov (United States)

    Sugano, Kokichi; Nakajima, Takeshi; Sekine, Shigeki; Taniguchi, Hirokazu; Saito, Shinya; Takahashi, Masahiro; Ushiama, Mineko; Sakamoto, Hiromi; Yoshida, Teruhiko

    2016-11-01

    Germline PMS2 gene mutations were detected by RT-PCR/direct sequencing of total RNA extracted from puromycin-treated peripheral blood lymphocytes (PBL) and multiplex ligation-dependent probe amplification (MLPA) analyses of Japanese patients with colorectal cancer (CRC) fulfilling either the revised Bethesda Guidelines or being an age at disease onset of younger than 70 years, and screened by mismatch repair protein immunohistochemistry of formalin-fixed paraffin embedded sections. Of the 501 subjects examined, 7 (1.40%) showed the downregulated expression of the PMS2 protein alone and were referred to the genetic counseling clinic. Germline PMS2 mutations were detected in 6 (85.7%), including 3 nonsense and 1 frameshift mutations by RT-PCR/direct sequencing and 2 genomic deletions by MLPA. No mutations were identified in the other MMR genes (i.e. MSH2, MLH1 and MSH6). The prevalence of the downregulated expression of the PMS2 protein alone was 1.40% among the subjects examined and IHC results predicted the presence of PMS2 germline mutations. RT-PCR from puromycin-treated PBL and MLPA may be employed as the first screening step to detect PMS2 mutations without pseudogene interference, followed by the long-range PCR/nested PCR validation using genomic DNA. © 2016 The Authors. Cancer Science published by John Wiley & Sons Australia, Ltd on behalf of Japanese Cancer Association.

  4. Proven germline mosaicism in a father of two children with CHARGE syndrome.

    Science.gov (United States)

    Pauli, S; Pieper, L; Häberle, J; Grzmil, P; Burfeind, P; Steckel, M; Lenz, U; Michelmann, H W

    2009-05-01

    CHARGE syndrome is an autosomal dominant malformation syndrome caused by mutations in the CHD7 gene. The majority of cases are sporadic and only few familial cases have been reported. In these families, mosaicism in one parent, as well as parent- to-child transmission of a CHD7 mutation, has been described. In some further cases, germline mosaicism has been suggested. Here, we report the first case in which germline mosaicism could be demonstrated in a father of two affected children with CHARGE syndrome. The truncating mutation c.7302dupA in exon 34 of the CHD7 gene was found in both affected children but was not detected in parental lymphocytes. However, in DNA extracted from the father's spermatozoa, the c.7302dupA mutation could be identified. Furthermore, mutation analysis of DNA isolated from 59 single spermatozoa revealed that the c.7302dupA mutation occurs in 16 spermatozoa, confirming germline mosaicism in the father of the affected children. This result has a high impact for genetic counselling of the family and for their recurrence risk in further pregnancies.

  5. Exposure to the BPA-Substitute Bisphenol S Causes Unique Alterations of Germline Function.

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    Yichang Chen

    2016-07-01

    Full Text Available Concerns about the safety of Bisphenol A, a chemical found in plastics, receipts, food packaging and more, have led to its replacement with substitutes now found in a multitude of consumer products. However, several popular BPA-free alternatives, such as Bisphenol S, share a high degree of structural similarity with BPA, suggesting that these substitutes may disrupt similar developmental and reproductive pathways. We compared the effects of BPA and BPS on germline and reproductive functions using the genetic model system Caenorhabditis elegans. We found that, similarly to BPA, BPS caused severe reproductive defects including germline apoptosis and embryonic lethality. However, meiotic recombination, targeted gene expression, whole transcriptome and ontology analyses as well as ToxCast data mining all indicate that these effects are partly achieved via mechanisms distinct from BPAs. These findings therefore raise new concerns about the safety of BPA alternatives and the risk associated with human exposure to mixtures.

  6. Prospective Evaluation of Germline Alterations in Patients With Exocrine Pancreatic Neoplasms.

    Science.gov (United States)

    Lowery, Maeve A; Wong, Winston; Jordan, Emmet J; Lee, Jonathan W; Kemel, Yelena; Vijai, Joseph; Mandelker, Diana; Zehir, Ahmet; Capanu, Marinela; Salo-Mullen, Erin; Arnold, Angela G; Yu, Kenneth H; Varghese, Anna M; Kelsen, David P; Brenner, Robin; Kaufmann, Erica; Ravichandran, Vignesh; Mukherjee, Semanti; Berger, Michael F; Hyman, David M; Klimstra, David S; Abou-Alfa, Ghassan K; Tjan, Catherine; Covington, Christina; Maynard, Hannah; Allen, Peter J; Askan, Gokce; Leach, Steven D; Iacobuzio-Donahue, Christine A; Robson, Mark E; Offit, Kenneth; Stadler, Zsofia K; O'Reilly, Eileen M

    2018-02-28

    Identification of pathogenic germline alterations (PGAs) has important clinical and therapeutic implications in pancreas cancer. We performed comprehensive germline testing (GT) in an unselected prospective cohort of patients with exocrine pancreatic neoplasms with genotype and phenotype association to facilitate identification of prognostic and/or predictive biomarkers and examine potential therapeutic implications. Six hundred fifteen unselected patients with exocrine pancreatic neoplasms were prospectively consented for somatic tumor and matched sample profiling for 410-468 genes. GT for PGAs in 76 genes associated with cancer susceptibility was performed in an "identified" manner in 356 (57.9%) patients and in an "anonymized" manner in 259 (42.1%) patients, using an institutional review board-approved protocol. Detailed clinical and pathological features, response to platinum, and overall survival (OS) were collected for the identified cohort. OS was analyzed with Kaplan-Meier curves. PGAs were present in 122 (19.8%) of 615 patients involving 24 different genes, including BRCA1/2, ATM, PALB2, and multiple additional genes associated with the DNA damage response pathway. Of 122 patients with germline alterations, 41.8% did not meet current guidelines for GT. The difference in median OS was not statistically significant between patients with and without PGA (50.8 months, 95% confidence interval = 34.5 to not reached, two-sided P = .94). Loss of heterozygosity was found in 60.0% of BRCA1/2. PGAs frequently occur in pancreas exocrine neoplasms and involve multiple genes beyond those previously associated with hereditary pancreatic cancer. These PGAs are therapeutically actionable in about 5% to 10% of patients. These data support routinely offering GT in all pancreatic ductal adenocarcimona patients with a broad panel of known hereditary cancer predisposition genes.

  7. Transgenic mice produced by retroviral transduction of male germ-line stem cells

    OpenAIRE

    Nagano, Makoto; Brinster, Clayton J.; Orwig, Kyle E.; Ryu, Buom-Yong; Avarbock, Mary R.; Brinster, Ralph L.

    2001-01-01

    Male germ-line stem cells are the only cell type in postnatal mammals that have the capability to self-renew and to contribute genes to the next generation. Genetic modification of these cells would provide an opportunity to study the biology of their complex self-renewal and differentiation processes, as well as enable the generation of transgenic animals in a wide range of species. Although retroviral vectors have been used as an efficient method to introduce genes into a variety of cell ty...

  8. Prevalence of pathogenic germline variants detected by multigene sequencing in unselected Japanese patients with ovarian cancer.

    Science.gov (United States)

    Hirasawa, Akira; Imoto, Issei; Naruto, Takuya; Akahane, Tomoko; Yamagami, Wataru; Nomura, Hiroyuki; Masuda, Kiyoshi; Susumu, Nobuyuki; Tsuda, Hitoshi; Aoki, Daisuke

    2017-12-22

    Pathogenic germline BRCA1 , BRCA2 ( BRCA1/2 ), and several other gene variants predispose women to primary ovarian, fallopian tube, and peritoneal carcinoma (OC), although variant frequency and relevance information is scarce in Japanese women with OC. Using targeted panel sequencing, we screened 230 unselected Japanese women with OC from our hospital-based cohort for pathogenic germline variants in 75 or 79 OC-associated genes. Pathogenic variants of 11 genes were identified in 41 (17.8%) women: 19 (8.3%; BRCA1 ), 8 (3.5%; BRCA2 ), 6 (2.6%; mismatch repair genes), 3 (1.3%; RAD51D ), 2 (0.9%; ATM ), 1 (0.4%; MRE11A ), 1 ( FANCC ), and 1 ( GABRA6 ). Carriers of BRCA1/2 or any other tested gene pathogenic variants were more likely to be diagnosed younger, have first or second-degree relatives with OC, and have OC classified as high-grade serous carcinoma (HGSC). After adjustment for these variables, all 3 features were independent predictive factors for pathogenic variants in any tested genes whereas only the latter two remained for variants in BRCA1/2 . Our data indicate similar variant prevalence in Japanese patients with OC and other ethnic groups and suggest that HGSC and OC family history may facilitate genetic predisposition prediction in Japanese patients with OC and referring high-risk patients for genetic counseling and testing.

  9. Association of the germline TP53 R72P and MDM2 SNP309 variants with breast cancer survival in specific breast tumor subgroups

    NARCIS (Netherlands)

    van den Broek, Alexandra J.; Broeks, Annegien; Horlings, Hugo M.; Canisius, Sander V. M.; Braaf, Linde M.; Langerød, Anita; van't Veer, Laura J.; Schmidt, Marjanka K.

    2011-01-01

    The tumor suppressor gene TP53 and its regulator MDM2 are both important players in the DNA-damage repair "TP53 response pathway". Common germline polymorphisms in these genes may affect outcome in patients with tumors characterized by additional somatic changes in the same or a related pathway. To

  10. Stem cells are units of natural selection for tissue formation, for germline development, and in cancer development.

    Science.gov (United States)

    Weissman, Irving L

    2015-07-21

    It is obvious that natural selection operates at the level of individuals and collections of individuals. Nearly two decades ago we showed that in multi-individual colonies of protochordate colonial tunicates sharing a blood circulation, there exists an exchange of somatic stem cells and germline stem cells, resulting in somatic chimeras and stem cell competitions for gonadal niches. Stem cells are unlike other cells in the body in that they alone self-renew, so that they form clones that are perpetuated for the life of the organism. Stem cell competitions have allowed the emergence of competitive somatic and germline stem cell clones. Highly successful germline stem cells usually outcompete less successful competitors both in the gonads of the genotype partner from which they arise and in the gonads of the natural parabiotic partners. Therefore, natural selection also operates at the level of germline stem cell clones. In the colonial tunicate Botryllus schlosseri the formation of natural parabionts is prevented by a single-locus highly polymorphic histocompatibility gene called Botryllus histocompatibility factor. This limits germline stem cell predation to kin, as the locus has hundreds of alleles. We show that in mice germline stem cells compete for gonad niches, and in mice and humans, blood-forming stem cells also compete for bone marrow niches. We show that the clonal progression from blood-forming stem cells to acute leukemias by successive genetic and epigenetic events in blood stem cells also involves competition and selection between clones and propose that this is a general theme in cancer.

  11. Lack of GNAQ and GNA11 germ-line mutations in familial melanoma pedigrees with uveal melanoma or blue nevi

    Directory of Open Access Journals (Sweden)

    Jason Ezra Hawkes

    2013-06-01

    Full Text Available Approximately 10% of melanoma cases are familial, but only 25-40% of familial melanoma cases can be attributed to germ-line mutations in the CDKN2A - the most significant high-risk melanoma susceptibility locus identified to date. The pathogenic mutation(s in most of the remaining familial melanoma pedigrees have not yet been identified. The most common mutations in nevi and sporadic melanoma are found in BRAF and NRAS, both of which result in constitutive activation of the MAPK pathway. However, these mutations are not found in uveal melanomas or the intradermal melanocytic proliferations known as blue nevi. Rather, multiple studies report a strong association between these lesions and somatic mutations in Guanine nucleotide-binding protein G(q subunit alpha (GNAQ, Guanine nucleotide-binding protein G(q subunit alpha-11 (GNA11 and BRCA1 associated protein-1 (BAP1. Recently, germ-line mutations in BAP1, the gene encoding a tumor suppressing deubiquitinating enzyme, have been associated with predisposition to a variety of cancers including uveal melanoma, but no studies have examined the association of germ-line mutations in GNAQ and GNA11 with uveal melanoma and blue nevi. We have now done so by sequencing exon 5 of both of these genes in 13 unique familial melanoma pedigrees, members of which have had either uveal or cutaneous melanoma and/or blue nevi. Germ-line DNA from a total of 22 individuals was used for sequencing; however no deleterious mutations were detected. Nevertheless, such candidate gene studies and the discovery of novel germ-line mutations associated with an increased MM susceptibility can lead to a better understanding of the pathways involved in melanocyte transformation, formulation of risk assessment, and the development of specific drug therapies.

  12. Solid renal tumor severity in von Hippel Lindau disease is related to germline deletion length and location.

    Science.gov (United States)

    Maranchie, Jodi K; Afonso, Anoushka; Albert, Paul S; Kalyandrug, Sivaram; Phillips, John L; Zhou, Shubo; Peterson, James; Ghadimi, Bijan M; Hurley, Katheen; Riss, Joseph; Vasselli, James R; Ried, Thomas; Zbar, Berton; Choyke, Peter; Walther, McClellan M; Klausner, Richard D; Linehan, W Marston

    2004-01-01

    von Hippel Lindau disease (VHL) is an autosomal dominant familial cancer syndrome linked to alteration of the VHL tumor suppressor gene. Affected patients are predisposed to develop pheochromocytomas and cystic and solid tumors of the kidney, CNS, pancreas, retina, and epididymis. However, organ involvement varies considerably among families and has been shown to correlate with the underlying germline alteration. Clinically, we observed a paradoxically lower prevalence of renal cell carcinoma (RCC) in patients with complete germline deletion of VHL. To determine if a relationship existed between the type of VHL deletion and disease, we retrospectively evaluated 123 patients from 55 families with large germline VHL deletions, including 42 intragenic partial deletions and 13 complete VHL deletions, by history and radiographic imaging. Each individual and family was scored for cystic or solid involvement of CNS, pancreas, and kidney, and for pheochromocytoma. Germline deletions were mapped using a combination of fluorescent in situ hybridization (FISH) and quantitative Southern and Southern blot analysis. An age-adjusted comparison demonstrated a higher prevalence of RCC in patients with partial germline VHL deletions relative to complete deletions (48.9 vs. 22.6%, p=0.007). This striking phenotypic dichotomy was not seen for cystic renal lesions or for CNS (p=0.22), pancreas (p=0.72), or pheochromocytoma (p=0.34). Deletion mapping revealed that development of RCC had an even greater correlation with retention of HSPC300 (C3orf10), located within the 30-kb region of chromosome 3p, immediately telomeric to VHL (52.3 vs. 18.9%, p <0.001), suggesting the presence of a neighboring gene or genes critical to the development and maintenance of RCC. Careful correlation of genotypic data with objective phenotypic measures will provide further insight into the mechanisms of tumor formation. Copyright 2003 Wiley-Liss, Inc.

  13. Human Germline: A New Research Frontier

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    M. Azim Surani

    2015-06-01

    Full Text Available We recently elucidated the mechanism of human primordial germ cell (hPGC specification and resetting of the epigenome for totipotency. The regulators of hPGC specification also initiate resetting of the epigenome, leading to a comprehensive erasure of DNA methylation, erasure of imprints and X reactivation in early hPGCs in vivo. These studies reveal differences with the mouse model, which are probably due to differences in the regulation of human pluripotency, and in postimplantation development at gastrulation, which indicates the importance of non-rodent models for investigations. Within the extreme hypomethylated environment of the early human germline are loci that are resistant to DNA demethylation, with subsequent predominant expression in neural cells. These loci provide a model for studies on the mechanism of transgenerational epigenetic inheritance, and their response to environmental factors. Such epigenetic mechanism of inheritance could potentially provide greater phenotypic plasticity, with significant consequences for human development and disease.

  14. Methods in Molecular Biology: Germline Stem Cells | Center for Cancer Research

    Science.gov (United States)

    The protocols in Germline Stem Cells are intended to present selected genetic, molecular, and cellular techniques used in germline stem cell research. The book is divided into two parts. Part I covers germline stem cell identification and regulation in model organisms. Part II covers current techniques used in in vitro culture and applications of germline stem cells.

  15. Neurotransmitter Transporter-Like: a male germline-specific SLC6 transporter required for Drosophila spermiogenesis.

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    Nabanita Chatterjee

    2011-01-01

    Full Text Available The SLC6 class of membrane transporters, known primarily as neurotransmitter transporters, is increasingly appreciated for its roles in nutritional uptake of amino acids and other developmentally specific functions. A Drosophila SLC6 gene, Neurotransmitter transporter-like (Ntl, is expressed only in the male germline. Mobilization of a transposon inserted near the 3' end of the Ntl coding region yields male-sterile mutants defining a single complementation group. Germline transformation with Ntl cDNAs under control of male germline-specific control elements restores Ntl/Ntl homozygotes to normal fertility, indicating that Ntl is required only in the germ cells. In mutant males, sperm morphogenesis appears normal, with elongated, individualized and coiled spermiogenic cysts accumulating at the base of the testes. However, no sperm are transferred to the seminal vesicle. The level of polyglycylation of Ntl mutant sperm tubulin appears to be significantly lower than that of wild type controls. Glycine transporters are the most closely related SLC6 transporters to Ntl, suggesting that Ntl functions as a glycine transporter in developing sperm, where augmentation of the cytosolic pool of glycine may be required for the polyglycylation of the massive amounts of tubulin in the fly's giant sperm. The male-sterile phenotype of Ntl mutants may provide a powerful genetic system for studying the function of an SLC6 transporter family in a model organism.

  16. A Mononucleotide Markers Panel to Identify hMLH1/hMSH2 Germline Mutations

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    M. Pedroni

    2007-01-01

    Full Text Available Hereditary NonPolyposis Colorectal Cancer (Lynch syndrome is an autosomal dominant disease caused by germline mutations in a class of genes deputed to maintain genomic integrity during cell replication, mutations result in a generalized genomic instability, particularly evident at microsatellite loci (Microsatellite Instability, MSI. MSI is present in 85–90% of colorectal cancers that occur in Lynch Syndrome. To standardize the molecular diagnosis of MSI, a panel of 5 microsatellite markers was proposed (known as the “Bethesda panel”. Aim of our study is to evaluate if MSI testing with two mononucleotide markers, such as BAT25 and BAT26, was sufficient to identify patients with hMLH1/hMSH2 germline mutations. We tested 105 tumours for MSI using both the Bethesda markers and the two mononucleotide markers BAT25 and BAT26. Moreover, immunohistochemical evaluation of MLH1 and MSH2 proteins was executed on the tumours with at least one unstable microsatellite, whereas germline hMLH1/hMSH2 mutations were searched for all cases showing two or more unstable microsatellites.

  17. The Landscape of Somatic Genetic Alterations in Breast Cancers From ATM Germline Mutation Carriers.

    Science.gov (United States)

    Weigelt, Britta; Bi, Rui; Kumar, Rahul; Blecua, Pedro; Mandelker, Diana L; Geyer, Felipe C; Pareja, Fresia; James, Paul A; Couch, Fergus J; Eccles, Diana M; Blows, Fiona; Pharoah, Paul; Li, Anqi; Selenica, Pier; Lim, Raymond S; Jayakumaran, Gowtham; Waddell, Nic; Shen, Ronglai; Norton, Larry; Wen, Hannah Y; Powell, Simon N; Riaz, Nadeem; Robson, Mark E; Reis-Filho, Jorge S; Chenevix-Trench, Georgia

    2018-02-28

    Pathogenic germline variants in ataxia-telangiectasia mutated (ATM), a gene that plays a role in DNA damage response and cell cycle checkpoints, confer an increased breast cancer (BC) risk. Here, we investigated the phenotypic characteristics and landscape of somatic genetic alterations in 24 BCs from ATM germline mutation carriers by whole-exome and targeted sequencing. ATM-associated BCs were consistently hormone receptor positive and largely displayed minimal immune infiltrate. Although 79.2% of these tumors exhibited loss of heterozygosity of the ATM wild-type allele, none displayed high activity of mutational signature 3 associated with defective homologous recombination DNA (HRD) repair. No TP53 mutations were found in the ATM-associated BCs. Analysis of an independent data set confirmed that germline ATM variants and TP53 somatic mutations are mutually exclusive. Our findings indicate that ATM-associated BCs often harbor bi-allelic inactivation of ATM, are phenotypically distinct from BRCA1/2-associated BCs, lack HRD-related mutational signatures, and that TP53 and ATM genetic alterations are likely epistatic.

  18. Hermes (Rbpms is a Critical Component of RNP Complexes that Sequester Germline RNAs during Oogenesis

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    Tristan Aguero

    2016-01-01

    Full Text Available The germ cell lineage in Xenopus is specified by the inheritance of germ plasm that assembles within the mitochondrial cloud or Balbiani body in stage I oocytes. Specific RNAs, such as nanos1, localize to the germ plasm. nanos1 has the essential germline function of blocking somatic gene expression and thus preventing Primordial Germ Cell (PGC loss and sterility. Hermes/Rbpms protein and nanos RNA co-localize within germinal granules, diagnostic electron dense particles found within the germ plasm. Previous work indicates that nanos accumulates within the germ plasm through a diffusion/entrapment mechanism. Here we show that Hermes/Rbpms interacts with nanos through sequence specific RNA localization signals found in the nanos-3′UTR. Importantly, Hermes/Rbpms specifically binds nanos, but not Vg1 RNA in the nucleus of stage I oocytes. In vitro binding data show that Hermes/Rbpms requires additional factors that are present in stage I oocytes in order to bind nanos1. One such factor may be hnRNP I, identified in a yeast-2-hybrid screen as directly interacting with Hermes/Rbpms. We suggest that Hermes/Rbpms functions as part of a RNP complex in the nucleus that facilitates selection of germline RNAs for germ plasm localization. We propose that Hermes/Rbpms is required for nanos RNA to form within the germinal granules and in this way, participates in the germline specific translational repression and sequestration of nanos RNA.

  19. Hermes (Rbpms) is a Critical Component of RNP Complexes that Sequester Germline RNAs during Oogenesis.

    Science.gov (United States)

    Aguero, Tristan; Zhou, Yi; Kloc, Malgorzata; Chang, Patrick; Houliston, Evelyn; King, Mary Lou

    2016-03-01

    The germ cell lineage in Xenopus is specified by the inheritance of germ plasm that assembles within the mitochondrial cloud or Balbiani body in stage I oocytes. Specific RNAs, such as nanos1 , localize to the germ plasm. nanos1 has the essential germline function of blocking somatic gene expression and thus preventing Primordial Germ Cell (PGC) loss and sterility. Hermes/Rbpms protein and nanos RNA co-localize within germinal granules, diagnostic electron dense particles found within the germ plasm. Previous work indicates that nanos accumulates within the germ plasm through a diffusion/entrapment mechanism. Here we show that Hermes/Rbpms interacts with nanos through sequence specific RNA localization signals found in the nanos -3'UTR. Importantly, Hermes/Rbpms specifically binds nanos , but not Vg1 RNA in the nucleus of stage I oocytes. In vitro binding data show that Hermes/Rbpms requires additional factors that are present in stage I oocytes in order to bind nanos1 . One such factor may be hnRNP I, identified in a yeast-2-hybrid screen as directly interacting with Hermes/Rbpms. We suggest that Hermes/Rbpms functions as part of a RNP complex in the nucleus that facilitates selection of germline RNAs for germ plasm localization. We propose that Hermes/Rbpms is required for nanos RNA to form within the germinal granules and in this way, participates in the germline specific translational repression and sequestration of nanos RNA .

  20. Production of germline transgenic prairie voles (Microtus ochrogaster) using lentiviral vectors.

    Science.gov (United States)

    Donaldson, Zoe R; Yang, Shang-Hsun; Chan, Anthony W S; Young, Larry J

    2009-12-01

    The study of alternative model organisms has yielded tremendous insights into the regulation of behavioral and physiological traits not displayed by more widely used animal models, such as laboratory rats and mice. In particular, comparative approaches often exploit species ideally suited for investigating specific phenomenon. For instance, comparative studies of socially monogamous prairie voles and polygamous meadow voles have been instrumental toward gaining an understanding of the genetic and neurobiological basis of social bonding. However, laboratory studies of less commonly used organisms, such as prairie voles, have been limited by a lack of genetic tools, including the ability to manipulate the genome. Here, we show that lentiviral vector-mediated transgenesis is a rapid and efficient approach for creating germline transgenics in alternative laboratory rodents. Injection of a green fluorescent protein (GFP)-expressing lentiviral vector into the perivitelline space of 23 single-cell embryos yielded three live offspring (13 %), one of which (33%) contained germline integration of a GFP transgene driven by the human ubiquitin-C promoter. In comparison, transfer of 23 uninjected embryos yielded six live offspring (26%). Green fluorescent protein is present in all tissues examined and is expressed widely in the brain. The GFP transgene is heritable and stably expressed until at least the F(2) generation. This technology has the potential to allow investigation of specific gene candidates in prairie voles and provides a general protocol to pursue germline transgenic manipulation in many different rodent species.

  1. Clinical Assessment and Diagnosis of Germline Predisposition to Hematopoietic Malignancies: The University of Chicago Experience

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    Ami V. Desai

    2017-12-01

    Full Text Available With the increasing use of clinical genomics to guide cancer treatment and management, there is a rise in the identification of germline cancer predisposition syndromes and a critical need for patients with germline findings to be referred for surveillance and care. The University of Chicago Hematopoietic Malignancies Cancer Risk Team has established a unique approach to patient care for individuals with hereditary hematologic malignancies through close communication and coordination between our pediatric and adult programs. Dedicated program members, including physicians, nurses, genetic counselors, and clinical research assistants, screen individuals for cancer predisposition at initial diagnosis through survivorship, in addition to testing individuals with an established family history of a cancer predisposition syndrome. Sample procurement, such as a skin biopsy at the time of bone marrow aspirate/biopsy in individuals with a positive screen, has facilitated timely identification of clinical germline findings or has served as a pipeline for translational research. Our integrated translational research program has led to the identification of novel syndromes in collaboration with other investigators, which have been incorporated iteratively into our clinical pipeline. Individuals are referred for clinical assessment based on personal and family history, identification of variants in susceptibility genes via molecular tumor testing, and during evaluation for matched related allogeneic stem cell transplantation. Upon referral, genetic counseling incorporates education with mindfulness of the psychosocial issues surrounding germline testing at different ages. The training and role of genetic counselors continues to grow, with the discovery of new predisposition syndromes, in the age of improved molecular diagnostics and new models for service delivery, such as telemedicine. With the identification of new syndromes that may predispose individuals

  2. Minisatellite germline mutation rate in the Techa River population

    Energy Technology Data Exchange (ETDEWEB)

    Dubrova, Yuri E. [Department of Genetics, University of Leicester, University Road, Leicester LE1 7RH (United Kingdom)]. E-mail: yed2@le.ac.uk; Ploshchanskaya, Olga G. [Urals Research Centre for Radiation Medicine, Medgorodok, Chelyabinsk 454076 (Russian Federation); Department of Radiobiology, Chelyabinsk State University, Chelyabinsk 454021 (Russian Federation); Kozionova, Olga S. [Urals Research Centre for Radiation Medicine, Medgorodok, Chelyabinsk 454076 (Russian Federation); Department of Radiobiology, Chelyabinsk State University, Chelyabinsk 454021 (Russian Federation); Akleyev, Alexander V. [Urals Research Centre for Radiation Medicine, Medgorodok, Chelyabinsk 454076 (Russian Federation); Department of Radiobiology, Chelyabinsk State University, Chelyabinsk 454021 (Russian Federation)

    2006-12-01

    Germline mutation at eight minisatellite loci has been studied among the irradiated families from the Techa River population and non-exposed families from the rural area of the Chelyabinsk and Kurgan Oblasts. The groups were matched by ethnicity, parental age, occupation and smoking habit. A statistically significant 1.7-fold increase in mutation rate was found in the germline of irradiated fathers, whereas maternal germline mutation rate in the exposed families was not elevated. Most of the minisatellite loci showed an elevated paternal mutation rate in the exposed group, indicating a generalised increase in minisatellite germline mutation rate in the Techa River population. These data suggest that the elevated minisatellite mutation rate can be attributed to radioactive exposure. The spectra of paternal mutation seen in the unexposed and exposed families were indistinguishable.

  3. Minisatellite germline mutation rate in the Techa River population

    International Nuclear Information System (INIS)

    Dubrova, Yuri E.; Ploshchanskaya, Olga G.; Kozionova, Olga S.; Akleyev, Alexander V.

    2006-01-01

    Germline mutation at eight minisatellite loci has been studied among the irradiated families from the Techa River population and non-exposed families from the rural area of the Chelyabinsk and Kurgan Oblasts. The groups were matched by ethnicity, parental age, occupation and smoking habit. A statistically significant 1.7-fold increase in mutation rate was found in the germline of irradiated fathers, whereas maternal germline mutation rate in the exposed families was not elevated. Most of the minisatellite loci showed an elevated paternal mutation rate in the exposed group, indicating a generalised increase in minisatellite germline mutation rate in the Techa River population. These data suggest that the elevated minisatellite mutation rate can be attributed to radioactive exposure. The spectra of paternal mutation seen in the unexposed and exposed families were indistinguishable

  4. Performance of Lynch syndrome predictive models in quantifying the likelihood of germline mutations in patients with abnormal MLH1 immunoexpression.

    Science.gov (United States)

    Cabreira, Verónica; Pinto, Carla; Pinheiro, Manuela; Lopes, Paula; Peixoto, Ana; Santos, Catarina; Veiga, Isabel; Rocha, Patrícia; Pinto, Pedro; Henrique, Rui; Teixeira, Manuel R

    2017-01-01

    Lynch syndrome (LS) accounts for up to 4 % of all colorectal cancers (CRC). Detection of a pathogenic germline mutation in one of the mismatch repair genes is the definitive criterion for LS diagnosis, but it is time-consuming and expensive. Immunohistochemistry is the most sensitive prescreening test and its predictive value is very high for loss of expression of MSH2, MSH6, and (isolated) PMS2, but not for MLH1. We evaluated if LS predictive models have a role to improve the molecular testing algorithm in this specific setting by studying 38 individuals referred for molecular testing and who were subsequently shown to have loss of MLH1 immunoexpression in their tumors. For each proband we calculated a risk score, which represents the probability that the patient with CRC carries a pathogenic MLH1 germline mutation, using the PREMM 1,2,6 and MMRpro predictive models. Of the 38 individuals, 18.4 % had a pathogenic MLH1 germline mutation. MMRpro performed better for the purpose of this study, presenting a AUC of 0.83 (95 % CI 0.67-0.9; P < 0.001) compared with a AUC of 0.68 (95 % CI 0.51-0.82, P = 0.09) for PREMM 1,2,6 . Considering a threshold of 5 %, MMRpro would eliminate unnecessary germline mutation analysis in a significant proportion of cases while keeping very high sensitivity. We conclude that MMRpro is useful to correctly predict who should be screened for a germline MLH1 gene mutation and propose an algorithm to improve the cost-effectiveness of LS diagnosis.

  5. Molecular Background of Colorectal Tumors From Patients with Lynch Syndrome Associated With Germline Variants in PMS2.

    Science.gov (United States)

    Ten Broeke, S W; van Bavel, T C; Jansen, A M L; Gómez-García, E; Hes, F J; van Hest, L P; Letteboer, T G W; Olderode-Berends, M J W; Ruano, D; Spruijt, L; Suerink, M; Tops, C M; van Eijk, R; Morreau, H; van Wezel, T; Nielsen, M

    2018-05-11

    Germline variants in the mismatch repair genes MLH1, MSH2 (EPCAM), MSH6, or PMS2 cause Lynch syndrome. Patients with these variants have an increased risk of developing colorectal cancers (CRCs) that differ from sporadic CRCs in genetic and histologic features. It has been a challenge to study CRCs associated with PMS2 variants (PMS2-associated CRCs) because these develop less frequently and in patients of older ages than colorectal tumors with variants in the other mismatch repair genes. We analyzed 20 CRCs associated with germline variants in PMS2, 22 sporadic CRCs, 18 CRCs with germline variants in MSH2, and 24 CRCs from patients with germline variants in MLH1. Tumor tissue blocks were collected from Dutch pathology departments in 2017. After extraction of tumor DNA, we used a platform designed to detect approximately 3000 somatic hotspot variants in 55 genes (including KRAS, APC, CTNNB1, and TP53). Somatic variant frequencies were compared using the Fisher's exact test. None of the PMS2-associated CRCs contained any somatic variants in the catenin beta 1 gene (CTNNB1), which encodes β-catenin, whereas 14/24 MLH1-associated CRCs (58%) contained variants in CTNNB1. Half of PMS2-associated CRCs contained KRAS variants, but only 20% of these were in hotspots that encoded G12D or G13D. These hotspot variants occurred more frequently in CRCs associated with variants in MLH1 (37.5%, P=.44) and MSH2 (and 71.4%, P=.035) than with variants in PMS2. In a genetic analysis of 84 colorectal tumors, we found tumors from patients with PMS2-associated Lynch syndrome to be distinct from colorectal tumors associated with defects in other mismatch repair genes. This might account for differences in development and less frequent occurrence. Copyright © 2018 AGA Institute. Published by Elsevier Inc. All rights reserved.

  6. Genetic and genomic analysis modeling of germline c-MYC overexpression and cancer susceptibility

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    Nunes Virginia

    2008-01-01

    Full Text Available Abstract Background Germline genetic variation is associated with the differential expression of many human genes. The phenotypic effects of this type of variation may be important when considering susceptibility to common genetic diseases. Three regions at 8q24 have recently been identified to independently confer risk of prostate cancer. Variation at 8q24 has also recently been associated with risk of breast and colorectal cancer. However, none of the risk variants map at or relatively close to known genes, with c-MYC mapping a few hundred kilobases distally. Results This study identifies cis-regulators of germline c-MYC expression in immortalized lymphocytes of HapMap individuals. Quantitative analysis of c-MYC expression in normal prostate tissues suggests an association between overexpression and variants in Region 1 of prostate cancer risk. Somatic c-MYC overexpression correlates with prostate cancer progression and more aggressive tumor forms, which was also a pathological variable associated with Region 1. Expression profiling analysis and modeling of transcriptional regulatory networks predicts a functional association between MYC and the prostate tumor suppressor KLF6. Analysis of MYC/Myc-driven cell transformation and tumorigenesis substantiates a model in which MYC overexpression promotes transformation by down-regulating KLF6. In this model, a feedback loop through E-cadherin down-regulation causes further transactivation of c-MYC. Conclusion This study proposes that variation at putative 8q24 cis-regulator(s of transcription can significantly alter germline c-MYC expression levels and, thus, contribute to prostate cancer susceptibility by down-regulating the prostate tumor suppressor KLF6 gene.

  7. Clinical and genetic spectrum of Birt–Hogg–Dubé syndrome patients in whom pneumothorax and/or multiple lung cysts are the presenting feature

    Science.gov (United States)

    Kunogi, Makiko; Kurihara, Masatoshi; Ikegami, Takako Shigihara; Kobayashi, Toshiyuki; Shindo, Noriko; Kumasaka, Toshio; Gunji, Yoko; Kikkawa, Mika; Iwakami, Shin-ichiro; Hino, Okio; Takahashi, Kazuhisa

    2010-01-01

    Background Birt–Hogg–Dubé syndrome (BHDS) is an inherited autosomal genodermatosis characterised by fibrofolliculomas of the skin, renal tumours and multiple lung cysts. Genetic studies have disclosed that the clinical picture as well as responsible germline FLCN mutations are diverse. Objectives BHDS may be caused by a germline deletion which cannot be detected by a conventional genetic approach. Real-time quantitative polymerase chain reaction (qPCR) may be able to identify such a mutation and thus provide us with a more accurate clinical picture of BHDS. Methods This study analysed 36 patients with multiple lung cysts of undetermined causes. Denaturing high performance liquid chromatography (DHPLC) was applied for mutation screening. If no abnormality was detected by DHPLC, the amount of each FLCN exon in genome was quantified by qPCR. Results An FLCN germline mutation was found in 23 (63.9%) of the 36 patients by DHPLC and direct sequencing (13 unique small nucleotide alterations which included 11 novel mutations). A large genomic deletion was identified in two of the remaining 13 patients by qPCR (one patient with exon 14 deletion and one patient with a deletion encompassing exons 9 to 14). Mutations including genomic deletions were most frequently identified in the 3′-end of the FLCN gene including exons 12 and 13 (13/25=52.0%). The BHDS patients whose multiple cysts prompted the diagnosis in this study showed a very low incidence of skin and renal involvement. Conclusions BHDS is due to large deletions as well as small nucleotide alterations. Racial differences may occur between Japanese and patients of European decent in terms of FLCN mutations and clinical manifestations. PMID:20413710

  8. EPHB2 germline variants in patients with colorectal cancer or hyperplastic polyposis

    International Nuclear Information System (INIS)

    Kokko, Antti; Tomlinson, Ian PM; Vahteristo, Pia; Aaltonen, Lauri A; Laiho, Päivi; Lehtonen, Rainer; Korja, Sanna; Carvajal-Carmona, Luis G; Järvinen, Heikki; Mecklin, Jukka-Pekka; Eng, Charis; Schleutker, Johanna

    2006-01-01

    Ephrin receptor B2 (EPHB2) has recently been proposed as a novel tumor suppressor gene in colorectal cancer (CRC). Inactivation of the gene has been shown to correlate with progression of colorectal tumorigenesis, and somatic mutations have been reported in both colorectal and prostate tumors. Here we have analyzed the EPHB2 gene for germline alterations in 101 individuals either with 1) CRC and a personal or family history of prostate cancer (PC), or 2) intestinal hyperplastic polyposis (HPP), a condition associated with malignant degeneration such as serrated adenoma and CRC. Four previously unknown missense alterations were observed, which may be associated with the disease phenotype. Two of the changes, I361V and R568W, were identified in Finnish CRC patients, but not in over 300 Finnish familial CRC or PC patients or more than 200 population-matched healthy controls. The third change, D861N, was observed in a UK HPP patient, but not in additional 40 UK HPP patients or in 200 UK healthy controls. The fourth change R80H, originally identified in a Finnish CRC patient, was also found in 1/106 familial CRC patients and in 9/281 healthy controls and is likely to be a neutral polymorphism. We detected novel germline EPHB2 alterations in patients with colorectal tumors. The results suggest a limited role for these EPHB2 variants in colon tumor predisposition. Further studies including functional analyses are needed to confirm this

  9. Germline Mutations of Inhibins in Early-Onset Ovarian Epithelial Tumors

    Science.gov (United States)

    Tournier, Isabelle; Marlin, Régine; Walton, Kelly; Charbonnier, Françoise; Coutant, Sophie; Théry, Jean-Christophe; Charbonnier, Camille; Spurrell, Cailyn; Vezain, Myriam; Ippolito, Lorena; Bougeard, Gaëlle; Roman, Horace; Tinat, Julie; Sabourin, Jean-Christophe; Stoppa-Lyonnet, Dominique; Caron, Olivier; Bressac-de Paillerets, Brigitte; Vaur, Dominique; King, Mary-Claire; Harrison, Craig; Frebourg, Thierry

    2014-01-01

    To identify novel genetic bases of early-onset epithelial ovarian tumors, we used the trio exome sequencing strategy in a patient without familial history of cancer who presented metastatic serous ovarian adenocarcinomas at 21 years of age. We identified a single de novo mutation (c.1157A>G/p.Asn386Ser) within the INHBA gene encoding the βA-subunit of inhibins/activins, which play a key role in ovarian development. In vitro, this mutation alters the ratio of secreted activins and inhibins. In a second patient with early-onset serous borderline papillary cystadenoma, we identified an unreported germline mutation (c.179G>T/p.Arg60Leu) of the INHA gene encoding the α-subunit, the partner of the βA-subunit. This mutation also alters the secreted activin/inhibin ratio, by disrupting both inhibin A and inhibin B biosynthesis. In a cohort of 62 cases, we detected an additional unreported germline mutation of the INHBA gene (c.839G>A/p.Gly280Glu). Our results strongly suggest that inhibin mutations contribute to the genetic determinism of epithelial ovarian tumors. PMID:24302632

  10. Pitfalls in molecular analysis for mismatch repair deficiency in a family with biallelic pms2 germline mutations.

    Science.gov (United States)

    Leenen, C H M; Geurts-Giele, W R R; Dubbink, H J; Reddingius, R; van den Ouweland, A M; Tops, C M J; van de Klift, H M; Kuipers, E J; van Leerdam, M E; Dinjens, W N M; Wagner, A

    2011-12-01

    Heterozygous germline mutations in the mismatch repair (MMR) genes MLH1, MSH2, MSH6 and PMS2 cause Lynch syndrome. Biallelic mutations in the MMR genes are associated with a childhood cancer syndrome [constitutional mismatch repair deficiency (CMMR-D)]. This is predominantly characterized by hematological malignancies and tumors of the bowel and brain, often associated with signs of neurofibromatosis type 1 (NF1). Diagnostic strategies for selection of patients for MMR gene analysis include analysis of microsatellite instability (MSI) and immunohistochemical (IHC) analysis of MMR proteins in tumor tissue. We report the clinical characterization and molecular analyses of tumor specimens from a family with biallelic PMS2 germline mutations. This illustrates the pitfalls of present molecular screening strategies. Tumor tissues of five family members were analyzed for MSI and IHC. MSI was observed in only one of the analyzed tissues. However, IHC analysis of brain tumor tissue of the index patient and his sister showed absence of PMS2 expression, and germline mutation analyses showed biallelic mutations in PMS2: p.Ser46IIe and p.Pro246fs. The same heterozygous mutations were confirmed in the father and mother, respectively. These data support the conclusion that in case of a clinical phenotype of CMMR-D, it is advisable to routinely combine MSI analysis with IHC analysis for the expression of MMR proteins. With inconclusive or conflicting results, germline mutation analysis of the MMR genes should be considered after thorough counselling of the patients and/or their relatives. © 2011 John Wiley & Sons A/S.

  11. Germline but macrophage-tropic CYBB mutations in kindreds with X-linked predisposition to tuberculous mycobacterial diseases

    OpenAIRE

    2011-01-01

    Abstract Germline mutations in the human CYBB gene, encoding the gp91phox subunit of the phagocyte NADPH oxidase, impair the respiratory burst of phagocytes and result in X-linked chronic granulomatous disease. We report two kindreds in which otherwise healthy male adults show X-linked recessive Mendelian susceptibility to mycobacterial diseases. These patients harbor mutations in CYBB that profoundly reduce the respiratory burst in monocyte-derived macrophages, but not in monocyte...

  12. RAD50 germline mutations are associated with poor survival in BRCA1/2-negative breast cancer patients.

    Science.gov (United States)

    Fan, Cong; Zhang, Juan; Ouyang, Tao; Li, Jinfeng; Wang, Tianfeng; Fan, Zhaoqing; Fan, Tie; Lin, Benyao; Xie, Yuntao

    2018-05-04

    RAD50 is a highly conserved DNA double-strand break (DSB) repair gene. However, the associations between RAD50 germline mutations and the survival and risk of breast cancer have not been fully elucidated. Here, we aimed to investigate the clinical impact of RAD50 germline mutations in a large cohort of unselected breast cancer patients. In this study, RAD50 germline mutations were determined using next-generation sequencing in 7657 consecutive unselected breast cancer patients without BRCA1/2 mutations. We also screened for RAD50 recurrent mutations (L719fs, K994fs, and H1269fs) in 5000 healthy controls using Sanger sequencing. We found that 26 out of 7657 (0.34%) patients had RAD50 pathogenic mutations, and 16 patients carried one of the three recurrent mutations (L719fs, n=6 cases; K994fs, n=5 cases; and H1269fs, n=5 cases); the recurrent mutation rate was 0.21%. The frequency of the three recurrent mutations in the 5000 healthy controls was 0.18% (9/5000). These mutations did not confer an increased risk of breast cancer in the studied patients [odds ratios (OR), 1.16; 95% confidence interval (CI), 0.51-2.63; P = 0.72]. Nevertheless, multivariate analysis revealed that RAD50 pathogenic mutations were an independent unfavourable predictor of recurrence-free survival (RFS) [adjusted hazard ratio (HR) 2.66; 95% CI, 1.18-5.98; P=0.018] and disease-specific survival (DSS) (adjusted HR 4.36; 95% CI, 1.58-12.03; P=0.004) in the entire study cohort. Our study suggested that RAD50 germline mutations are not associated with an increased risk of breast cancer, but patients with RAD50 germline mutations have unfavourable survival compared with patients without these mutations. This article is protected by copyright. All rights reserved. © 2018 UICC.

  13. Cell surface fucosylation does not affect development of colon tumors in mice with germline Smad3 mutation

    Science.gov (United States)

    Domino, Steven E.; Karnak, David M.; Hurd, Elizabeth A.

    2006-01-01

    Background/Aims: Neoplasia-related alterations in cell surface α(1,2)fucosylated glycans have been reported in multiple tumors including colon, pancreas, endometrium, cervix, bladder, lung, and choriocarcinoma. Spontaneous colorectal tumors from mice with a germline null mutation of transforming growth factor-β signaling gene Smad3 (Madh3) were tested for α(1,2)fucosylated glycan expression. Methods: Ulex Europaeus Agglutinin-I lectin staining, fucosyltransferase gene northern blot analysis, and a cross of mutant mice with Fut2 and Smad3 germline mutations were performed. Results: Spontaneous colorectal tumors from Smad3 (-/-) homozygous null mice were found to express α(1,2)fucosylated glycans in an abnormal pattern compared to adjacent nonneoplastic colon. Northern blot analysis of α(1,2)fucosyltransferase genes Fut1 and Fut2 revealed that Fut2, but not Fut1, steady-state mRNA levels were significantly increased in tumors relative to adjacent normal colonic mucosa. Mutant mice with a Fut2-inactivating germline mutation were crossed with Smad3 targeted mice. In Smad3 (-/-)/Fut2 (-/-) double knock-out mice, UEA-I lectin staining was eliminated from colon and colon tumors, however, the number and size of tumors present by 24 weeks of age did not vary regardless of the Fut2 genotype. Conclusions: In this model of colorectal cancer, cell surface α(1,2)fucosylation does not affect development of colon tumors. PMID:17264540

  14. Spontaneous germline excision of Tol1, a DNA-based transposable element naturally occurring in the medaka fish genome.

    Science.gov (United States)

    Watanabe, Kohei; Koga, Hajime; Nakamura, Kodai; Fujita, Akiko; Hattori, Akimasa; Matsuda, Masaru; Koga, Akihiko

    2014-04-01

    DNA-based transposable elements are ubiquitous constituents of eukaryotic genomes. Vertebrates are, however, exceptional in that most of their DNA-based elements appear to be inactivated. The Tol1 element of the medaka fish, Oryzias latipes, is one of the few elements for which copies containing an undamaged gene have been found. Spontaneous transposition of this element in somatic cells has previously been demonstrated, but there is only indirect evidence for its germline transposition. Here, we show direct evidence of spontaneous excision in the germline. Tyrosinase is the key enzyme in melanin biosynthesis. In an albino laboratory strain of medaka fish, which is homozygous for a mutant tyrosinase gene in which a Tol1 copy is inserted, we identified de novo reversion mutations related to melanin pigmentation. The gamete-based reversion rate was as high as 0.4%. The revertant fish carried the tyrosinase gene from which the Tol1 copy had been excised. We previously reported the germline transposition of Tol2, another DNA-based element that is thought to be a recent invader of the medaka fish genome. Tol1 is an ancient resident of the genome. Our results indicate that even an old element can contribute to genetic variation in the host genome as a natural mutator.

  15. Correlation of FCGR3A and EGFR germline polymorphisms with the efficacy of cetuximab in KRAS wild-type metastatic colorectal cancer

    NARCIS (Netherlands)

    Pander, Jan; Gelderblom, Hans; Antonini, Ninja F.; Tol, Jolien; van Krieken, Johan H. J. M.; van der Straaten, Tahar; Punt, Cornelis J. A.; Guchelaar, Henk-Jan

    2010-01-01

    Next to KRAS mutation status, additional predictive markers are needed for the response to cetuximab in patients with metastatic colorectal cancer (mCRC). Previous studies indicated that germline polymorphisms in specific genes may predict efficacy and toxicity of cetuximab in mCRC patients.

  16. The signature of somatic hypermutation appears to be written into the germline IgV segment repertoire.

    Science.gov (United States)

    Blanden, R V; Rothenfluh, H S; Zylstra, P; Weiller, G F; Steele, E J

    1998-04-01

    We present here a unifying hypothesis for the molecular mechanism of somatic hypermutation and somatic gene conversion in IgV genes involving reverse transcription using RNA templates from the V-gene loci to produce cDNA which undergoes homologous recombination with chromosomal V(D)J DNA. Experimental evidence produced over the last 20 years is essentially consistent with this hypothesis. We also review evidence suggesting that somatically generated IgV sequences from B lymphocytes have been fed back to germline DNA over evolutionary time.

  17. Germline variants in the ATM gene and breast cancer susceptibility ...

    African Journals Online (AJOL)

    Chaymaa Marouf

    2017-03-06

    Mar 6, 2017 ... Results: We did not detect the ATM c.7271T > G and c.1066–6T > G (IVS10–6T > G) ..... [8] Lavin MF, Shiloh Y. The genetic defect in ataxia-telangiectasia. .... [36] Inskip HM, Kinlen LJ, Taylor AM, Woods CG, Arlett CF. Risk of ...

  18. Simple detection of germline microsatellite instability for diagnosis of constitutional mismatch repair cancer syndrome.

    Science.gov (United States)

    Ingham, Danielle; Diggle, Christine P; Berry, Ian; Bristow, Claire A; Hayward, Bruce E; Rahman, Nazneen; Markham, Alexander F; Sheridan, Eamonn G; Bonthron, David T; Carr, Ian M

    2013-06-01

    Heterozygous mutations in DNA mismatch repair (MMR) genes result in predisposition to colorectal cancer (hereditary nonpolyposis colorectal cancer or Lynch syndrome). Patients with biallelic mutations in these genes, however, present earlier, with constitutional mismatch repair deficiency cancer syndrome (CMMRD), which is characterized by a spectrum of rare childhood malignancies and café-au-lait skin patches. The hallmark of MMR deficiency, microsatellite instability (MSI), is readily detectable in tumor DNA in Lynch syndrome, but is also present in constitutional DNA of CMMRD patients. However, detection of constitutional or germline MSI (gMSI) has hitherto relied on technically difficult assays that are not routinely applicable for clinical diagnosis. Consequently, we have developed a simple high-throughput screening methodology to detect gMSI in CMMRD patients based on the presence of stutter peaks flanking a dinucleotide repeat allele when amplified from patient blood DNA samples. Using the three different microsatellite markers, the gMSI ratio was determined in a cohort of normal individuals and 10 CMMRD patients, with biallelic germline mutations in PMS2 (seven patients), MSH2 (one patient), or MSH6 (two patients). Subjects with either PMS2 or MSH2 mutations were easily identified; however, this measure was not altered in patients with CMMRD due to MSH6 mutation. © 2013 Wiley Periodicals, Inc.

  19. A novel molecular diagnostics platform for somatic and germline precision oncology.

    Science.gov (United States)

    Cabanillas, Rubén; Diñeiro, Marta; Castillo, David; Pruneda, Patricia C; Penas, Cristina; Cifuentes, Guadalupe A; de Vicente, Álvaro; Durán, Noelia S; Álvarez, Rebeca; Ordóñez, Gonzalo R; Cadiñanos, Juan

    2017-07-01

    Next-generation sequencing (NGS) opens new options in clinical oncology, from therapy selection to genetic counseling. However, realization of this potential not only requires succeeding in the bioinformatics and interpretation of the results, but also in their integration into the clinical practice. We have developed a novel NGS diagnostic platform aimed at detecting (1) somatic genomic alterations associated with the response to approved targeted cancer therapies and (2) germline mutations predisposing to hereditary malignancies. Next-generation sequencing libraries enriched in the exons of 215 cancer genes (97 for therapy selection and 148 for predisposition, with 30 informative for both applications), as well as selected introns from 17 genes involved in drug-related rearrangements, were prepared from 39 tumors (paraffin-embedded tissues/cytologies), 36 germline samples (blood) and 10 cell lines using hybrid capture. Analysis of NGS results was performed with specifically developed bioinformatics pipelines. The platform detects single-nucleotide variants (SNVs) and insertions/deletions (indels) with sensitivity and specificity >99.5% (allelic frequency ≥0.1), as well as copy-number variants (CNVs) and rearrangements. Somatic testing identified tailored approved targeted drugs in 35/39 tumors (89.74%), showing a diagnostic yield comparable to that of leading commercial platforms. A somatic EGFR p.E746_S752delinsA mutation in a mediastinal metastasis from a breast cancer prompted its anatomopathologic reassessment, its definite reclassification as a lung cancer and its treatment with gefitinib (partial response sustained for 15 months). Testing of 36 germline samples identified two pathogenic mutations (in CDKN2A and BRCA2 ). We propose a strategy for interpretation and reporting of results adaptable to the aim of the request, the availability of tumor and/or normal samples and the scope of the informed consent. With an adequate methodology, it is possible to

  20. Impact of germline and somatic missense variations on drug binding sites.

    Science.gov (United States)

    Yan, C; Pattabiraman, N; Goecks, J; Lam, P; Nayak, A; Pan, Y; Torcivia-Rodriguez, J; Voskanian, A; Wan, Q; Mazumder, R

    2017-03-01

    Advancements in next-generation sequencing (NGS) technologies are generating a vast amount of data. This exacerbates the current challenge of translating NGS data into actionable clinical interpretations. We have comprehensively combined germline and somatic nonsynonymous single-nucleotide variations (nsSNVs) that affect drug binding sites in order to investigate their prevalence. The integrated data thus generated in conjunction with exome or whole-genome sequencing can be used to identify patients who may not respond to a specific drug because of alterations in drug binding efficacy due to nsSNVs in the target protein's gene. To identify the nsSNVs that may affect drug binding, protein-drug complex structures were retrieved from Protein Data Bank (PDB) followed by identification of amino acids in the protein-drug binding sites using an occluded surface method. Then, the germline and somatic mutations were mapped to these amino acids to identify which of these alter protein-drug binding sites. Using this method we identified 12 993 amino acid-drug binding sites across 253 unique proteins bound to 235 unique drugs. The integration of amino acid-drug binding sites data with both germline and somatic nsSNVs data sets revealed 3133 nsSNVs affecting amino acid-drug binding sites. In addition, a comprehensive drug target discovery was conducted based on protein structure similarity and conservation of amino acid-drug binding sites. Using this method, 81 paralogs were identified that could serve as alternative drug targets. In addition, non-human mammalian proteins bound to drugs were used to identify 142 homologs in humans that can potentially bind to drugs. In the current protein-drug pairs that contain somatic mutations within their binding site, we identified 85 proteins with significant differential gene expression changes associated with specific cancer types. Information on protein-drug binding predicted drug target proteins and prevalence of both somatic and

  1. Male germline recombination of a conditional allele by the widely used Dermo1-cre (Twist2-cre) transgene.

    Science.gov (United States)

    He, Yun; Sun, Xiumei; Wang, Li; Mishina, Yuji; Guan, Jun-Lin; Liu, Fei

    2017-09-01

    Conditional gene knockout using the Cre/loxP system is instrumental in advancing our understanding of the function of genes in a wide range of disciplines. It is becoming increasingly apparent in the literature that recombination mediated by some Cre transgenes can occur in unexpected tissues. Dermo1-Cre (Twist2-Cre) has been widely used to target skeletal lineage cells as well as other mesoderm-derived cells. Here we report that Dermo1-Cre exhibits spontaneous male germline recombination activity leading to a Cre-mediated recombination of a floxed Ptk2 (Protein tyrosine kinase 2, also known as Fak [Focal adhesion kinase]) allele but not a floxed Rb1cc1 (RB1 inducible coiled-coil 1, also known as Fip200 [FAK-family Interacting Protein of 200 kDa]) allele at high frequency. This ectopic germline activity of Dermo1-Cre occurred in all or none manner in a given litter. We demonstrated that the occurrence of germline recombination activity of Dermo1-Cre transgene can be avoided by using female mice as parental Dermo1-Cre carriers. © 2017 Wiley Periodicals, Inc.

  2. Co-option of the piRNA pathway for germline-specific alternative splicing of C. elegans TOR.

    Science.gov (United States)

    Barberán-Soler, Sergio; Fontrodona, Laura; Ribó, Anna; Lamm, Ayelet T; Iannone, Camilla; Cerón, Julián; Lehner, Ben; Valcárcel, Juan

    2014-09-25

    Many eukaryotic genes contain embedded antisense transcripts and repetitive sequences of unknown function. We report that male germline-specific expression of an antisense transcript contained in an intron of C. elegans Target of Rapamycin (TOR, let-363) is associated with (1) accumulation of endo-small interfering RNAs (siRNAs) against an embedded Helitron transposon and (2) activation of an alternative 3' splice site of TOR. The germline-specific Argonaute proteins PRG-1 and CSR-1, which participate in self/nonself RNA recognition, antagonistically regulate the generation of these endo-siRNAs, TOR mRNA levels, and 3' splice-site selection. Supply of exogenous double-stranded RNA against the region of sense/antisense overlap reverses changes in TOR expression and splicing and suppresses the progressive multigenerational sterility phenotype of prg-1 mutants. We propose that recognition of a "nonself" intronic transposon by endo-siRNAs/the piRNA system provides physiological regulation of expression and alternative splicing of a host gene that, in turn, contributes to the maintenance of germline function across generations. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  3. Germline mutations in lysine specific demethylase 1 (LSD1/KDM1A) confer susceptibility to multiple myeloma.

    Science.gov (United States)

    Wei, Xiaomu; Calvo-Vidal, M Nieves; Chen, Siwei; Wu, Gang; Revuelta, Maria V; Sun, Jian; Zhang, Jinghui; Walsh, Michael F; Nichols, Kim E; Joseph, Vijai; Snyder, Carrie; Vachon, Celine M; McKay, James D; Wang, Shu-Ping; Jayabalan, David S; Jacobs, Lauren M; Becirovic, Dina; Waller, Rosalie G; Artomov, Mykyta; Viale, Agnes; Patel, Jayeshkumar; Phillip, Jude M; Chen-Kiang, Selina; Curtin, Karen; Salama, Mohamed; Atanackovic, Djordje; Niesvizky, Ruben; Landgren, Ola; Slager, Susan L; Godley, Lucy A; Churpek, Jane; Garber, Judy E; Anderson, Kenneth C; Daly, Mark J; Roeder, Robert G; Dumontet, Charles; Lynch, Henry T; Mullighan, Charles G; Camp, Nicola J; Offit, Kenneth; Klein, Robert J; Yu, Haiyuan; Cerchietti, Leandro; Lipkin, Steven M

    2018-03-20

    Given the frequent and largely incurable occurrence of multiple myeloma (MM), identification of germline genetic mutations that predispose cells to MM may provide insight into disease etiology and the developmental mechanisms of its cell of origin, the plasma cell. Here we identified familial and early-onset MM kindreds with truncating mutations in lysine-specific demethylase 1 (LSD1/KDM1A), an epigenetic transcriptional repressor that primarily demethylates histone H3 on lysine 4 and regulates hematopoietic stem cell self-renewal. Additionally, we found higher rates of germline truncating and predicted deleterious missense KDM1A mutations in MM patients unselected for family history compared to controls. Both monoclonal gammopathy of unknown significance (MGUS) and MM cells have significantly lower KDM1A transcript levels compared with normal plasma cells. Transcriptome analysis of MM cells from KDM1A mutation carriers shows enrichment of pathways and MYC target genes previously associated with myeloma pathogenesis. In mice, antigen challenge followed by pharmacological inhibition of KDM1A promoted plasma cell expansion, enhanced secondary immune response, elicited appearance of serum paraprotein, and mediated upregulation of MYC transcriptional targets. These changes are consistent with the development of MGUS. Collectively, our findings show KDM1A is the first autosomal dominant MM germline predisposition gene, providing new insights into its mechanistic roles as a tumor suppressor during post-germinal center B cell differentiation. Copyright ©2018, American Association for Cancer Research.

  4. Sex chromosome-specific regulation in the Drosophila male germline but little evidence for chromosomal dosage compensation or meiotic inactivation.

    Directory of Open Access Journals (Sweden)

    Colin D Meiklejohn

    2011-08-01

    Full Text Available The evolution of heteromorphic sex chromosomes (e.g., XY in males or ZW in females has repeatedly elicited the evolution of two kinds of chromosome-specific regulation: dosage compensation--the equalization of X chromosome gene expression in males and females--and meiotic sex chromosome inactivation (MSCI--the transcriptional silencing and heterochromatinization of the X during meiosis in the male (or Z in the female germline. How the X chromosome is regulated in the Drosophila melanogaster male germline is unclear. Here we report three new findings concerning gene expression from the X in Drosophila testes. First, X chromosome-wide dosage compensation appears to be absent from most of the Drosophila male germline. Second, microarray analysis provides no evidence for X chromosome-specific inactivation during meiosis. Third, we confirm the previous discovery that the expression of transgene reporters driven by autosomal spermatogenesis-specific promoters is strongly reduced when inserted on the X chromosome versus the autosomes; but we show that this chromosomal difference in expression is established in premeiotic cells and persists in meiotic cells. The magnitude of the X-autosome difference in transgene expression cannot be explained by the absence of dosage compensation, suggesting that a previously unrecognized mechanism limits expression from the X during spermatogenesis in Drosophila. These findings help to resolve several previously conflicting reports and have implications for patterns of genome evolution and speciation in Drosophila.

  5. Germline CYBB mutations that selectively affect macrophages in kindreds with X-linked predisposition to tuberculous mycobacterial disease

    Science.gov (United States)

    Bustamante, Jacinta; Arias, Andres A; Vogt, Guillaume; Picard, Capucine; Galicia, Lizbeth Blancas; Prando, Carolina; Grant, Audrey V; Marchal, Christophe C; Hubeau, Marjorie; Chapgier, Ariane; de Beaucoudrey, Ludovic; Puel, Anne; Feinberg, Jacqueline; Valinetz, Ethan; Jannière, Lucile; Besse, Céline; Boland, Anne; Brisseau, Jean-Marie; Blanche, Stéphane; Lortholary, Olivier; Fieschi, Claire; Emile, Jean-François; Boisson-Dupuis, Stéphanie; Al-Muhsen, Saleh; Woda, Bruce; Newburger, Peter E; Condino-Neto, Antonio; Dinauer, Mary C; Abel, Laurent; Casanova, Jean-Laurent

    2011-01-01

    Germline mutations in CYBB, the human gene encoding the gp91phox subunit of the phagocyte NADPH oxidase, impair the respiratory burst of all types of phagocytes and result in X-linked chronic granulomatous disease (CGD). We report here two kindreds in which otherwise healthy male adults developed X-linked recessive Mendelian susceptibility to mycobacterial disease (MSMD) syndromes. These patients had previously unknown mutations in CYBB that resulted in an impaired respiratory burst in monocyte-derived macrophages but not in monocytes or granulocytes. The macrophage-specific functional consequences of the germline mutation resulted from cell-specific impairment in the assembly of the NADPH oxidase. This ‘experiment of nature’ indicates that CYBB is associated with MSMD and demonstrates that the respiratory burst in human macrophages is a crucial mechanism for protective immunity to tuberculous mycobacteria. PMID:21278736

  6. Synchronous lung tumours in a patient with metachronous colorectal carcinoma and a germline MSH2 mutation.

    LENUS (Irish Health Repository)

    Canney, A

    2012-02-01

    Mutations of DNA mismatch repair genes are characterised by microsatellite instability and are implicated in carcinogenesis. This mutation susceptible phenotype has been extensively studied in patients with hereditary non-polyposis colon carcinoma, but little is known of the contribution of such mutations in other tumour types, particularly non-small-cell lung carcinoma. This report describes the occurrence of two synchronous lung tumours, one mimicking a metastatic colon carcinoma, in a male patient with a history of metachronous colonic carcinoma. Immunohistochemistry supported a pulmonary origin for both lesions. Mismatch repair protein immunohistochemistry showed loss of MSH2 and MSH6 expression in both colonic tumours and in one lung tumour showing enteric differentiation. Subsequent mutational analysis demonstrated a deleterious germline mutation of the MSH2 mismatch repair gene. The significance of these findings and the practical diagnostic difficulties encountered in this case are discussed.

  7. Cancer-germline antigen vaccines and epigenetic enhancers

    DEFF Research Database (Denmark)

    Gjerstorff, Morten Frier; Burns, Jorge; Ditzel, Henrik Jorn

    2010-01-01

    IMPORTANCE OF THE FIELD: Immunotherapy holds great potential for disseminated cancer, and cancer-germline (CG) antigens are among the most promising tumor targets. They are widely expressed in different cancer types and are essentially tumor-specific, since their expression in normal tissues is l...

  8. Germline CDKN2A/P16INK4A mutations contribute to genetic determinism of sarcoma.

    Science.gov (United States)

    Jouenne, Fanélie; Chauvot de Beauchene, Isaure; Bollaert, Emeline; Avril, Marie-Françoise; Caron, Olivier; Ingster, Olivier; Lecesne, Axel; Benusiglio, Patrick; Terrier, Philippe; Caumette, Vincent; Pissaloux, Daniel; de la Fouchardière, Arnaud; Cabaret, Odile; N'Diaye, Birama; Velghe, Amélie; Bougeard, Gaelle; Mann, Graham J; Koscielny, Serge; Barrett, Jennifer H; Harland, Mark; Newton-Bishop, Julia; Gruis, Nelleke; Van Doorn, Remco; Gauthier-Villars, Marion; Pierron, Gaelle; Stoppa-Lyonnet, Dominique; Coupier, Isabelle; Guimbaud, Rosine; Delnatte, Capucine; Scoazec, Jean-Yves; Eggermont, Alexander M; Feunteun, Jean; Tchertanov, Luba; Demoulin, Jean-Baptiste; Frebourg, Thierry; Bressac-de Paillerets, Brigitte

    2017-09-01

    Sarcomas are rare mesenchymal malignancies whose pathogenesis is poorly understood; both environmental and genetic risk factors could contribute to their aetiology. We performed whole-exome sequencing (WES) in a familial aggregation of three individuals affected with soft-tissue sarcoma (STS) without TP53 mutation (Li-Fraumeni-like, LFL) and found a shared pathogenic mutation in CDKN2A tumour suppressor gene. We searched for individuals with sarcoma among 474 melanoma-prone families with a CDKN2A -/+ genotype and for CDKN2A mutations in 190 TP53 -negative LFL families where the index case was a sarcoma. Including the initial family, eight independent sarcoma cases carried a germline mutation in the CDKN2A /p16 INK4A gene. In five out of seven formalin-fixed paraffin-embedded sarcomas, heterozygosity was lost at germline CDKN2A mutations sites demonstrating complete loss of function. As sarcomas are rare in CDKN2A /p16 INK4A carriers, we searched in constitutional WES of nine carriers for potential modifying rare variants and identified three in platelet-derived growth factor receptor ( PDGFRA ) gene. Molecular modelling showed that two never-described variants could impact the PDGFRA extracellular domain structure. Germline mutations in CDKN2A /P16 INK4A , a gene known to predispose to hereditary melanoma, pancreatic cancer and tobacco-related cancers, account also for a subset of hereditary sarcoma. In addition, we identified PDGFRA as a candidate modifier gene. © Article author(s) (or their employer(s) unless otherwise stated in the text of the article) 2017. All rights reserved. No commercial use is permitted unless otherwise expressly granted.

  9. Spliced DNA Sequences in the Paramecium Germline: Their Properties and Evolutionary Potential

    Science.gov (United States)

    Catania, Francesco; McGrath, Casey L.; Doak, Thomas G.; Lynch, Michael

    2013-01-01

    Despite playing a crucial role in germline-soma differentiation, the evolutionary significance of developmentally regulated genome rearrangements (DRGRs) has received scant attention. An example of DRGR is DNA splicing, a process that removes segments of DNA interrupting genic and/or intergenic sequences. Perhaps, best known for shaping immune-system genes in vertebrates, DNA splicing plays a central role in the life of ciliated protozoa, where thousands of germline DNA segments are eliminated after sexual reproduction to regenerate a functional somatic genome. Here, we identify and chronicle the properties of 5,286 sequences that putatively undergo DNA splicing (i.e., internal eliminated sequences [IESs]) across the genomes of three closely related species of the ciliate Paramecium (P. tetraurelia, P. biaurelia, and P. sexaurelia). The study reveals that these putative IESs share several physical characteristics. Although our results are consistent with excision events being largely conserved between species, episodes of differential IES retention/excision occur, may have a recent origin, and frequently involve coding regions. Our findings indicate interconversion between somatic—often coding—DNA sequences and noncoding IESs, and provide insights into the role of DNA splicing in creating potentially functional genetic innovation. PMID:23737328

  10. Identification of Two Novel HOXB13 Germline Mutations in Portuguese Prostate Cancer Patients

    Science.gov (United States)

    Maia, Sofia; Cardoso, Marta; Pinto, Pedro; Pinheiro, Manuela; Santos, Catarina; Peixoto, Ana; Bento, Maria José; Oliveira, Jorge; Henrique, Rui; Jerónimo, Carmen; Teixeira, Manuel R.

    2015-01-01

    The HOXB13 germline variant G84E (rs138213197) was recently described in men of European descent, with the highest prevalence in Northern Europe. The G84E mutation has not been found in patients of African or Asian ancestry, which may carry other HOXB13 variants, indicating allelic heterogeneity depending on the population. In order to gain insight into the full scope of coding HOXB13 mutations in Portuguese prostate cancer patients, we decided to sequence the entire coding region of the HOXB13 gene in 462 early-onset or familial/hereditary cases. Additionally, we searched for somatic HOXB13 mutations in 178 prostate carcinomas to evaluate their prevalence in prostate carcinogenesis. Three different patients were found to carry in their germline DNA two novel missense variants, which were not identified in 132 control subjects. Both variants are predicted to be deleterious by different in silico tools. No somatic mutations were found. These findings further support the hypothesis that different rare HOXB13 mutations may be found in different ethnic groups. Detection of mutations predisposing to prostate cancer may require re-sequencing rather than genotyping, as appropriate to the population under investigation. PMID:26176944

  11. Remobilization of Sleeping Beauty transposons in the germline of Xenopus tropicalis

    Directory of Open Access Journals (Sweden)

    Yergeau Donald A

    2011-11-01

    Full Text Available Abstract Background The Sleeping Beauty (SB transposon system has been used for germline transgenesis of the diploid frog, Xenopus tropicalis. Injecting one-cell embryos with plasmid DNA harboring an SB transposon substrate together with mRNA encoding the SB transposase enzyme resulted in non-canonical integration of small-order concatemers of the transposon. Here, we demonstrate that SB transposons stably integrated into the frog genome are effective substrates for remobilization. Results Transgenic frogs that express the SB10 transposase were bred with SB transposon-harboring animals to yield double-transgenic 'hopper' frogs. Remobilization events were observed in the progeny of the hopper frogs and were verified by Southern blot analysis and cloning of the novel integrations sites. Unlike the co-injection method used to generate founder lines, transgenic remobilization resulted in canonical transposition of the SB transposons. The remobilized SB transposons frequently integrated near the site of the donor locus; approximately 80% re-integrated with 3 Mb of the donor locus, a phenomenon known as 'local hopping'. Conclusions In this study, we demonstrate that SB transposons integrated into the X. tropicalis genome are effective substrates for excision and re-integration, and that the remobilized transposons are transmitted through the germline. This is an important step in the development of large-scale transposon-mediated gene- and enhancer-trap strategies in this highly tractable developmental model system.

  12. Germ-line origins of mutation in families with hemophilia B: The sex ratio varies with the type of mutation

    Energy Technology Data Exchange (ETDEWEB)

    Ketterling, R.P.; Vielhaber, E.; Bottema, C.D.K.; Schaid, D.J.; Sommer, S.S. (Mayo Clinic/Foundation, Rochester, MN (United States)); Cohen, M.P. (Vanderbilt Univ., Nashville, TN (United States)); Sexauer, C.L. (Children' s Hospital, Oklahoma City, OK (United States))

    1993-01-01

    Previous epidemiological and biochemical studies have generated conflicting estimates of the sex ratio of mutation. Direct genomic sequencing in combination with haplotype analysis extends previous analyses by allowing the precise mutation to be determined in a given family. From analysis of the factor IX gene of 260 consecutive families with hemophilia B, the authors report the germ-line origin of mutation in 25 families. When combined with 14 origins of mutation reported by others and with 4 origins previously reported by them, a total of 25 occur in the female germ line, and 18 occur in the male germ line. The excess of germ-line origins in females does not imply an overall excess mutation rate per base pair in the female germ line. Bayesian analysis of the data indicates that the sex ratio varies with the type of mutation. The aggregate of single-base substitutions shows a male predominance of germ-line mutations (P < .002). The maximum-likelihood estimate of the male predominance is 3.5-fold. Of the single-base substitutions, deletions display a sex ratio of unity. Analysis of the parental age at transmission of a new mutation suggests that germ-line mutations are associated with a small increase in parental age in females but little, if any, increase in males. Although direct genomic sequencing offers a general method for defining the origin of mutation in specific families, accurate estimates of the sex ratios of different mutational classes require large sample sizes and careful correction for multiple biases of ascertainment. The biases in the present data result in an underestimate of the enhancement of mutation in males. 62 refs., 1 fig., 5 tabs.

  13. The Paramecium germline genome provides a niche for intragenic parasitic DNA: evolutionary dynamics of internal eliminated sequences.

    Science.gov (United States)

    Arnaiz, Olivier; Mathy, Nathalie; Baudry, Céline; Malinsky, Sophie; Aury, Jean-Marc; Denby Wilkes, Cyril; Garnier, Olivier; Labadie, Karine; Lauderdale, Benjamin E; Le Mouël, Anne; Marmignon, Antoine; Nowacki, Mariusz; Poulain, Julie; Prajer, Malgorzata; Wincker, Patrick; Meyer, Eric; Duharcourt, Sandra; Duret, Laurent; Bétermier, Mireille; Sperling, Linda

    2012-01-01

    Insertions of parasitic DNA within coding sequences are usually deleterious and are generally counter-selected during evolution. Thanks to nuclear dimorphism, ciliates provide unique models to study the fate of such insertions. Their germline genome undergoes extensive rearrangements during development of a new somatic macronucleus from the germline micronucleus following sexual events. In Paramecium, these rearrangements include precise excision of unique-copy Internal Eliminated Sequences (IES) from the somatic DNA, requiring the activity of a domesticated piggyBac transposase, PiggyMac. We have sequenced Paramecium tetraurelia germline DNA, establishing a genome-wide catalogue of -45,000 IESs, in order to gain insight into their evolutionary origin and excision mechanism. We obtained direct evidence that PiggyMac is required for excision of all IESs. Homology with known P. tetraurelia Tc1/mariner transposons, described here, indicates that at least a fraction of IESs derive from these elements. Most IES insertions occurred before a recent whole-genome duplication that preceded diversification of the P. aurelia species complex, but IES invasion of the Paramecium genome appears to be an ongoing process. Once inserted, IESs decay rapidly by accumulation of deletions and point substitutions. Over 90% of the IESs are shorter than 150 bp and present a remarkable size distribution with a -10 bp periodicity, corresponding to the helical repeat of double-stranded DNA and suggesting DNA loop formation during assembly of a transpososome-like excision complex. IESs are equally frequent within and between coding sequences; however, excision is not 100% efficient and there is selective pressure against IES insertions, in particular within highly expressed genes. We discuss the possibility that ancient domestication of a piggyBac transposase favored subsequent propagation of transposons throughout the germline by allowing insertions in coding sequences, a fraction of the

  14. The Paramecium germline genome provides a niche for intragenic parasitic DNA: evolutionary dynamics of internal eliminated sequences.

    Directory of Open Access Journals (Sweden)

    Olivier Arnaiz

    Full Text Available Insertions of parasitic DNA within coding sequences are usually deleterious and are generally counter-selected during evolution. Thanks to nuclear dimorphism, ciliates provide unique models to study the fate of such insertions. Their germline genome undergoes extensive rearrangements during development of a new somatic macronucleus from the germline micronucleus following sexual events. In Paramecium, these rearrangements include precise excision of unique-copy Internal Eliminated Sequences (IES from the somatic DNA, requiring the activity of a domesticated piggyBac transposase, PiggyMac. We have sequenced Paramecium tetraurelia germline DNA, establishing a genome-wide catalogue of -45,000 IESs, in order to gain insight into their evolutionary origin and excision mechanism. We obtained direct evidence that PiggyMac is required for excision of all IESs. Homology with known P. tetraurelia Tc1/mariner transposons, described here, indicates that at least a fraction of IESs derive from these elements. Most IES insertions occurred before a recent whole-genome duplication that preceded diversification of the P. aurelia species complex, but IES invasion of the Paramecium genome appears to be an ongoing process. Once inserted, IESs decay rapidly by accumulation of deletions and point substitutions. Over 90% of the IESs are shorter than 150 bp and present a remarkable size distribution with a -10 bp periodicity, corresponding to the helical repeat of double-stranded DNA and suggesting DNA loop formation during assembly of a transpososome-like excision complex. IESs are equally frequent within and between coding sequences; however, excision is not 100% efficient and there is selective pressure against IES insertions, in particular within highly expressed genes. We discuss the possibility that ancient domestication of a piggyBac transposase favored subsequent propagation of transposons throughout the germline by allowing insertions in coding sequences, a

  15. Two co-existing germline mutations P53 V157D and PMS2 R20Q promote tumorigenesis in a familial cancer syndrome.

    Science.gov (United States)

    Wang, Zuoyun; Sun, Yihua; Gao, Bin; Lu, Yi; Fang, Rong; Gao, Yijun; Xiao, Tian; Liu, Xin-Yuan; Pao, William; Zhao, Yun; Chen, Haiquan; Ji, Hongbin

    2014-01-01

    Germline mutations are responsible for familial cancer syndromes which account for approximately 5-10% of all types of cancers. These mutations mainly occur at tumor suppressor genes or genome stability genes, such as DNA repair genes. Here we have identified a cancer predisposition family, in which eight members were inflicted with a wide spectrum of cancer including one diagnosed with lung cancer at 22years old. Sequencing analysis of tumor samples as well as histologically normal specimens identified two germline mutations co-existing in the familial cancer syndrome, the mutation of tumor suppressor gene P53 V157D and mismatch repair gene PMS2 R20Q. We further demonstrate that P53 V157D and/or PMS2 R20Q mutant promotes lung cancer cell proliferation. These two mutants are capable of promoting colony formation in soft agar as well as tumor formation in transgenic drosophila system. Collectively, these data have uncovered the important role of co-existing germline P53 and PMS2 mutations in the familial cancer syndrome development. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  16. Development of germline manipulation technologies in livestock

    International Nuclear Information System (INIS)

    Whitelaw, C.B.A.

    2005-01-01

    Genetic improvement by conventional breeding is restricted to those genetic loci present in the parental breeding individuals. Gene addition through transgenic technology offers a route to overcome this restriction. The transgene can be introduced into the germ cells or the fertilized zygote, using viral vectors, by simple co-culture or direct micro-injection. Alternatively, the transgene can be incorporated into a somatic cell, which is then incorporated into a developing embryo. This latter approach allows gene-targeting strategies to be employed. Using pronuclear injection methods, transgenic livestock have been generated with the aim of enhancing breeding traits of agricultural importance, or for biomedical applications. Neither has been taken beyond the development phase. Before they are, in addition to issues of commercial development, basic technological issues addressing inefficiency and complexity of the methodology need to be overcome, and appropriate gene targets identified. At the moment, perhaps the most encouraging development involves the use of viral vectors that offer increased simplicity and efficiency. By combining this new technology with transgenes that evoke the powerful intracellular machinery involved in RNA interference, pioneering applications to generate animals that are less susceptible to infectious disease may be possible. (author)

  17. Gene Expression Architecture of Mouse Dorsal and Tail Skin Reveals Functional Differences in Inflammation and Cancer | Office of Cancer Genomics

    Science.gov (United States)

    Inherited germline polymorphisms can cause gene expression levels in normal tissues to differ substantially between individuals. We present an analysis of the genetic architecture of normal adult skin from 470 genetically unique mice, demonstrating the effect of germline variants, skin tissue location, and perturbation by exogenous inflammation or tumorigenesis on gene signaling pathways.

  18. Germline Genetic Modification and Identity: the Mitochondrial and Nuclear Genomes.

    Science.gov (United States)

    Scott, Rosamund; Wilkinson, Stephen

    2017-12-01

    In a legal 'first', the UK removed a prohibition against modifying embryos in human reproduction, to enable mitochondrial replacement techniques (MRTs), a move the Government distanced from 'germline genetic modification', which it aligned with modifying the nuclear genome. This paper (1) analyzes the uses and meanings of this term in UK/US legal and policy debates; and (2) evaluates related ethical concerns about identity. It shows that, with respect to identity, MRTs and nuclear genome editing techniques such as CRISPR/Cas-9 (now a policy topic), are not as different as has been supposed. While it does not follow that the two should be treated exactly alike, one of the central reasons offered for treating MRTs more permissively than nuclear genetic modification, and for not regarding MRTs as 'germline genetic modification', is thereby in doubt. Identity cannot, by itself, do the work thus far assigned to it, explicitly or otherwise, in law and policy.

  19. Familial adenomatous polyposis patients without an identified APC germline mutation have a severe phenotype

    DEFF Research Database (Denmark)

    Bisgaard, M L; Ripa, R; Knudsen, Anne Louise

    2004-01-01

    BACKGROUND: Development of more than 100 colorectal adenomas is diagnostic of the dominantly inherited autosomal disease familial adenomatous polyposis (FAP). Germline mutations can be identified in the adenomatous polyposis coli (APC) gene in approximately 80% of patients. The APC protein...... comprises several regions and domains for interaction with other proteins, and specific clinical manifestations are associated with the mutation assignment to one of these regions or domains. AIMS: The phenotype in patients without an identified causative APC mutation was compared with the phenotype...... in patients with a known APC mutation and with the phenotypes characteristic of patients with mutations in specific APC regions and domains. PATIENTS: Data on 121 FAP probands and 149 call up patients from 70 different families were extracted from the Danish Polyposis register. METHODS: Differences in 16...

  20. Association Between Germline Mutation in VSIG10L and Familial Barrett Neoplasia

    Science.gov (United States)

    Fecteau, Ryan E.; Kong, Jianping; Kresak, Adam; Brock, Wendy; Song, Yeunjoo; Fujioka, Hisashi; Elston, Robert; Willis, Joseph E.; Lynch, John P.; Markowitz, Sanford D.; Guda, Kishore; Chak, Amitabh

    2016-01-01

    IMPORTANCE Esophageal adenocarcinoma and its precursor lesion Barrett esophagus have seen a dramatic increase in incidence over the past 4 decades yet marked genetic heterogeneity of this disease has precluded advances in understanding its pathogenesis and improving treatment. OBJECTIVE To identify novel disease susceptibility variants in a familial syndrome of esophageal adenocarcinoma and Barrett esophagus, termed familial Barrett esophagus, by using high-throughput sequencing in affected individuals from a large, multigenerational family. DESIGN, SETTING, AND PARTICIPANTS We performed whole exome sequencing (WES) from peripheral lymphocyte DNA on 4 distant relatives from our multiplex, multigenerational familial Barrett esophagus family to identify candidate disease susceptibility variants. Gene variants were filtered, verified, and segregation analysis performed to identify a single candidate variant. Gene expression analysis was done with both quantitative real-time polymerase chain reaction and in situ RNA hybridization. A 3-dimensional organotypic cell culture model of esophageal maturation was utilized to determine the phenotypic effects of our gene variant. We used electron microscopy on esophageal mucosa from an affected family member carrying the gene variant to assess ultrastructural changes. MAIN OUTCOMES AND MEASURES Identification of a novel, germline disease susceptibility variant in a previously uncharacterized gene. RESULTS A multiplex, multigenerational family with 14 members affected (3 members with esophageal adenocarcinoma and 11 with Barrett esophagus) was identified, and whole-exome sequencing identified a germline mutation (S631G) at a highly conserved serine residue in the uncharacterized gene VSIG10L that segregated in affected members. Transfection of S631G variant into a 3-dimensional organotypic culture model of normal esophageal squamous cells dramatically inhibited epithelial maturation compared with the wild-type. VSIG10L exhibited

  1. Association Between Germline Mutation in VSIG10L and Familial Barrett Neoplasia.

    Science.gov (United States)

    Fecteau, Ryan E; Kong, Jianping; Kresak, Adam; Brock, Wendy; Song, Yeunjoo; Fujioka, Hisashi; Elston, Robert; Willis, Joseph E; Lynch, John P; Markowitz, Sanford D; Guda, Kishore; Chak, Amitabh

    2016-10-01

    Esophageal adenocarcinoma and its precursor lesion Barrett esophagus have seen a dramatic increase in incidence over the past 4 decades yet marked genetic heterogeneity of this disease has precluded advances in understanding its pathogenesis and improving treatment. To identify novel disease susceptibility variants in a familial syndrome of esophageal adenocarcinoma and Barrett esophagus, termed familial Barrett esophagus, by using high-throughput sequencing in affected individuals from a large, multigenerational family. We performed whole exome sequencing (WES) from peripheral lymphocyte DNA on 4 distant relatives from our multiplex, multigenerational familial Barrett esophagus family to identify candidate disease susceptibility variants. Gene variants were filtered, verified, and segregation analysis performed to identify a single candidate variant. Gene expression analysis was done with both quantitative real-time polymerase chain reaction and in situ RNA hybridization. A 3-dimensional organotypic cell culture model of esophageal maturation was utilized to determine the phenotypic effects of our gene variant. We used electron microscopy on esophageal mucosa from an affected family member carrying the gene variant to assess ultrastructural changes. Identification of a novel, germline disease susceptibility variant in a previously uncharacterized gene. A multiplex, multigenerational family with 14 members affected (3 members with esophageal adenocarcinoma and 11 with Barrett esophagus) was identified, and whole-exome sequencing identified a germline mutation (S631G) at a highly conserved serine residue in the uncharacterized gene VSIG10L that segregated in affected members. Transfection of S631G variant into a 3-dimensional organotypic culture model of normal esophageal squamous cells dramatically inhibited epithelial maturation compared with the wild-type. VSIG10L exhibited high expression in normal squamous esophagus with marked loss of expression in Barrett

  2. Germline MLH1, MSH2 and MSH6 variants in Brazilian patients with colorectal cancer and clinical features suggestive of Lynch Syndrome.

    Science.gov (United States)

    Schneider, Nayê Balzan; Pastor, Tatiane; Paula, André Escremim de; Achatz, Maria Isabel; Santos, Ândrea Ribeiro Dos; Vianna, Fernanda Sales Luiz; Rosset, Clévia; Pinheiro, Manuela; Ashton-Prolla, Patricia; Moreira, Miguel Ângelo Martins; Palmero, Edenir Inêz

    2018-05-01

    Lynch syndrome (LS) is the most common hereditary colorectal cancer syndrome, caused by germline mutations in one of the major genes involved in mismatch repair (MMR): MLH1, MSH2, MSH6 and more rarely, PMS2. Recently, germline deletions in EPCAM have been also associated to the syndrome. Most of the pathogenic MMR mutations found in LS families occur in MLH1 or MSH2. Gene variants include missense, nonsense, frameshift mutations, large genomic rearrangements and splice-site variants and most of the studies reporting the molecular characterization of LS families have been conducted outside South America. In this study, we analyzed 60 unrelated probands diagnosed with colorectal cancer and LS criteria. Testing for germline mutations and/or rearrangements in the most commonly affected MMR genes (MLH1, MSH2, EPCAM and MSH6) was done by Sanger sequencing and MLPA. Pathogenic or likely pathogenic variants were identified in MLH1 or MSH2 in 21 probands (35.0%). Of these, approximately one-third were gene rearrangements. In addition, nine variants of uncertain significance (VUS) were identified in 10 (16.6%) of the sixty probands analyzed. Other four novel variants were identified, only in MLH1. Our results suggest that MSH6 pathogenic variants are not common among Brazilian LS probands diagnosed with CRC and that MMR gene rearrangements account for a significant proportion of the germline variants in this population underscoring the need to include rearrangement analysis in the molecular testing of Brazilian individuals with suspected Lynch syndrome. © 2018 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  3. In Vitro Cytotoxicity of Nanoparticles in Mammalian Germline Stem Cells

    OpenAIRE

    Braydich-Stolle, Laura; Hussain, Saber; Schlager, John J.; Hofmann, Marie-Claude

    2005-01-01

    Gametogenesis is a complex biological process that is particularly sensitive to environmental insults such as chemicals. Many chemicals have a negative impact on the germline, either by directly affecting the germ cells, or indirectly through their action on the somatic nursing cells. Ultimately, these effects can inhibit fertility, and they may have negative consequences for the development of the offspring. Recently, nanomaterials such as nanotubes, nanowires, fullerene derivatives (buckyba...

  4. Male germline stem cells in non-human primates

    Directory of Open Access Journals (Sweden)

    S. Sharma

    2017-09-01

    Full Text Available Over the past few decades, several studies have attempted to decipher the biology of mammalian germline stem cells (GSCs. These studies provide evidence that regulatory mechanisms for germ cell specification and migration are evolutionarily conserved across species. The characteristics and functions of primate GSCs are highly distinct from rodent species; therefore the findings from rodent models cannot be extrapolated to primates. Due to limited availability of human embryonic and testicular samples for research purposes, two non-human primate models (marmoset and macaque monkeys are extensively employed to understand human germline development and differentiation. This review provides a broader introduction to the in vivo and in vitro germline stem cell terminology from primordial to differentiating germ cells. Primordial germ cells (PGCs are the most immature germ cells colonizing the gonad prior to sex differentiation into testes or ovaries. PGC specification and migratory patterns among different primate species are compared in the review. It also reports the distinctions and similarities in expression patterns of pluripotency markers (OCT4A, NANOG, SALL4 and LIN28 during embryonic developmental stages, among marmosets, macaques and humans. This review presents a comparative summary with immunohistochemical and molecular evidence of germ cell marker expression patterns during postnatal developmental stages, among humans and non-human primates. Furthermore, it reports findings from the recent literature investigating the plasticity behavior of germ cells and stem cells in other organs of humans and monkeys. The use of non-human primate models would enable bridging the knowledge gap in primate GSC research and understanding the mechanisms involved in germline development. Reported similarities in regulatory mechanisms and germ cell expression profile in primates demonstrate the preclinical significance of monkey models for development of

  5. The roles of DNA damage-dependent signals and MAPK cascades in tributyltin-induced germline apoptosis in Caenorhabditis elegans.

    Science.gov (United States)

    Wang, Yun; Wang, Shunchang; Luo, Xun; Yang, Yanan; Jian, Fenglei; Wang, Xuemin; Xie, Lucheng

    2014-08-01

    The induction of apoptosis is recognized to be a major mechanism of tributyltin (TBT) toxicity. However, the underlying signaling pathways for TBT-induced apoptosis remain unclear. In this study, using the nematode Caenorhabditis elegans, we examined whether DNA damage response (DDR) pathway and mitogen-activated protein kinase (MAPK) signaling cascades are involved in TBT-induced germline apoptosis and cell cycle arrest. Our results demonstrated that exposing worms to TBT at the dose of 10nM for 6h significantly increased germline apoptosis in N2 strain. Germline apoptosis was absent in strains that carried ced-3 or ced-4 loss-of-function alleles, indicating that both caspase protein CED-3 and Apaf-1 protein CED-4 were required for TBT-induced apoptosis. TBT-induced apoptosis was blocked in the Bcl-2 gain-of-function strain ced-9(n1950), whereas TBT induced a minor increase in the BH3-only protein EGL-1 mutated strain egl-1(n1084n3082). Checkpoint proteins HUS-1 and CLK-2 exerted proapoptotic effects, and the null mutation of cep-1, the homologue of tumor suppressor gene p53, significantly inhibited TBT-induced apoptosis. Apoptosis in the loss-of-function strains of ERK, JNK and p38 MAPK signaling pathways were completely or mildly suppressed under TBT stress. These results were supported by the results of mRNA expression levels of corresponding genes. The present study indicated that TBT-induced apoptosis required the core apoptotic machinery, and that DDR genes and MAPK pathways played essential roles in signaling the processes. Copyright © 2014 Elsevier Ltd. All rights reserved.

  6. Germline genetic variants in the Wnt/beta-catenin pathway as predictors of colorectal cancer risk

    Science.gov (United States)

    Hildebrandt, Michelle A.T.; Reyes, Monica E.; Lin, Moubin; He, Yonggang; Nguyen, Son V.; Hawk, Ernest T.; Wu, Xifeng

    2016-01-01

    Background The Wnt/beta-catenin signaling pathway plays a key role in stem cell maintenance in the colorectum. Rare high penetrance genetic mutations in components of this pathway result in familial colorectal cancer, yet the impact of common, germline variants remains unknown. Methods We assessed 172 variants in 26 genes from the Wnt/beta-catenin pathway in 809 CRC cases and 814 healthy controls, followed by replication of the top findings in another 691 cases and 775 controls. In silico informatic tools were used to predict functional effects of variants. Results Eighteen SNPs in the pathway were significantly associated with CRC risk (P <0.05) in the discovery phase. We observed a significant dose-response increase in CRC risk by number of risk genotypes carried (P = 4.19 × 10−8). Gene-based analysis implicated CSNK1D (P = 0.014), FZD3 (P = 0.023), and APC (P = 0.027) as significant for CRC risk. In the replication phase, FZD3:rs11775139 remained significantly associated with reduced risk with a pooled OR of 0.85 (95% CI: 0.76–0.94, P = 0.001). Although borderline significant in the replication population, APC:rs2545162 was highly significant in the pooled analysis - OR: 1.42, 95% CI: 1.16–1.74, P =0.00085. Functional assessment identified several potential biological mechanisms underlying these associations. Conclusions Our findings suggest that common germline variants in the Wnt/beta-catenin pathway maybe involved in CRC development. Impact These variants may be informative in CRC risk assessment to identify individuals at increased risk who would be candidates for screening. PMID:26809274

  7. Pharmacogenetic characterization of naturally occurring germline NT5C1A variants to chemotherapeutic nucleoside analogs

    Science.gov (United States)

    Saliba, Jason; Zabriskie, Ryan; Ghosh, Rajarshi; Powell, Bradford C; Hicks, Stephanie; Kimmel, Marek; Meng, Qingchang; Ritter, Deborah I; Wheeler, David A; Gibbs, Richard A; Tsai, Francis T F; Plon, Sharon E

    2016-01-01

    Background Mutations or alteration in expression of the 5’ nucleotidase gene family can confer altered responses to treatment with nucleoside analogs. While investigating leukemia susceptibility genes, we discovered a very rare p.L254P NT5C1A missense variant in the substrate recognition motif. Given the paucity of cellular drug response data from NT5C1A germline variation, we characterized p.L254P and eight rare variants of NT5C1A from genomic databases. Methods Through lentiviral infection, we created HEK293 cell lines that stably overexpress wildtype NT5C1A, p.L254P, or eight NT5C1A variants reported in the NHLBI Exome Variant server (one truncating and seven missense). IC50 values were determined by cytotoxicity assays after exposure to chemotherapeutic nucleoside analogs (Cladribine, Gemcitabine, 5-Fluorouracil). In addition, we used structure-based homology modeling to generate a 3D model for the C-terminal region of NT5C1A. Results The p.R180X (truncating), p.A214T, and p.L254P missense changes were the only variants that significantly impaired protein function across all nucleotide analogs tested (>5-fold difference versus WT; p<.05). Several of the remaining variants individually displayed differential effects (both more and less resistant) across the analogs tested. The homology model provided a structural framework to understand the impact of NT5C1A mutants on catalysis and drug processing. The model predicted active site residues within NT5C1A motif III and we experimentally confirmed that p.K314 (not p.K320) is required for NT5C1A activity. Conclusion We characterized germline variation and predicted protein structures of NT5C1A. Individual missense changes showed substantial variation in response to the different nucleoside analogs tested, which may impact patients’ responses to treatment. PMID:26906009

  8. Cell lineage analysis of the mammalian female germline.

    Directory of Open Access Journals (Sweden)

    Yitzhak Reizel

    Full Text Available Fundamental aspects of embryonic and post-natal development, including maintenance of the mammalian female germline, are largely unknown. Here we employ a retrospective, phylogenetic-based method for reconstructing cell lineage trees utilizing somatic mutations accumulated in microsatellites, to study female germline dynamics in mice. Reconstructed cell lineage trees can be used to estimate lineage relationships between different cell types, as well as cell depth (number of cell divisions since the zygote. We show that, in the reconstructed mouse cell lineage trees, oocytes form clusters that are separate from hematopoietic and mesenchymal stem cells, both in young and old mice, indicating that these populations belong to distinct lineages. Furthermore, while cumulus cells sampled from different ovarian follicles are distinctly clustered on the reconstructed trees, oocytes from the left and right ovaries are not, suggesting a mixing of their progenitor pools. We also observed an increase in oocyte depth with mouse age, which can be explained either by depth-guided selection of oocytes for ovulation or by post-natal renewal. Overall, our study sheds light on substantial novel aspects of female germline preservation and development.

  9. Detection and precise mapping of germline rearrangements in BRCA1, BRCA2, MSH2, and MLH1 using zoom-in array comparative genomic hybridization (aCGH)

    DEFF Research Database (Denmark)

    Staaf, Johan; Törngren, Therese; Rambech, Eva

    2008-01-01

    Disease-predisposing germline mutations in cancer susceptibility genes may consist of large genomic rearrangements that are challenging to detect and characterize using standard PCR-based mutation screening methods. Here, we describe a custom-made zoom-in microarray comparative genomic hybridizat......Disease-predisposing germline mutations in cancer susceptibility genes may consist of large genomic rearrangements that are challenging to detect and characterize using standard PCR-based mutation screening methods. Here, we describe a custom-made zoom-in microarray comparative genomic...... deletions or duplications occurring in BRCA1 (n=11), BRCA2 (n=2), MSH2 (n=7), or MLH1 (n=9). Additionally, we demonstrate its applicability for uncovering complex somatic rearrangements, exemplified by zoom-in analysis of the PTEN and CDKN2A loci in breast cancer cells. The sizes of rearrangements ranged...... from several 100 kb, including large flanking regions, to rearrangements, allowing convenient design...

  10. Probing the germline-dependence of epigenetic inheritance using artificial insemination in mice

    Science.gov (United States)

    Bohacek, Johannes; von Werdt, Sarah; Mansuy, Isabelle M.

    2016-01-01

    Abstract We developed a simple, noninvasive artificial insemination technique to study epigenetic germline inheritance in mice. This technique avoids interfering factors introduced by superovulation, surgery, in vitro culture or mating that can confound the transmission of acquired epigenetic information through the germline. Using a stress model, we demonstrate that our method is suited to test the causal involvement of the male germline in transmitting acquired information from father to offspring. PMID:29492284

  11. Genome engineering through CRISPR/Cas9 technology in the human germline and pluripotent stem cells.

    Science.gov (United States)

    Vassena, R; Heindryckx, B; Peco, R; Pennings, G; Raya, A; Sermon, K; Veiga, A

    2016-06-01

    With the recent development of CRISPR (clustered regularly interspaced short palindromic repeats)/Cas9 genome editing technology, the possibility to genetically manipulate the human germline (gametes and embryos) has become a distinct technical possibility. Although many technical challenges still need to be overcome in order to achieve adequate efficiency and precision of the technology in human embryos, the path leading to genome editing has never been simpler, more affordable, and widespread. In this narrative review we seek to understand the possible impact of CRISR/Cas9 technology on human reproduction from the technical and ethical point of view, and suggest a course of action for the scientific community. This non-systematic review was carried out using Medline articles in English, as well as technical documents from the Human Fertilisation and Embryology Authority and reports in the media. The technical possibilities of the CRISPR/Cas9 technology with regard to human reproduction are analysed based on results obtained in model systems such as large animals and laboratory rodents. Further, the possibility of CRISPR/Cas9 use in the context of human reproduction, to modify embryos, germline cells, and pluripotent stem cells is reviewed based on the authors' expert opinion. Finally, the possible uses and consequences of CRISPR/cas9 gene editing in reproduction are analysed from the ethical point of view. We identify critical technical and ethical issues that should deter from employing CRISPR/Cas9 based technologies in human reproduction until they are clarified. Overcoming the numerous technical limitations currently associated with CRISPR/Cas9 mediated editing of the human germline will depend on intensive research that needs to be transparent and widely disseminated. Rather than a call to a generalized moratorium, or banning, of this type of research, efforts should be placed on establishing an open, international, collaborative and regulated research

  12. Prospective Genomic Profiling of Prostate Cancer Across Disease States Reveals Germline and Somatic Alterations That May Affect Clinical Decision Making.

    Science.gov (United States)

    Abida, Wassim; Armenia, Joshua; Gopalan, Anuradha; Brennan, Ryan; Walsh, Michael; Barron, David; Danila, Daniel; Rathkopf, Dana; Morris, Michael; Slovin, Susan; McLaughlin, Brigit; Curtis, Kristen; Hyman, David M; Durack, Jeremy C; Solomon, Stephen B; Arcila, Maria E; Zehir, Ahmet; Syed, Aijazuddin; Gao, Jianjiong; Chakravarty, Debyani; Vargas, Hebert Alberto; Robson, Mark E; Joseph, Vijai; Offit, Kenneth; Donoghue, Mark T A; Abeshouse, Adam A; Kundra, Ritika; Heins, Zachary J; Penson, Alexander V; Harris, Christopher; Taylor, Barry S; Ladanyi, Marc; Mandelker, Diana; Zhang, Liying; Reuter, Victor E; Kantoff, Philip W; Solit, David B; Berger, Michael F; Sawyers, Charles L; Schultz, Nikolaus; Scher, Howard I

    2017-07-01

    A long natural history and a predominant osseous pattern of metastatic spread are impediments to the adoption of precision medicine in patients with prostate cancer. To establish the feasibility of clinical genomic profiling in the disease, we performed targeted deep sequencing of tumor and normal DNA from patients with locoregional, metastatic non-castrate, and metastatic castration-resistant prostate cancer (CRPC). Patients consented to genomic analysis of their tumor and germline DNA. A hybridization capture-based clinical assay was employed to identify single nucleotide variations, small insertions and deletions, copy number alterations and structural rearrangements in over 300 cancer-related genes in tumors and matched normal blood. We successfully sequenced 504 tumors from 451 patients with prostate cancer. Potentially actionable alterations were identified in DNA damage repair (DDR), PI3K, and MAP kinase pathways. 27% of patients harbored a germline or a somatic alteration in a DDR gene that may predict for response to PARP inhibition. Profiling of matched tumors from individual patients revealed that somatic TP53 and BRCA2 alterations arose early in tumors from patients who eventually developed metastatic disease. In contrast, comparative analysis across disease states revealed that APC alterations were enriched in metastatic tumors, while ATM alterations were specifically enriched in CRPC. Through genomic profiling of prostate tumors representing the disease clinical spectrum, we identified a high frequency of potentially actionable alterations and possible drivers of disease initiation, metastasis and castration-resistance. Our findings support the routine use of tumor and germline DNA profiling for patients with advanced prostate cancer, for the purpose of guiding enrollment in targeted clinical trials and counseling families at increased risk of malignancy.

  13. A role of BRCA1 and BRCA2 germline mutations in breast cancer susceptibility within Sardinian population

    International Nuclear Information System (INIS)

    Palomba, Grazia; Tanda, Francesco; Farris, Antonio; Orrù, Sandra; Floris, Carlo; Pisano, Marina; Lovicu, Mario; Santona, Maria Cristina; Landriscina, Gennaro; Crisponi, Laura; Palmieri, Giuseppe; Loi, Angela; Monne, Maria; Uras, Antonella; Fancello, Patrizia; Piras, Giovanna; Gabbas, Attilio; Cossu, Antonio; Budroni, Mario; Contu, Antonio

    2009-01-01

    In recent years, numerous studies have assessed the prevalence of germline mutations in BRCA1 and BRCA2 genes in various cohorts. We here extensively investigated the prevalence and geographical distribution of BRCA1-2 mutations in the entire genetically-homogeneous Sardinian population. The occurrence of phenotypic characteristics which may be predictive for the presence of BRCA1-2 germline mutations was also evaluated. Three hundred and forty-eight breast cancer patients presenting a familial recurrence of invasive breast or ovarian carcinoma with at least two affected family members were screened for BRCA1-2 mutations by DHPLC analysis and DNA sequencing. Association of BRCA1 and BRCA2 mutational status with clinical and pathological parameters was evaluated by Pearson's Chi-Squared test. Overall, 8 BRCA1 and 5 BRCA2 deleterious mutations were detected in 35/348 (10%) families; majority (23/35;66%) of mutations was found in BRCA2 gene. The geographical distribution of BRCA1-2 mutations was related to three specific large areas of Sardinia, reflecting its ancient history: a) the Northern area, linguistically different from the rest of the island (where a BRCA2 c.8764-8765delAG mutation with founder effect was predominant); b) the Middle area, land of the ancient Sardinian population (where BRCA2 mutations are still more common than BRCA1 mutations); and c) the South-Western area, with many Phoenician and Carthaginian locations (where BRCA1 mutations are prevalent). We also found that phenotypic features such as high tumor grading and lack of expression of estrogen/progesterone receptors together with age at diagnosis and presence of ovarian cancer in the family may be predictive for the presence of BRCA1-2 germline mutations

  14. Germline disruption of Pten localization causes enhanced sex-dependent social motivation and increased glial production.

    Science.gov (United States)

    Tilot, Amanda K; Gaugler, Mary K; Yu, Qi; Romigh, Todd; Yu, Wanfeng; Miller, Robert H; Frazier, Thomas W; Eng, Charis

    2014-06-15

    PTEN Hamartoma Tumor Syndrome (PHTS) is an autosomal-dominant genetic condition underlying a subset of autism spectrum disorder (ASD) with macrocephaly. Caused by germline mutations in PTEN, PHTS also causes increased risks of multiple cancers via dysregulation of the PI3K and MAPK signaling pathways. Conditional knockout models have shown that neural Pten regulates social behavior, proliferation and cell size. Although much is known about how the intracellular localization of PTEN regulates signaling in cancer cell lines, we know little of how PTEN localization influences normal brain physiology and behavior. To address this, we generated a germline knock-in mouse model of cytoplasm-predominant Pten and characterized its behavioral and cellular phenotypes. The homozygous Pten(m3m4) mice have decreased total Pten levels including a specific drop in nuclear Pten and exhibit region-specific increases in brain weight. The Pten(m3m4) model displays sex-specific increases in social motivation, poor balance and normal recognition memory-a profile reminiscent of some individuals with high functioning ASD. The cytoplasm-predominant protein caused cellular hypertrophy limited to the soma and led to increased NG2 cell proliferation and accumulation of glia. The animals also exhibit significant astrogliosis and microglial activation, indicating a neuroinflammatory phenotype. At the signaling level, Pten(m3m4) mice show brain region-specific differences in Akt activation. These results demonstrate that differing alterations to the same autism-linked gene can cause distinct behavioral profiles. The Pten(m3m4) model is the first murine model of inappropriately elevated social motivation in the context of normal cognition and may expand the range of autism-related behaviors replicated in animal models. © The Author 2014. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  15. Effect of ATM heterozygosity on heritable DNA damage in mice following paternal F0 germline irradiation

    International Nuclear Information System (INIS)

    Baulch, Janet E.; Li, M.-W.; Raabe, Otto G.

    2007-01-01

    The ataxia telangiectasia mutated (ATM) gene product maintains genome integrity and initiates cellular DNA repair pathways following exposures to genotoxic agents. ATM also plays a significant role in meiotic recombination during spermatogenesis. Fertilization with sperm carrying damaged DNA could lead to adverse effects in offspring including developmental defects or increased cancer susceptibility. Currently, there is little information regarding the effect of ATM heterozygosity on germline DNA repair and heritable effects of paternal germline-ionizing irradiation. We used neutral pH comet assays to evaluate spermatozoa 45 days after acute whole-body irradiation of male mice (0.1 Gy, attenuated 137 Cs γ rays) to determine the effect of ATM heterozygosity on delayed DNA damage effects of Type A/B spermatogonial irradiation. Using the neutral pH sperm comet assay, significant irradiation-related differences were found in comet tail length, percent tail DNA and tail extent moment, but there were no observed differences in effect between wild-type and ATM +/- mice. However, evaluation of spermatozoa from third generation descendants of irradiated male mice for heritable chromatin effects revealed significant differences in DNA electrophoretic mobility in the F 3 descendants that were based upon the irradiated F 0 sire's genotype. In this study, radiation-induced chromatin alterations to Type A/B spermatogonia, detected in mature sperm 45 days post-irradiation, led to chromatin effects in mature sperm three generations later. The early cellular response to and repair of DNA damage is critical and appears to be affected by ATM zygosity. Our results indicate that there is potential for heritable genetic or epigenetic changes following Type A/B spermatogonial irradiation and that ATM heterozygosity increases this effect

  16. Partial pleural covering for intractable pneumothorax in patients with Birt-Hogg-Dubé Syndrome.

    Science.gov (United States)

    Okada, Akira; Hirono, Tatsuhiko; Watanabe, Takehiro; Hasegawa, Go; Tanaka, Reiko; Furuya, Mitsuko

    2017-03-01

    Birt-Hogg-Dubé syndrome (BHD) is an inherited disorder associated with a germline mutation of the folliculin (FLCN) gene. Most patients with BHD have multiple pulmonary cysts, and are at high risk of repeated pneumothorax. Although an increasing number of patients are diagnosed with BHD by genetic testing, therapeutic approaches for intractable pneumothorax have not yet been described. We treated three patients who had repeated episodes of pneumothorax. All had multiple pulmonary cysts in the lower lobes, and two had a family history of pneumothorax. Video-assisted thoracic surgery was used to perform wedge resections and partial pleural covering of the cystic lesions. The partial pleural covering technique used sheets of polyglycolic acid felt or regenerative oxidized cellulose mesh. The resected tissues underwent histopathological evaluation, and peripheral blood leukocytes were tested for FLCN mutations. The operative times were less than 2 h, and there were no complications. The resected cysts had histopathological features characteristic of BHD lung. All patients were found to have FLCN germline mutations; thus their repeated pneumothoraces were a manifestation of BHD. None of the patients developed respiratory problems after undergoing the partial pleural covering procedure, and they have all been well without pneumothorax for 30 months or more. Partial pleural covering combined with resection of protruding cysts should be a safe and effective therapeutic approach for BHD patients with intractable pneumothorax. Further investigation is needed to establish a detailed protocol for treatment of pneumothorax that results in minimal functional impairment. © 2015 John Wiley & Sons Ltd.

  17. Selecting one of several mating types through gene segment joining and deletion in Tetrahymena thermophila.

    Directory of Open Access Journals (Sweden)

    Marcella D Cervantes

    Full Text Available The unicellular eukaryote Tetrahymena thermophila has seven mating types. Cells can mate only when they recognize cells of a different mating type as non-self. As a ciliate, Tetrahymena separates its germline and soma into two nuclei. During growth the somatic nucleus is responsible for all gene transcription while the germline nucleus remains silent. During mating, a new somatic nucleus is differentiated from a germline nucleus and mating type is decided by a stochastic process. We report here that the somatic mating type locus contains a pair of genes arranged head-to-head. Each gene encodes a mating type-specific segment and a transmembrane domain that is shared by all mating types. Somatic gene knockouts showed both genes are required for efficient non-self recognition and successful mating, as assessed by pair formation and progeny production. The germline mating type locus consists of a tandem array of incomplete gene pairs representing each potential mating type. During mating, a complete new gene pair is assembled at the somatic mating type locus; the incomplete genes of one gene pair are completed by joining to gene segments at each end of germline array. All other germline gene pairs are deleted in the process. These programmed DNA rearrangements make this a fascinating system of mating type determination.

  18. Parallel germline infiltration of a lentivirus in two Malagasy lemurs.

    Directory of Open Access Journals (Sweden)

    Clément Gilbert

    2009-03-01

    Full Text Available Retroviruses normally infect the somatic cells of their host and are transmitted horizontally, i.e., in an exogenous way. Occasionally, however, some retroviruses can also infect and integrate into the genome of germ cells, which may allow for their vertical inheritance and fixation in a given species; a process known as endogenization. Lentiviruses, a group of mammalian retroviruses that includes HIV, are known to infect primates, ruminants, horses, and cats. Unlike many other retroviruses, these viruses have not been demonstrably successful at germline infiltration. Here, we report on the discovery of endogenous lentiviral insertions in seven species of Malagasy lemurs from two different genera -- Cheirogaleus and Microcebus. Combining molecular clock analyses and cross-species screening of orthologous insertions, we show that the presence of this endogenous lentivirus in six species of Microcebus is the result of one endogenization event that occurred about 4.2 million years ago. In addition, we demonstrate that this lentivirus independently infiltrated the germline of Cheirogaleus and that the two endogenization events occurred quasi-simultaneously. Using multiple proviral copies, we derive and characterize an apparently full length and intact consensus for this lentivirus. These results provide evidence that lentiviruses have repeatedly infiltrated the germline of prosimian species and that primates have been exposed to lentiviruses for a much longer time than what can be inferred based on sequence comparison of circulating lentiviruses. The study sets the stage for an unprecedented opportunity to reconstruct an ancestral primate lentivirus and thereby advance our knowledge of host-virus interactions.

  19. Identification of ALK germline mutation (3605delG) in pediatric anaplastic medulloblastoma.

    Science.gov (United States)

    Coco, Simona; De Mariano, Marilena; Valdora, Francesca; Servidei, Tiziana; Ridola, Vita; Andolfo, Immacolata; Oberthuer, André; Tonini, Gian Paolo; Longo, Luca

    2012-10-01

    The anaplastic lymphoma kinase (ALK) gene has been found either rearranged or mutated in several neoplasms such as anaplastic large-cell lymphoma, non-small-cell lung cancer, neuroblastoma and anaplastic thyroid cancer. Medulloblastoma (MB) is an embryonic pediatric cancer arising from nervous system, a tissue in which ALK is expressed during embryonic development. We performed an ALK mutation screening in 52 MBs and we found a novel heterozygous germline deletion of a single base in exon 23 (3605delG) in a case with marked anaplasia. This G deletion results in a frameshift mutation producing a premature stop codon in exon 25 of ALK tyrosine kinase domain. We also screened three human MB cell lines without finding any mutation of ALK gene. Quantitative expression analysis of 16 out of 52 samples showed overexpression of ALK mRNA in three MBs. In the present study, we report the first mutation of ALK found in MB. Moreover, a deletion of ALK gene producing a stop codon has not been detected in human tumors up to now. Further investigations are now required to elucidate whether the truncated form of ALK may have a role in signal transduction.

  20. Germline Mutations and Polymorphisms in the Origins of Cancers in Women

    Directory of Open Access Journals (Sweden)

    Kim M. Hirshfield

    2010-01-01

    Full Text Available Several female malignancies including breast, ovarian, and endometrial cancers can be characterized based on known somatic and germline mutations. Initiation and propagation of tumors reflect underlying genomic alterations such as mutations, polymorphisms, and copy number variations found in genes of multiple cellular pathways. The contributions of any single genetic variation or mutation in a population depend on its frequency and penetrance as well as tissue-specific functionality. Genome wide association studies, fluorescence in situ hybridization, comparative genomic hybridization, and candidate gene studies have enumerated genetic contributors to cancers in women. These include p53, BRCA1, BRCA2, STK11, PTEN, CHEK2, ATM, BRIP1, PALB2, FGFR2, TGFB1, MDM2, MDM4 as well as several other chromosomal loci. Based on the heterogeneity within a specific tumor type, a combination of genomic alterations defines the cancer subtype, biologic behavior, and in some cases, response to therapeutics. Consideration of tumor heterogeneity is therefore important in the critical analysis of gene associations in cancer.

  1. SMARCB1/INI1 germline mutations contribute to 10% of sporadic schwannomatosis

    Directory of Open Access Journals (Sweden)

    Bourdon Violaine

    2011-01-01

    Full Text Available Abstract Background Schwannomatosis is a disease characterized by multiple non-vestibular schwannomas. Although biallelic NF2 mutations are found in schwannomas, no germ line event is detected in schwannomatosis patients. In contrast, germline mutations of the SMARCB1 (INI1 tumor suppressor gene were described in familial and sporadic schwannomatosis patients. Methods To delineate the SMARCB1 gene contribution, the nine coding exons were sequenced in a series of 56 patients affected with a variable number of non-vestibular schwannomas. Results Nine variants scattered along the sequence of SMARCB1 were identified. Five of them were classified as deleterious. All five patients carrying a SMARCB1 mutation had more multiple schwannomas, corresponding to 10.2% of patients with schwannomatosis. They were also diagnosed before 35 years of age. Conclusions These results suggest that patients with schwannomas have a significant probability of carrying a SMARCB1 mutation. Combined with data available from other studies, they confirm the clinical indications for genetic screening of the SMARCB1 gene.

  2. Germline Cas9 expression yields highly efficient genome engineering in a major worldwide disease vector, Aedes aegypti.

    Science.gov (United States)

    Li, Ming; Bui, Michelle; Yang, Ting; Bowman, Christian S; White, Bradley J; Akbari, Omar S

    2017-12-05

    The development of CRISPR/Cas9 technologies has dramatically increased the accessibility and efficiency of genome editing in many organisms. In general, in vivo germline expression of Cas9 results in substantially higher activity than embryonic injection. However, no transgenic lines expressing Cas9 have been developed for the major mosquito disease vector Aedes aegypti Here, we describe the generation of multiple stable, transgenic Ae. aegypti strains expressing Cas9 in the germline, resulting in dramatic improvements in both the consistency and efficiency of genome modifications using CRISPR. Using these strains, we disrupted numerous genes important for normal morphological development, and even generated triple mutants from a single injection. We have also managed to increase the rates of homology-directed repair by more than an order of magnitude. Given the exceptional mutagenic efficiency and specificity of the Cas9 strains we engineered, they can be used for high-throughput reverse genetic screens to help functionally annotate the Ae. aegypti genome. Additionally, these strains represent a step toward the development of novel population control technologies targeting Ae. aegypti that rely on Cas9-based gene drives. Copyright © 2017 the Author(s). Published by PNAS.

  3. Transposon Invasion of the Paramecium Germline Genome Countered by a Domesticated PiggyBac Transposase and the NHEJ Pathway

    Science.gov (United States)

    Dubois, Emeline; Bischerour, Julien; Marmignon, Antoine; Mathy, Nathalie; Régnier, Vinciane; Bétermier, Mireille

    2012-01-01

    Sequences related to transposons constitute a large fraction of extant genomes, but insertions within coding sequences have generally not been tolerated during evolution. Thanks to their unique nuclear dimorphism and to their original mechanism of programmed DNA elimination from their somatic nucleus (macronucleus), ciliates are emerging model organisms for the study of the impact of transposable elements on genomes. The germline genome of the ciliate Paramecium, located in its micronucleus, contains thousands of short intervening sequences, the IESs, which interrupt 47% of genes. Recent data provided support to the hypothesis that an evolutionary link exists between Paramecium IESs and Tc1/mariner transposons. During development of the macronucleus, IESs are excised precisely thanks to the coordinated action of PiggyMac, a domesticated piggyBac transposase, and of the NHEJ double-strand break repair pathway. A PiggyMac homolog is also required for developmentally programmed DNA elimination in another ciliate, Tetrahymena. Here, we present an overview of the life cycle of these unicellular eukaryotes and of the developmentally programmed genome rearrangements that take place at each sexual cycle. We discuss how ancient domestication of a piggyBac transposase might have allowed Tc1/mariner elements to spread throughout the germline genome of Paramecium, without strong counterselection against insertion within genes. PMID:22888464

  4. Germline Hypermethylation of MLH1 and EPCAM Deletions Are a Frequent Cause of Lynch Syndrome

    NARCIS (Netherlands)

    Niessen, Renee C.; Hofstra, Robert M. W.; Westers, Helga; Ligtenberg, Marjolijn J. L.; Kooi, Krista; Jager, Paul O. J.; de Groote, Marloes L.; Dijkhuizen, Trijnie; Olderode-Berends, Maran J. W.; Hollema, Harry; Kleibeuker, Jan H.; Sijmons, Rolf H.

    It was shown that Lynch syndrome can be caused by germline hypermethylation of the MLH1 and MSH2 promoters. Furthermore, it has been demonstrated very recently that germline deletions of the 3' region of EPCAM cause transcriptional read-through which results in silencing of MSH2 by hypermethylation.

  5. Germline hypermethylation of MLH1 and EPCAM deletions are a frequent cause of Lynch syndrome.

    NARCIS (Netherlands)

    Niessen, R.C.; Hofstra, R.M.; Westers, H.; Ligtenberg, M.J.L.; Kooi, K.; Jager, P.O.; Groote, M.L. de; Dijkhuizen, T.; Olderode-Berends, M.J.; Hollema, H.; Kleibeuker, J.H.; Sijmons, R.H.

    2009-01-01

    It was shown that Lynch syndrome can be caused by germline hypermethylation of the MLH1 and MSH2 promoters. Furthermore, it has been demonstrated very recently that germline deletions of the 3' region of EPCAM cause transcriptional read-through which results in silencing of MSH2 by hypermethylation.

  6. C. elegans germline-deficient mutants respond to pathogen infection using shared and distinct mechanisms.

    Directory of Open Access Journals (Sweden)

    Michael TeKippe

    2010-07-01

    Full Text Available Reproduction extracts a cost in resources that organisms are then unable to utilize to deal with a multitude of environmental stressors. In the nematode C. elegans, development of the germline shortens the lifespan of the animal and increases its susceptibility to microbial pathogens. Prior studies have demonstrated germline-deficient nematodes to have increased resistance to gram negative bacteria. We show that germline-deficient strains display increased resistance across a broad range of pathogens including gram positive and gram negative bacteria, and the fungal pathogen Cryptococcus neoformans. Furthermore, we show that the FOXO transcription factor DAF-16, which regulates longevity and immunity in C. elegans, appears to be crucial for maintaining longevity in both wild-type and germline-deficient backgrounds. Our studies indicate that germline-deficient mutants glp-1 and glp-4 respond to pathogen infection using common and different mechanisms that involve the activation of DAF-16.

  7. Infrequent detection of germline allele-specific expression of TGFBR1 in lymphoblasts and tissues of colon cancer patients.

    LENUS (Irish Health Repository)

    Guda, Kishore

    2009-06-15

    Recently, germline allele-specific expression (ASE) of the gene encoding for transforming growth factor-beta type I receptor (TGFBR1) has been proposed to be a major risk factor for cancer predisposition in the colon. Germline ASE results in a lowered expression of one of the TGFBR1 alleles (>1.5-fold), and was shown to occur in approximately 20% of informative familial and sporadic colorectal cancer (CRC) cases. In the present study, using the highly quantitative pyrosequencing technique, we estimated the frequency of ASE in TGFBR1 in a cohort of affected individuals from familial clusters of advanced colon neoplasias (cancers and adenomas with high-grade dysplasia), and also from a cohort of individuals with sporadic CRCs. Cases were considered positive for the presence of ASE if demonstrating an allelic expression ratio <0.67 or >1.5. Using RNA derived from lymphoblastoid cell lines, we find that of 46 informative Caucasian advanced colon neoplasia cases with a family history, only 2 individuals display a modest ASE, with allelic ratios of 1.65 and 1.73, respectively. Given that ASE of TGFBR1, if present, would likely be more pronounced in the colon compared with other tissues, we additionally determined the allele ratios of TGFBR1 in the RNA derived from normal-appearing colonic mucosa of sporadic CRC cases. We, however, found no evidence of ASE in any of 44 informative sporadic cases analyzed. Taken together, we find that germline ASE of TGFBR1, as assayed in lymphoblastoid and colon epithelial cells of colon cancer patients, is a relatively rare event.

  8. Genetic Basis for Developmental Homeostasis of Germline Stem Cell Niche Number: A Network of Tramtrack-Group Nuclear BTB Factors

    Science.gov (United States)

    Chalvet, Fabienne; Netter, Sophie; Dos Santos, Nicolas; Poisot, Emilie; Paces-Fessy, Mélanie; Cumenal, Delphine; Peronnet, Frédérique; Pret, Anne-Marie; Théodore, Laurent

    2012-01-01

    The potential to produce new cells during adult life depends on the number of stem cell niches and the capacity of stem cells to divide, and is therefore under the control of programs ensuring developmental homeostasis. However, it remains generally unknown how the number of stem cell niches is controlled. In the insect ovary, each germline stem cell (GSC) niche is embedded in a functional unit called an ovariole. The number of ovarioles, and thus the number of GSC niches, varies widely among species. In Drosophila, morphogenesis of ovarioles starts in larvae with the formation of terminal filaments (TFs), each made of 8–10 cells that pile up and sort in stacks. TFs constitute organizers of individual germline stem cell niches during larval and early pupal development. In the Drosophila melanogaster subgroup, the number of ovarioles varies interspecifically from 8 to 20. Here we show that pipsqueak, Trithorax-like, batman and the bric-à-brac (bab) locus, all encoding nuclear BTB/POZ factors of the Tramtrack Group, are involved in limiting the number of ovarioles in D. melanogaster. At least two different processes are differentially perturbed by reducing the function of these genes. We found that when the bab dose is reduced, sorting of TF cells into TFs was affected such that each TF contains fewer cells and more TFs are formed. In contrast, psq mutants exhibited a greater number of TF cells per ovary, with a normal number of cells per TF, thereby leading to formation of more TFs per ovary than in the wild type. Our results indicate that two parallel genetic pathways under the control of a network of nuclear BTB factors are combined in order to negatively control the number of germline stem cell niches. PMID:23185495

  9. Expression of germline markers in three species of amphioxus supports a preformation mechanism of germ cell development in cephalochordates

    Science.gov (United States)

    2013-01-01

    Background In a previous study, we showed that the cephalochordate amphioxus Branchiostoma floridae has localized maternal transcripts of conserved germ cell markers Vasa and Nanos in its early embryos. These results provided strong evidence to support a preformation mechanism for primordial germ cell (PGC) development in B. floridae. Results In this study, we further characterize the expression of B. floridae homologs of Piwi and Tudor, which play important roles in germline development in diverse metazoan animals. We show that maternal mRNA of one of the identified Piwi-like homologs, Bf-Piwil1, also colocalizes with Vasa in the vegetal germ plasm and has zygotic expression in both the putative PGCs and the tail bud, suggesting it may function in both germline and somatic stem cells. More interestingly, one Tudor family gene, Bf-Tdrd7, is only expressed maternally and colocalizes with Vasa in germ plasm, suggesting that it may function exclusively in germ cell specification. To evaluate the conservation of the preformation mechanism among amphioxus species, we further analyze Vasa, Nanos, Piwil1, and Tdrd7 expression in two Asian amphioxus species, B. belcheri and B. japonicum. Their maternal transcripts all localize in similar patterns to those seen in B. floridae. In addition, we labeled putative PGCs with Vasa antibody to trace their dynamic distribution in developing larvae. Conclusions We identify additional germ plasm components in amphioxus and demonstrate the molecular distinction between the putative germline stem cells and somatic stem cells. Moreover, our results suggest that preformation may be a conserved mechanism for PGC specification among Branchiostoma species. Our Vasa antibody staining results suggest that after the late neurula stage, amphioxus PGCs probably proliferate with the tail bud cells during posterior elongation and are deposited near the forming myomere boundaries. Subsequently, these PGCs would concentrate at the ventral tip of the

  10. Germline variant FGFR4  p.G388R exposes a membrane-proximal STAT3 binding site.

    Science.gov (United States)

    Ulaganathan, Vijay K; Sperl, Bianca; Rapp, Ulf R; Ullrich, Axel

    2015-12-24

    Variant rs351855-G/A is a commonly occurring single-nucleotide polymorphism of coding regions in exon 9 of the fibroblast growth factor receptor FGFR4 (CD334) gene (c.1162G>A). It results in an amino-acid change at codon 388 from glycine to arginine (p.Gly388Arg) in the transmembrane domain of the receptor. Despite compelling genetic evidence for the association of this common variant with cancers of the bone, breast, colon, prostate, skin, lung, head and neck, as well as soft-tissue sarcomas and non-Hodgkin lymphoma, the underlying biological mechanism has remained elusive. Here we show that substitution of the conserved glycine 388 residue to a charged arginine residue alters the transmembrane spanning segment and exposes a membrane-proximal cytoplasmic signal transducer and activator of transcription 3 (STAT3) binding site Y(390)-(P)XXQ(393). We demonstrate that such membrane-proximal STAT3 binding motifs in the germline of type I membrane receptors enhance STAT3 tyrosine phosphorylation by recruiting STAT3 proteins to the inner cell membrane. Remarkably, such germline variants frequently co-localize with somatic mutations in the Catalogue of Somatic Mutations in Cancer (COSMIC) database. Using Fgfr4 single nucleotide polymorphism knock-in mice and transgenic mouse models for breast and lung cancers, we validate the enhanced STAT3 signalling induced by the FGFR4 Arg388-variant in vivo. Thus, our findings elucidate the molecular mechanism behind the genetic association of rs351855 with accelerated cancer progression and suggest that germline variants of cell-surface molecules that recruit STAT3 to the inner cell membrane are a significant risk for cancer prognosis and disease progression.

  11. Immunoregulatory effect of evodiamine in mice of various germlines.

    Science.gov (United States)

    Hu, Hai-Yan; Song, Zhao-Yang; Deng, Lan; Zhang, Mei-Xia

    2008-08-01

    The aim of this study was to investigate the effect of evodiamine on the proliferation and the immune function of thymocytes and splenocyte of mice from three germlines, which were 8 weeks old masculinity BALB/c, C57BL/6 and F1 hybridization mice. Cells of thymus and spleen were harvested and prepared as unicellular suspension. Cell proliferation was detected by MTT method, while the concentration of IL-2 was detected by ELISA, mRNA levels of bcl-2 and cdk2 in cells treated with evodiamine were detected by RT-PCR, the apoptosis rate and intracellular reactive oxygen species (ROS) concentration were analyzed by FCM, and the protein levels of BCL-2, CDK2 and BAX were determined by fluorescence microscope. The results indicated that at 0.5, 0.75 and 1 micromol/L, evodiamine inhibited the proliferation and externalization of thymocytes and splenocytes stimulated with ConA (p rate increased at a prolong period of time. After treatment with evodiamine for 24 and 48 hours, the cells were divided into two groups, one of which was negatively stained by 2 7-dichlorofluorescein (DCF), which indicated that ROS level decreased significantly in the dying cells. It is concluded that evodiamine inhibits proliferation and induces apoptosis of thymocytes and splenocytes from different germline mice, and at the same time decreases secretion of IL-2 through down-regulating bcl-2 and cdk2 levels.

  12. In vitro cytotoxicity of nanoparticles in mammalian germline stem cells.

    Science.gov (United States)

    Braydich-Stolle, Laura; Hussain, Saber; Schlager, John J; Hofmann, Marie-Claude

    2005-12-01

    Gametogenesis is a complex biological process that is particularly sensitive to environmental insults such as chemicals. Many chemicals have a negative impact on the germline, either by directly affecting the germ cells, or indirectly through their action on the somatic nursing cells. Ultimately, these effects can inhibit fertility, and they may have negative consequences for the development of the offspring. Recently, nanomaterials such as nanotubes, nanowires, fullerene derivatives (buckyballs), and quantum dots have received enormous national attention in the creation of new types of analytical tools for biotechnology and the life sciences. Despite the wide application of nanomaterials, there is a serious lack of information concerning their impact on human health and the environment. Thus, there are limited studies available on toxicity of nanoparticles for risk assessment of nanomaterials. The purpose of this study was to assess the suitability of a mouse spermatogonial stem cell line as a model to assess nanotoxicity in the male germline in vitro. The effects of different types of nanoparticles on these cells were evaluated by light microscopy, and by cell proliferation and standard cytotoxicity assays. Our results demonstrate a concentration-dependent toxicity for all types of particles tested, whereas the corresponding soluble salts had no significant effect. Silver nanoparticles were the most toxic while molybdenum trioxide (MoO(3)) nanoparticles were the least toxic. Our results suggest that this cell line provides a valuable model with which to assess the cytotoxicity of nanoparticles in the germ line in vitro.

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  19. Germline contamination and leakage in whole genome somatic single nucleotide variant detection.

    Science.gov (United States)

    Sendorek, Dorota H; Caloian, Cristian; Ellrott, Kyle; Bare, J Christopher; Yamaguchi, Takafumi N; Ewing, Adam D; Houlahan, Kathleen E; Norman, Thea C; Margolin, Adam A; Stuart, Joshua M; Boutros, Paul C

    2018-01-31

    The clinical sequencing of cancer genomes to personalize therapy is becoming routine across the world. However, concerns over patient re-identification from these data lead to questions about how tightly access should be controlled. It is not thought to be possible to re-identify patients from somatic variant data. However, somatic variant detection pipelines can mistakenly identify germline variants as somatic ones, a process called "germline leakage". The rate of germline leakage across different somatic variant detection pipelines is not well-understood, and it is uncertain whether or not somatic variant calls should be considered re-identifiable. To fill this gap, we quantified germline leakage across 259 sets of whole-genome somatic single nucleotide variant (SNVs) predictions made by 21 teams as part of the ICGC-TCGA DREAM Somatic Mutation Calling Challenge. The median somatic SNV prediction set contained 4325 somatic SNVs and leaked one germline polymorphism. The level of germline leakage was inversely correlated with somatic SNV prediction accuracy and positively correlated with the amount of infiltrating normal cells. The specific germline variants leaked differed by tumour and algorithm. To aid in quantitation and correction of leakage, we created a tool, called GermlineFilter, for use in public-facing somatic SNV databases. The potential for patient re-identification from leaked germline variants in somatic SNV predictions has led to divergent open data access policies, based on different assessments of the risks. Indeed, a single, well-publicized re-identification event could reshape public perceptions of the values of genomic data sharing. We find that modern somatic SNV prediction pipelines have low germline-leakage rates, which can be further reduced, especially for cloud-sharing, using pre-filtering software.

  20. Germline mutations in MAP3K6 are associated with familial gastric cancer.

    Directory of Open Access Journals (Sweden)

    Daniel Gaston

    2014-10-01

    Full Text Available Gastric cancer is among the leading causes of cancer-related deaths worldwide. While heritable forms of gastric cancer are relatively rare, identifying the genes responsible for such cases can inform diagnosis and treatment for both hereditary and sporadic cases of gastric cancer. Mutations in the E-cadherin gene, CDH1, account for 40% of the most common form of familial gastric cancer (FGC, hereditary diffuse gastric cancer (HDGC. The genes responsible for the remaining forms of FGC are currently unknown. Here we examined a large family from Maritime Canada with FGC without CDH1 mutations, and identified a germline coding variant (p.P946L in mitogen-activated protein kinase kinase kinase 6 (MAP3K6. Based on conservation, predicted pathogenicity and a known role of the gene in cancer predisposition, MAP3K6 was considered a strong candidate and was investigated further. Screening of an additional 115 unrelated individuals with non-CDH1 FGC identified the p.P946L MAP3K6 variant, as well as four additional coding variants in MAP3K6 (p.F849Sfs*142, p.P958T, p.D200Y and p.V207G. A somatic second-hit variant (p.H506Y was present in DNA obtained from one of the tumor specimens, and evidence of DNA hypermethylation within the MAP3K6 gene was observed in DNA from the tumor of another affected individual. These findings, together with previous evidence from mouse models that MAP3K6 acts as a tumor suppressor, and studies showing the presence of somatic mutations in MAP3K6 in non-hereditary gastric cancers and gastric cancer cell lines, point towards MAP3K6 variants as a predisposing factor for FGC.

  1. Mutation analysis of SDHB and SDHC: novel germline mutations in sporadic head and neck paraganglioma and familial paraganglioma and/or pheochromocytoma

    Directory of Open Access Journals (Sweden)

    Wong Nora

    2006-01-01

    Full Text Available Abstract Background Germline mutations of the SDHD, SDHB and SDHC genes, encoding three of the four subunits of succinate dehydrogenase, are a major cause of hereditary paraganglioma and pheochromocytoma, and demonstrate that these genes are classic tumor suppressors. Succinate dehydrogenase is a heterotetrameric protein complex and a component of both the Krebs cycle and the mitochondrial respiratory chain (succinate:ubiquinone oxidoreductase or complex II. Methods Using conformation sensitive gel electrophoresis (CSGE and direct DNA sequencing to analyse genomic DNA from peripheral blood lymphocytes, here we describe the mutation analysis of the SDHB and SDHC genes in 37 patients with sporadic (i.e. no known family history head and neck paraganglioma and five pheochromocytoma and/or paraganglioma families. Results Two sporadic patients were found to have a SDHB splice site mutation in intron 4, c.423+1G>A, which produces a mis-spliced transcript with a 54 nucleotide deletion, resulting in an 18 amino acid in-frame deletion. A third patient was found to carry the c.214C>T (p.Arg72Cys missense mutation in exon 4 of SDHC, which is situated in a highly conserved protein motif that constitutes the quinone-binding site of the succinate: ubiquinone oxidoreductase (SQR complex in E. coli. Together with our previous results, we found 27 germline mutations of SDH genes in 95 cases (28% of sporadic head and neck paraganglioma. In addition all index patients of five families showing hereditary pheochromocytoma-paraganglioma were found to carry germline mutations of SDHB: four of which were novel, c.343C>T (p.Arg115X, c.141G>A (p.Trp47X, c.281G>A (p.Arg94Lys, and c.653G>C (p.Trp218Ser, and one reported previously, c.136C>T, p.Arg46X. Conclusion In conclusion, these data indicate that germline mutations of SDHB and SDHC play a minor role in sporadic head and neck paraganglioma and further underline the importance of germline SDHB mutations in cases of

  2. Inherited germline ATRX mutation in two brothers with ATR-X syndrome and osteosarcoma.

    Science.gov (United States)

    Ji, Jianling; Quindipan, Catherine; Parham, David; Shen, Lishuang; Ruble, David; Bootwalla, Moiz; Maglinte, Dennis T; Gai, Xiaowu; Saitta, Sulagna C; Biegel, Jaclyn A; Mascarenhas, Leo

    2017-05-01

    We report a family in which two brothers had an undiagnosed genetic disorder comprised of dysmorphic features, microcephaly, severe intellectual disability (non-verbal), mild anemia, and cryptorchidism. Both developed osteosarcoma. Trio exome sequencing (using blood samples from the younger brother and both parents) was performed and a nonsense NM_000489.4:c.7156C>T (p.Arg2386*) mutation in the ATRX gene was identified in the proband (hemizygous) and in the mother's peripheral blood DNA (heterozygous). The mother is healthy, does not exhibit any clinical manifestations of ATR-X syndrome and there was no family history of cancer. The same hemizygous pathogenic variant was confirmed in the affected older brother's skin tissue by subsequent Sanger sequencing. Chromosomal microarray studies of both brothers' osteosarcomas revealed complex copy number alterations consistent with the clinical diagnosis of osteosarcoma. Recently, somatic mutations in the ATRX gene have been observed as recurrent alterations in both osteosarcoma and brain tumors. However, it is unclear if there is any association between osteosarcoma and germline ATRX mutations, specifically in patients with constitutional ATR-X syndrome. This is the first report of osteosarcoma diagnosed in two males with ATR-X syndrome, suggesting a potential increased risk for cancer in patients with this disorder. © 2017 Wiley Periodicals, Inc.

  3. Germline stem cells and neo-oogenesis in the adult human ovary.

    Science.gov (United States)

    Liu, Yifei; Wu, Chao; Lyu, Qifeng; Yang, Dongzi; Albertini, David F; Keefe, David L; Liu, Lin

    2007-06-01

    It remains unclear whether neo-oogenesis occurs in postnatal ovaries of mammals, based on studies in mice. We thought to test whether adult human ovaries contain germline stem cells (GSCs) and undergo neo-oogenesis. Rather than using genetic manipulation which is unethical in humans, we took the approach of analyzing the expression of meiotic marker genes and genes for germ cell proliferation, which are required for neo-oogenesis, in adult human ovaries covering an age range from 28 to 53 years old, compared to testis and fetal ovaries served as positive controls. We show that active meiosis, neo-oogenesis and GSCs are unlikely to exist in normal, adult, human ovaries. No early meiotic-specific or oogenesis-associated mRNAs for SPO11, PRDM9, SCP1, TERT and NOBOX were detectable in adult human ovaries using RT-PCR, compared to fetal ovary and adult testis controls. These findings are further corroborated by the absence of early meiocytes and proliferating germ cells in adult human ovarian cortex probed with markers for meiosis (SCP3), oogonium (OCT3/4, c-KIT), and cell cycle progression (Ki-67, PCNA), in contrast to fetal ovary controls. If postnatal oogenesis is confirmed in mice, then this species would represent an exception to the rule that neo-oogenesis does not occur in adults.

  4. Autosomal dominant polycystic kidney disease caused by somatic and germline mosaicism.

    Science.gov (United States)

    Tan, A Y; Blumenfeld, J; Michaeel, A; Donahue, S; Bobb, W; Parker, T; Levine, D; Rennert, H

    2015-04-01

    Autosomal dominant polycystic kidney disease (ADPKD) is a heterogeneous genetic disorder caused by loss of function mutations of PKD1 or PKD2 genes. Although PKD1 is highly polymorphic and the new mutation rate is relatively high, the role of mosaicism is incompletely defined. Herein, we describe the molecular analysis of ADPKD in a 19-year-old female proband and her father. The proband had a PKD1 truncation mutation c.10745dupC (p.Val3584ArgfsX43), which was absent in paternal peripheral blood lymphocytes (PBL). However, very low quantities of this mutation were detected in the father's sperm DNA, but not in DNA from his buccal cells or urine sediment. Next generation sequencing (NGS) analysis determined the level of this mutation in the father's PBL, buccal cells and sperm to be ∼3%, 4.5% and 10%, respectively, consistent with somatic and germline mosaicism. The PKD1 mutation in ∼10% of her father's sperm indicates that it probably occurred early in embryogenesis. In ADPKD cases where a de novo mutation is suspected because of negative PKD gene testing of PBL, additional evaluation with more sensitive methods (e.g. NGS) of the proband PBL and paternal sperm can enhance detection of mosaicism and facilitate genetic counseling. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  5. Germline mutations of regulator of telomere elongation helicase 1, RTEL1, in Dyskeratosis congenita.

    Science.gov (United States)

    Ballew, Bari J; Yeager, Meredith; Jacobs, Kevin; Giri, Neelam; Boland, Joseph; Burdett, Laurie; Alter, Blanche P; Savage, Sharon A

    2013-04-01

    Dyskeratosis congenita (DC) is an inherited bone marrow failure and cancer predisposition syndrome caused by aberrant telomere biology. The classic triad of dysplastic nails, abnormal skin pigmentation, and oral leukoplakia is diagnostic of DC, but substantial clinical heterogeneity exists; the clinically severe variant Hoyeraal Hreidarsson syndrome (HH) also includes cerebellar hypoplasia, severe immunodeficiency, enteropathy, and intrauterine growth retardation. Germline mutations in telomere biology genes account for approximately one-half of known DC families. Using exome sequencing, we identified mutations in RTEL1, a helicase with critical telomeric functions, in two families with HH. In the first family, two siblings with HH and very short telomeres inherited a premature stop codon from their mother who has short telomeres. The proband from the second family has HH and inherited a premature stop codon in RTEL1 from his father and a missense mutation from his mother, who also has short telomeres. In addition, inheritance of only the missense mutation led to very short telomeres in the proband's brother. Targeted sequencing identified a different RTEL1 missense mutation in one additional DC proband who has bone marrow failure and short telomeres. Both missense mutations affect the helicase domain of RTEL1, and three in silico prediction algorithms suggest that they are likely deleterious. The nonsense mutations both cause truncation of the RTEL1 protein, resulting in loss of the PIP box; this may abrogate an important protein-protein interaction. These findings implicate a new telomere biology gene, RTEL1, in the etiology of DC.

  6. Sex-lethal enables germline stem cell differentiation by down-regulating Nanos protein levels during Drosophila oogenesis.

    Science.gov (United States)

    Chau, Johnnie; Kulnane, Laura Shapiro; Salz, Helen K

    2012-06-12

    Drosophila ovarian germ cells require Sex-lethal (Sxl) to exit from the stem cell state and to enter the differentiation pathway. Sxl encodes a female-specific RNA binding protein and in somatic cells serves as the developmental switch gene for somatic sex determination and X-chromosome dosage compensation. None of the known Sxl target genes are required for germline differentiation, leaving open the question of how Sxl promotes the transition from stem cell to committed daughter cell. We address the mechanism by which Sxl regulates this transition through the identification of nanos as one of its target genes. Previous studies have shown that Nanos protein is necessary for GSC self-renewal and is rapidly down-regulated in the daughter cells fated to differentiate in the adult ovary. We find that this dynamic expression pattern is limited to female germ cells and is under Sxl control. In the absence of Sxl, or in male germ cells, Nanos protein is continuously expressed. Furthermore, this female-specific expression pattern is dependent on the presence of canonical Sxl binding sites located in the nanos 3' untranslated region. These results, combined with the observation that nanos RNA associates with the Sxl protein in ovarian extracts and loss and gain of function studies, suggest that Sxl enables the switch from germline stem cell to committed daughter cell by posttranscriptional down-regulation of nanos expression. These findings connect sexual identity to the stem cell self-renewal/differentiation decision and highlight the importance of posttranscriptional gene regulatory networks in controlling stem cell behavior.

  7. A computational approach to distinguish somatic vs. germline origin of genomic alterations from deep sequencing of cancer specimens without a matched normal.

    Directory of Open Access Journals (Sweden)

    James X Sun

    2018-02-01

    Full Text Available A key constraint in genomic testing in oncology is that matched normal specimens are not commonly obtained in clinical practice. Thus, while well-characterized genomic alterations do not require normal tissue for interpretation, a significant number of alterations will be unknown in whether they are germline or somatic, in the absence of a matched normal control. We introduce SGZ (somatic-germline-zygosity, a computational method for predicting somatic vs. germline origin and homozygous vs. heterozygous or sub-clonal state of variants identified from deep massively parallel sequencing (MPS of cancer specimens. The method does not require a patient matched normal control, enabling broad application in clinical research. SGZ predicts the somatic vs. germline status of each alteration identified by modeling the alteration's allele frequency (AF, taking into account the tumor content, tumor ploidy, and the local copy number. Accuracy of the prediction depends on the depth of sequencing and copy number model fit, which are achieved in our clinical assay by sequencing to high depth (>500x using MPS, covering 394 cancer-related genes and over 3,500 genome-wide single nucleotide polymorphisms (SNPs. Calls are made using a statistic based on read depth and local variability of SNP AF. To validate the method, we first evaluated performance on samples from 30 lung and colon cancer patients, where we sequenced tumors and matched normal tissue. We examined predictions for 17 somatic hotspot mutations and 20 common germline SNPs in 20,182 clinical cancer specimens. To assess the impact of stromal admixture, we examined three cell lines, which were titrated with their matched normal to six levels (10-75%. Overall, predictions were made in 85% of cases, with 95-99% of variants predicted correctly, a significantly superior performance compared to a basic approach based on AF alone. We then applied the SGZ method to the COSMIC database of known somatic variants

  8. Natural Gene Therapy in Dystrophic Epidermolysis Bullosa

    NARCIS (Netherlands)

    van den Akker, Peter C.; Nijenhuis, Albertine; Hofstra, Robert M. W.; Jonkman, Marcel F.; Pasmooij, Anna M. G.; Meijer, G.

    Background: Dystrophic epidermolysis bullosa is a genetic blistering disorder caused by mutations in the type VII collagen gene, COL7A1. In revertant mosaicism, germline mutations are corrected by somatic events resulting in a mosaic disease distribution. This "natural gene therapy" phenomenon long

  9. Germline mutations in ABL1 cause an autosomal dominant syndrome characterized by congenital heart defects and skeletal malformations.

    Science.gov (United States)

    Wang, Xia; Charng, Wu-Lin; Chen, Chun-An; Rosenfeld, Jill A; Al Shamsi, Aisha; Al-Gazali, Lihadh; McGuire, Marianne; Mew, Nicholas Ah; Arnold, Georgianne L; Qu, Chunjing; Ding, Yan; Muzny, Donna M; Gibbs, Richard A; Eng, Christine M; Walkiewicz, Magdalena; Xia, Fan; Plon, Sharon E; Lupski, James R; Schaaf, Christian P; Yang, Yaping

    2017-04-01

    ABL1 is a proto-oncogene well known as part of the fusion gene BCR-ABL1 in the Philadelphia chromosome of leukemia cancer cells. Inherited germline ABL1 changes have not been associated with genetic disorders. Here we report ABL1 germline variants cosegregating with an autosomal dominant disorder characterized by congenital heart disease, skeletal abnormalities, and failure to thrive. The variant c.734A>G (p.Tyr245Cys) was found to occur de novo or cosegregate with disease in five individuals (families 1-3). Additionally, a de novo c.1066G>A (p.Ala356Thr) variant was identified in a sixth individual (family 4). We overexpressed the mutant constructs in HEK 293T cells and observed increased tyrosine phosphorylation, suggesting increased ABL1 kinase activities associated with both the p.Tyr245Cys and p.Ala356Thr substitutions. Our clinical and experimental findings, together with previously reported teratogenic effects of selective BCR-ABL inhibitors in humans and developmental defects in Abl1 knockout mice, suggest that ABL1 has an important role during organismal development.

  10. Germline mutations in ABL1 cause an autosomal dominant syndrome characterized by congenital heart defects and skeletal malformations

    Science.gov (United States)

    Wang, Xia; Charng, Wu-Lin; Chen, Chun-An; Rosenfeld, Jill A.; Shamsi, Aisha Al; Al-Gazali, Lihadh; McGuire, Marianne; Mew, Nicholas Ah; Arnold, Georgianne L.; Qu, Chunjing; Ding, Yan; Muzny, Donna M.; Gibbs, Richard A.; Eng, Christine M.; Walkiewicz, Magdalena; Xia, Fan; Plon, Sharon E.; Lupski, James R.; Schaaf, Christian P.; Yang, Yaping

    2017-01-01

    ABL1 is a proto-oncogene well known as part of the fusion gene BCR-ABL in the Philadelphia chromosome of leukemia cancer cells1. Inherited germline ABL1 changes have not been associated with genetic disorders. Here we report ABL1 germline variants co-segregating with an autosomal dominant disorder characterized by congenital heart disease, skeletal abnormalities, and failure to thrive. The variant c.734A>G (p.Tyr245Cys) was found as de novo or co-segregating with disease in five individuals (families 1-3). Additionally, a de novo c.1066G>A (p.Ala356Thr) variant was identified in the sixth individual (family 4). We overexpressed the mutant constructs in HEK 293T cells and observed increased tyrosine phosphorylation, suggesting increased ABL1 kinase activities associated with both p.Tyr245Cys and p.Ala356Thr substitutions. Our clinical and laboratory findings, together with previously reported teratogenic effects of selective BCR-ABL inhibitors in humans2-5 and developmental defects in Abl1 knock-out mice6,7, suggest ABL1 plays an important role during organismal development. PMID:28288113

  11. Germ-line PHD1 and PHD2 mutations detected in patients with pheochromocytoma/paraganglioma-polycythemia.

    Science.gov (United States)

    Yang, Chunzhang; Zhuang, Zhengping; Fliedner, Stephanie M J; Shankavaram, Uma; Sun, Michael G; Bullova, Petra; Zhu, Roland; Elkahloun, Abdel G; Kourlas, Peter J; Merino, Maria; Kebebew, Electron; Pacak, Karel

    2015-01-01

    We have investigated genetic/pathogenetic factors associated with a new clinical entity in patients presenting with pheochromocytoma/paraganglioma (PHEO/PGL) and polycythemia. Two patients without hypoxia-inducible factor 2α (HIF2A) mutations, who presented with similar clinical manifestations, were analyzed for other gene mutations, including prolyl hydroxylase (PHD) mutations. We have found for the first time a germ-line mutation in PHD1 in one patient and a novel germ-line PHD2 mutation in a second patient. Both mutants exhibited reduced protein stability with substantial quantitative protein loss and thus compromised catalytic activities. Due to the unique association of patients' polycythemia with borderline or mildly elevated erythropoietin (EPO) levels, we also performed an in vitro sensitivity assay of erythroid progenitors to EPO and for EPO receptor (EPOR) expression. The results show inappropriate hypersensitivity of erythroid progenitors to EPO in these patients, indicating increased EPOR expression/activity. In addition, the present study indicates that HIF dysregulation due to PHD mutations plays an important role in the pathogenesis of these tumors and associated polycythemia. The PHD1 mutation appears to be a new member contributing to the genetic landscape of this novel clinical entity. Our results support the existence of a specific PHD1- and PHD2-associated PHEO/PGL-polycythemia disorder. • A novel germ-l i n e PHD1 mutation causing heochromocytoma/paraganglioma and polycythemia. • Increased EPOR activity and inappropriate hypersensitivity of erythroid progenitors to EPO.

  12. Germline Mutations in Mtap Cooperate with Myc to Accelerate Tumorigenesis in Mice.

    Directory of Open Access Journals (Sweden)

    Yuwaraj Kadariya

    Full Text Available The gene encoding the methionine salvage pathway methylthioadenosine phosphorylase (MTAP is a tumor suppressor gene that is frequently inactivated in a wide variety of human cancers. In this study, we have examined if heterozygosity for a null mutation in Mtap (Mtap(lacZ could accelerate tumorigenesis development in two different mouse cancer models, Eμ-myc transgenic and Pten(+/- .Mtap Eμ-myc and Mtap Pten mice were generated and tumor-free survival was monitored over time. Tumors were also examined for a variety of histological and protein markers. In addition, microarray analysis was performed on the livers of Mtap(lacZ/+ and Mtap (+/+ mice.Survival in both models was significantly decreased in Mtap(lacZ/+ compared to Mtap(+/+ mice. In Eµ-myc mice, Mtap mutations accelerated the formation of lymphomas from cells in the early pre-B stage, and these tumors tended to be of higher grade and have higher expression levels of ornithine decarboxylase compared to those observed in control Eµ-myc Mtap(+/+ mice. Surprisingly, examination of Mtap status in lymphomas in Eµ-myc Mtap(lacZ/+ and Eµ-myc Mtap(+/+ animals did not reveal significant differences in the frequency of loss of Mtap protein expression, despite having shorter latency times, suggesting that haploinsufficiency of Mtap may be playing a direct role in accelerating tumorigenesis. Consistent with this idea, microarray analysis on liver tissue from age and sex matched Mtap(+/+ and Mtap(lacZ/+ animals found 363 transcripts whose expression changed at least 1.5-fold (P<0.01. Functional categorization of these genes reveals enrichments in several pathways involved in growth control and cancer.Our findings show that germline inactivation of a single Mtap allele alters gene expression and enhances lymphomagenesis in Eµ-myc mice.

  13. Wnt6 maintains anterior escort cells as an integral component of the germline stem cell niche.

    Science.gov (United States)

    Wang, Xiaoxi; Page-McCaw, Andrea

    2018-02-07

    Stem cells reside in a niche, a local environment whose cellular and molecular complexity is still being elucidated. In Drosophila ovaries, germline stem cells depend on cap cells for self-renewing signals and physical attachment. Germline stem cells also contact the anterior escort cells, and here we report that anterior escort cells are absolutely required for germline stem cell maintenance. When escort cells die from impaired Wnt signaling or hid expression, the loss of anterior escort cells causes loss of germline stem cells. Anterior escort cells function as an integral niche component by promoting DE-cadherin anchorage and by transiently expressing the Dpp ligand to promote full-strength BMP signaling in germline stem cells. Anterior escort cells are maintained by Wnt6 ligands produced by cap cells; without Wnt6 signaling, anterior escort cells die leaving vacancies in the niche, leading to loss of germline stem cells. Our data identify anterior escort cells as constituents of the germline stem cell niche, maintained by a cap cell-produced Wnt6 survival signal. © 2018. Published by The Company of Biologists Ltd.

  14. Common germline polymorphisms associated with breast cancer-specific survival

    DEFF Research Database (Denmark)

    Pirie, Ailith; Guo, Qi; Kraft, Peter

    2015-01-01

    in the meta-analysis. Fifty-four of these were evaluated in the full set of 37,954 breast cancer cases with 2,900 events and the two additional variants were evaluated in a reduced sample size of 30,000 samples in order to ensure independence from the previously published studies. Five variants reached...... evaluated in the pooled analysis of over 37,000 breast cancer cases for association with breast cancer-specific survival. Previous associations were evaluated using a one-sided test based on the reported direction of effect. RESULTS: Fifty-six variants from 45 previous publications were evaluated......-specific survival using data from a pooled analysis of eight breast cancer survival genome-wide association studies (GWAS) from the Breast Cancer Association Consortium. METHODS: A literature review was conducted of all previously published associations between common germline variants and three survival outcomes...

  15. Impact of functional germline variants and a deletion polymorphism in APOBEC3A and APOBEC3B on breast cancer risk and survival in a Swedish study population.

    Science.gov (United States)

    Göhler, Stella; Da Silva Filho, Miguel Inacio; Johansson, Robert; Enquist-Olsson, Kerstin; Henriksson, Roger; Hemminki, Kari; Lenner, Per; Försti, Asta

    2016-01-01

    The C → T mutation signature caused by APOBEC family members contributes to the development of breast cancer (BC). Also overexpression of APOBEC3B and a ~29.5-kb deletion polymorphism between APOBEC3A and APOBEC3B have been associated with increased BC risk. We investigated in a population-based study, with 782 Swedish BC cases and 1559 controls, associations between potentially functional germline variants in APOBEC3A or APOBEC3B gene and BC risk and survival. Additionally, we identified deletion polymorphism carriers and explored possible associations with BC. No evidence of association between any germline variant, including the deletion polymorphism, and BC risk or survival was observed. Only APOBEC3A promoter polymorphism rs5757402 was associated with low stage (OR = 0.69, 95 % CI 0.50-0.96, dominant model). The reported association between the deletion polymorphism and BC risk was not confirmed in the Swedish population, nor did any genotyped germline variant show any association with BC risk or survival.

  16. Parent-of-origin and trans-generational germline influences on behavioral development: the interacting roles of mothers, fathers, and grandparents.

    Science.gov (United States)

    Curley, J P; Mashoodh, R

    2010-05-01

    Mothers and fathers do not contribute equally to the development of their offspring. In addition to the differential investment of mothers versus fathers in the rearing of offspring, there are also a number of germline factors that are transmitted unequally from one parent or the other that contribute significantly to offspring development. This article shall review four major sources of such parent-of-origin effects. Firstly, there is increasing evidence that genes inherited on the sex chromosomes including the nonpseudoautosomal part of the Y chromosome that is only inherited from fathers to sons, contribute to brain development and behavior independently of the organizing effects of sex hormones. Secondly, recent work has demonstrated that mitochondrial DNA that is primarily inherited only from mothers may play a much greater than anticipated role in neurobehavioral development. Thirdly, there exists a class of genes known as imprinted genes that are epigenetically silenced when passed on in a parent-of-origin specific manner and have been shown to regulate brain development and a variety of behaviors. Finally, there is converging evidence from several disciplines that environmental variations experienced by mothers and fathers may lead to plasticity in the development and behavior of offspring and that this phenotypic inheritance can be solely transmitted through the germline. Mechanistically, this may be achieved through altered programming within germ cells of the epigenetic status of particular genes such as retrotransposons and imprinted genes or potentially through altered expression of RNAs within gametes.

  17. Contribution of germline TP53 variants and assessment of HER-2 status among young breast cancer patients in Malaysia

    Directory of Open Access Journals (Sweden)

    Shao Yan Lau

    2017-12-01

    Full Text Available Background: Li-Fraumeni Syndrome (LFS is caused by a mutation in the TP53 tumour suppressor gene. This rare hereditary condition predisposes individuals to an increased risk of cancers including breast cancer in women at a relatively young age, which accounts for nearly 25%–30% of all LFS‑associated cancers. Studies have shown that breast tumours in women with a germline TP53 deleterious variants are associated with a human epidermal growth factor receptor 2 (HER2-positive phenotype. Taken together, this study aimed to investigate the contribution of germline TP53 variants and its association with tumour HER-2 status in a cohort of young women with breast cancer. Methods: From 2002 to 2017, 4048 women with breast cancer treated at University Malaya Medical Centre or Sime Darby Medical Centre participated in the Malaysian Breast Cancer Genetics Study. Of which, 87 patients were diagnosed before 30 years of age. All patients were analysed for germline TP53 single nucleotide variants, small insertions or deletions by amplicon‑based targeted sequencing and validated by Sanger sequencing. DNA from patients who tested negative for sequencing were subsequently evaluated for the presence of TP53 exon deletions or duplications by multiplex ligation‑dependent probe amplification. HER-2 status of breast tumours was defined by immunohistochemistry, fluorescence in situ hybridisation and/or silver in situ hybridisation. Results: 5 distinct TP53 variants were detected in 5 individuals. 3 out of 5 TP53 variants were classified as frameshift mutations, one nonsense mutation and one in-frame duplication. Variants in other genes were detected in 17 individuals. No large genomic rearrangements were detected in the remaining 65 sequencing-negative patients. The assessment of HER-2 status will be presented. Conclusions: Our results suggest that alterations in TP53 gene were identified in approximately 5.7% (5/87 of this cohort of young women with breast

  18. ZTF-8 interacts with the 9-1-1 complex and is required for DNA damage response and double-strand break repair in the C. elegans germline.

    Directory of Open Access Journals (Sweden)

    Hyun-Min Kim

    2014-10-01

    Full Text Available Germline mutations in DNA repair genes are linked to tumor progression. Furthermore, failure in either activating a DNA damage checkpoint or repairing programmed meiotic double-strand breaks (DSBs can impair chromosome segregation. Therefore, understanding the molecular basis for DNA damage response (DDR and DSB repair (DSBR within the germline is highly important. Here we define ZTF-8, a previously uncharacterized protein conserved from worms to humans, as a novel factor involved in the repair of both mitotic and meiotic DSBs as well as in meiotic DNA damage checkpoint activation in the C. elegans germline. ztf-8 mutants exhibit specific sensitivity to γ-irradiation and hydroxyurea, mitotic nuclear arrest at S-phase accompanied by activation of the ATL-1 and CHK-1 DNA damage checkpoint kinases, as well as accumulation of both mitotic and meiotic recombination intermediates, indicating that ZTF-8 functions in DSBR. However, impaired meiotic DSBR progression partially fails to trigger the CEP-1/p53-dependent DNA damage checkpoint in late pachytene, also supporting a role for ZTF-8 in meiotic DDR. ZTF-8 partially co-localizes with the 9-1-1 DDR complex and interacts with MRT-2/Rad1, a component of this complex. The human RHINO protein rescues the phenotypes observed in ztf-8 mutants, suggesting functional conservation across species. We propose that ZTF-8 is involved in promoting repair at stalled replication forks and meiotic DSBs by transducing DNA damage checkpoint signaling via the 9-1-1 pathway. Our findings define a conserved function for ZTF-8/RHINO in promoting genomic stability in the germline.

  19. Universal screening for Lynch syndrome in endometrial cancers: frequency of germline mutations and identification of patients with Lynch-like syndrome.

    Science.gov (United States)

    Dillon, Jessica L; Gonzalez, Jorge L; DeMars, Leslie; Bloch, Katarzyna J; Tafe, Laura J

    2017-12-01

    Lynch syndrome (LS) is an inherited clinical syndrome characterized by a high risk of colorectal, endometrial (lifetime risk of up to 60%), ovarian, and urinary tract cancers. The diagnosis is confirmed by identification of germline mutations in the DNA mismatch repair genes MLH1, PMS2, MSH2, MSH6, or EPCAM. In 2015, our institution implemented universal screening of endometrial cancer (EC) hysterectomy specimens by mismatch repair immunohistochemistry (IHC) with reflex MLH1 promoter hypermethylation analysis for tumors with loss of MLH1/PMS2 expression. Patients with tumors negative for MLH1 methylation and those with a loss of the heterodimer pair MSH2 and MSH6, or isolated loss of either PMS2 or MSH6 were referred to the Familial Cancer Program for genetic counseling and consideration of germline testing. Between May 2015 to Dec 2016, 233 EC patients were screened by IHC for LS with a median age of 63 years. Sixty tumors (27%) had abnormal IHC staining results. Fifty-one (22%) harbored heterodimeric loss of MLH1 and PMS2, 49 of which showed MLH1 promoter methylation (1 failure, 1 negative). One showed loss of MLH1/PMS2 and MSH6, 2 showed loss of MSH2/MSH6, and 6 had isolated loss of MSH6 only. Ten patients underwent genetic counseling, and germline testing was performed in 8; LS was confirmed in 5 patients (2.1%). In addition, 3 patients with negative germline testing and presumed Lynch-like syndrome were identified and offered additional somatic testing. Universal screening for LS in EC patients has yielded positive results for identification of patients at risk for this inherited syndrome. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Screening for germline phosphatase and tensin homolog-mutations in suspected Cowden syndrome and Cowden syndrome-like families among uterine cancer patients

    Science.gov (United States)

    TZORTZATOS, GERASIMOS; ARAVIDIS, CHRISTOS; LINDBLOM, ANNIKA; MINTS, MIRIAM; THAM, EMMA

    2015-01-01

    Cowden syndrome (CS) is an autosomal dominant disorder characterized by multiple hamartomas in the breast, thyroid and endometrium, with a prevalence of 1 per 250,000. Females with CS have a 21–28% lifetime risk of developing uterine cancer. Germline mutations in the phosphatase and tensin homolog (PTEN) gene, a tumor suppressor gene, are responsible for 30–80% of CS cases. PTEN is a nine-exon gene, located on chromosome 10q23.3, which encodes the 403 amino acid PTEN protein. It negatively regulates the phosphoinositide 3-kinase/protein kinase B/mammalian target of rapamycin pathway, affecting various cellular processes and signaling pathways. The present study examined whether PTEN mutations are present in CS-like families with uterine cancer (UC). UC patients underwent surgery at Karolinska University Hospital, Stockholm, Sweden (2008–2012). Pedigrees were analyzed and 54 unrelated CS-like families were identified. CS-like families were defined as having at least one occurrence of uterine cancer and one of breast cancer, as well as at least one additional Cowden-associated tumor (uterine, breast, thyroid, colon or kidney cancer) in the same individual or in first-degree relatives. Genomic DNA was amplified using polymerase chain reaction, and DNA sequencing analysis of all nine exons of the PTEN gene was conducted. No germline PTEN mutations or polymorphisms were identified. Germline PTEN mutations are rare in CS-like families with uterine cancer, therefore, genetic screening must be restricted to patients that meet the strict National Comprehensive Cancer Network criteria. Gynecologists must be aware of the CS criteria and identify potential cases of CS in females where uterine cancer is the sentinel cancer. PMID:25789042

  1. AluY-mediated germline deletion, duplication and somatic stem cell reversion in UBE2T defines a new subtype of Fanconi anemia.

    Science.gov (United States)

    Virts, Elizabeth L; Jankowska, Anna; Mackay, Craig; Glaas, Marcel F; Wiek, Constanze; Kelich, Stephanie L; Lottmann, Nadine; Kennedy, Felicia M; Marchal, Christophe; Lehnert, Erik; Scharf, Rüdiger E; Dufour, Carlo; Lanciotti, Marina; Farruggia, Piero; Santoro, Alessandra; Savasan, Süreyya; Scheckenbach, Kathrin; Schipper, Jörg; Wagenmann, Martin; Lewis, Todd; Leffak, Michael; Farlow, Janice L; Foroud, Tatiana M; Honisch, Ellen; Niederacher, Dieter; Chakraborty, Sujata C; Vance, Gail H; Pruss, Dmitry; Timms, Kirsten M; Lanchbury, Jerry S; Alpi, Arno F; Hanenberg, Helmut

    2015-09-15

    Fanconi anemia (FA) is a rare inherited disorder clinically characterized by congenital malformations, progressive bone marrow failure and cancer susceptibility. At the cellular level, FA is associated with hypersensitivity to DNA-crosslinking genotoxins. Eight of 17 known FA genes assemble the FA E3 ligase complex, which catalyzes monoubiquitination of FANCD2 and is essential for replicative DNA crosslink repair. Here, we identify the first FA patient with biallelic germline mutations in the ubiquitin E2 conjugase UBE2T. Both mutations were aluY-mediated: a paternal deletion and maternal duplication of exons 2-6. These loss-of-function mutations in UBE2T induced a cellular phenotype similar to biallelic defects in early FA genes with the absence of FANCD2 monoubiquitination. The maternal duplication produced a mutant mRNA that could encode a functional protein but was degraded by nonsense-mediated mRNA decay. In the patient's hematopoietic stem cells, the maternal allele with the duplication of exons 2-6 spontaneously reverted to a wild-type allele by monoallelic recombination at the duplicated aluY repeat, thereby preventing bone marrow failure. Analysis of germline DNA of 814 normal individuals and 850 breast cancer patients for deletion or duplication of UBE2T exons 2-6 identified the deletion in only two controls, suggesting aluY-mediated recombinations within the UBE2T locus are rare and not associated with an increased breast cancer risk. Finally, a loss-of-function germline mutation in UBE2T was detected in a high-risk breast cancer patient with wild-type BRCA1/2. Cumulatively, we identified UBE2T as a bona fide FA gene (FANCT) that also may be a rare cancer susceptibility gene. © The Author 2015. Published by Oxford University Press.

  2. Identification of germline susceptibility loci in ETV6-RUNX1-rearranged childhood acute lymphoblastic leukemia

    Science.gov (United States)

    Ellinghaus, E; Stanulla, M; Richter, G; Ellinghaus, D; te Kronnie, G; Cario, G; Cazzaniga, G; Horstmann, M; Panzer Grümayer, R; Cavé, H; Trka, J; Cinek, O; Teigler-Schlegel, A; ElSharawy, A; Häsler, R; Nebel, A; Meissner, B; Bartram, T; Lescai, F; Franceschi, C; Giordan, M; Nürnberg, P; Heinzow, B; Zimmermann, M; Schreiber, S; Schrappe, M; Franke, A

    2012-01-01

    Acute lymphoblastic leukemia (ALL) is a malignant disease of the white blood cells. The etiology of ALL is believed to be multifactorial and likely to involve an interplay of environmental and genetic variables. We performed a genome-wide association study of 355 750 single-nucleotide polymorphisms (SNPs) in 474 controls and 419 childhood ALL cases characterized by a t(12;21)(p13;q22) — the most common chromosomal translocation observed in childhood ALL — which leads to an ETV6–RUNX1 gene fusion. The eight most strongly associated SNPs were followed-up in 951 ETV6-RUNX1-positive cases and 3061 controls from Germany/Austria and Italy, respectively. We identified a novel, genome-wide significant risk locus at 3q28 (TP63, rs17505102, PCMH=8.94 × 10−9, OR=0.65). The separate analysis of the combined German/Austrian sample only, revealed additional genome-wide significant associations at 11q11 (OR8U8, rs1945213, P=9.14 × 10−11, OR=0.69) and 8p21.3 (near INTS10, rs920590, P=6.12 × 10−9, OR=1.36). These associations and another association at 11p11.2 (PTPRJ, rs3942852, P=4.95 × 10−7, OR=0.72) remained significant in the German/Austrian replication panel after correction for multiple testing. Our findings demonstrate that germline genetic variation can specifically contribute to the risk of ETV6–RUNX1-positive childhood ALL. The identification of TP63 and PTPRJ as susceptibility genes emphasize the role of the TP53 gene family and the importance of proteins regulating cellular processes in connection with tumorigenesis. PMID:22076464

  3. Estimating Exceptionally Rare Germline and Somatic Mutation Frequencies via Next Generation Sequencing.

    Directory of Open Access Journals (Sweden)

    Jordan Eboreime

    Full Text Available We used targeted next generation deep-sequencing (Safe Sequencing System to measure ultra-rare de novo mutation frequencies in the human male germline by attaching a unique identifier code to each target DNA molecule. Segments from three different human genes (FGFR3, MECP2 and PTPN11 were studied. Regardless of the gene segment, the particular testis donor or the 73 different testis pieces used, the frequencies for any one of the six different mutation types were consistent. Averaging over the C>T/G>A and G>T/C>A mutation types the background mutation frequency was 2.6x10-5 per base pair, while for the four other mutation types the average background frequency was lower at 1.5x10-6 per base pair. These rates far exceed the well documented human genome average frequency per base pair (~10-8 suggesting a non-biological explanation for our data. By computational modeling and a new experimental procedure to distinguish between pre-mutagenic lesion base mismatches and a fully mutated base pair in the original DNA molecule, we argue that most of the base-dependent variation in background frequency is due to a mixture of deamination and oxidation during the first two PCR cycles. Finally, we looked at a previously studied disease mutation in the PTPN11 gene and could easily distinguish true mutations from the SSS background. We also discuss the limits and possibilities of this and other methods to measure exceptionally rare mutation frequencies, and we present calculations for other scientists seeking to design their own such experiments.

  4. Whole Gene Capture Analysis of 15 CRC Susceptibility Genes in Suspected Lynch Syndrome Patients.

    Science.gov (United States)

    Jansen, Anne M L; Geilenkirchen, Marije A; van Wezel, Tom; Jagmohan-Changur, Shantie C; Ruano, Dina; van der Klift, Heleen M; van den Akker, Brendy E W M; Laros, Jeroen F J; van Galen, Michiel; Wagner, Anja; Letteboer, Tom G W; Gómez-García, Encarna B; Tops, Carli M J; Vasen, Hans F; Devilee, Peter; Hes, Frederik J; Morreau, Hans; Wijnen, Juul T

    2016-01-01

    Lynch Syndrome (LS) is caused by pathogenic germline variants in one of the mismatch repair (MMR) genes. However, up to 60% of MMR-deficient colorectal cancer cases are categorized as suspected Lynch Syndrome (sLS) because no pathogenic MMR germline variant can be identified, which leads to difficulties in clinical management. We therefore analyzed the genomic regions of 15 CRC susceptibility genes in leukocyte DNA of 34 unrelated sLS patients and 11 patients with MLH1 hypermethylated tumors with a clear family history. Using targeted next-generation sequencing, we analyzed the entire non-repetitive genomic sequence, including intronic and regulatory sequences, of 15 CRC susceptibility genes. In addition, tumor DNA from 28 sLS patients was analyzed for somatic MMR variants. Of 1979 germline variants found in the leukocyte DNA of 34 sLS patients, one was a pathogenic variant (MLH1 c.1667+1delG). Leukocyte DNA of 11 patients with MLH1 hypermethylated tumors was negative for pathogenic germline variants in the tested CRC susceptibility genes and for germline MLH1 hypermethylation. Somatic DNA analysis of 28 sLS tumors identified eight (29%) cases with two pathogenic somatic variants, one with a VUS predicted to pathogenic and LOH, and nine cases (32%) with one pathogenic somatic variant (n = 8) or one VUS predicted to be pathogenic (n = 1). This is the first study in sLS patients to include the entire genomic sequence of CRC susceptibility genes. An underlying somatic or germline MMR gene defect was identified in ten of 34 sLS patients (29%). In the remaining sLS patients, the underlying genetic defect explaining the MMRdeficiency in their tumors might be found outside the genomic regions harboring the MMR and other known CRC susceptibility genes.

  5. Lynch syndrome caused by germline PMS2 mutations: delineating the cancer risk.

    Science.gov (United States)

    ten Broeke, Sanne W; Brohet, Richard M; Tops, Carli M; van der Klift, Heleen M; Velthuizen, Mary E; Bernstein, Inge; Capellá Munar, Gabriel; Gomez Garcia, Encarna; Hoogerbrugge, Nicoline; Letteboer, Tom G W; Menko, Fred H; Lindblom, Annika; Mensenkamp, Arjen R; Moller, Pal; van Os, Theo A; Rahner, Nils; Redeker, Bert J W; Sijmons, Rolf H; Spruijt, Liesbeth; Suerink, Manon; Vos, Yvonne J; Wagner, Anja; Hes, Frederik J; Vasen, Hans F; Nielsen, Maartje; Wijnen, Juul T

    2015-02-01

    The clinical consequences of PMS2 germline mutations are poorly understood compared with other Lynch-associated mismatch repair gene (MMR) mutations. The aim of this European cohort study was to define the cancer risk faced by PMS2 mutation carriers. Data were collected from 98 PMS2 families ascertained from family cancer clinics that included a total of 2,548 family members and 377 proven mutation carriers. To adjust for potential ascertainment bias, a modified segregation analysis model was used to calculate colorectal cancer (CRC) and endometrial cancer (EC) risks. Standardized incidence ratios (SIRs) were calculated to estimate risks for other Lynch syndrome-associated cancers. The cumulative risk (CR) of CRC for male mutation carriers by age 70 years was 19%. The CR among female carriers was 11% for CRC and 12% for EC. The mean age of CRC development was 52 years, and there was a significant difference in mean age of CRC between the probands (mean, 47 years; range, 26 to 68 years) and other family members with a PMS2 mutation (mean, 58 years; range, 31 to 86 years; P PMS2 mutation, and it should be noted that we observed a substantial variation in cancer phenotype within and between families, suggesting the influence of genetic modifiers and lifestyle factors on cancer risks. © 2014 by American Society of Clinical Oncology.

  6. Doxycyclin ameliorates a starvation-induced germline tumor in C. elegans daf-18/PTEN mutant background.

    Science.gov (United States)

    Wolf, Tim; Qi, Wenjing; Schindler, Verena; Runkel, Eva Diana; Baumeister, Ralf

    2014-08-01

    Managing available resources is a key necessity of each organism to cope with the environment. The nematode C. elegans responds to nutritional deprivation or harsh environmental conditions with a multitude of developmental adaptations, among them a starvation-induced quiescence at early larval development (L1). daf-18, the C. elegans homolog of the human tumor suppressor gene PTEN, is essential for the maintenance of survival and germline stem cell arrest during the L1 diapause. We show here that daf-18 mutants, independently to their failure to maintain G2 arrest of the primordial germ cells, develop a gonad phenotype after refeeding. This highly penetrant gonadal phenotype is further enhanced by a mutation in shc-1, encoding a protein homologous to the human adaptor ShcA. Features of this phenotype are a tumor-like phenotype encompassing hyper-proliferation of germ cell nuclei and disruption/invasion of the basement membrane surrounding the gonad. The penetrance of this phenotype is reduced by decreasing starvation temperature. In addition, it is also ameliorated in a dose-dependent way by exposure to the antibiotic doxycyclin either during starvation or during subsequent refeeding. Since, in eukaryotic cells, doxycyclin specifically blocks mitochondrial translation, our results suggest that daf-18 and shc-1;daf-18 mutants fail to adapt mitochondrial activity to reduced nutritional availability during early larval developing. Copyright © 2014 Elsevier Inc. All rights reserved.

  7. Gene therapy: theoretical and bioethical concepts.

    Science.gov (United States)

    Smith, Kevin R

    2003-01-01

    Gene therapy holds great promise. Somatic gene therapy has the potential to treat a wide range of disorders, including inherited conditions, cancers, and infectious diseases. Early progress has already been made in the treatment of a range of disorders. Ethical issues surrounding somatic gene therapy are primarily those concerned with safety. Germline gene therapy is theoretically possible but raises serious ethical concerns concerning future generations.

  8. A Model of Evolution of Development Based on Germline Penetration of New “No-Junk” DNA

    Directory of Open Access Journals (Sweden)

    Borys Wróbel

    2012-08-01

    Full Text Available There is a mounting body of evidence that somatic transposition may be involved in normal development of multicellular organisms and in pathology, especially cancer. Epigenetic Tracking (ET is an abstract model of multicellular development, able to generate complex 3-dimensional structures. Its aim is not to model the development of a particular organism nor to merely summarise mainstream knowledge on genetic regulation of development. Rather, the goal of ET is to provide a theoretical framework to test new postulated genetic mechanisms, not fully established yet in mainstream biology. The first proposal is that development is orchestrated through a subset of cells which we call driver cells. In these cells, the cellular state determines a specific pattern of gene activation which leads to the occurrence of developmental events. The second proposal is that evolution of development is affected by somatic transposition events. We postulate that when the genome of a driver cell does not specify what developmental event should be undertaken when the cell is in a particular cellular state, somatic transposition events can reshape the genome, build new regulatory regions, and lead to a new pattern of gene activation in the cell. Our third hypothesis, not supported yet by direct evidence, but consistent with some experimental observations, is that these new “no-junk” sequences—regulatory regions created by transposable elements at new positions in the genome—can exit the cell and enter the germline, to be incorporated in the genome of the progeny. We call this mechanism germline penetration. This process allows heritable incorporation of novel developmental events in the developmental trajectory. In this paper we will present the model and link these three postulated mechanisms to biological observations.

  9. A novel germ-line point mutation in RET exon 8 (Gly(533)Cys) in a large kindred with familial medullary thyroid carcinoma

    OpenAIRE

    Silva, Adriana Madeira Alvares da [UNIFESP; Maciel, Rui Monteiro de Barros [UNIFESP; Dias-da-Silva, Magnus Régios [UNIFESP; Toledo, Silvia Regina Caminada de [UNIFESP; De Carvalho, Marcos B.; Cerutti, Janete Maria [UNIFESP

    2003-01-01

    Familial medullary thyroid carcinoma is related to germ-line mutations in the RET oncogene, mainly in cysteine codon 10 or 11, whereas noncysteine mutations in codons 13 - 15 are rare. We now report a new missense point mutation in exon 8 of the RET gene (1597G-->T) corresponding to a Gly(533)Cys substitution in the cystein-rich domain of RET protein in 76 patients from a 6-generation Brazilian family with 229 subjects, with ascendants from Spain. It is likely that the mutation causes familia...

  10. Molecular and phenotypic abnormalities in individuals with germline heterozygous PTEN mutations and autism.

    Science.gov (United States)

    Frazier, T W; Embacher, R; Tilot, A K; Koenig, K; Mester, J; Eng, C

    2015-09-01

    PTEN is a tumor suppressor associated with an inherited cancer syndrome and an important regulator of ongoing neural connectivity and plasticity. The present study examined molecular and phenotypic characteristics of individuals with germline heterozygous PTEN mutations and autism spectrum disorder (ASD) (PTEN-ASD), with the aim of identifying pathophysiologic markers that specifically associate with PTEN-ASD and that may serve as targets for future treatment trials. PTEN-ASD patients (n=17) were compared with idiopathic (non-PTEN) ASD patients with (macro-ASD, n=16) and without macrocephaly (normo-ASD, n=38) and healthy controls (n=14). Group differences were evaluated for PTEN pathway protein expression levels, global and regional structural brain volumes and cortical thickness measures, neurocognition and adaptive behavior. RNA expression patterns and brain characteristics of a murine model of Pten mislocalization were used to further evaluate abnormalities observed in human PTEN-ASD patients. PTEN-ASD had a high proportion of missense mutations and showed reduced PTEN protein levels. Compared with the other groups, prominent white-matter and cognitive abnormalities were specifically associated with PTEN-ASD patients, with strong reductions in processing speed and working memory. White-matter abnormalities mediated the relationship between PTEN protein reductions and reduced cognitive ability. The Pten(m3m4) murine model had differential expression of genes related to myelination and increased corpus callosum. Processing speed and working memory deficits and white-matter abnormalities may serve as useful features that signal clinicians that PTEN is etiologic and prompting referral to genetic professionals for gene testing, genetic counseling and cancer risk management; and could reveal treatment targets in trials of treatments for PTEN-ASD.

  11. Germline transformation of the western corn rootworm, Diabrotica virgifera virgifera.

    Science.gov (United States)

    Chu, F; Klobasa, W; Wu, P; Pinzi, S; Grubbs, N; Gorski, S; Cardoza, Y; Lorenzen, M D

    2017-08-01

    The western corn rootworm (WCR), a major pest of maize, is notorious for rapidly adapting biochemically, behaviourally and developmentally to a variety of control methods. Despite much effort, the genetic basis of WCR adaptation remains a mystery. Since transformation-based applications such as transposon tagging and enhancer trapping have facilitated genetic dissection of model species such as Drosophila melanogaster, we developed a germline-transformation system for WCR in an effort to gain a greater understanding of the basic biology of this economically important insect. Here we report the use of a fluorescent-marked Minos element to create transgenic WCR. We demonstrate that the transgenic strains express both an eye-specific fluorescent marker and piggyBac transposase. We identified insertion-site junction sequences via inverse PCR and assessed insertion copy number using digital droplet PCR (ddPCR). Interestingly, most WCR identified as transgenic via visual screening for DsRed fluorescence proved to carry multiple Minos insertions when tested via ddPCR. A total of eight unique insertion strains were created by outcrossing the initial transgenic strains to nontransgenic WCR mates. Establishing transgenic technologies for this beetle is the first step towards bringing a wide range of transformation-based tools to bear on understanding WCR biology. © 2017 The Royal Entomological Society.

  12. In vitro propagation of male germline stem cells from piglets.

    Science.gov (United States)

    Zheng, Yi; Tian, Xiue; Zhang, Yaqing; Qin, Jinzhou; An, Junhui; Zeng, Wenxian

    2013-07-01

    To study the effects of serum and growth factors on propagation of porcine male germline stem cells (MGSCs) in vitro and develop a culture system for these stem cells. Fresh testicular cells from neonatal piglets were obtained by mechanical dissociation and collagenase-trypsin digestion. After differential plating, non-adhering cells were cultured in media supplemented with different concentrations of serum (0, 1 %, 2 %, 5 %, 10 %). After 10 days of primary culture, the cells were maintained in media supplemented with different concentrations of growth factors (basic fibroblast growth factor and epidermal growth factor at 1, 5, 10 ng/ml). The number of MGSC-derived colonies with different sizes was determined in each treatment to assess the effects of serum concentrations and growth factors. The number of MGSC-derived colonies was significantly higher in the presence of 1 % rather than 10 % fetal bovine serum (FBS). Basic fibroblast growth factor (bFGF) at 1, 5 ng/ml and epidermal growth factor (EGF) at 5, 10 ng/ml significantly promoted colony formation. Immunocytochemistry, reverse transcriptase-polymerase chain reaction (RT-PCR) and xenotransplantation assays demonstrated the presence of functional stem cells in cultured cell population. In vitro propagation of porcine MGSCs could be maintained in the presence of 1 % FBS and supplementation of growth factors for 1 month.

  13. Germline-specific H1 variants: the "sexy" linker histones.

    Science.gov (United States)

    Pérez-Montero, Salvador; Carbonell, Albert; Azorín, Fernando

    2016-03-01

    The eukaryotic genome is packed into chromatin, a nucleoprotein complex mainly formed by the interaction of DNA with the abundant basic histone proteins. The fundamental structural and functional subunit of chromatin is the nucleosome core particle, which is composed by 146 bp of DNA wrapped around an octameric protein complex formed by two copies of each core histone H2A, H2B, H3, and H4. In addition, although not an intrinsic component of the nucleosome core particle, linker histone H1 directly interacts with it in a monomeric form. Histone H1 binds nucleosomes near the exit/entry sites of linker DNA, determines nucleosome repeat length and stabilizes higher-order organization of nucleosomes into the ∼30 nm chromatin fiber. In comparison to core histones, histone H1 is less well conserved through evolution. Furthermore, histone H1 composition in metazoans is generally complex with most species containing multiple variants that play redundant as well as specific functions. In this regard, a characteristic feature is the presence of specific H1 variants that replace somatic H1s in the germline and during early embryogenesis. In this review, we summarize our current knowledge about their structural and functional properties.

  14. BRCA1 and BRCA2 Germline Mutations in Asian and European Populations

    Directory of Open Access Journals (Sweden)

    Ute Hamann

    2017-02-01

    Full Text Available Women who carry a pathogenic mutation in the breast cancer susceptibility genes BRCA1 or BRCA2 (BRCA have markedly increased risks of developing breast and ovarian cancers during their lifetime. It has been estimated that their breast and ovarian cancer risks are in the range of 46-87% and 15-68%, respectively. Therefore it is of utmost clinical importance to identify BRCA mutation carriers in order to target unaffected women for prevention and/or close surveillance and to help affected women choose the best chemotherapy regimen. Genetic testing for BRCA germline mutations is expanding in clinical oncology centers worldwide. Given the high costs of complete BRCA gene screens, a lot of effort has been expended on deciding upon whom to test. Relevant issues involved in decision making include the prior probability of a woman having a BRCA mutation, which is a function of her age and her disease status, her ethnic group, and her family history of breast or ovarian cancer. The frequency and spectrum of mutations in these genes show considerable variation by ethnic groups and by geographic regions. Most studies have been conducted in European and North American populations, while studies in Asian, Hispanic, and African populations are fewer. In most populations, many BRCA mutations were identified, which were distributed all over the genes. However, in some populations, a relatively small number of specific BRCA mutations are recurrent and account for the majority of all mutations in that population. Many of the recurrent mutations are founder mutations, which were derived from a common ancestor. Founder mutations are present in Ashkenazi Jewish, European, and Islander (Faroe, Easter, and Pitcairn populations. Such mutations have also been identified in patients from several Asian, South American, and African countries. Population-specific genetic risk assessment and genetic mutation screening have been facilitated at low costs. Given that mutations

  15. Gene

    Data.gov (United States)

    U.S. Department of Health & Human Services — Gene integrates information from a wide range of species. A record may include nomenclature, Reference Sequences (RefSeqs), maps, pathways, variations, phenotypes,...

  16. High-resolution melting (HRM) assay for the detection of recurrent BRCA1/BRCA2 germline mutations in Tunisian breast/ovarian cancer families.

    Science.gov (United States)

    Riahi, Aouatef; Kharrat, Maher; Lariani, Imen; Chaabouni-Bouhamed, Habiba

    2014-12-01

    Germline deleterious mutations in the BRCA1/BRCA2 genes are associated with an increased risk for the development of breast and ovarian cancer. Given the large size of these genes the detection of such mutations represents a considerable technical challenge. Therefore, the development of cost-effective and rapid methods to identify these mutations became a necessity. High resolution melting analysis (HRM) is a rapid and efficient technique extensively employed as high-throughput mutation scanning method. The purpose of our study was to assess the specificity and sensitivity of HRM for BRCA1 and BRCA2 genes scanning. As a first step we estimate the ability of HRM for detection mutations in a set of 21 heterozygous samples harboring 8 different known BRCA1/BRCA2 variations, all samples had been preliminarily investigated by direct sequencing, and then we performed a blinded analysis by HRM in a set of 68 further sporadic samples of unknown genotype. All tested heterozygous BRCA1/BRCA2 variants were easily identified. However the HRM assay revealed further alteration that we initially had not searched (one unclassified variant). Furthermore, sequencing confirmed all the HRM detected mutations in the set of unknown samples, including homozygous changes, indicating that in this cohort, with the optimized assays, the mutations detections sensitivity and specificity were 100 %. HRM is a simple, rapid and efficient scanning method for known and unknown BRCA1/BRCA2 germline mutations. Consequently the method will allow for the economical screening of recurrent mutations in Tunisian population.

  17. Detection of induced male germline mutation: Correlations and comparisons between traditional germline mutation assays, transgenic rodent assays and expanded simple tandem repeat instability assays

    Energy Technology Data Exchange (ETDEWEB)

    Singer, Timothy M. [Mutagenesis Section, Environmental and Occupational Toxicology Division, Safe Environments Programme, 0803A, Health Canada, Ottawa, Ont., K1A 0K9 (Canada); Department of Biology, Carleton University, 1125 Colonel By Drive, Ottawa, Ont., K1S 5B6 (Canada); Lambert, Iain B. [Department of Biology, Carleton University, 1125 Colonel By Drive, Ottawa, Ont., K1S 5B6 (Canada); Williams, Andrew [Biostatistics and Epidemiology Division, Safe Environments Programme, 6604B, Health Canada, Ottawa, Ont., K1A 0K9 (Canada); Douglas, George R. [Mutagenesis Section, Environmental and Occupational Toxicology Division, Safe Environments Programme, 0803A, Health Canada, Ottawa, Ont., K1A 0K9 (Canada); Yauk, Carole L. [Mutagenesis Section, Environmental and Occupational Toxicology Division, Safe Environments Programme, 0803A, Health Canada, Ottawa, Ont., K1A 0K9 (Canada)]. E-mail: carole_yauk@hc-sc.gc.ca

    2006-06-25

    Several rodent assays are capable of monitoring germline mutation. These include traditional assays, such as the dominant lethal (DL) assay, the morphological specific locus (SL) test and the heritable translocation (HT) assay, and two assays that have been developed more recently-the expanded simple tandem repeat (ESTR) and transgenic rodent (TGR) mutation assays. In this paper, we have compiled the limited amount of experimental data that are currently available to make conclusions regarding the comparative ability of the more recently developed assays to detect germline mutations induced by chemical and radiological agents. The data suggest that ESTR and TGR assays are generally comparable with SL in detecting germline mutagenicity induced by alkylating agents and radiation, though TGR offered less sensitivity than ESTR in some cases. The DL and HT assays detect clastogenic events and are most susceptible to mutations arising in post-spermatogonial cells, and they may not provide the best comparisons with TGR and ESTR instability. The measurement of induced ESTR instability represents a relatively sensitive method of identifying agents causing germline mutation in rodents, and may also be useful for bio-monitoring exposed individuals in the human population. Any future use of the TGR and ESTR germline mutation assays in a regulatory testing context will entail more robust and extensive characterization of assay performance. This will require substantially more data, including experiments measuring multiple endpoints, a greatly expanded database of chemical agents and a focus on characterizing stage-specific activity of mutagens in these assays, preferably by sampling epididymal sperm exposed at defined pre-meiotic, meiotic and post-meiotic stages of development.

  18. Detection of induced male germline mutation: Correlations and comparisons between traditional germline mutation assays, transgenic rodent assays and expanded simple tandem repeat instability assays

    International Nuclear Information System (INIS)

    Singer, Timothy M.; Lambert, Iain B.; Williams, Andrew; Douglas, George R.; Yauk, Carole L.

    2006-01-01

    Several rodent assays are capable of monitoring germline mutation. These include traditional assays, such as the dominant lethal (DL) assay, the morphological specific locus (SL) test and the heritable translocation (HT) assay, and two assays that have been developed more recently-the expanded simple tandem repeat (ESTR) and transgenic rodent (TGR) mutation assays. In this paper, we have compiled the limited amount of experimental data that are currently available to make conclusions regarding the comparative ability of the more recently developed assays to detect germline mutations induced by chemical and radiological agents. The data suggest that ESTR and TGR assays are generally comparable with SL in detecting germline mutagenicity induced by alkylating agents and radiation, though TGR offered less sensitivity than ESTR in some cases. The DL and HT assays detect clastogenic events and are most susceptible to mutations arising in post-spermatogonial cells, and they may not provide the best comparisons with TGR and ESTR instability. The measurement of induced ESTR instability represents a relatively sensitive method of identifying agents causing germline mutation in rodents, and may also be useful for bio-monitoring exposed individuals in the human population. Any future use of the TGR and ESTR germline mutation assays in a regulatory testing context will entail more robust and extensive characterization of assay performance. This will require substantially more data, including experiments measuring multiple endpoints, a greatly expanded database of chemical agents and a focus on characterizing stage-specific activity of mutagens in these assays, preferably by sampling epididymal sperm exposed at defined pre-meiotic, meiotic and post-meiotic stages of development

  19. Complete Genome Sequence of Germline Chromosomally Integrated Human Herpesvirus 6A and Analyses Integration Sites Define a New Human Endogenous Virus with Potential to Reactivate as an Emerging Infection.

    OpenAIRE

    Tweedy, J; Spyrou, MA; Pearson, M; Lassner, D; Kuhl, U; Gompels, UA

    2016-01-01

    Human herpesvirus-6A and B (HHV-6A, HHV-6B) have recently defined endogenous genomes, resulting from integration into the germline: chromosomally-integrated "CiHHV-6A/B". These affect approximately 1.0% of human populations, giving potential for virus gene expression in every cell. We previously showed that CiHHV-6A was more divergent than CiHHV-6B by examining four genes in 44 European CiHHV-6A/B cardiac/haematology patients. There was evidence for gene expression/reactivation, imp...

  20. RAD51C germline mutations in breast and ovarian cancer cases from high-risk families.

    Directory of Open Access Journals (Sweden)

    Jessica Clague

    Full Text Available BRCA1 and BRCA2 are the most well-known breast cancer susceptibility genes. Additional genes involved in DNA repair have been identified as predisposing to breast cancer. One such gene, RAD51C, is essential for homologous recombination repair. Several likely pathogenic RAD51C mutations have been identified in BRCA1- and BRCA2-negative breast and ovarian cancer families. We performed complete sequencing of RAD51C in germline DNA of 286 female breast and/or ovarian cancer cases with a family history of breast and ovarian cancers, who had previously tested negative for mutations in BRCA1 and BRCA2. We screened 133 breast cancer cases, 119 ovarian cancer cases, and 34 with both breast and ovarian cancers. Fifteen DNA sequence variants were identified; including four intronic, one 5' UTR, one promoter, three synonymous, and six non-synonymous variants. None were truncating. The in-silico SIFT and Polyphen programs were used to predict possible pathogenicity of the six non-synonomous variants based on sequence conservation. G153D and T287A were predicted to be likely pathogenic. Two additional variants, A126T and R214C alter amino acids in important domains of the protein such that they could be pathogenic. Two-hybrid screening and immunoblot analyses were performed to assess the functionality of these four non-synonomous variants in yeast. The RAD51C-G153D protein displayed no detectable interaction with either XRCC3 or RAD51B, and RAD51C-R214C displayed significantly decreased interaction with both XRCC3 and RAD51B (p<0.001. Immunoblots of RAD51C-Gal4 activation domain fusion peptides showed protein levels of RAD51C-G153D and RAD51C-R214C that were 50% and 60% of the wild-type, respectively. Based on these data, the RAD51C-G153D variant is likely to be pathogenic, while the RAD51C- R214C variant is hypomorphic of uncertain pathogenicity. These results provide further support that RAD51C is a rare breast and ovarian cancer susceptibility gene.

  1. A germline mutation in the BRCA1 3'UTR predicts Stage IV breast cancer.

    Science.gov (United States)

    Dorairaj, Jemima J; Salzman, David W; Wall, Deirdre; Rounds, Tiffany; Preskill, Carina; Sullivan, Catherine A W; Lindner, Robert; Curran, Catherine; Lezon-Geyda, Kim; McVeigh, Terri; Harris, Lyndsay; Newell, John; Kerin, Michael J; Wood, Marie; Miller, Nicola; Weidhaas, Joanne B

    2014-06-10

    A germline, variant in the BRCA1 3'UTR (rs8176318) was previously shown to predict breast and ovarian cancer risk in women from high-risk families, as well as increased risk of triple negative breast cancer. Here, we tested the hypothesis that this variant predicts tumor biology, like other 3'UTR mutations in cancer. The impact of the BRCA1-3'UTR-variant on BRCA1 gene expression, and altered response to external stimuli was tested in vitro using a luciferase reporter assay. Gene expression was further tested in vivo by immunoflourescence staining on breast tumor tissue, comparing triple negative patient samples with the variant (TG or TT) or non-variant (GG) BRCA1 3'UTR. To determine the significance of the variant on clinically relevant endpoints, a comprehensive collection of West-Irish breast cancer patients were tested for the variant. Finally, an association of the variant with breast screening clinical phenotypes was evaluated using a cohort of women from the High Risk Breast Program at the University of Vermont. Luciferase reporters with the BRCA1-3'UTR-variant (T allele) displayed significantly lower gene expression, as well as altered response to external hormonal stimuli, compared to the non-variant 3'UTR (G allele) in breast cancer cell lines. This was confirmed clinically by the finding of reduced BRCA1 gene expression in triple negative samples from patients carrying the homozygous TT variant, compared to non-variant patients. The BRCA1-3'UTR-variant (TG or TT) also associated with a modest increased risk for developing breast cancer in the West-Irish cohort (OR=1.4, 95% CI 1.1-1.8, p=0.033). More importantly, patients with the BRCA1-3'UTR-variant had a 4-fold increased risk of presenting with Stage IV disease (p=0.018, OR=3.37, 95% CI 1.3-11.0). Supporting that this finding is due to tumor biology, and not difficulty screening, obese women with the BRCA1-3'UTR-variant had significantly less dense breasts (p=0.0398) in the Vermont cohort. A variant in

  2. Novel somatic and germline mutations in intracranial germ cell tumours.

    Science.gov (United States)

    Wang, Linghua; Yamaguchi, Shigeru; Burstein, Matthew D; Terashima, Keita; Chang, Kyle; Ng, Ho-Keung; Nakamura, Hideo; He, Zongxiao; Doddapaneni, Harshavardhan; Lewis, Lora; Wang, Mark; Suzuki, Tomonari; Nishikawa, Ryo; Natsume, Atsushi; Terasaka, Shunsuke; Dauser, Robert; Whitehead, William; Adekunle, Adesina; Sun, Jiayi; Qiao, Yi; Marth, Gábor; Muzny, Donna M; Gibbs, Richard A; Leal, Suzanne M; Wheeler, David A; Lau, Ching C

    2014-07-10

    Intracranial germ cell tumours (IGCTs) are a group of rare heterogeneous brain tumours that are clinically and histologically similar to the more common gonadal GCTs. IGCTs show great variation in their geographical and gender distribution, histological composition and treatment outcomes. The incidence of IGCTs is historically five- to eightfold greater in Japan and other East Asian countries than in Western countries, with peak incidence near the time of puberty. About half of the tumours are located in the pineal region. The male-to-female incidence ratio is approximately 3-4:1 overall, but is even higher for tumours located in the pineal region. Owing to the scarcity of tumour specimens available for research, little is currently known about this rare disease. Here we report the analysis of 62 cases by next-generation sequencing, single nucleotide polymorphism array and expression array. We find the KIT/RAS signalling pathway frequently mutated in more than 50% of IGCTs, including novel recurrent somatic mutations in KIT, its downstream mediators KRAS and NRAS, and its negative regulator CBL. Novel somatic alterations in the AKT/mTOR pathway included copy number gains of the AKT1 locus at 14q32.33 in 19% of patients, with corresponding upregulation of AKT1 expression. We identified loss-of-function mutations in BCORL1, a transcriptional co-repressor and tumour suppressor. We report significant enrichment of novel and rare germline variants in JMJD1C, which codes for a histone demethylase and is a coactivator of the androgen receptor, among Japanese IGCT patients. This study establishes a molecular foundation for understanding the biology of IGCTs and suggests potentially promising therapeutic strategies focusing on the inhibition of KIT/RAS activation and the AKT1/mTOR pathway.

  3. Novel somatic and germline mutations in intracranial germ cell tumors

    Science.gov (United States)

    Wang, Linghua; Yamaguchi, Shigeru; Burstein, Matthew D.; Terashima, Keita; Chang, Kyle; Ng, Ho-Keung; Nakamura, Hideo; He, Zongxiao; Doddapaneni, Harshavardhan; Lewis, Lora; Wang, Mark; Suzuki, Tomonari; Nishikawa, Ryo; Natsume, Atsushi; Terasaka, Shunsuke; Dauser, Robert; Whitehead, William; Adekunle, Adesina; Sun, Jiayi; Qiao, Yi; Marth, Gábor; Muzny, Donna M.; Gibbs, Richard A.; Leal, Suzanne M.; Wheeler, David A.; Lau, Ching C.

    2015-01-01

    Intracranial germ cell tumors (IGCTs) are a group of rare heterogeneous brain tumors which are clinically and histologically similar to the more common gonadal GCTs. IGCTs show great variation in their geographic and gender distribution, histological composition and treatment outcomes. The incidence of IGCTs is historically 5–8 fold greater in Japan and other East Asian countries than in Western countries1 with peak incidence near the time of puberty2. About half of the tumors are located in the pineal region. The male-to-female incidence ratio is approximately 3–4:1 overall but even higher for tumors located in the pineal region3. Due to the scarcity of tumor specimens available for research, little is currently known about this rare disease. Here we report the analysis of 62 cases by next generation sequencing, SNP array and expression array. We find the KIT/RAS signaling pathway frequently mutated in over 50% of IGCTs including novel recurrent somatic mutations in KIT, its downstream mediators KRAS and NRAS, and its negative regulator CBL. Novel somatic alterations in the AKT/mTOR pathway included copy number gain of the AKT1 locus at 14q32.33 in 19% of patients, with corresponding upregulation of AKT1 expression. We identified loss-of-function mutations in BCORL1, a transcriptional corepressor and tumor suppressor. We report significant enrichment of novel and rare germline variants in JMJD1C, a histone demethylase and coactivator of the androgen receptor, among Japanese IGCT patients. This study establishes a molecular foundation for understanding the biology of IGCTs and suggests potentially promising therapeutic strategies focusing on the inhibition of KIT/RAS activation and the AKT1/mTOR pathway. PMID:24896186

  4. THE GERMLINE STEM CELL NICHE UNIT IN MAMMALIAN TESTES

    Science.gov (United States)

    Oatley, Jon M.; Brinster, Ralph L.

    2014-01-01

    This review addresses current understanding of the germline stem cell niche unit in mammalian testes. Spermatogenesis is a classic model of tissue-specific stem cell function relying on self-renewal and differentiation of spermatogonial stem cells (SSCs). These fate decisions are influenced by a niche microenvironment composed of a growth factor milieu that is provided by several testis somatic support cell populations. Investigations over the last two decades have identified key determinants of the SSC niche including cytokines that regulate SSC functions and support cells providing these factors, adhesion molecules that influence SSC homing, and developmental heterogeneity of the niche during postnatal aging. Emerging evidence suggests that Sertoli cells are a key support cell population influencing the formation and function of niches by secreting soluble factors and possibly orchestrating contributions of other support cells. Investigations with mice have shown that niche influence on SSC proliferation differs during early postnatal development and adulthood. Moreover, there is mounting evidence of an age-related decline in niche function, which is likely influenced by systemic factors. Defining the attributes of stem cell niches is key to developing methods to utilize these cells for regenerative medicine. The SSC population and associated niche comprise a valuable model system for study that provides fundamental knowledge about the biology of tissue-specific stem cells and their capacity to sustain homeostasis of regenerating tissue lineages. While the stem cell is essential for maintenance of all self-renewing tissues and has received considerable attention, the role of niche cells is at least as important and may prove to be more receptive to modification in regenerative medicine. PMID:22535892

  5. Deep sequencing of atrial fibrillation patients with mitral valve regurgitation shows no evidence of mosaicism but reveals novel rare germline variants

    DEFF Research Database (Denmark)

    Gregers, Emilie; Ahlberg, Gustav; Christensen, Thea

    2017-01-01

    the HaloPlex Target Enrichment System. MuTect software was used for identification of somatic point variants. We functionally characterized selected variants using electrophysiologic techniques. RESULTS: No somatic variants were identified in the cardiac tissue. Thirty-three patients (75%) had a rare...... patient population undergoing surgery for mitral valve regurgitation (MVR) to determine whether these patients are genetically predisposed to AF. METHODS: DNA was extracted from blood and left atrial tissue from 44 AF patients with MVR. Using next-generation sequencing, we investigated 110 genes using...... germline variation in ≥1 candidate genes. Fourteen variants were novel. Fifteen variants were predicted damaging or likely damaging in ≥6 in silico predictions. We identified rare variants in genes never directly associated with AF: KCNE4, SCN4B, NEURL1, and CAND2. Interestingly, 7 patients (16%) had...

  6. Pan-cancer analysis reveals technical artifacts in TCGA germline variant calls.

    Science.gov (United States)

    Buckley, Alexandra R; Standish, Kristopher A; Bhutani, Kunal; Ideker, Trey; Lasken, Roger S; Carter, Hannah; Harismendy, Olivier; Schork, Nicholas J

    2017-06-12

    Cancer research to date has largely focused on somatically acquired genetic aberrations. In contrast, the degree to which germline, or inherited, variation contributes to tumorigenesis remains unclear, possibly due to a lack of accessible germline variant data. Here we called germline variants on 9618 cases from The Cancer Genome Atlas (TCGA) database representing 31 cancer types. We identified batch effects affecting loss of function (LOF) variant calls that can be traced back to differences in the way the sequence data were generated both within and across cancer types. Overall, LOF indel calls were more sensitive to technical artifacts than LOF Single Nucleotide Variant (SNV) calls. In particular, whole genome amplification of DNA prior to sequencing led to an artificially increased burden of LOF indel calls, which confounded association analyses relating germline variants to tumor type despite stringent indel filtering strategies. The samples affected by these technical artifacts include all acute myeloid leukemia and practically all ovarian cancer samples. We demonstrate how technical artifacts induced by whole genome amplification of DNA can lead to false positive germline-tumor type associations and suggest TCGA whole genome amplified samples be used with caution. This study draws attention to the need to be sensitive to problems associated with a lack of uniformity in data generation in TCGA data.

  7. Pathological assessment of mismatch repair gene variants in Lynch syndrome

    DEFF Research Database (Denmark)

    Rasmussen, Lene Juel; Heinen, Christopher D; Royer-Pokora, Brigitte

    2012-01-01

    Lynch syndrome (LS) is caused by germline mutations in DNA mismatch repair (MMR) genes and is the most prevalent hereditary colorectal cancer syndrome. A significant proportion of variants identified in MMR and other common cancer susceptibility genes are missense or noncoding changes whose...

  8. APC gene mutations and extraintestinal phenotype of familial adenomatous polyposis

    NARCIS (Netherlands)

    Giardiello, F. M.; Petersen, G. M.; Piantadosi, S.; Gruber, S. B.; Traboulsi, E. I.; Offerhaus, G. J.; Muro, K.; Krush, A. J.; Booker, S. V.; Luce, M. C.; Laken, S. J.; Kinzler, K. W.; Vogelstein, B.; Hamilton, S. R.

    1997-01-01

    Familial adenomatous polyposis (FAP) is caused by germline mutation of the adenomatous polyposis coli (APC) gene on chromosome 5q. This study assessed genotype-phenotype correlations for extraintestinal lesions in FAP. Mutations of the APC gene were compared with the occurrence of seven

  9. Pediatric oncologist willingness to offer germline TP53 testing in osteosarcoma.

    Science.gov (United States)

    Shaul, Eliana; Roth, Michael; Lo, Yungtai; Geller, David S; Hoang, Bang; Yang, Rui; Malkin, David; Gorlick, Richard; Gill, Jonathan

    2018-03-15

    Li-Fraumeni syndrome (LFS) is a cancer predisposition syndrome caused by mutations in the tumor-suppressor gene TP53. Osteosarcoma is a sentinel cancer in LFS. Prior studies using Sanger sequencing platforms have demonstrated that 3% of individuals with osteosarcoma harbor a mutation in TP53. New data from next-generation sequencing have demonstrated that 3.8% of patients with osteosarcoma have a known pathogenic variant, and an additional 5.7% carry exonic variants of unknown significance in TP53. Pediatric oncologists were e-mailed an anonymous 18-question survey assessing their willingness to offer TP53 germline testing to a child with osteosarcoma with or without a family history, and they were evaluated for changes in their choices with the prior data and the new data. One hundred seventy-seven pediatric oncologists (22%) responded to the survey. Respondents were more likely to offer TP53 testing to a patient with a positive family history (77.4% vs 12.4%; P offer TP53 testing once they were provided with the new data (25.4% vs 12.4%; P = .0038). The proportion of providers who responded that they were unsure increased significantly when they were presented with the new data (25.4% vs 10.2%; P = .0002). Potential implications for other family members and the possibility that surveillance imaging would detect new malignancies at an earlier stage were important factors influencing a provider's decision to offer TP53 testing. Recent data increase the proportion of providers willing to offer testing, and this suggests concern on the part of pediatric oncologists that variants of unknown significance may be disease-defining in rare cancers. Cancer 2018;124:1242-50. © 2018 American Cancer Society. © 2018 American Cancer Society.

  10. The clinical phenotype of Lynch syndrome due to germline PMS2 mutations

    Science.gov (United States)

    Senter, Leigha; Clendenning, Mark; Sotamaa, Kaisa; Hampel, Heather; Green, Jane; Potter, John D.; Lindblom, Annika; Lagerstedt, Kristina; Thibodeau, Stephen N.; Lindor, Noralane M.; Young, Joanne; Winship, Ingrid; Dowty, James G.; White, Darren M.; Hopper, John L.; Baglietto, Laura; Jenkins, Mark A.; de la Chapelle, Albert

    2009-01-01

    Background and Aims Although the clinical phenotype of Lynch syndrome (also known as Hereditary Nonpolyposis Colorectal Cancer) has been well described, little is known about disease in PMS2 mutation carriers. Now that mutation detection methods can discern mutations in PMS2 from mutations in its pseudogenes, more mutation carriers have been identified. Information about the clinical significance of PMS2 mutations is crucial for appropriate counseling. Here, we report the clinical characteristics of a large series of PMS2 mutation carriers. Methods We performed PMS2 mutation analysis using long range PCR and MLPA for 99 probands diagnosed with Lynch syndrome-associated tumors showing isolated loss of PMS2 by immunohistochemistry. Penetrance was calculated using a modified segregation analysis adjusting for ascertainment. Results Germline PMS2 mutations were detected in 62% of probands (n = 55 monoallelic; 6 biallelic). Among families with monoallelic PMS2 mutations, 65.5% met revised Bethesda guidelines. Compared with the general population, in mutation carriers, the incidence of colorectal cancer was 5.2 fold higher and the incidence of endometrial cancer was 7.5 fold higher. In North America, this translates to a cumulative cancer risk to age 70 of 15–20% for colorectal cancer, 15% for endometrial cancer, and 25–32% for any Lynch syndrome-associated cancer. No elevated risk for non-Lynch syndrome-associated cancers was observed. Conclusions PMS2 mutations contribute significantly to Lynch syndrome but the penetrance for monoallelic mutation carriers appears to be lower than that for the other mismatch repair genes. Modified counseling and cancer surveillance guidelines for PMS2 mutation carriers are proposed. PMID:18602922

  11. The clinical phenotype of Lynch syndrome due to germ-line PMS2 mutations.

    Science.gov (United States)

    Senter, Leigha; Clendenning, Mark; Sotamaa, Kaisa; Hampel, Heather; Green, Jane; Potter, John D; Lindblom, Annika; Lagerstedt, Kristina; Thibodeau, Stephen N; Lindor, Noralane M; Young, Joanne; Winship, Ingrid; Dowty, James G; White, Darren M; Hopper, John L; Baglietto, Laura; Jenkins, Mark A; de la Chapelle, Albert

    2008-08-01

    Although the clinical phenotype of Lynch syndrome (also known as hereditary nonpolyposis colorectal cancer) has been well described, little is known about disease in PMS2 mutation carriers. Now that mutation detection methods can discern mutations in PMS2 from mutations in its pseudogenes, more mutation carriers have been identified. Information about the clinical significance of PMS2 mutations is crucial for appropriate counseling. Here, we report the clinical characteristics of a large series of PMS2 mutation carriers. We performed PMS2 mutation analysis using long-range polymerase chain reaction and multiplex ligation-dependent probe amplification for 99 probands diagnosed with Lynch syndrome-associated tumors showing isolated loss of PMS2 by immunohistochemistry. Penetrance was calculated using a modified segregation analysis adjusting for ascertainment. Germ-line PMS2 mutations were detected in 62% of probands (n = 55 monoallelic; 6 biallelic). Among families with monoallelic PMS2 mutations, 65.5% met revised Bethesda guidelines. Compared with the general population, in mutation carriers, the incidence of colorectal cancer was 5.2-fold higher, and the incidence of endometrial cancer was 7.5-fold higher. In North America, this translates to a cumulative cancer risk to age 70 years of 15%-20% for colorectal cancer, 15% for endometrial cancer, and 25%-32% for any Lynch syndrome-associated cancer. No elevated risk for non-Lynch syndrome-associated cancers was observed. PMS2 mutations contribute significantly to Lynch syndrome, but the penetrance for monoallelic mutation carriers appears to be lower than that for the other mismatch repair genes. Modified counseling and cancer surveillance guidelines for PMS2 mutation carriers are proposed.

  12. Autosomal dominant hypoparathyroidism caused by germline mutation in GNA11: phenotypic and molecular characterization.

    Science.gov (United States)

    Li, Dong; Opas, Evan E; Tuluc, Florin; Metzger, Daniel L; Hou, Cuiping; Hakonarson, Hakon; Levine, Michael A

    2014-09-01

    Most cases of autosomal dominant hypoparathyroidism (ADH) are caused by gain-of-function mutations in CASR or dominant inhibitor mutations in GCM2 or PTH. Our objectives were to identify the genetic basis for ADH in a multigenerational family and define the underlying disease mechanism. Here we evaluated a multigenerational family with ADH in which affected subjects had normal sequences in these genes and were shorter than unaffected family members. We collected clinical and biochemical data from 6 of 11 affected subjects and performed whole-exome sequence analysis on DNA from two affected sisters and their affected father. Functional studies were performed after expression of wild-type and mutant Gα11 proteins in human embryonic kidney-293-CaR cells that stably express calcium-sensing receptors. Whole-exome-sequencing followed by Sanger sequencing revealed a heterozygous mutation, c.179G>T; p.R60L, in GNA11, which encodes the α-subunit of G11, the principal heterotrimeric G protein that couples calcium-sensing receptors to signal activation in parathyroid cells. Functional studies of Gα11 R60L showed increased accumulation of intracellular concentration of free calcium in response to extracellular concentration of free calcium with a significantly decreased EC50 compared with wild-type Gα11. By contrast, R60L was significantly less effective than the oncogenic Q209L form of Gα11 as an activator of the MAPK pathway. Compared to subjects with CASR mutations, patients with GNA11 mutations lacked hypercalciuria and had normal serum magnesium levels. Our findings indicate that the germline gain-of-function mutation of GNA11 is a cause of ADH and implicate a novel role for GNA11 in skeletal growth.

  13. Association of the germline TP53 R337H mutation with breast cancer in southern Brazil

    Directory of Open Access Journals (Sweden)

    Srivastava Kumar

    2008-12-01

    Full Text Available Abstract Background The germline TP53-R337H mutation is strongly associated with pediatric adrenocortical tumors (ACT in southern Brazil; it has low penetrance and limited tissue specificity in most families and therefore is not associated with Li-Fraumeni syndrome. However, other tumor types, mainly breast cancer, have been observed in carriers of several unrelated kindreds, raising the possibility that the R337H mutation may also contribute to breast tumorigenesis in a genetic background-specific context. Methods We conducted a case-control study to determine the prevalence of the R337H mutation by sequencing TP53 exon 10 in 123 women with breast cancer and 223 age- and sex-matched control subjects from southern Brazil. Fisher's test was used to compare the prevalence of the R337H. Results The R337H mutation was found in three patients but in none of the controls (p = 0.0442. Among the carriers, two had familial history of cancer meeting the Li-Fraumeni-like criteria. Remarkably, tumors in each of these three cases underwent loss of heterozygosity by eliminating the mutant TP53 allele rather than the wild-type allele. Polymorphisms were identified within the TP53 (R72P and Ins16 and MDM2 (SNP309 genes that may further diminish TP53 tumor suppressor activity. Conclusion These results demonstrate that the R337H mutation can significantly increase the risk of breast cancer in carriers, which likely depends on additional cooperating genetic factors. These findings are also important for understanding how low-penetrant mutant TP53 alleles can differentially influence tumor susceptibility.

  14. Association of the germline TP53 R337H mutation with breast cancer in southern Brazil

    International Nuclear Information System (INIS)

    Assumpção, Juliana G; Zeferino, Luiz Carlos; Dufloth, Rozany M; Brandalise, Silvia Regina; Yunes, José Andres; Seidinger, Ana Luíza; Mastellaro, Maria José; Ribeiro, Raul C; Zambetti, Gerard P; Ganti, Ramapriya; Srivastava, Kumar; Shurtleff, Sheila; Pei, Deqing

    2008-01-01

    The germline TP53-R337H mutation is strongly associated with pediatric adrenocortical tumors (ACT) in southern Brazil; it has low penetrance and limited tissue specificity in most families and therefore is not associated with Li-Fraumeni syndrome. However, other tumor types, mainly breast cancer, have been observed in carriers of several unrelated kindreds, raising the possibility that the R337H mutation may also contribute to breast tumorigenesis in a genetic background-specific context. We conducted a case-control study to determine the prevalence of the R337H mutation by sequencing TP53 exon 10 in 123 women with breast cancer and 223 age- and sex-matched control subjects from southern Brazil. Fisher's test was used to compare the prevalence of the R337H. The R337H mutation was found in three patients but in none of the controls (p = 0.0442). Among the carriers, two had familial history of cancer meeting the Li-Fraumeni-like criteria. Remarkably, tumors in each of these three cases underwent loss of heterozygosity by eliminating the mutant TP53 allele rather than the wild-type allele. Polymorphisms were identified within the TP53 (R72P and Ins16) and MDM2 (SNP309) genes that may further diminish TP53 tumor suppressor activity. These results demonstrate that the R337H mutation can significantly increase the risk of breast cancer in carriers, which likely depends on additional cooperating genetic factors. These findings are also important for understanding how low-penetrant mutant TP53 alleles can differentially influence tumor susceptibility

  15. Thyroid cancer in a patient with a germline MSH2 mutation. Case report and review of the Lynch syndrome expanding tumour spectrum

    Directory of Open Access Journals (Sweden)

    Stulp Rein P

    2008-02-01

    Full Text Available Abstract Lynch syndrome (HNPCC is a dominantly inherited disorder characterized by germline defects in DNA mismatch repair (MMR genes and the development of a variety of cancers, predominantly colorectal and endometrial. We present a 44-year-old woman who was shown to carry the truncating MSH2 gene mutation that had previously been identified in her family. Recently, she had been diagnosed with an undifferentiated carcinoma of the thyroid and an adenoma of her coecum. Although the thyroid carcinoma was not MSI-high (1 out of 5 microsatellites instable, it did show complete loss of immunohistochemical expression for the MSH2 protein, suggesting that this tumour was not coincidental. Although the risks for some tumour types, including breast cancer, soft tissue sarcoma and prostate cancer, are not significantly increased in Lynch syndrome, MMR deficiency in the presence of a corresponding germline defect has been demonstrated in incidental cases of a growing range of tumour types, which is reviewed in this paper. Interestingly, the MSH2-associated tumour spectrum appears to be wider than that of MLH1 and generally the risk for most extra-colonic cancers appears to be higher for MSH2 than for MLH1 mutation carriers. Together with a previously reported case, our findings show that anaplastic thyroid carcinoma can develop in the setting of Lynch syndrome. Uncommon Lynch syndrome-associated tumour types might be useful in the genetic analysis of a Lynch syndrome suspected family if samples from typical Lynch syndrome tumours are unavailable.

  16. Experimental evidence showing that no mitotically active female germline progenitors exist in postnatal mouse ovaries.

    Science.gov (United States)

    Zhang, Hua; Zheng, Wenjing; Shen, Yan; Adhikari, Deepak; Ueno, Hiroo; Liu, Kui

    2012-07-31

    It has been generally accepted for more than half a century that, in most mammalian species, oocytes cannot renew themselves in postnatal or adult life, and that the number of oocytes is already fixed in fetal or neonatal ovaries. This assumption, however, has been challenged over the past decade. In this study, we have taken an endogenous genetic approach to this question and generated a multiple fluorescent Rosa26(rbw/+);Ddx4-Cre germline reporter mouse model for in vivo and in vitro tracing of the development of female germline cell lineage. Through live cell imaging and de novo folliculogenesis experiments, we show that the Ddx4-expressing cells from postnatal mouse ovaries did not enter mitosis, nor did they contribute to oocytes during de novo folliculogenesis. Our results provide evidence that supports the traditional view that no postnatal follicular renewal occurs in mammals, and no mitotically active Ddx4-expressing female germline progenitors exist in postnatal mouse ovaries.

  17. CSR-1 and P granules suppress sperm-specific transcription in the C. elegans germline.

    Science.gov (United States)

    Campbell, Anne C; Updike, Dustin L

    2015-05-15

    Germ granules (P granules) in C. elegans are required for fertility and function to maintain germ cell identity and pluripotency. Sterility in the absence of P granules is often accompanied by the misexpression of soma-specific proteins and the initiation of somatic differentiation in germ cells. To investigate whether this is caused by the accumulation of somatic transcripts, we performed mRNA-seq on dissected germlines with and without P granules. Strikingly, we found that somatic transcripts do not increase in the young adult germline when P granules are impaired. Instead, we found that impairing P granules causes sperm-specific mRNAs to become highly overexpressed. This includes the accumulation of major sperm protein (MSP) transcripts in germ cells, a phenotype that is suppressed by feminization of the germline. A core component of P granules, the endo-siRNA-binding Argonaute protein CSR-1, has recently been ascribed with the ability to license transcripts for germline expression. However, impairing CSR-1 has very little effect on the accumulation of its mRNA targets. Instead, we found that CSR-1 functions with P granules to prevent MSP and sperm-specific mRNAs from being transcribed in the hermaphrodite germline. These findings suggest that P granules protect germline integrity through two different mechanisms, by (1) preventing the inappropriate expression of somatic proteins at the level of translational regulation, and by (2) functioning with CSR-1 to limit the domain of sperm-specific expression at the level of transcription. © 2015. Published by The Company of Biologists Ltd.

  18. Development of Spontaneous Mammary Tumors in BALB/c-p53+-Mice: Detection of Early Genetic Alterations and the Mapping of BALB/c Susceptibility Genes

    National Research Council Canada - National Science Library

    Blackburn, Anneke

    2002-01-01

    The TP53 tumor suppressor gene is defective in the majority of sporadic breast cancers, and breast cancer is the most frequent tumor type in women with Li-Fraumeni syndrome who inherit germline mutations in TP53...

  19. Development of Spontaneous Mammary Tumors in BALB/c-p53+/-Mice: Detection of Early Genetic Alterations and the Mapping of BALB/c Susceptibility Genes

    National Research Council Canada - National Science Library

    Smith, Sallie

    2004-01-01

    The TP53 tumor suppressor gene is defective in the majority of sporadic breast cancers, and breast cancer is the most frequent tumor type in women with Li-Fraumeni syndrome and bear germline mutations in TP53...

  20. Identification of Germline Genetic Mutations in Pancreatic Cancer Patients

    Science.gov (United States)

    Salo-Mullen, Erin E.; O’Reilly, Eileen; Kelsen, David; Ashraf, Asad M.; Lowery, Maeve; Yu, Kenneth; Reidy, Diane; Epstein, Andrew S.; Lincoln, Anne; Saldia, Amethyst; Jacobs, Lauren M.; Rau-Murthy, Rohini; Zhang, Liying; Kurtz, Robert; Saltz, Leonard; Offit, Kenneth; Robson, Mark; Stadler, Zsofia K.

    2016-01-01

    Background Pancreatic adenocarcinoma (PAC) is part of several cancer predisposition syndromes; however, indications for genetic counseling/testing are not well-defined. We sought to determine mutation prevalence and characteristics that predict for inherited predisposition to PAC. Methods We identified 175 consecutive PAC patients who underwent clinical genetics assessment at Memorial Sloan Kettering between 2011–2014. Clinical data, family history, and germline results were evaluated. Results Among 159 PAC patients who pursued genetic testing, 24 pathogenic mutations were identified (15.1%; 95%CI, 9.5%–20.7%), including BRCA2(n=13), BRCA1(n=4), p16(n=2), PALB2(n=1), and Lynch syndrome(n=4). BRCA1/BRCA2 prevalence was 13.7% in Ashkenazi Jewish(AJ) (n=95) and 7.1% in non-AJ(n=56) patients. In AJ patients with strong, weak, or absent family history of BRCA-associated cancers, mutation prevalence was 16.7%, 15.8%, and 7.4%, respectively. Mean age at diagnosis in all mutation carriers was 58.5y(range 45–75y) compared to 64y(range 27–87y) in non-mutation carriers(P=0.02). Although BRCA2 was the most common mutation identified, no patients with early-onset PAC(≤50y) harbored a BRCA2 mutation and the mean age at diagnosis in BRCA2 carriers was equivalent to non-mutation carriers(P=0.34). Mutation prevalence in early-onset patients(n=21) was 28.6%, including BRCA1(n=2), p16(n=2), MSH2(n=1) and MLH1(n=1). Conclusion Mutations in BRCA2 account for over 50% of PAC patients with an identified susceptibility syndrome. AJ patients had high BRCA1/BRCA2 prevalence regardless of personal/family history, suggesting that ancestry alone indicates a need for genetic evaluation. With the exception of BRCA2-associated PAC, inherited predisposition to PAC is associated with earlier age at PAC diagnosis suggesting that this subset of patients may also represent a population warranting further evaluation. PMID:26440929

  1. Von Hippel-Lindau (VHL inactivation in sporadic clear cell renal cancer: associations with germline VHL polymorphisms and etiologic risk factors.

    Directory of Open Access Journals (Sweden)

    Lee E Moore

    2011-10-01

    Full Text Available Renal tumor heterogeneity studies have utilized the von Hippel-Lindau VHL gene to classify disease into molecularly defined subtypes to examine associations with etiologic risk factors and prognosis. The aim of this study was to provide a comprehensive analysis of VHL inactivation in clear cell renal tumors (ccRCC and to evaluate relationships between VHL inactivation subgroups with renal cancer risk factors and VHL germline single nucleotide polymorphisms (SNPs. VHL genetic and epigenetic inactivation was examined among 507 sporadic RCC/470 ccRCC cases using endonuclease scanning and using bisulfite treatment and Sanger sequencing across 11 CpG sites within the VHL promoter. Case-only multivariate analyses were conducted to identify associations between alteration subtypes and risk factors. VHL inactivation, either through sequence alterations or promoter methylation in tumor DNA, was observed among 86.6% of ccRCC cases. Germline VHL SNPs and a haplotype were associated with promoter hypermethylation in tumor tissue (OR = 6.10; 95% CI: 2.28-16.35, p = 3.76E-4, p-global = 8E-5. Risk of having genetic VHL inactivation was inversely associated with smoking due to a higher proportion of wild-type ccRCC tumors [former: OR = 0.70 (0.20-1.31 and current: OR = 0.56 (0.32-0.99; P-trend = 0.04]. Alteration prevalence did not differ by histopathologic characteristics or occupational exposure to trichloroethylene. ccRCC cases with particular VHL germline polymorphisms were more likely to have VHL inactivation through promoter hypermethylation than through sequence alterations in tumor DNA, suggesting that the presence of these SNPs may represent an example of facilitated epigenetic variation (an inherited propensity towards epigenetic variation in renal tissue. A proportion of tumors from current smokers lacked VHL alterations and may represent a biologically distinct clinical entity from inactivated cases.

  2. Localization of early germ cells in a stony coral, Euphyllia ancora: potential implications for a germline stem cell system in coral gametogenesis

    Science.gov (United States)

    Shikina, Shinya; Chung, Yi-Jou; Wang, Hsiang-Ming; Chiu, Yi-Ling; Shao, Zih-Fang; Lee, Yan-Horn; Chang, Ching-Fong

    2015-06-01

    Most corals exhibit annual or multiple gametogenic cycles. Thus far, coral gametogenesis has been studied in many species and locations during the past three decades; however, currently, only a few papers exist that describe the origin of germ cells, such as germline stem cells (GSCs), which support the continuous production of gametes in every reproductive cycle. To address this issue, in this study, we focused on and identified piwi gene, which has been used as a marker of germline cells, including GSCs, in various metazoans, in a scleractinian coral, Euphyllia ancora. Reverse-transcription PCR and Western blotting analyses revealed that E. ancora piwi-like ( Eapiwi) is expressed in mesentery tissues where the sites of gametogenesis are located for both sexes. Immunohistochemistry with a specific antibody against Eapiwi revealed strong immunoreactivity in the spermatogonia in males and in the oogonia and early oocytes in females, demonstrating that Eapiwi could be used as an early germ cell marker in E. ancora. Subsequent immunohistochemical analyses regarding the spatial and temporal distribution patterns of early germ cells in mesentery tissues revealed that early germ cells were present throughout the year in the mesentery tissue we examined, regardless of the sexual reproductive cycle. In particular, small numbers of early germ cells were observed in specific sites of mesentery tissues with fully matured gonads in both sexes. These early germ cells were not released together with mature gametes during the spawning period and remained in the mesentery tissues. These results suggested that these early germ cells most likely serve as a reservoir of germline cells and that some of these cells would produce differentiated germ cells for the upcoming sexual reproduction period; hence, these cells would function as GSCs. Our data provide new information for understanding continuous gamete production in corals.

  3. Late manifestation of subclinical hyperthyroidism after goitrogenesis in an index patient with a N670S TSH receptor germline mutation masquerading as TSH receptor antibody negative Graves' disease.

    Science.gov (United States)

    Schaarschmidt, J; Paschke, S; Özerden, M; Jäschke, H; Huth, S; Eszlinger, M; Meller, J; Paschke, R

    2012-12-01

    In 27 families with familial non-autoimmune hyperthyroidism (FNAH) reported up to date, the onset of hyperthyroidism varies from 18 months to 60 years. Also the manifestation of goitres is variable in these families. A 74-year-old woman first presented at the age of 69 years with tachyarrhythmia and hypertension. After initial treatment of her hypertension and oral anticoagulation for her intermittent atrial fibrillation, a thyroid workup revealed a suppressed TSH and normal fT3 and fT4. TPO, TSH receptor (TSHR), and thyroglobulin antibodies were negative. Thyroid ultrasound revealed a thyroid volume of 102 ml with several nodules with diameters of up to 2.6 cm right and up to 1.8 cm left. Scintigraphy showed a homogeneous Technetium-99 m ((99 m)Tc) uptake of 1.27%. She was subsequently treated with 1 GBq radioiodine ((131)I). At the age of 74, her thyroid function was normal and her thyroid volume decreased to 90 ml. Because of the diffuse (99 m)Tc uptake and the negative TPO, TSHR, and thyroglobulin antibodies, genetic analysis of her TSHR gene was performed, in spite of her negative family history for hyperthyroidism. Sequencing revealed a N670S TSHR germline mutation. Previous in vitro characterisation of this TSHR mutation suggests a weak constitutive activity, yet the experimental data are ambiguous. This case illustrates the necessity to analyse patients with hyperthyroidism accompanied by diffuse (99 m)Tc uptake and negative TPO, TSHR, and thyroglobulin antibodies for TSHR germline mutations. Moreover, it demonstrates that TSHR germline mutations may first lead to longstanding nodular goitrogenesis before the late manifestation of subclinical hyperthyroidism. © Georg Thieme Verlag KG Stuttgart · New York.

  4. Differential impact of the HEN1 homolog HENN-1 on 21U and 26G RNAs in the germline of Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Leonie M Kamminga

    Full Text Available RNA interference (RNAi-related pathways affect gene activity by sequence-specific recruitment of Ago proteins to mRNA target molecules. The sequence specificity of this process stems from small RNA (sRNA co-factors bound by the Ago protein. Stability of sRNA molecules in some pathways is in part regulated by Hen1-mediated methylation of their 3' ends. Here we describe the effects of the Caenorhabditis elegans HEN1 RNA-methyl-transferase homolog, HENN-1, on the different RNAi pathways in this nematode. We reveal differential effects of HENN-1 on the two pathways that are known to employ methylated sRNA molecules: the 26G and 21U pathways. Surprisingly, in the germline, stability of 21U RNAs, the C. elegans piRNAs, is only mildly affected by loss of methylation; and introduction of artificial 21U target RNA does not further destabilize non-methylated 21U RNAs. In contrast, most 26G RNAs display reduced stability and respond to loss of HENN-1 by displaying increased 3'-uridylation frequencies. Within the 26G RNA class, we find that specifically ERGO-1-bound 26G RNAs are modified by HENN-1, while ALG-3/ALG-4-bound 26G RNAs are not. Global gene expression analysis of henn-1 mutants reveals mild effects, including down-regulation of many germline-expressed genes. Our data suggest that, apart from direct effects of reduced 26G RNA levels of henn-1 on gene expression, most effects on global gene expression are indirect. These studies further refine our understanding of endogenous RNAi in C. elegans and the roles for Hen1 like enzymes in these pathways.

  5. Recurrence and Variability of Germline EPCAM Deletions in Lynch Syndrome

    NARCIS (Netherlands)

    Kuiper, Roland P.; Vissers, Lisenka E. L. M.; Venkatachalam, Ramprasath; Bodmer, Danielle; Hoenselaar, Eveline; Goossens, Monique; Haufe, Aline; Kamping, Eveline; Niessen, Renee C.; Hogervorst, Frans B. L.; Gille, Johan J. P.; Redeker, Bert; Tops, Carli M. J.; van Gijn, Marielle E.; van den Ouweland, Ans M. W.; Rahner, Nils; Steinke, Verena; Kahl, Philip; Holinski-Feder, Elke; Morak, Monika; Kloor, Matthias; Stemmler, Susanne; Betz, Beate; Hutter, Pierre; Bunyan, David J.; Syngal, Sapna; Culver, Julie O.; Graham, Tracy; Chan, Tsun L.; Nagtegaal, Iris D.; van Krieken, J. Han J. M.; Schackert, Hans K.; Hoogerbrugge, Nicoline; van Kessel, Ad Geurts; Ligtenberg, Marjolijn J. L.

    Recently, we identified 3' end deletions in the EPCAM gene as a novel cause of Lynch syndrome. These truncating EPCAM deletions cause allele-specific epigenetic silencing of the neighboring DNA mismatch repair gene MSH2 in tissues expressing EPCAM. Here we screened a cohort of unexplained Lynch-like

  6. Breast cancer in patients carrying a germ-line CHEK2 mutation: Outcome after breast conserving surgery and adjuvant radiotherapy

    International Nuclear Information System (INIS)

    Meyer, Andreas; Doerk, Thilo; Sohn, Christof; Karstens, Johann H.; Bremer, Michael

    2007-01-01

    Background and purpose: Women carrying mutations in the CHEK2 gene are at an increased breast cancer risk. Data about outcome and prognosis for these patients after standard multimodality treatment are scarce at present. Materials and methods: One-hundred and fifty (150) patients with non-metastasized early-stage breast cancer (T1-2) receiving postoperative radiotherapy following breast-conservative surgery at our department were included in this analysis. Carriers were identified using mutation-specific restriction enzyme-based screening assays in previous investigations. Twenty-five breast cancer patients were heterozygous for one of three CHEK2 gene mutations (I157T, n = 13; 1100delC, n = 10; IVS2+1G > A, n = 2). The comparison group consisted of 125 early-stage breast cancer patients without a CHEK2 gene mutation (non-carriers). Median follow-up was 87 months for the total cohort of patients. Results: Local recurrences occurred in 13 patients (carriers, 3 (12%); non-carriers, 10 (8%)) and distant metastases occurred in 27 patients (carriers, 8 (32%); non-carriers, 19 (15%)). Twenty-five patients had deceased (carriers, 8 (32%); non-carriers, 17 (14%)) with all but 3 deaths related to breast cancer. Actuarial 7-year local relapse-free survival was 86% in carriers versus 90% in non-carriers (p = 0.48). Actuarial metastasis-free, disease-free and overall survival at 7 years were 64% vs. 84% (p = 0.045), 59% vs. 78% (p = 0.07) and 69% vs. 87% (p = 0.10), respectively. In a multivariate step-wise Cox regression analysis presence of a CHEK2 mutation remained a borderline significant discriminator for metastasis-free survival (p = 0.048; OR = 0.4; 95% CI 0.2-1.0) next to T-stage (p = 0.001; OR 0.3; 95% CI 0.1-0.6). Conclusions: Heterozygosity for a germline CHEK2 mutation appears to represent an adverse prognostic factor in patients with early-stage breast cancer. If confirmed in larger studies these data may serve as a basis for future surveillance and treatment

  7. Germline or somatic GPR101 duplication leads to X-linked acrogigantism: a clinico-pathological and genetic study.

    Science.gov (United States)

    Iacovazzo, Donato; Caswell, Richard; Bunce, Benjamin; Jose, Sian; Yuan, Bo; Hernández-Ramírez, Laura C; Kapur, Sonal; Caimari, Francisca; Evanson, Jane; Ferraù, Francesco; Dang, Mary N; Gabrovska, Plamena; Larkin, Sarah J; Ansorge, Olaf; Rodd, Celia; Vance, Mary L; Ramírez-Renteria, Claudia; Mercado, Moisés; Goldstone, Anthony P; Buchfelder, Michael; Burren, Christine P; Gurlek, Alper; Dutta, Pinaki; Choong, Catherine S; Cheetham, Timothy; Trivellin, Giampaolo; Stratakis, Constantine A; Lopes, Maria-Beatriz; Grossman, Ashley B; Trouillas, Jacqueline; Lupski, James R; Ellard, Sian; Sampson, Julian R; Roncaroli, Federico; Korbonits, Márta

    2016-06-01

    Non-syndromic pituitary gigantism can result from AIP mutations or the recently identified Xq26.3 microduplication causing X-linked acrogigantism (XLAG). Within Xq26.3, GPR101 is believed to be the causative gene, and the c.924G > C (p.E308D) variant in this orphan G protein-coupled receptor has been suggested to play a role in the pathogenesis of acromegaly.We studied 153 patients (58 females and 95 males) with pituitary gigantism. AIP mutation-negative cases were screened for GPR101 duplication through copy number variation droplet digital PCR and high-density aCGH. The genetic, clinical and histopathological features of XLAG patients were studied in detail. 395 peripheral blood and 193 pituitary tumor DNA samples from acromegaly patients were tested for GPR101 variants.We identified 12 patients (10 females and 2 males; 7.8 %) with XLAG. In one subject, the duplicated region only contained GPR101, but not the other three genes in found to be duplicated in the previously reported patients, defining a new smallest region of overlap of duplications. While females presented with germline mutations, the two male patients harbored the mutation in a mosaic state. Nine patients had pituitary adenomas, while three had hyperplasia. The comparison of the features of XLAG, AIP-positive and GPR101&AIP-negative patients revealed significant differences in sex distribution, age at onset, height, prolactin co-secretion and histological features. The pathological features of XLAG-related adenomas were remarkably similar. These tumors had a sinusoidal and lobular architecture. Sparsely and densely granulated somatotrophs were admixed with lactotrophs; follicle-like structures and calcifications were commonly observed. Patients with sporadic of familial acromegaly did not have an increased prevalence of the c.924G > C (p.E308D) GPR101 variant compared to public databases.In conclusion, XLAG can result from germline or somatic duplication of GPR101. Duplication of GPR101

  8. PTCH1 Germline Mutations and the Basaloid Follicular Hamartoma Values in the Tumor Spectrum of Basal Cell Carcinoma Syndrome (NBCCS).

    Science.gov (United States)

    Ponti, Giovanni; Manfredini, Marco; Pastorino, Lorenza; Maccaferri, Monia; Tomasi, Aldo; Pellacani, Giovanni

    2018-01-01

    Nevoid basal cell carcinoma syndrome (NBCCS) is an autosomal dominantly inherited disorder characterized by multiple basal cell carcinomas (BCC), odontogenic tumors and various skeletal anomalies. Basaloid follicular hamartomas (BFHs) constitute rare neoplasms that can be detected in sporadic and familial settings as in the Basaloid Follicular Hamartoma Syndrome (BFHS). Although BFHS shares clinical, histopathological and genetic overlapping with the NBCCS, they are still considered two distinctive entities. The aim of our single-institution study was the analysis of a cohort of PTCH1-mutated patients in order to define clinical and biomolecular relationship between NBCCS and BFHs. In our study we evaluated PTCH1 gene-carrier probands affected by NBCCS to detect the incidence of BFHs and their correlation with this rare syndrome. Among probands we recognized 4 patients with BFHs. We found 15 germline PTCH1 mutations, uniformly distributed across the PTCH1 gene. Six of them had familial history of NBCCS, two of them were novel and have not been described previously. NBCCS and BFHS may be the same genetic entity and not two distinctive syndromes. The inclusion of BFH in the NBCCS cutaneous tumor spectrum might be useful for the recognition of misdiagnosed NBCCS cases that could benefit from tailored surveillance strategies. Copyright© 2018, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  9. Germline transgenic pigs by Sleeping Beauty transposition in porcine zygotes and targeted integration in the pig genome.

    Directory of Open Access Journals (Sweden)

    Wiebke Garrels

    Full Text Available Genetic engineering can expand the utility of pigs for modeling human diseases, and for developing advanced therapeutic approaches. However, the inefficient production of transgenic pigs represents a technological bottleneck. Here, we assessed the hyperactive Sleeping Beauty (SB100X transposon system for enzyme-catalyzed transgene integration into the embryonic porcine genome. The components of the transposon vector system were microinjected as circular plasmids into the cytoplasm of porcine zygotes, resulting in high frequencies of transgenic fetuses and piglets. The transgenic animals showed normal development and persistent reporter gene expression for >12 months. Molecular hallmarks of transposition were confirmed by analysis of 25 genomic insertion sites. We demonstrate germ-line transmission, segregation of individual transposons, and continued, copy number-dependent transgene expression in F1-offspring. In addition, we demonstrate target-selected gene insertion into transposon-tagged genomic loci by Cre-loxP-based cassette exchange in somatic cells followed by nuclear transfer. Transposase-catalyzed transgenesis in a large mammalian species expands the arsenal of transgenic technologies for use in domestic animals and will facilitate the development of large animal models for human diseases.

  10. Germline rearrangements in families with strong family history of glioma and malignant melanoma, colon, and breast cancer

    Science.gov (United States)

    Andersson, Ulrika; Wibom, Carl; Cederquist, Kristina; Aradottir, Steina; Borg, Åke; Armstrong, Georgina N.; Shete, Sanjay; Lau, Ching C.; Bainbridge, Matthew N.; Claus, Elizabeth B.; Barnholtz-Sloan, Jill; Lai, Rose; Il'yasova, Dora; Houlston, Richard S.; Schildkraut, Joellen; Bernstein, Jonine L.; Olson, Sara H.; Jenkins, Robert B.; Lachance, Daniel H.; Wrensch, Margaret; Davis, Faith G.; Merrell, Ryan; Johansen, Christoffer; Sadetzki, Siegal; Bondy, Melissa L.; Melin, Beatrice S.; Adatto, Phyllis; Morice, Fabian; Payen, Sam; McQuinn, Lacey; McGaha, Rebecca; Guerra, Sandra; Paith, Leslie; Roth, Katherine; Zeng, Dong; Zhang, Hui; Yung, Alfred; Aldape, Kenneth; Gilbert, Mark; Weinberger, Jeffrey; Colman, Howard; Conrad, Charles; de Groot, John; Forman, Arthur; Groves, Morris; Levin, Victor; Loghin, Monica; Puduvalli, Vinay; Sawaya, Raymond; Heimberger, Amy; Lang, Frederick; Levine, Nicholas; Tolentino, Lori; Saunders, Kate; Thach, Thu-Trang; Iacono, Donna Dello; Sloan, Andrew; Gerson, Stanton; Selman, Warren; Bambakidis, Nicholas; Hart, David; Miller, Jonathan; Hoffer, Alan; Cohen, Mark; Rogers, Lisa; Nock, Charles J; Wolinsky, Yingli; Devine, Karen; Fulop, Jordonna; Barrett, Wendi; Shimmel, Kristen; Ostrom, Quinn; Barnett, Gene; Rosenfeld, Steven; Vogelbaum, Michael; Weil, Robert; Ahluwalia, Manmeet; Peereboom, David; Staugaitis, Susan; Schilero, Cathy; Brewer, Cathy; Smolenski, Kathy; McGraw, Mary; Naska, Theresa; Rosenfeld, Steven; Ram, Zvi; Blumenthal, Deborah T.; Bokstein, Felix; Umansky, Felix; Zaaroor, Menashe; Cohen, Avi; Tzuk-Shina, Tzeela; Voldby, Bo; Laursen, René; Andersen, Claus; Brennum, Jannick; Henriksen, Matilde Bille; Marzouk, Maya; Davis, Mary Elizabeth; Boland, Eamon; Smith, Marcel; Eze, Ogechukwu; Way, Mahalia; Lada, Pat; Miedzianowski, Nancy; Frechette, Michelle; Paleologos, Nina; Byström, Gudrun; Svedberg, Eva; Huggert, Sara; Kimdal, Mikael; Sandström, Monica; Brännström, Nikolina; Hayat, Amina; Tihan, Tarik; Zheng, Shichun; Berger, Mitchel; Butowski, Nicholas; Chang, Susan; Clarke, Jennifer; Prados, Michael; Rice, Terri; Sison, Jeannette; Kivett, Valerie; Duo, Xiaoqin; Hansen, Helen; Hsuang, George; Lamela, Rosito; Ramos, Christian; Patoka, Joe; Wagenman, Katherine; Zhou, Mi; Klein, Adam; McGee, Nora; Pfefferle, Jon; Wilson, Callie; Morris, Pagan; Hughes, Mary; Britt-Williams, Marlin; Foft, Jessica; Madsen, Julia; Polony, Csaba; McCarthy, Bridget; Zahora, Candice; Villano, John; Engelhard, Herbert; Borg, Ake; Chanock, Stephen K; Collins, Peter; Elston, Robert; Kleihues, Paul; Kruchko, Carol; Petersen, Gloria; Plon, Sharon; Thompson, Patricia; Johansen, C.; Sadetzki, S.; Melin, B.; Bondy, Melissa L.; Lau, Ching C.; Scheurer, Michael E.; Armstrong, Georgina N.; Liu, Yanhong; Shete, Sanjay; Yu, Robert K.; Aldape, Kenneth D.; Gilbert, Mark R.; Weinberg, Jeffrey; Houlston, Richard S.; Hosking, Fay J.; Robertson, Lindsay; Papaemmanuil, Elli; Claus, Elizabeth B.; Claus, Elizabeth B.; Barnholtz-Sloan, Jill; Sloan, Andrew E.; Barnett, Gene; Devine, Karen; Wolinsky, Yingli; Lai, Rose; McKean-Cowdin, Roberta; Il'yasova, Dora; Schildkraut, Joellen; Sadetzki, Siegal; Yechezkel, Galit Hirsh; Bruchim, Revital Bar-Sade; Aslanov, Lili; Sadetzki, Siegal; Johansen, Christoffer; Kosteljanetz, Michael; Broholm, Helle; Bernstein, Jonine L.; Olson, Sara H.; Schubert, Erica; DeAngelis, Lisa; Jenkins, Robert B.; Yang, Ping; Rynearson, Amanda; Andersson, Ulrika; Wibom, Carl; Henriksson, Roger; Melin, Beatrice S.; Cederquist, Kristina; Aradottir, Steina; Borg, Åke; Merrell, Ryan; Lada, Patricia; Wrensch, Margaret; Wiencke, John; Wiemels, Joe; McCoy, Lucie; McCarthy, Bridget J.; Davis, Faith G.

    2014-01-01

    Background Although familial susceptibility to glioma is known, the genetic basis for this susceptibility remains unidentified in the majority of glioma-specific families. An alternative approach to identifying such genes is to examine cancer pedigrees, which include glioma as one of several cancer phenotypes, to determine whether common chromosomal modifications might account for the familial aggregation of glioma and other cancers. Methods Germline rearrangements in 146 glioma families (from the Gliogene Consortium; http://www.gliogene.org/) were examined using multiplex ligation-dependent probe amplification. These families all had at least 2 verified glioma cases and a third reported or verified glioma case in the same family or 2 glioma cases in the family with at least one family member affected with melanoma, colon, or breast cancer.The genomic areas covering TP53, CDKN2A, MLH1, and MSH2 were selected because these genes have been previously reported to be associated with cancer pedigrees known to include glioma. Results We detected a single structural rearrangement, a deletion of exons 1-6 in MSH2, in the proband of one family with 3 cases with glioma and one relative with colon cancer. Conclusions Large deletions and duplications are rare events in familial glioma cases, even in families with a strong family history of cancers that may be involved in known cancer syndromes. PMID:24723567

  11. Global transcriptional repression in C. elegans germline precursors by regulated sequestration of TFIID component TAF-4

    Science.gov (United States)

    Guven-Ozkan, Tugba; Nishi, Yuichi; Robertson, Scott M.; Lin, Rueyling

    2008-01-01

    In C. elegans, four asymmetric divisions, beginning with the zygote (P0), generate transcriptionally repressed germline blastomeres (P1–P4) and somatic sisters that become transcriptionally active. The protein PIE-1 represses transcription in the later germline blastomeres, but not in the earlier germline blastomeres P0 and P1. We show here that OMA-1 and OMA-2, previously shown to regulate oocyte maturation, repress transcription in P0 and P1 by binding to and sequestering in the cytoplasm TAF-4, a component critical for assembly of TFIID and the pol II preinitiation complex. OMA-1/2 binding to TAF-4 is developmentally regulated, requiring phosphorylation by the DYRK kinase MBK-2, which is activated at meiosis II following fertilization. OMA-1/2 are normally degraded after the first mitosis, but ectopic expression of wildtype OMA-1 is sufficient to repress transcription in both somatic and later germline blastomeres. We propose that phosphorylation by MBK-2 serves as a developmental switch, converting OMA-1/2 from oocyte to embryo regulators. PMID:18854162

  12. Global transcriptional repression in C. elegans germline precursors by regulated sequestration of TAF-4.

    Science.gov (United States)

    Guven-Ozkan, Tugba; Nishi, Yuichi; Robertson, Scott M; Lin, Rueyling

    2008-10-03

    In C. elegans, four asymmetric divisions, beginning with the zygote (P0), generate transcriptionally repressed germline blastomeres (P1-P4) and somatic sisters that become transcriptionally active. The protein PIE-1 represses transcription in the later germline blastomeres but not in the earlier germline blastomeres P0 and P1. We show here that OMA-1 and OMA-2, previously shown to regulate oocyte maturation, repress transcription in P0 and P1 by binding to and sequestering in the cytoplasm TAF-4, a component critical for assembly of TFIID and the pol II preinitiation complex. OMA-1/2 binding to TAF-4 is developmentally regulated, requiring phosphorylation by the DYRK kinase MBK-2, which is activated at meiosis II after fertilization. OMA-1/2 are normally degraded after the first mitosis, but ectopic expression of wild-type OMA-1 is sufficient to repress transcription in both somatic and later germline blastomeres. We propose that phosphorylation by MBK-2 serves as a developmental switch, converting OMA-1/2 from oocyte to embryo regulators.

  13. A germline RET proto-oncogene mutation in multiple members of an ...

    African Journals Online (AJOL)

    Background: Multiple endocrine neoplasia type 2A (MEN2A) is a rare cancer associated-syndrome, inherited in an autosomal dominant fashion and caused by germline mutation in RET proto-oncogene. Clinical diagnosis depends on the manifestation of two or more certain endocrine tumors in an individual, such as ...

  14. TGF-β superfamily signaling in testis formation and early male germline development.

    Science.gov (United States)

    Young, Julia C; Wakitani, Shoichi; Loveland, Kate L

    2015-09-01

    The TGF-β ligand superfamily contains at least 40 members, many of which are produced and act within the mammalian testis to facilitate formation of sperm. Their progressive expression at key stages and in specific cell types determines the fertility of adult males, influencing testis development and controlling germline differentiation. BMPs are essential for the interactive instructions between multiple cell types in the early embryo that drive initial specification of gamete precursors. In the nascent foetal testis, several ligands including Nodal, TGF-βs, Activins and BMPs, serve as key masculinizing switches by regulating male germline pluripotency, somatic and germline proliferation, and testicular vascularization and architecture. In postnatal life, local production of these factors determine adult testis size by regulating Sertoli cell multiplication and differentiation, in addition to specifying germline differentiation and multiplication. Because TGF-β superfamily signaling is integral to testis formation, it affects processes that underlie testicular pathologies, including testicular cancer, and its potential to contribute to subfertility is beginning to be understood. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Promyelocytic leukaemia zinc finger maintains self-renewal of male germline stem cells (mGSCs) and its expression pattern in dairy goat testis.

    Science.gov (United States)

    Song, W; Zhu, H; Li, M; Li, N; Wu, J; Mu, H; Yao, X; Han, W; Liu, W; Hua, J

    2013-08-01

    Previous studies have shown that promyelocytic leukaemia zinc finger (PLZF) is a spermatogonia-specific transcription factor in the testis, required to regulate self-renewal and maintenance of the spermatogonia stem cell. Up to now, expression and function of PLZF in the goat testis has not been known. The objectives of this study were to investigate PLZF expression pattern in the dairy goat and its effect on male goat germline stem cell (mGSC) self-renewal and differentiation. Testis development and expression patterns of PLZF in the dairy goat were analysed by haematoxylin and eosin staining, immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR). Furthermore, effects of PLZF overexpression on mGSC self-renewal and differentiation were evaluated by quantitative RT-PCR (QRT-PCR), immunofluorescence and BrdU incorporation assay. Promyelocytic leukaemia zinc finger was essential for dairy goat testis development and expression of several proliferation and pluripotency-associated proteins including OCT4, C-MYC were upregulated by PLZF overexpression. The study demonstrated that PLZF played a key role in maintaining self-renewal of mGSCs and its overexpression enhanced expression of proliferation-associated genes. Promyelocytic leukaemia zinc finger could function in the dairy goat as well as in other species in maintaining self-renewal of germline stem cells and this study provides a model to study the mechanism on self-renewal and differentiation of mGSCs in livestock. © 2013 John Wiley & Sons Ltd.

  16. Oocyte maturation and origin of the germline as revealed by the expression of Nanos-like in the Pacific oyster Crassostrea gigas.

    Science.gov (United States)

    Xu, Rui; Li, Qi; Yu, Hong; Kong, Lingfeng

    2018-04-13

    Nanos gene plays an important role in germline development in animals. However, the molecular mechanisms involved in germline development in Mollusca, the second largest animal phylum, are still poorly understood. Here we identified the Nanos orthologue from the Pacific oyster Crassostrea gigas (Cg-Nanos-like), and investigated the expression patterns of Nanos during gametogenesis and embryogenesis in C. gigas. Tissue expression analysis showed that Cg-Nanos-like was specifically expressed in female gonads. During the reproductive cycle, the expression of Cg-Nanos-like mRNA increased matching the seasonal development of the ovarian tissues in diploids, while the expression levels were significantly lower in the ovaries of sterile triploids compared to diploids. High expression of Cg-Nanos-like transcripts were detected in early embryonic stages, while the expression significantly dropped at gastrulation and was barely detectable in veliger stages. In situ hybridization showed that Cg-Nanos-like was expressed at different stages of developing oocytes, whereas positive signals were detected only in spermatogonia during the spermatogenic cycle. These findings indicated that Cg-Nanos-like was involved in the development of germ cells, and maintenance of oocyte maturation. In early embryogenesis, the transcripts were broadly expressed; following gastrulation, the expression was restricted to two cell clumps, which might be the putative primordial germ cells (PGCs) or their precursors. Based on the results, the formation of the PGCs in C. gigas was consistent with the model of transition from epigenesis to preformation. Copyright © 2017. Published by Elsevier B.V.

  17. Familial isolated primary hyperparathyroidism associated with germline GCM2 mutations is more aggressive and has a lesser rate of biochemical cure.

    Science.gov (United States)

    El Lakis, Mustapha; Nockel, Pavel; Guan, Bin; Agarwal, Sunita; Welch, James; Simonds, William F; Marx, Stephen; Li, Yulong; Nilubol, Naris; Patel, Dhaval; Yang, Lily; Merkel, Roxanne; Kebebew, Electron

    2018-01-01

    Hereditary primary hyperparathyroidism may be syndromic or nonsyndromic (familial isolated hyperparathyroidism). Recently, germline activating mutations in the GCM2 gene were identified in a subset of familial isolated hyperparathyroidism. This study examined the clinical and biochemical characteristics and the treatment outcomes of GCM2 mutation-positive familial isolated hyperparathyroidism as compared to sporadic primary hyperparathyroidism. We performed a retrospective analysis of clinical features, parathyroid pathology, and operative outcomes in 18 patients with GCM2 germline mutations and 457 patients with sporadic primary hyperparathyroidism. Age at diagnosis, sex distribution, race/ethnicity, and preoperative serum calcium concentrations were similar between the 2 groups. The preoperative serum levels of intact parathyroid hormone was greater in patients with GCM2-associated primary hyperparathyroidism (239 ± 394 vs 136 ± 113, P = .005) as were rates of multigland disease and parathyroid carcinoma in the GCM2 group (78% vs 14.3%, P hyperparathyroidism patients have greater preoperative parathyroid hormone levels, a greater rate of multigland disease, a lesser rate of biochemical cure, and a substantial risk of parathyroid carcinoma. Knowledge of these clinical characteristics could optimize the surgical management of GCM2-associated familial isolated hyperparathyroidism. Published by Elsevier Inc.

  18. Expression and knockdown of zebrafish folliculin suggests requirement for embryonic brain morphogenesis.

    Science.gov (United States)

    Kenyon, Emma J; Luijten, Monique N H; Gill, Harmeet; Li, Nan; Rawlings, Matthew; Bull, James C; Hadzhiev, Yavor; van Steensel, Maurice A M; Maher, Eamonn; Mueller, Ferenc

    2016-07-08

    Birt-Hogg-Dubé syndrome (BHD) is a dominantly inherited familial cancer syndrome characterised by the development of benign skin fibrofolliculomas, multiple lung and kidney cysts, spontaneous pneumothorax and susceptibility to renal cell carcinoma. BHD is caused by mutations in the gene encoding Folliculin (FLCN). Little is known about what FLCN does in a healthy individual and how best to treat those with BHD. As a first approach to developing a vertebrate model for BHD we aimed to identify the temporal and spatial expression of flcn transcripts in the developing zebrafish embryo. To gain insights into the function of flcn in a whole organism system we generated a loss of function model of flcn by the use of morpholino knockdown in zebrafish. flcn is expressed broadly and upregulated in the fin bud, somites, eye and proliferative regions of the brain of the Long-pec stage zebrafish embryos. Together with knockdown phenotypes, expression analysis suggest involvement of flcn in zebrafish embryonic brain development. We have utilised the zFucci system, an in vivo, whole organism cell cycle assay to study the potential role of flcn in brain development. We found that at the 18 somite stage there was a significant drop in cells in the S-M phase of the cell cycle in flcn morpholino injected embryos with a corresponding increase of cells in the G1 phase. This was particularly evident in the brain, retina and somites of the embryo. Timelapse analysis of the head region of flcn morpholino injected and mismatch control embryos shows the temporal dynamics of cell cycle misregulation during development. In conclusion we show that zebrafish flcn is expressed in a non-uniform manner and is likely required for the maintenance of correct cell cycle regulation during embryonic development. We demonstrate the utilisation of the zFucci system in testing the role of flcn in cell proliferation and suggest a function for flcn in regulating cell proliferation in vertebrate embryonic

  19. Germline mutation in RNASEL predicts increased risk of head and neck, uterine cervix and breast cancer.

    Directory of Open Access Journals (Sweden)

    Bo Eskerod Madsen

    Full Text Available UNLABELLED: THE BACKGROUND: Ribonuclease L (RNASEL, encoding the 2'-5'-oligoadenylate (2-5A-dependent RNase L, is a key enzyme in the interferon induced antiviral and anti-proliferate pathway. Mutations in RNASEL segregate with the disease in prostate cancer families and specific genotypes are associated with an increased risk of prostate cancer. Infection by human papillomavirus (HPV is the major risk factor for uterine cervix cancer and for a subset of head and neck squamous cell carcinomas (HNSCC. HPV, Epstein Barr virus (EBV and sequences from mouse mammary tumor virus (MMTV have been detected in breast tumors, and the presence of integrated SV40 T/t antigen in breast carcinomas correlates with an aggressive phenotype and poor prognosis. A genetic predisposition could explain why some viral infections persist and induce cancer, while others disappear spontaneously. This points at RNASEL as a strong susceptibility gene. METHODOLOGY/PRINCIPAL FINDINGS: To evaluate the implication of an abnormal activity of RNase L in the onset and development of viral induced cancers, the study was initiated by searching for germline mutations in patients diagnosed with uterine cervix cancer. The rationale behind is that close to 100% of the cervix cancer patients have a persistent HPV infection, and if a defective RNase L were responsible for the lack of ability to clear the HPV infection, we would expect to find a wide spectrum of mutations in these patients, leading to a decreased RNase L activity. The HPV genotype was established in tumor DNA from 42 patients diagnosed with carcinoma of the uterine cervix and somatic tissue from these patients was analyzed for mutations by direct sequencing of all coding and regulatory regions of RNASEL. Fifteen mutations, including still uncharacterized, were identified. The genotype frequencies of selected single nucleotide polymorphisms (SNPs established in the cervix cancer patients were compared between 382 patients

  20. Expansion of B-1a cells with germline heavy chain sequence in lupus mice

    Directory of Open Access Journals (Sweden)

    Nichol E Holodick

    2016-03-01

    Full Text Available B6.Sle1.Sle2.Sle3 (B6.TC lupus-prone mice carrying the NZB allele of Cdkn2c, encoding for the cyclin-dependent kinase inhibitor P18INK4, accumulate B-1a cells due to a higher rate of proliferative self-renewal. However, it is unclear whether this affects primarily early appearing B-1a cells of fetal origin or later appearing B-1a cells that emerge from bone marrow. B-1a cells are the major source of natural autoantibodies, and it has been shown that their protective nature is associated with a germline-like sequence, which is characterized by few N-nucleotide insertions and a repertoire skewed towards rearrangements predominated during fetal life, VH11 and VH12. To determine the nature of B-1a cells expanded in B6.TC mice, we amplified immunoglobulin genes by PCR from single cells in mice. Sequencing showed a significantly higher proportion of B-1a cell antibodies display fewer N-additions in B6.TC mice than in B6 control mice. Following this lower number of N-insertions within the CDR-H3 region, the B6.TC B-1a cells display shorter CDR-H3 length than B6 B-1a cells. The absence of N-additions is a surrogate for fetal origin, as TdT expression starts after birth in mice. Therefore, our results suggest that the B-1a cell population is not only expanded in autoimmune B6.TC mice but also qualitatively different with the majority of cells from fetal origin. Accordingly, our sequencing results also demonstrated overuse of VH11 and VH12 in autoimmune B6.TC mice as compared to B6 controls. These results suggest that the development of lupus autoantibodies in these mice is coupled with skewing of the B-1a cell repertoire and possible retention of protective natural antibodies.

  1. MSH6 and PMS2 germ-line pathogenic variants implicated in Lynch syndrome are associated with breast cancer.

    Science.gov (United States)

    Roberts, Maegan E; Jackson, Sarah A; Susswein, Lisa R; Zeinomar, Nur; Ma, Xinran; Marshall, Megan L; Stettner, Amy R; Milewski, Becky; Xu, Zhixiong; Solomon, Benjamin D; Terry, Mary Beth; Hruska, Kathleen S; Klein, Rachel T; Chung, Wendy K

    2018-01-18

    PurposeAn association of Lynch syndrome (LS) with breast cancer has been long suspected; however, there have been insufficient data to address this question for each of the LS genes individually.MethodsWe conducted a retrospective review of personal and family history in 423 women with pathogenic or likely pathogenic germ-line variants in MLH1 (N = 65), MSH2 (N = 94), MSH6 (N = 140), or PMS2 (N = 124) identified via clinical multigene hereditary cancer testing. Standard incidence ratios (SIRs) of breast cancer were calculated by comparing breast cancer frequencies in our study population with those in the general population (Surveillance, Epidemiology, and End Results 18 data).ResultsWhen evaluating by gene, the age-standardized breast cancer risks for MSH6 (SIR = 2.11; 95% confidence interval (CI), 1.56-2.86) and PMS2 (SIR = 2.92; 95% CI, 2.17-3.92) were associated with a statistically significant risk for breast cancer whereas no association was observed for MLH1 (SIR = 0.87; 95% CI, 0.42-1.83) or MSH2 (SIR = 1.22; 95% CI, 0.72-2.06).ConclusionOur data demonstrate that two LS genes, MSH6 and PMS2, are associated with an increased risk for breast cancer and should be considered when ordering genetic testing for individuals who have a personal and/or family history of breast cancer.GENETICS in MEDICINE advance online publication, 18 January 2018; doi:10.1038/gim.2017.254.

  2. BRCA1 and BRCA2 germline mutations in Malaysian women with early-onset breast cancer without a family history.

    Directory of Open Access Journals (Sweden)

    Gaik Theng Toh

    Full Text Available BACKGROUND: In Asia, breast cancer is characterised by an early age of onset: In Malaysia, approximately 50% of cases occur in women under the age of 50 years. A proportion of these cases may be attributable, at least in part, to genetic components, but to date, the contribution of genetic components to breast cancer in many of Malaysia's ethnic groups has not been well-characterised. METHODOLOGY: Given that hereditary breast carcinoma is primarily due to germline mutations in one of two breast cancer susceptibility genes, BRCA1 and BRCA2, we have characterised the spectrum of BRCA mutations in a cohort of 37 individuals with early-onset disease (

  3. Whole-genome sequencing of spermatocytic tumors provides insights into the mutational processes operating in the male germline

    DEFF Research Database (Denmark)

    Giannoulatou, Eleni; Maher, Geoffrey J; Ding, Zhihao

    2017-01-01

    Adult male germline stem cells (spermatogonia) proliferate by mitosis and, after puberty, generate spermatocytes that undertake meiosis to produce haploid spermatozoa. Germ cells are under evolutionary constraint to curtail mutations and maintain genome integrity. Despite constant turnover...

  4. Germline single nucleotide polymorphisms associated with response of urothelial carcinoma to platinum-based therapy: the role of the host.

    LENUS (Irish Health Repository)

    Gallagher, D J

    2013-09-01

    Variations in urothelial carcinoma (UC) response to platinum chemotherapy are common and frequently attributed to genetic and epigenetic variations of somatic DNA. We hypothesized that variations in germline DNA may contribute to UC chemosensitivity.

  5. Production of germline ablated male pigs via Crispr/Cas editing of the NANOS2 gene

    Science.gov (United States)

    The availability of alternative models to flies, worms, and mice for studying germ cell biology is important for translating findings to higher order mammals. In this context, investigations in pigs and other livestock species can also serve to find applications for both basic biomedical research ...

  6. Activation of germline-specific genes is required for limb regeneration in the Mexican axolotl

    OpenAIRE

    Zhu, Wei; Pao, Gerald M; Satoh, Akira; Cummings, Gillian; Monaghan, James R; Harkins, Timothy T; Bryant, Susan V; Voss, S Randal; Gardiner, David M; Hunter, Tony

    2012-01-01

    The capacity for tissue and organ regeneration in humans is dwarfed by comparison to that of salamanders. Emerging evidence suggests that mechanisms learned from the early phase of salamander limb regeneration-wound healing, cellular dedifferentiation and blastemal formation-will reveal therapeutic approaches for tissue regeneration in humans. Here we describe a unique transcriptional fingerprint of regenerating limb tissue in the Mexican axolotl (Ambystoma mexicanum) that is indicative of ce...

  7. Biased Immunoglobulin Light Chain Gene Usage in the Shark.

    Science.gov (United States)

    Iacoangeli, Anna; Lui, Anita; Naik, Ushma; Ohta, Yuko; Flajnik, Martin; Hsu, Ellen

    2015-10-15

    This study of a large family of κ L chain clusters in nurse shark completes the characterization of its classical Ig gene content (two H chain isotypes, μ and ω, and four L chain isotypes, κ, λ, σ, and σ-2). The shark κ clusters are minigenes consisting of a simple VL-JL-CL array, where V to J recombination occurs over an ~500-bp interval, and functional clusters are widely separated by at least 100 kb. Six out of ~39 κ clusters are prerearranged in the germline (germline joined). Unlike the complex gene organization and multistep assembly process of Ig in mammals, each shark Ig rearrangement, somatic or in the germline, appears to be an independent event localized to the minigene. This study examined the expression of functional, nonproductive, and sterile transcripts of the κ clusters compared with the other three L chain isotypes. κ cluster usage was investigated in young sharks, and a skewed pattern of split gene expression was observed, one similar in functional and nonproductive rearrangements. These results show that the individual activation of the spatially distant κ clusters is nonrandom. Although both split and germline-joined κ genes are expressed, the latter are prominent in young animals and wane with age. We speculate that, in the shark, the differential activation of the multiple isotypes can be advantageously used in receptor editing. Copyright © 2015 by The American Association of Immunologists, Inc.

  8. Approaches to diagnose DNA mismatch repair gene defects in cancer

    DEFF Research Database (Denmark)

    Peña-Diaz, Javier; Rasmussen, Lene Juel

    2016-01-01

    development was first observed in colorectal cancer patients that carried inactivating germline mutations in MMR genes and the disease was named as hereditary non-polyposis colorectal cancer (HNPCC). Currently, a growing list of cancers is found to be MMR defective and HNPCC has been renamed Lynch syndrome...

  9. Paternal Age Explains a Major Portion of De Novo Germline Mutation Rate Variability in Healthy Individuals.

    Directory of Open Access Journals (Sweden)

    Simon L Girard

    Full Text Available De novo mutations (DNM are an important source of rare variants and are increasingly being linked to the development of many diseases. Recently, the paternal age effect has been the focus of a number of studies that attempt to explain the observation that increasing paternal age increases the risk for a number of diseases. Using disease-free familial quartets we show that there is a strong positive correlation between paternal age and germline DNM in healthy subjects. We also observed that germline CNVs do not follow the same trend, suggesting a different mechanism. Finally, we observed that DNM were not evenly distributed across the genome, which adds support to the existence of DNM hotspots.

  10. Germline mutation rates in mice following in utero exposure to diesel exhaust particles by maternal inhalation

    DEFF Research Database (Denmark)

    Ritz, Caitlin; Ruminski, Wojciech; Hougaard, Karin S.

    2011-01-01

    (PAPs) from industrial environments cause DNA damage and mutations in the sperm of adult male mice. Effects on the female and male germline during critical stages of development (in utero) are unknown. In mice, previous studies have shown that expanded simple tandem repeat (ESTR) loci exhibit high rates......The induction of inherited DNA sequence mutations arising in the germline (i.e., sperm or egg) of mice exposed in utero to diesel exhaust particles (DEPs) via maternal inhalation compared to unexposed controls was investigated in this study. Previous work has shown that particulate air pollutants...... of spontaneous mutation, making this endpoint a valuable tool for studying inherited mutation and genomic instability. In the present study, pregnant C57Bl/6 mice were exposed to 19mg/m3 DEP from gestational day 7 through 19, alongside air exposed controls. Male and female F1 offspring were raised to maturity...

  11. Induction of atherosclerosis in mice and hamsters without germline genetic engineering

    DEFF Research Database (Denmark)

    Bjørklund, Martin Mæng; Hollensen, Anne Kruse; Hagensen, Mette Kallestrup

    2014-01-01

    RATIONALE: Atherosclerosis can be achieved in animals by germline genetic engineering, leading to hypercholesterolemia, but such models are constrained to few species and strains, and they are difficult to combine with other powerful techniques involving genetic manipulation or variation. OBJECTIVE......: To develop a method for induction of atherosclerosis without germline genetic engineering. METHODS AND RESULTS: Recombinant adeno-associated viral vectors were engineered to encode gain-of-function proprotein convertase subtilisin/kexin type 9 mutants, and mice were given a single intravenous vector...... injection followed by high-fat diet feeding. Plasma proprotein convertase subtilisin/kexin type 9 and total cholesterol increased rapidly and were maintained at high levels, and after 12 weeks, mice had atherosclerotic lesions in the aorta. Histology of the aortic root showed progression of lesions...

  12. Maternal Metabolic Syndrome Programs Mitochondrial Dysfunction via Germline Changes across Three Generations

    Directory of Open Access Journals (Sweden)

    Jessica L. Saben

    2016-06-01

    Full Text Available Maternal obesity impairs offspring health, but the responsible mechanisms are not fully established. To address this question, we fed female mice a high-fat/high-sugar diet from before conception until weaning and then followed the outcomes in the next three generations of offspring, all fed a control diet. We observed that female offspring born to obese mothers had impaired peripheral insulin signaling that was associated with mitochondrial dysfunction and altered mitochondrial dynamic and complex proteins in skeletal muscle. This mitochondrial phenotype persisted through the female germline and was passed down to the second and third generations. Our results indicate that maternal programming of metabolic disease can be passed through the female germline and that the transfer of aberrant oocyte mitochondria to subsequent generations may contribute to the increased risk for developing insulin resistance.

  13. Synchronous Onset of Breast and Pancreatic Cancers: Results of Germline and Somatic Genetic Analysis

    Directory of Open Access Journals (Sweden)

    Michael Castro

    2016-07-01

    Full Text Available Background: Synchronous cancers have occasionally been detected at initial diagnosis among patients with breast and ovarian cancer. However, simultaneous coexistence and diagnosis of breast and pancreas cancer has not previously been reported. Case Report: Paternal transmission of a germline BRCA2 mutation to a patient who was diagnosed at age 40 with locally advanced breast and pancreas cancer is presented. Somatic genomic analysis of both cancers with next-generation DNA sequencing confirmed the germline result and reported a variety of variants of unknown significance alterations, of which two were present in both the breast and pancreas cancers. Discussion: The possibility that genomic alterations could have been responsible for modulating the phenotypic or clinical expression of this rare presentation is considered. The authors call attention to the practice of privatizing the clinicogenetic information gained from genetic testing and call for health policy that will facilitate sharing in order to advance the outcomes of patients diagnosed with hereditary cancers.

  14. Sexual selection, germline mutation rate and sperm competition

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    Møller AP

    2003-04-01

    Full Text Available Abstract Background An important component of sexual selection arises because females obtain viability benefits for their offspring from their mate choice. Females choosing extra-pair fertilization generally favor males with exaggerated secondary sexual characters, and extra-pair paternity increases the variance in male reproductive success. Furthermore, females are assumed to benefit from 'good genes' from extra-pair sires. How additive genetic variance in such viability genes is maintained despite strong directional selection remains an evolutionary enigma. We propose that sexual selection is associated with elevated mutation rates, changing the balance between mutation and selection, thereby increasing variance in fitness and hence the benefits to be obtained from good genes sexual selection. Two hypotheses may account for such elevated mutation: (1 Increased sperm production associated with sperm competition may increase mutation rate. (2 Mutator alleles increase mutation rates that are revealed by the expression of condition-dependent secondary sexual characters used by choosy females during their mate choice. M Petrie has independently developed the idea that mutator alleles may account for the maintenance of genetic variation in viability despite strong directional selection. Results A comparative study of birds revealed a positive correlation between mutation rate at minisatellite loci and extra-pair paternity, but not between mutation rate and relative testes mass which is a measure of relative sperm production. Minisatellite mutation rates were not related to longevity, suggesting a meiotic rather than a mitotic origin of mutations. Conclusion We found evidence of increased mutation rate in species with more intense sexual selection. Increased mutation was not associated with increased sperm production, and we suggest that species with intense sexual selection may maintain elevated mutation rates because sexual selection continuously

  15. C. elegans AMPKs promote survival and arrest germline development during nutrient stress

    Directory of Open Access Journals (Sweden)

    Masamitsu Fukuyama

    2012-08-01

    Mechanisms controlling development, growth, and metabolism are coordinated in response to changes in environmental conditions, enhancing the likelihood of survival to reproductive maturity. Much remains to be learned about the molecular basis underlying environmental influences on these processes. C. elegans larvae enter a developmentally dormant state called L1 diapause when hatched into nutrient-poor conditions. The nematode pten homologue daf-18 is essential for maintenance of survival and germline stem cell quiescence during this period (Fukuyama et al., 2006; Sigmond et al., 2008, but the details of the signaling network(s in which it functions remain to be elucidated. Here, we report that animals lacking both aak-1 and aak-2, which encode the two catalytic α subunits of AMP-activated protein kinase (AMPK, show reduced viability and failure to maintain mitotic quiescence in germline stem cells during L1 diapause. Furthermore, failure to arrest germline proliferation has a long term consequence; aak double mutants that have experienced L1 diapause develop into sterile adults when returned to food, whereas their continuously fed siblings are fertile. Both aak and daf-18 appear to maintain germline quiescence by inhibiting activity of the common downstream target, TORC1 (TOR Complex 1. In contrast, rescue of the lethality phenotype indicates that aak-2 acts not only in the intestine, as does daf-18, but also in neurons, likely promoting survival by preventing energy deprivation during L1 diapause. These results not only provide evidence that AMPK contributes to survival during L1 diapause in a manner distinct from that by which it controls dauer diapause, but they also suggest that AMPK suppresses TORC1 activity to maintain stem cell quiescence.

  16. Isolation and characterization of string-forming female germline stem cells from ovaries of neonatal mice.

    Science.gov (United States)

    Liu, Jing; Shang, Dantong; Xiao, Yao; Zhong, Pei; Cheng, Hanhua; Zhou, Rongjia

    2017-09-29

    Germline stem cells are essential in the generation of both male and female gametes. In mammals, the male testis produces sperm throughout the entire lifetime, facilitated by testicular germline stem cells. Oocyte renewal ceases in postnatal or adult life in mammalian females, suggesting that germline stem cells are absent from the mammalian ovary. However, studies in mice, rats, and humans have recently provided evidence for ovarian female germline stem cells (FGSCs). A better understanding of the role of FGSCs in ovaries could help improve fertility treatments. Here, we developed a rapid and efficient method for isolating FGSCs from ovaries of neonatal mice. Notably, our FGSC isolation method could efficiently isolate on average 15 cell "strings" per ovary from mice at 1-3 days postpartum. FGSCs isolated from neonatal mice displayed the string-forming cell configuration at mitosis ( i.e. a "stringing" FGSC (sFGSC) phenotype) and a disperse phenotype in postnatal mice. We also found that sFGSCs undergo vigorous mitosis especially at 1-3 days postpartum. After cell division, the sFGSC membranes tended to be connected to form sFGSCs. Moreover, F-actin filaments exhibited a cell-cortex distribution in sFGSCs, and E-cadherin converged in cell-cell connection regions, resulting in the string-forming morphology. Our new method provides a platform for isolating FGSCs from the neonatal ovary, and our findings indicate that FGCSs exhibit string-forming features in neonatal mice. The sFGSCs represent a valuable resource for analysis of ovary function and an in vitro model for future clinical use to address ovarian dysfunction. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  17. Germline excision of transgenes in Aedes aegypti by homing endonucleases.

    Science.gov (United States)

    Aryan, Azadeh; Anderson, Michelle A E; Myles, Kevin M; Adelman, Zach N

    2013-01-01

    Aedes (Ae.) aegypti is the primary vector for dengue viruses (serotypes1-4) and chikungunya virus. Homing endonucleases (HEs) are ancient selfish elements that catalyze double-stranded DNA breaks (DSB) in a highly specific manner. In this report, we show that the HEs Y2-I-AniI, I-CreI and I-SceI are all capable of catalyzing the excision of genomic segments from the Ae. aegypti genome in a heritable manner. Y2-I-AniI demonstrated the highest efficiency at two independent genomic targets, with 20-40% of Y2-I-AniI-treated individuals producing offspring that had lost the target transgene. HE-induced DSBs were found to be repaired via the single-strand annealing (SSA) and non-homologous end-joining (NHEJ) pathways in a manner dependent on the availability of direct repeat sequences in the transgene. These results support the development of HE-based gene editing and gene drive strategies in Ae. aegypti, and confirm the utility of HEs in the manipulation and modification of transgenes in this important vector.

  18. Germline PMS2 and somatic POLE exonuclease mutations cause hypermutability of the leading DNA strand in biallelic mismatch repair deficiency syndrome brain tumours.

    Science.gov (United States)

    Andrianova, Maria A; Chetan, Ghati Kasturirangan; Sibin, Madathan Kandi; Mckee, Thomas; Merkler, Doron; Narasinga, Rao Kvl; Ribaux, Pascale; Blouin, Jean-Louis; Makrythanasis, Periklis; Seplyarskiy, Vladimir B; Antonarakis, Stylianos E; Nikolaev, Sergey I

    2017-11-01

    Biallelic mismatch repair deficiency (bMMRD) in tumours is frequently associated with somatic mutations in the exonuclease domains of DNA polymerases POLE or POLD1, and results in a characteristic mutational profile. In this article, we describe the genetic basis of ultramutated high-grade brain tumours in the context of bMMRD. We performed exome sequencing of two second-cousin patients from a large consanguineous family of Indian origin with early onset of high-grade glioblastoma and astrocytoma. We identified a germline homozygous nonsense variant, p.R802*, in the PMS2 gene. Additionally, by genome sequencing of these tumours, we found extremely high somatic mutation rates (237/Mb and 123/Mb), as well as somatic mutations in the proofreading domain of POLE polymerase (p.P436H and p.L424V), which replicates the leading DNA strand. Most interestingly, we found, in both cancers, that the vast majority of mutations were consistent with the signature of POLE exo - , i.e. an abundance of C>A and C>T mutations, particularly in special contexts, on the leading strand. We showed that the fraction of mutations under positive selection among mutations in tumour suppressor genes is more than two-fold lower in ultramutated tumours than in other glioblastomas. Genetic analyses enabled the diagnosis of the two consanguineous childhood brain tumours as being due to a combination of PMS2 germline and POLE somatic variants, and confirmed them as bMMRD/POLE exo - disorders. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd. Copyright © 2017 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  19. Lynch syndrome-associated endometrial carcinoma with MLH1 germline mutation and MLH1 promoter hypermethylation: a case report and literature review.

    Science.gov (United States)

    Yokoyama, Takanori; Takehara, Kazuhiro; Sugimoto, Nao; Kaneko, Keika; Fujimoto, Etsuko; Okazawa-Sakai, Mika; Okame, Shinichi; Shiroyama, Yuko; Yokoyama, Takashi; Teramoto, Norihiro; Ohsumi, Shozo; Saito, Shinya; Imai, Kazuho; Sugano, Kokichi

    2018-05-21

    Lynch syndrome is an autosomal dominant inherited disease caused by germline mutations in mismatch repair genes. Analysis for microsatellite instability (MSI) and immunohistochemistry (IHC) of protein expressions of disease-associated genes is used to screen for Lynch syndrome in endometrial cancer patients. When losses of both MLH1 and PMS2 proteins are observed by IHC, MLH1 promoter methylation analysis is conducted to distinguish Lynch syndrome-associated endometrial cancer from sporadic cancer. Here we report a woman who developed endometrial cancer at the age of 49 years. She had a family history of colorectal cancer (first-degree relative aged 52 years) and stomach cancer (second-degree relative with the age of onset unknown). No other family history was present, and she failed to meet the Amsterdam II criteria for the diagnosis of Lynch syndrome. Losses of MLH1 and PMS2, but not MSH2 and MSH6, proteins were observed by IHC in endometrial cancer tissues. Because MLH1 promoter hypermethylation was detected in endometrial cancer tissue samples, the epigenetic silencing of MLH1 was suspected as the cause of the protein loss. However, because of the early onset of endometrial cancer and the positive family history, a diagnosis of Lynch syndrome was also suspected. Therefore, we provided her with genetic counseling. After obtaining her consent, MLH1 promoter methylation testing and genetic testing of peripheral blood were performed. MLH1 promoter methylation was not observed in peripheral blood. However, genetic testing revealed a large deletion of exon 5 in MLH1; thus, we diagnosed the presence of Lynch syndrome. Both MLH1 germline mutation and MLH1 promoter hypermethylation may be observed in endometrial cancer. Therefore, even if MLH1 promoter hypermethylation is detected, a diagnosis of Lynch syndrome cannot be excluded.

  20. Fitness loss and germline mutations in barn swallows breeding in Chernobyl

    Energy Technology Data Exchange (ETDEWEB)

    Ellegren, Hans; Lindgren, Gabriella; Primmer, C.R. [Swedish Univ. of Agricultural Sciences, Animal Breeding and Genetics Dept., Uppsala (Sweden); Moeller, A.P. [Universite Pierre et Marie Curie. Lab. d`Ecologie, Paris, 75 (France)

    1997-10-09

    The severe nuclear accident at Chernobyl in 1986 resulted in the worst reported accidental exposure of radioactive material to free-living organisms. Short-term effects on human populations inhabiting polluted areas include increased incidence of thyroid cancer, infant leukaemia, and congenital malformations in newborns. Two recent studies have reported, although with some controversy, that germline mutation rates were increased in humans and voles living close to Chernobyl, but little is known about the viability of the organisms affected. Here we report an increased frequency of partial albinism, a morphological aberration associated with a loss of fitness, among barn swallows, Hirundo rustica, breeding close to Chernobyl. Heretability estimates indicate that mutations causing albinism were at least partly of germline origin. Furthermore, evidence for an increased germline mutation rate was obtained from segregation analysis at two hypervariable microsatellite loci, indicating that mutation events in barn swallows from Chernobyl were two- to tenfold higher than in birds from control areas in Ukraine and Italy. (author).

  1. Fitness loss and germline mutations in barn swallows breeding in Chernobyl

    International Nuclear Information System (INIS)

    Ellegren, Hans; Lindgren, Gabriella; Primmer, C.R.; Moeller, A.P.

    1997-01-01

    The severe nuclear accident at Chernobyl in 1986 resulted in the worst reported accidental exposure of radioactive material to free-living organisms. Short-term effects on human populations inhabiting polluted areas include increased incidence of thyroid cancer, infant leukaemia, and congenital malformations in newborns. Two recent studies have reported, although with some controversy, that germline mutation rates were increased in humans and voles living close to Chernobyl, but little is known about the viability of the organisms affected. Here we report an increased frequency of partial albinism, a morphological aberration associated with a loss of fitness, among barn swallows, Hirundo rustica, breeding close to Chernobyl. Heretability estimates indicate that mutations causing albinism were at least partly of germline origin. Furthermore, evidence for an increased germline mutation rate was obtained from segregation analysis at two hypervariable microsatellite loci, indicating that mutation events in barn swallows from Chernobyl were two- to tenfold higher than in birds from control areas in Ukraine and Italy. (author)

  2. The Canonical E2Fs Are Required for Germline Development in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Xiaozhen Yao

    2018-05-01

    Full Text Available A number of cell fate determinations, including cell division, cell differentiation, and programmed cell death, intensely occur during plant germline development. How these cell fate determinations are regulated remains largely unclear. The transcription factor E2F is a core cell cycle regulator. Here we show that the Arabidopsis canonical E2Fs, including E2Fa, E2Fb, and E2Fc, play a redundant role in plant germline development. The e2fa e2fb e2fc (e2fabc triple mutant is sterile, although its vegetative development appears normal. On the one hand, the e2fabc microspores undergo cell death during pollen mitosis. Microspores start to die at the bicellular stage. By the tricellular stage, the majority of the e2fabc microspores are degenerated. On the other hand, a wild type ovule often has one megaspore mother cell (MMC, whereas the majority of e2fabc ovules have two to three MMCs. The subsequent female gametogenesis of e2fabc mutant is aborted and the vacuole is severely impaired in the embryo sac. Analysis of transmission efficiency showed that the canonical E2Fs from both male and female gametophyte are essential for plant gametogenesis. Our study reveals that the canonical E2Fs are required for plant germline development, especially the pollen mitosis and the archesporial cell (AC-MMC transition.

  3. Germline minisatellite mutations in workers occupationally exposed to radiation at the Sellafield nuclear facility

    International Nuclear Information System (INIS)

    Tawn, E Janet; Curwen, Gillian B; Rees, Gwen S; Jonas, Patricia

    2015-01-01

    Germline minisatellite mutation rates were investigated in male workers occupationally exposed to radiation at the Sellafield nuclear facility. DNA samples from 160 families with 255 offspring were analysed for mutations at eight hypervariable minisatellite loci (B6.7, CEB1, CEB15, CEB25, CEB36, MS1, MS31, MS32) by Southern hybridisation. No significant difference was observed between the paternal mutation rate of 5.0% (37 mutations in 736 alleles) for control fathers with a mean preconceptional testicular dose of 9 mSv and that of 5.8% (66 in 1137 alleles) for exposed fathers with a mean preconceptional testicular dose of 194 mSv. Subgrouping the exposed fathers into two dose groups with means of 111 mSv and 274 mSv revealed paternal mutation rates of 6.0% (32 mutations in 536 alleles) and 5.7% (34 mutations in 601 alleles), respectively, neither of which was significantly different in comparisons with the rate for the control fathers. Maternal mutation rates of 1.6% (12 mutations in 742 alleles) for the partners of control fathers and 1.7% (19 mutations in 1133 alleles) for partners of exposed fathers were not significantly different. This study provides evidence that paternal preconceptional occupational radiation exposure does not increase the germline minisatellite mutation rate and therefore refutes suggestions that such exposure could result in a destabilisation of the germline that can be passed on to future generations. (paper)

  4. Germline cytotoxic lymphocytes defective mutations in Chinese patients with lymphoma.

    Science.gov (United States)

    Chen, Xue; Zhang, Yang; Wang, Fang; Wang, Mangju; Teng, Wen; Lin, Yuehui; Han, Xiangping; Jin, Fangyuan; Xu, Yuanli; Cao, Panxiang; Fang, Jiancheng; Zhu, Ping; Tong, Chunrong; Liu, Hongxing

    2017-11-01

    Certain patients with lymphoma may harbor mutations in perforin 1 (PRF1), unc-13 homolog D (UNC13D), syntaxin 11 (STX11), STXBP2 (syntaxin binding protein 2) or SH2 domain containing 1A (SH2D1A), which causes functional defects of cytotoxic lymphocytes. Data regarding the association between genetic defects and the development of lymphoma in Chinese patients are limited to date. In the present study, 90 patients with lymphoma were analyzed for UNC13D, PRF1, STXBP2, STX11, SH2D1A and X-linked inhibitor of apoptosis. Mutations were observed in 24 (26.67%) patients; 16 patients exhibited mutations in UNC13D, 7 exhibited PRF1 mutations, and 1 exhibited monoallelic mutation in STX11. UNC13D c.2588G>A/p.G863D mutation was detected in 9 patients (10.00%) and in 4/210 controls (1.90%). This mutation was predicted to be pathogenic and it predominantly existed in the Chinese population. These findings suggest that impaired cytotoxic machinery may represent a predisposing factor for the development of lymphoma. Furthermore, these data describe a distinct mutation spectrum in Chinese patients with lymphoma, whereby UNC13D is the most frequently mutated gene. In addition, these findings suggest UNC13D c.2588G>A mutation is a founder mutation in Chinese patients.

  5. Analysis of APOBEC3A/3B germline deletion polymorphism in breast, cervical and oral cancers from South India and its impact on miRNA regulation.

    Science.gov (United States)

    Revathidevi, Sundaramoorthy; Manikandan, Mayakannan; Rao, Arunagiri Kuha Deva Magendhra; Vinothkumar, Vilvanathan; Arunkumar, Ganesan; Rajkumar, Kottayasamy Seenivasagam; Ramani, Rajendran; Rajaraman, Ramamurthy; Ajay, Chandrasekar; Munirajan, Arasambattu Kannan

    2016-09-01

    Breast cancer and cervical cancer are the leading causes of death in women worldwide as well as in India, whilst oral cancer is the top most common cancer among Asian especially in Indian men in terms of both incidence and mortality rate. Genetic factors determining the predisposition to cancer are being explored to identify the signature genetic variations associated with these cancers. Recently, a germline deletion polymorphism in APOBEC3 gene cluster which completely deletes APOBEC3B coding region has been studied for its association with cancer risk. We screened the germline deletion polymorphism in 409 cancer patients (224 breast cancer, 88 cervical cancer and 97 oral cancer samples), 478 controls and 239 cervical cancer tissue DNAs of South Indian origin. The results suggest that the APOBEC3A/3B deletion polymorphism is not significantly associated with cancer risk in our study population (OR 0.739, 95 % CI, p value 0.91457). Considering the viral restriction property of APOBEC3s, we also screened cervical cancer tissue DNAs for the human papilloma virus infection. We observed a gradual increase in the frequency of HPV16 infection from AA/BB cases (66.86 %) to AA/-- cases (71.43) which signifies the impact of this deletion polymorphism in HPV infection. In addition, we performed in silico analysis to understand the effect of this polymorphism on miRNA regulation of the APOBEC3A/3B fusion transcript. Only 8 APOBEC3B targeting miRNAs were observed to regulate the fusion transcript of which miR-34b-3p and miR-138-5p were found to be frequently downregulated in cancers suggesting miRNA-mediated deregulation of APOBEC3A expression in cancer patients harbouring this particular deletion polymorphism.

  6. Formation of germline chimera Gaok chicken used circulation primordial germ cells (circulation PGCs fresh and thawed

    Directory of Open Access Journals (Sweden)

    Kostaman T

    2014-03-01

    Full Text Available Formation of germline chimeras by transfer of chicken primordial germ cells (PGCs is one of the effective techniques for preservation and regeneration of genetic resources in chickens. This study attempted to form germline chimeras of Gaok chicken buy purifying circulated PGCs of donor embryo before it is transferred to the recipient (White Leghorn chickens=WL and studied the ability of recipient embryo on survival in incubators, and hatchability. This study used 200 fertile eggs of Gaok and 90 fertile WL breed all of the eggs was incubated at 380C and 60% humidity in a portable incubator. PGCs-circulation of the blood collected Gaok embryos at stage 14-16 were taken from the dorsal aorta, and then purified by centrifugation method using nycodenz. PGCs-circulation results further purification frozen in liquid nitrogen before being transferred to the recipient embryo. The results showed that for the development of embryos transferred to the fresh circulation of PGCs-circulation as many as 25 cells can survive up to day 14, while one of the transferred of 50 and 100 cells into recipient embryos was hatched (10%. On the contrari recipient embryos that are transferred to the frozen PGCs-circulation the embryos development was shorter, and only survived until day 10th (treatment 25 cells, day 14th (treatment of 50 cells and day 17th (treatment of 100 cells. It is concluded that the amount of PGCs-circulation embryos transferred to the recipient is one factor that influence the success of the development germline chimeras.

  7. Intraovarian Transplantation of Female Germline Stem Cells Rescue Ovarian Function in Chemotherapy-Injured Ovaries.

    Directory of Open Access Journals (Sweden)

    Jiaqiang Xiong

    Full Text Available Early menopause and infertility often occur in female cancer patients after chemotherapy (CTx. For these patients, oocyte/embryo cryopreservation or ovarian tissue cryopreservation is the current modality for fertility preservation. However, the above methods are limited in the long-term protection of ovarian function, especially for fertility preservation (very few females with cancer have achieved pregnancy with cryopreserved ovarian tissue or eggs until now. In addition, the above methods are subject to their scope (females with no husband or prepubertal females with no mature oocytes. Thus, many females who suffer from cancers would not adopt the above methods pre- and post-CTx due to their uncertainty, safety and cost-effectiveness. Therefore, millions of women have achieved long-term survival after thorough CTx treatment and have desired to rescue their ovarian function and fertility with economic, durable and reliable methods. Recently, some studies showed that mice with infertility caused by CTx can produce normal offspring through intraovarian injection of exogenous female germline stem cells (FGSCs. Though exogenous FGSC can be derived from mice without immune rejection in the same strain, it is difficult to obtain human female germline stem cells (hFGSCs, and immune rejection could occur between different individuals. In this study, infertility in mice was caused by CTx, and the ability of FGSCs to restore ovarian function or even produce offspring was assessed. We had successfully isolated and purified the FGSCs from adult female mice two weeks after CTx. After infection with GFP-carrying virus, the FGSCs were transplanted into ovaries of mice with infertility caused by CTx. Finally, ovarian function was restored and the recipients produced offspring long-term. These findings showed that mice with CTx possessed FGSCs, restoring ovarian function and avoiding immune rejection from exogenous germline stem cells.

  8. Intraovarian Transplantation of Female Germline Stem Cells Rescue Ovarian Function in Chemotherapy-Injured Ovaries.

    Science.gov (United States)

    Xiong, Jiaqiang; Lu, Zhiyong; Wu, Meng; Zhang, Jinjin; Cheng, Jing; Luo, Aiyue; Shen, Wei; Fang, Li; Zhou, Su; Wang, Shixuan

    2015-01-01

    Early menopause and infertility often occur in female cancer patients after chemotherapy (CTx). For these patients, oocyte/embryo cryopreservation or ovarian tissue cryopreservation is the current modality for fertility preservation. However, the above methods are limited in the long-term protection of ovarian function, especially for fertility preservation (very few females with cancer have achieved pregnancy with cryopreserved ovarian tissue or eggs until now). In addition, the above methods are subject to their scope (females with no husband or prepubertal females with no mature oocytes). Thus, many females who suffer from cancers would not adopt the above methods pre- and post-CTx due to their uncertainty, safety and cost-effectiveness. Therefore, millions of women have achieved long-term survival after thorough CTx treatment and have desired to rescue their ovarian function and fertility with economic, durable and reliable methods. Recently, some studies showed that mice with infertility caused by CTx can produce normal offspring through intraovarian injection of exogenous female germline stem cells (FGSCs). Though exogenous FGSC can be derived from mice without immune rejection in the same strain, it is difficult to obtain human female germline stem cells (hFGSCs), and immune rejection could occur between different individuals. In this study, infertility in mice was caused by CTx, and the ability of FGSCs to restore ovarian function or even produce offspring was assessed. We had successfully isolated and purified the FGSCs from adult female mice two weeks after CTx. After infection with GFP-carrying virus, the FGSCs were transplanted into ovaries of mice with infertility caused by CTx. Finally, ovarian function was restored and the recipients produced offspring long-term. These findings showed that mice with CTx possessed FGSCs, restoring ovarian function and avoiding immune rejection from exogenous germline stem cells.

  9. Germline HVR-II mitochondrial polymorphisms associated with breast cancer in Tunisian women.

    Science.gov (United States)

    Yacoubi Loueslati, B; Troudi, W; Cherni, L; Rhomdhane, K B; Mota-Vieira, L

    2010-08-31

    A high incidence of somatic mtDNA polymorphisms has been reported in a wide variety of human cancers; some of them have been proposed as markers for the early detection of breast cancer. However, little attention has been paid to the potential of germline mitochondrial sequence variations as genetic risk factors for cancer. We performed a case-control study of 70 unrelated Tunisian women with breast cancer and 80 healthy age- and gender-matched blood donors, taking into account clinicopathological data, to evaluate germline polymorphism of mitochondrial HVR-II region as a genetic risk factor for breast cancer. Through direct sequencing, we detected 351 polymorphisms in controls and 248 variants in patients, with 47 and 39 segregating sites, respectively. In both groups, more than 50% of the polymorphisms were due to four variants: 315 ins C, 309 ins C, 263 A>G, and 73 A>G. The HVR-II sequences were also classified into haplotypes on the basis of the polymorphisms. Fifty-nine different haplotypes were found, 20 of them shared between patients and controls. Both groups had specific haplotypes, 18 in breast cancer patients and 21 in controls. Statistical analysis revealed a weak protective effect against breast cancer risk for two mitochondrial polymorphisms - 152 T>C (odds ratio (OR) = 0.33, 95% confidence interval (CI) = 0.12-0.91) and 263 A>G (OR = 0.17, 95%CI = 0.06-0.47). In contrast, an increased risk of breast cancer was detected for the 315+C haplotype (OR = 11.66, 95%CI = 1.44-252.23). We conclude that mitochondrial variants can affect breast cancer risk. More extensive studies, involving different types of cancer and patients with different genetic makeup, will be required to improve our understanding of the effects of germline mtDNA polymorphisms on carcinogenesis.

  10. Understanding the role of p53 in adaptive response to radiation-induced germline mutations

    International Nuclear Information System (INIS)

    Langlois, N.L.; Quinn, J.S.; Somers, C.M.; Boreham, D.R.; Mitchel, R.E.J.

    2003-01-01

    Full text: Radiation-induced adaptive response is now a widely studied area of radiation biology. Studies have demonstrated reduced levels of radiation-induced biological damage when an 'adaptive dose' is given before a higher 'challenge dose' compared to when the challenge dose is given alone. It has been shown in some systems to be a result of inducible cellular repair systems. The adaptive response has been clearly demonstrated in many model systems, however its impact on heritable effects in the mammalian germline has never been studied. Expanded Simple Tandem Repeat (ESTR) loci have been used as markers demonstrating that induced heritable mutations in mice follow a dose-response relationship. Recent data in our laboratory show preliminary evidence of radiation-induced adaptive response suppressing germline mutations at ESTR loci in wild type mice. The frequency of heritable mutations was significantly reduced when a priming dose of 0.1 Gy was given 24 hours prior to a 1 Gy acute challenging dose. We are now conducting a follow-up study to attempt to understand the mechanism of this adaptive response. P53 is known to play a significant role in governing apoptosis, DNA repair and cancer induction. In order to determine what function p53 has in the adaptive response for heritable mutations, we have mated radiation treated Trp53+/- male mice (C57Bl) to untreated, normal females (C57Bl). Using DNA fingerprinting, we are investigating the rate of inherited radiation-induced mutations on pre- and post-meiotic radiation-treated gametocytes by examining mutation frequencies in offspring DNA. If p53 is integral in the mechanism of adaptive response, we should not see an adaptive response in radiation-induced heritable mutations in these mice. This research is significant in that it will provide insight to understanding the mechanism behind radiation-induced adaptive response in the mammalian germline

  11. Use of Germline Polymorphisms in Predicting Concurrent Chemoradiotherapy Response in Esophageal Cancer

    International Nuclear Information System (INIS)

    Chen, Pei-Chun; Chen, Yen-Ching; Lai, Liang-Chuan; Tsai, Mong-Hsun; Chen, Shin-Kuang; Yang, Pei-Wen; Lee, Yung-Chie; Hsiao, Chuhsing K.; Lee, Jang-Ming; Chuang, Eric Y.

    2012-01-01

    Purpose: To identify germline polymorphisms to predict concurrent chemoradiation therapy (CCRT) response in esophageal cancer patients. Materials and Methods: A total of 139 esophageal cancer patients treated with CCRT (cisplatin-based chemotherapy combined with 40 Gy of irradiation) and subsequent esophagectomy were recruited at the National Taiwan University Hospital between 1997 and 2008. After excluding confounding factors (i.e., females and patients aged ≥70 years), 116 patients were enrolled to identify single nucleotide polymorphisms (SNPs) associated with specific CCRT responses. Genotyping arrays and mass spectrometry were used sequentially to determine germline polymorphisms from blood samples. These polymorphisms remain stable throughout disease progression, unlike somatic mutations from tumor tissues. Two-stage design and additive genetic models were adopted in this study. Results: From the 26 SNPs identified in the first stage, 2 SNPs were found to be significantly associated with CCRT response in the second stage. Single nucleotide polymorphism rs16863886, located between SGPP2 and FARSB on chromosome 2q36.1, was significantly associated with a 3.93-fold increase in pathologic complete response to CCRT (95% confidence interval 1.62–10.30) under additive models. Single nucleotide polymorphism rs4954256, located in ZRANB3 on chromosome 2q21.3, was associated with a 3.93-fold increase in pathologic complete response to CCRT (95% confidence interval 1.57–10.87). The predictive accuracy for CCRT response was 71.59% with these two SNPs combined. Conclusions: This is the first study to identify germline polymorphisms with a high accuracy for predicting CCRT response in the treatment of esophageal cancer.

  12. Two Sets of Piwi Proteins Are Involved in Distinct sRNA Pathways Leading to Elimination of Germline-Specific DNA

    Directory of Open Access Journals (Sweden)

    Dominique I. Furrer

    2017-07-01

    Full Text Available Piwi proteins and piRNAs protect eukaryotic germlines against the spread of transposons. During development in the ciliate Paramecium, two Piwi-dependent sRNA classes are involved in the elimination of transposons and transposon-derived DNA: scan RNAs (scnRNAs, associated with Ptiwi01 and Ptiwi09, and iesRNAs, whose binding partners we now identify as Ptiwi10 and Ptiwi11. scnRNAs derive from the maternal genome and initiate DNA elimination during development, whereas iesRNAs continue DNA targeting until the removal process is complete. Here, we show that scnRNAs and iesRNAs are processed by distinct Dicer-like proteins and bind Piwi proteins in a mutually exclusive manner, suggesting separate biogenesis pathways. We also demonstrate that the PTIWI10 gene is transcribed from the developing nucleus and that its transcription depends on prior DNA excision, suggesting a mechanism of gene expression control triggered by the removal of short DNA segments interrupting the gene.

  13. Refining the role of PMS2 in Lynch syndrome: germline mutational analysis improved by comprehensive assessment of variants.

    Science.gov (United States)

    Borràs, Ester; Pineda, Marta; Cadiñanos, Juan; Del Valle, Jesús; Brieger, Angela; Hinrichsen, Inga; Cabanillas, Ruben; Navarro, Matilde; Brunet, Joan; Sanjuan, Xavier; Musulen, Eva; van der Klift, Helen; Lázaro, Conxi; Plotz, Guido; Blanco, Ignacio; Capellá, Gabriel

    2013-08-01

    The majority of mismatch repair (MMR) gene mutations causing Lynch syndrome (LS) occur either in MLH1 or MSH2. However, the relative contribution of PMS2 is less well defined. The aim of this study was to evaluate the role of PMS2 in LS by assessing the pathogenicity of variants of unknown significance (VUS) detected in the mutational analysis of PMS2 in a series of Spanish patients. From a cohort of 202 LS suspected patients, 13 patients showing loss of PMS2 expression in tumours were screened for germline mutations in PMS2, using a long range PCR based strategy and multiplex ligation dependent probe amplification (MLPA). Pathogenicity assessment of PMS2 VUS was performed evaluating clinicopathological data, frequency in control population and in silico and in vitro analyses at the RNA and protein level. Overall 25 different PMS2 DNA variants were detected. Fourteen were classified as polymorphisms. Nine variants were classified as pathogenic: seven alterations based on their molecular nature and two after demonstrating a functional defect (c.538-3C>G affected mRNA processing and c.137G>T impaired MMR activity). The c.1569C>G variant was classified as likely neutral while the c.384G>A remained as a VUS. We have also shown that the polymorphic variant c.59G>A is MMR proficient. Pathogenic PMS2 mutations were detected in 69% of patients harbouring LS associated tumours with loss of PMS2 expression. In all, PMS2 mutations account for 6% of the LS cases identified. The comprehensive functional analysis shown here has been useful in the classification of PMS2 VUS and contributes to refining the role of PMS2 in LS.

  14. Disease evolution and outcomes in familial AML with germline CEBPA mutations

    DEFF Research Database (Denmark)

    Tawana, Kiran; Wang, Jun; Renneville, Aline

    2015-01-01

    collected from 10 CEBPA-mutated families, representing 24 members with acute myeloid leukemia (AML). Whole-exome (WES) and deep sequencing were performed to genetically profile tumors and define patterns of clonal evolution. Germline CEBPA mutations clustered within the N-terminal and were highly penetrant......, with AML presenting at a median age of 24.5 years (range, 1.75-46 years). In all diagnostic tumors tested (n = 18), double CEBPA mutations (CEBPAdm) were detected, with acquired (somatic) mutations preferentially targeting the C-terminal. Somatic CEBPA mutations were unstable throughout the disease course...

  15. Low Base-Substitution Mutation Rate in the Germline Genome of the Ciliate Tetrahymena thermophila

    Science.gov (United States)

    2016-09-15

    Tetrahymena thermophila, a model eukaryote. PLoS Biol. 4:e286. Farlow A, et al. 2015. The spontaneous mutation rate in the fission yeast Schizosaccharomyces...spontane- ous mutations in yeast . Proc Natl Acad Sci U S A. 105:9272–9277. Lynn DH, Doerder FP. 2012. The life and times of Tetrahymena. Methods Cell...Low Base-Substitution Mutation Rate in the Germline Genome of the Ciliate Tetrahymena thermophila Hongan Long1,2,y, David J. Winter3,*,y, Allan Y.-C

  16. A germline chromothripsis event stably segregating in 11 individuals through three generations

    DEFF Research Database (Denmark)

    Bertelsen, Birgitte; Nazaryan-Petersen, Lusine; Sun, Wei

    2016-01-01

    PURPOSE: Parentally transmitted germ-line chromothripsis (G-CTH) has been identified in only a few cases. Most of these rearrangements were stably transmitted, in an unbalanced form, from a healthy mother to her child with congenital abnormalities probably caused by de novo copy-number changes...... of the DNA damage response, may be related to G-CTH formation. CONCLUSION: G-CTH rearrangements are not always associated with abnormal phenotypes and may be misinterpreted as balanced two-way translocations, suggesting that G-CTH is an underdiagnosed phenomenon.Genet Med 18 5, 494-500....

  17. Combining molecular and immunohistochemical analyses of key drivers in primary melanomas: interplay between germline and somatic variations.

    Science.gov (United States)

    Bruno, William; Martinuzzi, Claudia; Dalmasso, Bruna; Andreotti, Virginia; Pastorino, Lorenza; Cabiddu, Francesco; Gualco, Marina; Spagnolo, Francesco; Ballestrero, Alberto; Queirolo, Paola; Grillo, Federica; Mastracci, Luca; Ghiorzo, Paola

    2018-01-19

    Due to the high mutational somatic burden of Cutaneous Malignant Melanoma (CMM) a thorough profiling of the driver mutations and their interplay is necessary to explain the timing of tumorigenesis or for the identification of actionable genetic events. The aim of this study was to establish the mutation rate of some of the key drivers in melanoma tumorigenesis combining molecular analyses and/or immunohistochemistry in 93 primary CMMs from an Italian cohort also characterized for germline status, and to investigate an interplay between germline and somatic variants. BRAF mutations were present in 68% of cases, while CDKN2A germline mutations were found in 16 % and p16 loss in tissue was found in 63%. TERT promoter somatic mutations were detected in 38% of cases while the TERT -245T>C polymorphism was found in 51% of cases. NRAS mutations were found in 39% of BRAF negative or undetermined cases. NF1 was expressed in all cases analysed. MC1R variations were both considered as a dichotomous variable or scored. While a positive, although not significant association between CDKN2A germline mutations, but not MC1R variants, and BRAF somatic mutation was found, we did not observe other associations between germline and somatic events. A yet undescribed inverse correlation between TERT -245T>C polymorphism and the presence of BRAF mutation was found. It is possible to hypothesize that -245T>C polymorphism could be included in those genotypes which may influence the occurrence of BRAF mutations. Further studies are needed to investigate the role of -245T>C polymorphism as a germline predictor of BRAF somatic mutation status.

  18. TP53 germline mutation testing in 180 families suspected of Li-Fraumeni syndrome: mutation detection rate and relative frequency of cancers in different familial phenotypes

    NARCIS (Netherlands)

    Ruijs, M.W.G.; Verhoef, S.; Rookus, M.A.; Pruntel, R.; van der Hout, A.H.; Hogervorst, F.B.L.; Kluijt, I.; Sijmons, R.H.; Aalfs, C.M.; Wagner, A.; Ausems, M.G.E.M.; Hoogerbrugge, N.; van Asperen, C.J.; Gómez García, E.B.; Meijers-Heijboer, H.; ten Kate, L.P.; Menko, F.H.; van 't Veer, L.J.

    2010-01-01

    Background Li-Fraumeni syndrome (LFS) is a rare autosomal dominant cancer predisposition syndrome. Most families fulfilling the classical diagnostic criteria harbour TP53 germline mutations. However, TP53 germline mutations may also occur in less obvious phenotypes. As a result, different criteria

  19. Birt-Hogg-Dubé syndrom

    DEFF Research Database (Denmark)

    Rehfeld, Anders Aagaard; van Steensel, Maurice A M; Friis-Hansen, Lennart

    2010-01-01

    Birt-Hogg-Dubé (BHD) is a rare autosomal dominant genodermatosis, characterized by cutaneous hamartomas, pulmonary cysts, spontaneous pneumothorax and kidney tumours. BHD is caused by mutation in the gene which codes for folliculin (FLCN). FLCN is part of the mTOR-AMPK signal transduction pathway...

  20. Germline mutation rates at tandem repeat loci in DNA-repair deficient mice

    International Nuclear Information System (INIS)

    Barber, Ruth C.; Miccoli, Laurent; Buul, Paul P.W. van; Burr, Karen L.-A.; Duyn-Goedhart, Annemarie van; Angulo, Jaime F.; Dubrova, Yuri E.

    2004-01-01

    Mutation rates at two expanded simple tandem repeat (ESTR) loci were studied in the germline of non-exposed and irradiated severe combined immunodeficient (scid) and poly(ADP-ribose) polymerase (PARP-1 -/- ) deficient male mice. Non-exposed scid and PARP -/- male mice showed considerably elevated ESTR mutation rates, far higher than those in wild-type isogenic mice and other inbred strains. The irradiated scid and PARP-1 -/- male mice did not show any detectable increases in their mutation rate, whereas significant ESTR mutation induction was observed in the irradiated wild-type isogenic males. ESTR mutation spectra in the scid and PARP-1 -/- strains did not differ from those in the isogenic wild-type strains. Considering these data and the results of previous studies, we propose that a delay in repair of DNA damage in scid and PARP-1 -/- mice could result in replication fork pausing which, in turn, may affect ESTR mutation rate in the non-irradiated males. The lack of mutation induction in irradiated scid and PARP-1 -/- can be explained by the high cell killing effects of irradiation on the germline of deficient mice

  1. Single genome retrieval of context-dependent variability in mutation rates for human germline.

    Science.gov (United States)

    Sahakyan, Aleksandr B; Balasubramanian, Shankar

    2017-01-13

    Accurate knowledge of the core components of substitution rates is of vital importance to understand genome evolution and dynamics. By performing a single-genome and direct analysis of 39,894 retrotransposon remnants, we reveal sequence context-dependent germline nucleotide substitution rates for the human genome. The rates are characterised through rate constants in a time-domain, and are made available through a dedicated program (Trek) and a stand-alone database. Due to the nature of the method design and the imposed stringency criteria, we expect our rate constants to be good estimates for the rates of spontaneous mutations. Benefiting from such data, we study the short-range nucleotide (up to 7-mer) organisation and the germline basal substitution propensity (BSP) profile of the human genome; characterise novel, CpG-independent, substitution prone and resistant motifs; confirm a decreased tendency of moieties with low BSP to undergo somatic mutations in a number of cancer types; and, produce a Trek-based estimate of the overall mutation rate in human. The extended set of rate constants we report may enrich our resources and help advance our understanding of genome dynamics and evolution, with possible implications for the role of spontaneous mutations in the emergence of pathological genotypes and neutral evolution of proteomes.

  2. Ovarian Germline Stem Cells (OGSCs and the Hippo Signaling Pathway Association with Physiological and Pathological Ovarian Aging in Mice

    Directory of Open Access Journals (Sweden)

    Jia Li

    2015-07-01

    Full Text Available Background: The Hippo signaling pathway plays fundamental roles in stem cell maintenance in a variety of tissues and has thus implications for stem cell biology. Key components of this recently discovered pathway have been shown to be associated with primordial follicle activation. However, whether the Hippo signaling pathway plays a role in the development of Ovarian Germline Stem Cells (OGSCs during physiological and pathological ovarian aging in mice is unknown. Methods: Mice at the age of 7 days (7D, or of 2, 10, or 20 months (2M, 10M, 20M and mice at 2M treated with TPT and CY/BUS drugs were selected as physiological and pathological ovarian aging models, respectively. Immunohistochemistry was used to assess the development of follicles, and the co-localization of genes characteristic of OGSCs with MST1, LATS2 and YAP1 was assessed by immunofluorescence, western blotting and real-time PCR methods. Results: The Hippo signal pathway and MVH/OCT4 genes were co-expressed in the mouse ovarian cortex. The level and co-localization of LATS2, MST1, MVH, and OCT4 were significantly decreased with increased age, but YAP1 was more prevalent in the mouse ovarian cortex of 2M mice than 7D mice and was not observed in 20M mice. Furthermore, YAP1, MVH, and OCT4 were gradually decreased after TPT and CY/BUS treatment, and LATS2 mRNA and protein up-regulation persisted in TPT- and CY/BUS-treated mice. However, the expression of MST1 was lower in the TPT and CY/BUS groups compared with the control group. In addition, pYAP1 protein showed the highest expression in the ovarian cortexes of 7D mice compared with 20M mice, and the value of pYAP1/YAP1 decreased from 7D to 20M. Moreover, pYAP1 decreased in the TPT- and CY/BUS-treated groups, but the value of pYAP1/YAP1 increased in these groups. Conclusion: Taken together, our results show that the Hippo signaling pathway is associated with the changes that take place in OGSCs during physiological and pathological

  3. A Database to Support the Interpretation of Human Mismatch Repair Gene Variants

    NARCIS (Netherlands)

    Ou, Jianghua; Niessen, Renee C.; Vonk, Jan; Westers, Helga; Hofstra, Robert M. W.; Sijmons, Rolf H.

    Germline mutations in the mismatch repair (MMR) genes MLH1, MSH2, MSH6, or PMS2 can cause Lynch syndrome. This syndrome, also known as hereditary nonpolyposis colorectal cancer (HNPCC), is an autosomal dominantly-inherited disorder predominantly characterized by colorectal and endometrial cancer.

  4. Isolation and characterization of a PUF-domain of pumilio gene from ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-03-20

    Mar 20, 2009 ... study, a partial pumilio gene with complete PUF-domain in Bombyx mori has been ... Key words: Bombyx mori, pumilio, PUF-domain, RACE, germline stem cell. .... The first-strand cDNA was synthesized from 2 ug of total. RNA.

  5. Novel nonsense mutation of BRCA2 gene in a Moroccan man with ...

    African Journals Online (AJOL)

    Background: Breast cancer is the most common cancer in women worldwide. About 5 to 10% of cases are due to an inherited predisposition in two major genes, BRCA1 and BRCA2, transmitted as an autosomal dominant form. Male breast cancer is rare and is mainly due to BRCA2 than BRCA1 germline mutations.

  6. The Constrained Maximal Expression Level Owing to Haploidy Shapes Gene Content on the Mammalian X Chromosome

    NARCIS (Netherlands)

    Hurst, Laurence D; Ghanbarian, Avazeh T; Forrest, Alistair R R; Huminiecki, Lukasz; Clevers, J.C.; van de Wetering, M.L.

    2015-01-01

    X chromosomes are unusual in many regards, not least of which is their nonrandom gene content. The causes of this bias are commonly discussed in the context of sexual antagonism and the avoidance of activity in the male germline. Here, we examine the notion that, at least in some taxa, functionally

  7. Determining the functional significance of mismatch repair gene missense variants using biochemical and cellular assays

    DEFF Research Database (Denmark)

    Heinen, Christopher D; Juel Rasmussen, Lene

    2012-01-01

    ABSTRACT: With the discovery that the hereditary cancer susceptibility disease Lynch syndrome (LS) is caused by deleterious germline mutations in the DNA mismatch repair (MMR) genes nearly 20 years ago, genetic testing can now be used to diagnose this disorder in patients. A definitive diagnosis...

  8. Germline Variation at CDKN2A and Associations with Nevus Phenotypes among Members of Melanoma Families

    DEFF Research Database (Denmark)

    Taylor, Nicholas J; Mitra, Nandita; Goldstein, Alisa M

    2017-01-01

    Germline mutations in CDKN2A are frequently identified among melanoma kindreds and are associated with increased atypical nevus counts. However, a clear relationship between pathogenic CDKN2A mutation carriage and other nevus phenotypes including counts of common acquired nevi has not yet been es...

  9. Differential RPA-1 and RAD-51 recruitment in vivo throughout the C. elegans germline, as revealed by laser microirradiation.

    Science.gov (United States)

    Koury, Emily; Harrell, Kailey; Smolikove, Sarit

    2018-01-25

    Studies of the repair pathways associated with DNA double strand breaks (DSBs) are numerous, and provide evidence for cell-cycle specific regulation of homologous recombination (HR) by the regulation of its associated proteins. Laser microirradiation is a well-established method to examine in vitro kinetics of repair and allows for live-imaging of DSB repair from the moment of induction. Here we apply this method to whole, live organisms, introducing an effective system to analyze exogenous, microirradiation-induced breaks in the Caenorhabditis elegans germline. Through this method we observed the sequential kinetics of the recruitment of ssDNA binding proteins RPA-1 and RAD-51 in vivo. We analyze these kinetics throughout different regions of the germline, and thus throughout a range of developmental stages of mitotic and meiotic nuclei. Our analysis demonstrates a largely conserved timing of recruitment of ssDNA binding proteins to DSBs throughout the germline, with a delay of RAD-51 recruitment at mid-pachytene nuclei. Microirradiated nuclei are viable and undergo a slow kinetics of resolution. We observe RPA-1 and RAD-51 colocalization for hours post-microirradiation throughout the germline, suggesting that there are mixed RPA-1/RAD-51 filaments. Finally, through live imaging analysis we observed RAD-51 foci movement with low frequency of coalescence. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  10. A recurrent germline BAP1 mutation and extension of the BAP1 tumor predisposition spectrum to include basal cell carcinoma

    DEFF Research Database (Denmark)

    Wadt, Karin Anna Wallentin; Aoude, L G; Johansson, P

    2015-01-01

    ) and mesothelioma, as previously reported for germline BAP1 mutations. However, mutation carriers from three new families, and one previously reported family, developed basal cell carcinoma (BCC), thus suggesting inclusion of BCC in the phenotypic spectrum of the BAP1 tumor syndrome. This notion is supported...

  11. NanoTIO2 (UV-Titan) does not induce ESTR mutations in the germline of prenatally exposed female mice

    DEFF Research Database (Denmark)

    Boisen, Anne Mette Zenner; Shipley, Thomas; Hougaard, Karin Sørig

    2012-01-01

    Particulate air pollution has been linked to an increased risk of cardiovascular disease and cancer. Animal studies have shown that inhalation of air particulates induces mutations in the male germline. Expanded simple tandem repeat (ESTR) loci in mice are sensitive markers of mutagenic effects o...

  12. Reproductive and therapeutic cloning, germline therapy, and purchase of gametes and embryos: comments on Canadian legislation governing reproduction technologies

    Science.gov (United States)

    Bernier, L; Gregoire, D

    2004-01-01

    In this article the three main topics covered in the new legislation are commented on: cloning, germline therapy, and purchase of gametes and embryos. Some important issues also covered in the new legislation, such as privacy and access to information, data protection, identity of donors, and inspection, will not be addressed. PMID:15574437

  13. Germline Polymorphisms of the VEGF Pathway Predict Recurrence in Nonadvanced Differentiated Thyroid Cancer.

    Science.gov (United States)

    Marotta, Vincenzo; Sciammarella, Concetta; Capasso, Mario; Testori, Alessandro; Pivonello, Claudia; Chiofalo, Maria Grazia; Gambardella, Claudio; Grasso, Marica; Antonino, Antonio; Annunziata, Annamaria; Macchia, Paolo Emidio; Pivonello, Rosario; Santini, Luigi; Botti, Gerardo; Losito, Simona; Pezzullo, Luciano; Colao, Annamaria; Faggiano, Antongiulio

    2017-02-01

    Tumor angiogenesis is determined by host genetic background rather than environment. Germline single nucleotide polymorphisms (SNPs) of the vascular endothelial growth factor (VEGF) pathway have demonstrated prognostic value in different tumors. Our main objective was to test the prognostic value of germline SNPs of the VEGF pathway in nonadvanced differentiated thyroid cancer (DTC). Secondarily, we sought to correlate analyzed SNPs with microvessel density (MVD). Multicenter, retrospective, observational study. Four referral centers. Blood samples were obtained from consecutive DTC patients. Genotyping was performed according to the TaqMan protocol, including 4 VEGF-A (-2578C>A, -460T>C, +405G>C, and +936C>T) and 2 VEGFR-2 (+1192 C>T and +1719 T>A) SNPs. MVD was estimated by means of CD34 staining. Rate of recurrent structural disease/disease-free survival (DFS). Difference in MVD between tumors from patients with different genotype. Two hundred four patients with stage I-II DTC (mean follow-up, 73 ± 64 months) and 240 patients with low- to intermediate-risk DTC (mean follow-up, 70 ± 60 months) were enrolled. Two "risk" genotypes were identified by combining VEGF-A SNPs -2578 C>A, -460 T>C, and +405 G>C. The ACG homozygous genotype was protective in both stage I-II (odds ratio [OR], 0.08; 95% confidence interval [CI], 0.01 to 1.43; P = 0.018) and low- to intermediate-risk (OR, 0.14; 95% CI, 0.01 to 1.13; P = 0.035) patients. The CTG homozygous genotype was significantly associated with recurrence in stage I-II (OR, 5.47; 95% CI, 1.15 to 26.04; P = 0.018) and was slightly deleterious in low- to intermediate-risk (OR, 3.39; 95% CI, 0.8 to 14.33; P = 0.079) patients. MVD of primary tumors from patients harboring a protective genotype was significantly lower (median MVD, 76.5 ± 12.7 and 86.7 ± 27.9, respectively; P = 0.024). Analysis of germline VEGF-A SNPs could empower a prognostic approach to DTC. Copyright © 2017 by the Endocrine Society

  14. Temporal remodeling of the cell cycle accompanies differentiation in the Drosophila germline.

    Science.gov (United States)

    Hinnant, Taylor D; Alvarez, Arturo A; Ables, Elizabeth T

    2017-09-01

    Development of multicellular organisms relies upon the coordinated regulation of cellular differentiation and proliferation. Growing evidence suggests that some molecular regulatory pathways associated with the cell cycle machinery also dictate cell fate; however, it remains largely unclear how the cell cycle is remodeled in concert with cell differentiation. During Drosophila oogenesis, mature oocytes are created through a series of precisely controlled division and differentiation steps, originating from a single tissue-specific stem cell. Further, germline stem cells (GSCs) and their differentiating progeny remain in a predominantly linear arrangement as oogenesis proceeds. The ability to visualize the stepwise events of differentiation within the context of a single tissue make the Drosophila ovary an exceptional model for study of cell cycle remodeling. To describe how the cell cycle is remodeled in germ cells as they differentiate in situ, we used the Drosophila Fluorescence Ubiquitin-based Cell Cycle Indicator (Fly-FUCCI) system, in which degradable versions of GFP::E2f1 and RFP::CycB fluorescently label cells in each phase of the cell cycle. We found that the lengths of the G1, S, and G2 phases of the cell cycle change dramatically over the course of differentiation, and identified the 4/8-cell cyst as a key developmental transition state in which cells prepare for specialized cell cycles. Our data suggest that the transcriptional activator E2f1, which controls the transition from G1 to S phase, is a key regulator of mitotic divisions in the early germline. Our data support the model that E2f1 is necessary for proper GSC proliferation, self-renewal, and daughter cell development. In contrast, while E2f1 degradation by the Cullin 4 (Cul4)-containing ubiquitin E3 ligase (CRL4) is essential for developmental transitions in the early germline, our data do not support a role for E2f1 degradation as a mechanism to limit GSC proliferation or self-renewal. Taken

  15. Germline BAP1 inactivation is preferentially associated with metastatic ocular melanoma and cutaneous-ocular melanoma families.

    Directory of Open Access Journals (Sweden)

    Ching-Ni Jenny Njauw

    Full Text Available BAP1 has been shown to be a target of both somatic alteration in high-risk ocular melanomas (OM and germline inactivation in a few individuals from cancer-prone families. These findings suggest that constitutional BAP1 changes may predispose individuals to metastatic OM and that familial permeation of deleterious alleles could delineate a new cancer syndrome.To characterize BAP1's contribution to melanoma risk, we sequenced BAP1 in a set of 100 patients with OM, including 50 metastatic OM cases and 50 matched non-metastatic OM controls, and 200 individuals with cutaneous melanoma (CM including 7 CM patients from CM-OM families and 193 CM patients from CM-non-OM kindreds.Germline BAP1 mutations were detected in 4/50 patients with metastatic OM and 0/50 cases of non-metastatic OM (8% vs. 0%, p = 0.059. Since 2/4 of the BAP1 carriers reported a family history of CM, we analyzed 200 additional hereditary CM patients and found mutations in 2/7 CM probands from CM-OM families and 1/193 probands from CM-non-OM kindreds (29% vs. 0.52%, p = .003. Germline mutations co-segregated with both CM and OM phenotypes and were associated with the presence of unique nevoid melanomas and highly atypical nevoid melanoma-like melanocytic proliferations (NEMMPs. Interestingly, 7/14 germline variants identified to date reside in C-terminus suggesting that the BRCA1 binding domain is important in cancer predisposition.Germline BAP1 mutations are associated with a more aggressive OM phenotype and a recurrent phenotypic complex of cutaneous/ocular melanoma, atypical melanocytic proliferations and other internal neoplasms (ie. COMMON syndrome, which could be a useful clinical marker for constitutive BAP1 inactivation.

  16. Subset of Kappa and Lambda Germline Sequences Result in Light Chains with a Higher Molecular Mass Phenotype.

    Science.gov (United States)

    Barnidge, David R; Lundström, Susanna L; Zhang, Bo; Dasari, Surendra; Murray, David L; Zubarev, Roman A

    2015-12-04

    In our previous work, we showed that electrospray ionization of intact polyclonal kappa and lambda light chains isolated from normal serum generates two distinct, Gaussian-shaped, molecular mass distributions representing the light-chain repertoire. During the analysis of a large (>100) patient sample set, we noticed a low-intensity molecular mass distribution with a mean of approximately 24 250 Da, roughly 800 Da higher than the mean of the typical kappa molecular-mass distribution mean of 23 450 Da. We also observed distinct clones in this region that did not appear to contain any typical post-translational modifications that would account for such a large mass shift. To determine the origin of the high molecular mass clones, we performed de novo bottom-up mass spectrometry on a purified IgM monoclonal light chain that had a calculated molecular mass of 24 275.03 Da. The entire sequence of the monoclonal light chain was determined using multienzyme digestion and de novo sequence-alignment software and was found to belong to the germline allele IGKV2-30. The alignment of kappa germline sequences revealed ten IGKV2 and one IGKV4 sequences that contained additional amino acids in their CDR1 region, creating the high-molecular-mass phenotype. We also performed an alignment of lambda germline sequences, which showed additional amino acids in the CDR2 region, and the FR3 region of functional germline sequences that result in a high-molecular-mass phenotype. The work presented here illustrates the ability of mass spectrometry to provide information on the diversity of light-chain molecular mass phenotypes in circulation, which reflects the germline sequences selected by the immunoglobulin-secreting B-cell population.

  17. Sequences of the joining region genes for immunoglobulin heavy chains and their role in generation of antibody diversity.

    OpenAIRE

    Gough, N M; Bernard, O

    1981-01-01

    To assess the contribution to immunoglobulin heavy chain diversity made by recombination between variable region (VH) genes and joining region (JH) genes, we have determined the sequence of about 2000 nucleotides spanning the rearranged JH gene cluster associated with the VH gene expressed in plasmacytoma HPC76. The active VH76 gene has recombined with the second germ-line JH gene. The region we have studied contains two other JH genes, designated JH3 and JH4. No other JH gene was found withi...

  18. Improved injection needles facilitate germline transformation of the buckeye butterfly Junonia coenia.

    Science.gov (United States)

    Beaudette, Kahlia; Hughes, Tia M; Marcus, Jeffrey M

    2014-01-01

    Germline transformation with transposon vectors is an important tool for insect genetics, but progress in developing transformation protocols for butterflies has been limited by high post-injection ova mortality. Here we present an improved glass injection needle design for injecting butterfly ova that increases survival in three Nymphalid butterfly species. Using the needles to genetically transform the common buckeye butterfly Junonia coenia, the hatch rate for injected Junonia ova was 21.7%, the transformation rate was 3%, and the overall experimental efficiency was 0.327%, a substantial improvement over previous results in other butterfly species. Improved needle design and a higher efficiency of transformation should permit the deployment of transposon-based genetic tools in a broad range of less fecund lepidopteran species.

  19. Effects of prenatal exposure to nanoparticles titanium dioxide and carbon black on female germline DNA stability

    DEFF Research Database (Denmark)

    Boisen, Anne Mette Zenner

    Particulate air pollution has been associated with an increased risk of cardiovascular disease and cancer in humans. Air pollution may also adversely affect pregnancy outcome and the integrity of sperm cells DNA. Animal studies have shown that inhalation of air particulates can induce mutations...... in premeiotic sperm cells. The investigation of potential mutagenic risk is of outmost importance, as it may lead to cancer. Furthermore, heritable mutations may be passed on to descendents and thereby pose a permanent genetic risk to the population. The nanosized fraction of particulate air pollution has...... are needed. Expanded simple tandem repeat (ESTR) loci in mice are sensitive markers of mutagenic effects resulting from environmental exposures; Studies on adult mice have revealed that while particulate air pollution induced ESTR mutations in premeiotic sperm cells, the female germline was not affected...

  20. Derepression of the plant Chromovirus LORE1 induces germline transposition in regenerated plants.

    Directory of Open Access Journals (Sweden)

    Eigo Fukai

    2010-03-01

    Full Text Available Transposable elements represent a large proportion of the eukaryotic genomes. Long Terminal Repeat (LTR retrotransposons are very abundant and constitute the predominant family of transposable elements in plants. Recent studies have identified chromoviruses to be a widely distributed lineage of Gypsy elements. These elements contain chromodomains in their integrases, which suggests a preference for insertion into heterochromatin. In turn, this preference might have contributed to the patterning of heterochromatin observed in host genomes. Despite their potential importance for our understanding of plant genome dynamics and evolution, the regulatory mechanisms governing the behavior of chromoviruses and their activities remain largely uncharacterized. Here, we report a detailed analysis of the spatio-temporal activity of a plant chromovirus in the endogenous host. We examined LORE1a, a member of the endogenous chromovirus LORE1 family from the model legume Lotus japonicus. We found that this chromovirus is stochastically de-repressed in plant populations regenerated from de-differentiated cells and that LORE1a transposes in the male germline. Bisulfite sequencing of the 5' LTR and its surrounding region suggests that tissue culture induces a loss of epigenetic silencing of LORE1a. Since LTR promoter activity is pollen specific, as shown by the analysis of transgenic plants containing an LTR::GUS fusion, we conclude that male germline-specific LORE1a transposition in pollen grains is controlled transcriptionally by its own cis-elements. New insertion sites of LORE1a copies were frequently found in genic regions and show no strong insertional preferences. These distinctive novel features of LORE1 indicate that this chromovirus has considerable potential for generating genetic and epigenetic diversity in the host plant population. Our results also define conditions for the use of LORE1a as a genetic tool.

  1. Dietary regulation of developmental programming in ruminants: epigenetic modifications in the germline.

    Science.gov (United States)

    Sinclair, K D; Karamitri, A; Gardner, D S

    2010-01-01

    Ruminants have been utilised extensively to investigate the developmental origins of health and disease, with the sheep serving as the model species of choice to complement dietary studies in the rat and mouse. Surprisingly few studies, however, have investigated delayed effects of maternal undernutrition during pregnancy on adult offspring health and a consistent phenotype, together with underlying mechanistic pathways, has not emerged. Nevertheless, when broad consideration is given to all studies with ruminants it is apparent that interventions that are initiated very early in gestation, and/or prior to conception, lead to greater effects on adult physiology than those that are specifically targeted to late gestation. Effects induced following dietary interventions at the earliest stages of mammalian development have been shown to arise as a consequence of alterations to key epigenetic processes that occur in germ cells and pluripotent embryonic cells. Currently, our understanding of epigenetic programming in the germline is greatest for the mouse, and is considered in detail in this article together with what is known in ruminants. This species imbalance, however, looks set to change as fully annotated genomic maps are developed for domesticated large animal species, and with the advent of 'next-generation' DNA sequencing technologies that have the power to globally map the epigenome at single-base-pair resolution. These developments would help to address such issues as sexually dimorphic epigenetic alterations to DNA methylation that have been found to arise following dietary restrictions during the peri-conceptional period, the effects of paternal nutritional status on epigenetic programming through the germline, and transgenerational studies where, in future, greater emphasis in domesticated ruminants should be placed on traits of agricultural importance.

  2. Hereditary diffuse gastric cancer: updated clinical guidelines with an emphasis on germline CDH1 mutation carriers

    Science.gov (United States)

    van der Post, Rachel S; Vogelaar, Ingrid P; Carneiro, Fátima; Guilford, Parry; Huntsman, David; Hoogerbrugge, Nicoline; Caldas, Carlos; Schreiber, Karen E Chelcun; Hardwick, Richard H; Ausems, Margreet G E M; Bardram, Linda; Benusiglio, Patrick R; Bisseling, Tanya M; Blair, Vanessa; Bleiker, Eveline; Boussioutas, Alex; Cats, Annemieke; Coit, Daniel; DeGregorio, Lynn; Figueiredo, Joana; Ford, James M; Heijkoop, Esther; Hermens, Rosella; Humar, Bostjan; Kaurah, Pardeep; Keller, Gisella; Lai, Jennifer; Ligtenberg, Marjolijn J L; O'Donovan, Maria; Oliveira, Carla; Ragunath, Krish; Rasenberg, Esther; Richardson, Susan; Roviello, Franco; Schackert, Hans; Seruca, Raquel; Taylor, Amy; ter Huurne, Anouk; Tischkowitz, Marc; Joe, Sheena Tjon A; van Dijck, Benjamin; van Grieken, Nicole C T; van Hillegersberg, Richard; van Sandick, Johanna W; Vehof, Rianne; van Krieken, J Han; Fitzgerald, Rebecca C

    2015-01-01

    Germline CDH1 mutations confer a high lifetime risk of developing diffuse gastric (DGC) and lobular breast cancer (LBC). A multidisciplinary workshop was organised to discuss genetic testing, surgery, surveillance strategies, pathology reporting and the patient's perspective on multiple aspects, including diet post gastrectomy. The updated guidelines include revised CDH1 testing criteria (taking into account first-degree and second-degree relatives): (1) families with two or more patients with gastric cancer at any age, one confirmed DGC; (2) individuals with DGC before the age of 40 and (3) families with diagnoses of both DGC and LBC (one diagnosis before the age of 50). Additionally, CDH1 testing could be considered in patients with bilateral or familial LBC before the age of 50, patients with DGC and cleft lip/palate, and those with precursor lesions for signet ring cell carcinoma. Given the high mortality associated with invasive disease, prophylactic total gastrectomy at a centre of expertise is advised for individuals with pathogenic CDH1 mutations. Breast cancer surveillance with annual breast MRI starting at age 30 for women with a CDH1 mutation is recommended. Standardised endoscopic surveillance in experienced centres is recommended for those opting not to have gastrectomy at the current time, those with CDH1 variants of uncertain significance and those that fulfil hereditary DGC criteria without germline CDH1 mutations. Expert histopathological confirmation of (early) signet ring cell carcinoma is recommended. The impact of gastrectomy and mastectomy should not be underestimated; these can have severe consequences on a psychological, physiological and metabolic level. Nutritional problems should be carefully monitored. PMID:25979631

  3. TMC-SNPdb: an Indian germline variant database derived from whole exome sequences.

    Science.gov (United States)

    Upadhyay, Pawan; Gardi, Nilesh; Desai, Sanket; Sahoo, Bikram; Singh, Ankita; Togar, Trupti; Iyer, Prajish; Prasad, Ratnam; Chandrani, Pratik; Gupta, Sudeep; Dutt, Amit

    2016-01-01

    Cancer is predominantly a somatic disease. A mutant allele present in a cancer cell genome is considered somatic when it's absent in the paired normal genome along with public SNP databases. The current build of dbSNP, the most comprehensive public SNP database, however inadequately represents several non-European Caucasian populations, posing a limitation in cancer genomic analyses of data from these populations. We present the T: ata M: emorial C: entre-SNP D: ata B: ase (TMC-SNPdb), as the first open source, flexible, upgradable, and freely available SNP database (accessible through dbSNP build 149 and ANNOVAR)-representing 114 309 unique germline variants-generated from whole exome data of 62 normal samples derived from cancer patients of Indian origin. The TMC-SNPdb is presented with a companion subtraction tool that can be executed with command line option or using an easy-to-use graphical user interface with the ability to deplete additional Indian population specific SNPs over and above dbSNP and 1000 Genomes databases. Using an institutional generated whole exome data set of 132 samples of Indian origin, we demonstrate that TMC-SNPdb could deplete 42, 33 and 28% false positive somatic events post dbSNP depletion in Indian origin tongue, gallbladder, and cervical cancer samples, respectively. Beyond cancer somatic analyses, we anticipate utility of the TMC-SNPdb in several Mendelian germline diseases. In addition to dbSNP build 149 and ANNOVAR, the TMC-SNPdb along with the subtraction tool is available for download in the public domain at the following:Database URL: http://www.actrec.gov.in/pi-webpages/AmitDutt/TMCSNP/TMCSNPdp.html. © The Author(s) 2016. Published by Oxford University Press.

  4. Germline mutation of CBL is associated with moyamoya disease in a child with juvenile myelomonocytic leukemia and Noonan syndrome-like disorder.

    Science.gov (United States)

    Hyakuna, Nobuyuki; Muramatsu, Hideki; Higa, Takeshi; Chinen, Yasutsugu; Wang, Xinan; Kojima, Seiji

    2015-03-01

    Germline mutations in CBL have been identified in patients with Noonan syndrome-like phenotypes, while juvenile myelomonocytic leukemia (JMML) harbors duplication of a germline CBL, resulting in acquired isodisomy. The association between moyamoya disease and Noonan syndrome carrying a PTPN11 mutation has recently been reported. We present a patient with JMML who developed moyamoya disease and neovascular glaucoma. Our patient exhibited a Noonan syndrome-like phenotype. Genetic analysis revealed acquired isodisomy and a germline heterozygous mutation in CBL. This is a rare case of CBL mutation associated with moyamoya disease. Prolonged RAS pathway signaling may cause disruption of cerebrovascular development. © 2014 Wiley Periodicals, Inc.

  5. Germline TP53 mutations and single nucleotide polymorphisms in children Mutaciones y polimorfismos de un único nucleótido del gen TP53 en línea germinal en niños

    Directory of Open Access Journals (Sweden)

    Pamela Valva

    2009-02-01

    Full Text Available Mutations in the gene TP53, which codifies the tumor suppressor protein p53, are found in about 50% of tumors. These mutations can occur not only at somatic level, but also in germline. Pediatric cancer patients, mostly with additional family history of malignancy, should be considered as potential TP53 germline mutation carriers. Germline TP53 mutations and polymorphisms have been widely studied to determine their relation with different tumors' pathogenesis. Our aim was to analyze the occurrence frequency of germline TP53 mutations and polymorphisms and to relate these to tumor development in a pediatric series. Peripheral blood mononuclear cell samples from 26 children with solid tumors [PST] and 21 pediatric healthy donors [HD] were analyzed for germline mutations and polymorphisms in TP53 gene spanning from exon 5 to 8 including introns 5 and 7. These PCR amplified fragments were sequenced to determine variations. A heterozygous mutation at codon 245 was found in 1/26 PST and 0/21 HD. Comparative polymorphisms distribution, at position 14181 and 14201(intron 7, between HD and PST revealed a trend of association (p= 0.07 with cancer risk. HD group disclosed a similar polymorphism distribution as published data for Caucasian and Central/South American populations. This is the first study about TP53 variant frequency and distribution in healthy individuals and cancer patients in Argentina.El gen que codifica para la proteína supresora de tumor p53 (TP53 se encuentra mutado en aproximadamente el 50% de los tumores. Estas mutaciones pueden presentarse como somáticas o en línea germinal. Los niños con tumores, sobre todo aquellos con historia familiar de enfermedad oncológica, deben considerarse potenciales portadores de mutaciones en línea germinal. Las mutaciones de TP53 y los polimorfismos son estudiados para determinar su relación con la patogénesis de diferentes tumores. El objetivo del trabajo fue analizar la frecuencia de

  6. MicroRNA-212 post-transcriptionally regulates oocyte-specific basic-helix-loop-helix transcription factor, factor in the germline alpha (FIGLA, during bovine early embryogenesis.

    Directory of Open Access Journals (Sweden)

    Swamy K Tripurani

    Full Text Available Factor in the germline alpha (FIGLA is an oocyte-specific basic helix-loop-helix transcription factor essential for primordial follicle formation and expression of many genes required for folliculogenesis, fertilization and early embryonic survival. Here we report the characterization of bovine FIGLA gene and its regulation during early embryogenesis. Bovine FIGLA mRNA expression is restricted to gonads and is detected in fetal ovaries harvested as early as 90 days of gestation. FIGLA mRNA and protein are abundant in germinal vesicle and metaphase II stage oocytes, as well as in embryos from pronuclear to eight-cell stage but barely detectable at morula and blastocyst stages, suggesting that FIGLA might be a maternal effect gene. Recent studies in zebrafish and mice have highlighted the importance of non-coding small RNAs (microRNAs as key regulatory molecules targeting maternal mRNAs for degradation during embryonic development. We hypothesized that FIGLA, as a maternal transcript, is regulated by microRNAs during early embryogenesis. Computational predictions identified a potential microRNA recognition element (MRE for miR-212 in the 3' UTR of the bovine FIGLA mRNA. Bovine miR-212 is expressed in oocytes and tends to increase in four-cell and eight-cell stage embryos followed by a decline at morula and blastocyst stages. Transient transfection and reporter assays revealed that miR-212 represses the expression of FIGLA in a MRE dependent manner. In addition, ectopic expression of miR-212 mimic in bovine early embryos dramatically reduced the expression of FIGLA protein. Collectively, our results demonstrate that FIGLA is temporally regulated during bovine early embryogenesis and miR-212 is an important negative regulator of FIGLA during the maternal to zygotic transition in bovine embryos.

  7. Next-generation site-directed transgenesis in the malaria vector mosquito Anopheles gambiae: self-docking strains expressing germline-specific phiC31 integrase.

    Directory of Open Access Journals (Sweden)

    Janet M Meredith

    Full Text Available Diseases transmitted by mosquitoes have a devastating impact on global health and the situation is complicated due to difficulties with both existing control measures and the impact of climate change. Genetically modified mosquitoes that are refractory to disease transmission are seen as having great potential in the delivery of novel control strategies. The Streptomyces phage phiC31 integrase system has been successfully adapted for site-directed transgene integration in a range of insects, thus overcoming many limitations due to size constraints and random integration associated with transposon-mediated transformation. Using this technology, we previously published the first site-directed transformation of Anopheles gambiae, the principal vector of human malaria. Mosquitoes were initially engineered to incorporate the phiC31 docking site at a defined genomic location. A second phase of genetic modification then achieved site-directed integration of an anti-malarial effector gene. In the current publication we report improved efficiency and utility of the phiC31 integrase system following the generation of Anopheles gambiae self-docking strains. Four independent strains, with docking sites at known locations on three different chromosome arms, were engineered to express integrase under control of the regulatory regions of the nanos gene from Anopheles gambiae. The resulting protein accumulates in the posterior oocyte to provide integrase activity at the site of germline development. Two self-docking strains, exhibiting significantly different levels of integrase expression, were assessed for site-directed transgene integration and found to demonstrate greatly improved survival and efficiency of transformation. In the fight against malaria, it is imperative to establish a broad repertoire of both anti-malarial effector genes and tissue-specific promoters to regulate their expression, enabling those offering maximum effect with minimum fitness

  8. Detection of ATM germline variants by the p53 mitotic centrosomal localization test in BRCA1/2-negative patients with early-onset breast cancer.

    Science.gov (United States)

    Prodosmo, Andrea; Buffone, Amelia; Mattioni, Manlio; Barnabei, Agnese; Persichetti, Agnese; De Leo, Aurora; Appetecchia, Marialuisa; Nicolussi, Arianna; Coppa, Anna; Sciacchitano, Salvatore; Giordano, Carolina; Pinnarò, Paola; Sanguineti, Giuseppe; Strigari, Lidia; Alessandrini, Gabriele; Facciolo, Francesco; Cosimelli, Maurizio; Grazi, Gian Luca; Corrado, Giacomo; Vizza, Enrico; Giannini, Giuseppe; Soddu, Silvia

    2016-09-06

    Variant ATM heterozygotes have an increased risk of developing cancer, cardiovascular diseases, and diabetes. Costs and time of sequencing and ATM variant complexity make large-scale, general population screenings not cost-effective yet. Recently, we developed a straightforward, rapid, and inexpensive test based on p53 mitotic centrosomal localization (p53-MCL) in peripheral blood mononuclear cells (PBMCs) that diagnoses mutant ATM zygosity and recognizes tumor-associated ATM polymorphisms. Fresh PBMCs from 496 cancer patients were analyzed by p53-MCL: 90 cases with familial BRCA1/2-positive and -negative breast and/or ovarian cancer, 337 with sporadic cancers (ovarian, lung, colon, and post-menopausal breast cancers), and 69 with breast/thyroid cancer. Variants were confirmed by ATM sequencing. A total of seven individuals with ATM variants were identified, 5/65 (7.7 %) in breast cancer cases of familial breast and/or ovarian cancer and 2/69 (2.9 %) in breast/thyroid cancer. No variant ATM carriers were found among the other cancer cases. Excluding a single case in which both BRCA1 and ATM were mutated, no p53-MCL alterations were observed in BRCA1/2-positive cases. These data validate p53-MCL as reliable and specific test for germline ATM variants, confirm ATM as breast cancer susceptibility gene, and highlight a possible association with breast/thyroid cancers.

  9. Production of a germline-humanized cetuximab scFv and evaluation of its activity in recognizing EGFR- overexpressing cancer cells.

    Science.gov (United States)

    Banisadr, Arsham; Safdari, Yaghoub; Kianmehr, Anvarsadat; Pourafshar, Mahdieh

    2018-04-03

    The aim of this study was to produce a humanized single chain antibody (scFv) as a potential improved product design to target EGFR (Epidermal Growth Factor Receptor) overexpressing cancer cells. To this end, CDR loops of cetuximab (an FDA-approved anti-EGFR antibody) were grafted on framework regions derived from type 3 (VH3 and VL3 kappa) human germline sequences to obtain recombinant VH and VL domainslinked together with a flexible linker [(Gly 4 Ser) 3 ] to form a scFv. Codon optimized synthetic gene encoding the scFv (with NH2-VH-linker-VL-COOH orientation) was expressed in E. coli Origami™ 2(DE3) cells and the resultant scFv purified by using Ni-NTA affinity chromatography. The scFv, called cet.Hum scFv, was evaluated in ELISA and immunoblot to determine whether it can recognize EGFR. The scFv was able to recognize EGFR over-expressing cancer cells (A-431) but failed to detect cancer cells with low levels of EGFR (MCF-7 cells). Although the affinity of the scFv forA-431 cells was 9 fold lower than that of cetuximab, it was strong enough to recognize these cells. Considering its ability to bind EGFR molecules, the scFv may exhibit a potential application for the detection of EGFR-overexpressing cancer cells.

  10. Control of germline stem cell self-renewal and differentiation in the Drosophila ovary: concerted actions of niche signals and intrinsic factors.

    Science.gov (United States)

    Xie, Ting

    2013-01-01

    In the Drosophila ovary, germline stem cells (GSCs) physically interact with their niche composed of terminal filament cells, cap cells, and possibly GSC-contacting escort cells (ECs). A GSC divides to generate a self-renewing stem cell that remains in the niche and a differentiating daughter that moves away from the niche. The GSC niche provides a bone morphogenetic protein (BMP) signal that maintains GSC self-renewal by preventing stem cell differentiation via repression of the differentiation-promoting gene bag of marbles (bam). In addition, it expresses E-cadherin, which mediates cell adhesion for anchoring GSCs in the niche, enabling continuous self-renewal. GSCs themselves also express different classes of intrinsic factors, including signal transducers, transcription factors, chromatin remodeling factors, translation regulators, and miRNAs, which control self-renewal by strengthening interactions with the niche and repressing various differentiation pathways. Differentiated GSC daughters, known as cystoblasts (CBs), also express distinct classes of intrinsic factors to inhibit self-renewal and promote germ cell differentiation. Surprisingly, GSC progeny are also dependent on their surrounding ECs for proper differentiation at least partly by preventing BMP from diffusing to the differentiated germ cell zone and by repressing ectopic BMP expression. Therefore, both GSC self-renewal and CB differentiation are controlled by collaborative actions of extrinsic signals and intrinsic factors. Copyright © 2012 Wiley Periodicals, Inc.

  11. Minichromosome maintenance helicase paralog MCM9 is dispensible for DNA replication but functions in germ-line stem cells and tumor suppression.

    Science.gov (United States)

    Hartford, Suzanne A; Luo, Yunhai; Southard, Teresa L; Min, Irene M; Lis, John T; Schimenti, John C

    2011-10-25

    Effective DNA replication is critical to the health and reproductive success of organisms. The six MCM2-7 proteins, which form the replicative helicase, are essential for high-fidelity replication of the genome. Many eukaryotes have a divergent paralog, MCM9, that was reported to be essential for loading MCM2-7 onto replication origins in the Xenopus oocyte extract system. To address the in vivo role of mammalian MCM9, we created and analyzed the phenotypes of mice with various mutations in Mcm9 and an intronic DNA replication-related gene Asf1a. Ablation of Mcm9 was compatible with cell proliferation and mouse viability, showing that it is nonessential for MCM2-7 loading or DNA replication. Mcm9 mutants underwent p53-independent embryonic germ-cell depletion in both sexes, with males also exhibiting defective spermatogonial stem-cell renewal. MCM9-deficient cells had elevated genomic instability and defective cell cycle reentry following replication stress, and mutant animals were prone to sex-specific cancers, most notably hepatocellular carcinoma in males. The phenotypes of mutant mice and cells suggest that MCM9 evolved a specialized but nonessential role in DNA replication or replication-linked quality-control mechanisms that are especially important for germ-line stem cells, and also for tumor suppression and genome maintenance in the soma.

  12. Gene organization inside replication domains in mammalian genomes

    Science.gov (United States)

    Zaghloul, Lamia; Baker, Antoine; Audit, Benjamin; Arneodo, Alain

    2012-11-01

    We investigate the large-scale organization of human genes with respect to "master" replication origins that were previously identified as bordering nucleotide compositional skew domains. We separate genes in two categories depending on their CpG enrichment at the promoter which can be considered as a marker of germline DNA methylation. Using expression data in mouse, we confirm that CpG-rich genes are highly expressed in germline whereas CpG-poor genes are in a silent state. We further show that, whether tissue-specific or broadly expressed (housekeeping genes), the CpG-rich genes are over-represented close to the replication skew domain borders suggesting some coordination of replication and transcription. We also reveal that the transcription of the longest CpG-rich genes is co-oriented with replication fork progression so that the promoter of these transcriptionally active genes be located into the accessible open chromatin environment surrounding the master replication origins that border the replication skew domains. The observation of a similar gene organization in the mouse genome confirms the interplay of replication, transcription and chromatin structure as the cornerstone of mammalian genome architecture.

  13. Spontaneous Pneumothorax in Birt-Hogg-Dube' Syndrome: Two Case Reports

    Energy Technology Data Exchange (ETDEWEB)

    Bae, Hyoung Ju; Woo, Ok Hee; Yong, Hwan Seok; Kang, Eun Young; Kim, Hyun Koo; Choi, Young Ho; Shin, Bong Kyung; Kim, Yoon Kyung [Korea University School of Medicine, Korea University Guro Hospital, Seoul (Korea, Republic of)

    2011-01-15

    Birt-Hogg-Dube'(BHD) syndrome is a rare autosomal dominant inherited disorder that is characterized by skin fibrofolliculomas, renal tumors and multiple lung cysts with or without spontaneous pneumothorax. The disease is caused by germline mutations in the FLCN gene that codes for a protein of unknown function called folliculin. Patients with BHD syndrome do not always have all three manifestations of the skin, kidney and lung. To the best of our knowledge, there has been no case report of the radiologic findings of the lung manifestation in a patient with BHD syndrome in Korea. We report here on two cases of BHD syndrome that presented with spontaneous pneumothorax. The pulmonary abnormalities consisted of multiple thin-walled cysts of various sizes and shapes in both lungs

  14. Pulmonary cystic disease associated with integumentary and renal manifestations

    Science.gov (United States)

    Cayetano, Katherine S.; Albertson, Timothy E.; Chan, Andrew L.

    2013-01-01

    A 69-year-old man with multiple skin lesions on his face, neck and upper torso, which first appeared in the 3rd decade of his life, was admitted to our hospital. He had cystic changes in his lungs noted on chest computed tomography (CT) scanning, as well as a left kidney mass. This patient exhibited a rare complex of renal, cutaneous and pulmonary manifestations, eponymously named Birt-Hogg-Dube syndrome, with characteristic skin features (fibrofolliculomas, trichodiscomas and acrochordons). This syndrome is due to an autosomal dominant germ-line mutation of the folliculin (FLCN) gene located at chromosome 17p11.2. Diagnosis and differentiation from other disease complexes including the skin, kidneys and lungs are important in prognostication and management of potentially life-threatening complications such as renal cell carcinoma and pneumothoraces. PMID:24285950

  15. Insertion of an SVA-E retrotransposon into the CASP8 gene is associated with protection against prostate cancer

    NARCIS (Netherlands)

    Stacey, S.N.; Kehr, B.; Gudmundsson, J.; Zink, F.; Jonasdottir, A.; Gudjonsson, S.A.; Sigurdsson, A.; Halldorsson, B.V.; Agnarsson, B.A.; Benediktsdottir, K.R.; Aben, K.K.; Vermeulen, S.H.; Cremers, R.G.; Panadero, A.; Helfand, B.T.; Cooper, P.R.; Donovan, J.L.; Hamdy, F.C.; Jinga, V.; Okamoto, I.; Jonasson, J.G.; Tryggvadottir, L.; Johannsdottir, H.; Kristinsdottir, A.M.; Masson, G.; Magnusson, O.T.; Iordache, P.D.; Helgason, A.; Helgason, H.; Sulem, P.; Gudbjartsson, D.F.; Kong, A.; Jonsson, E.; Barkardottir, R.B.; Einarsson, G.V.; Rafnar, T.; Thorsteinsdottir, U.; Mates, I.N.; Neal, D.E.; Catalona, W.J.; Mayordomo, J.I.; Kiemeney, L.A.; Thorleifsson, G.; Stefansson, K.

    2016-01-01

    Transcriptional and splicing anomalies have been observed in intron 8 of the CASP8 gene (encoding procaspase-8) in association with cutaneous basal-cell carcinoma (BCC) and linked to a germline SNP rs700635. Here, we show that the rs700635[C] allele, which is associated with increased risk of BCC

  16. A novel germline POLE mutation causes an early onset cancer prone syndrome mimicking constitutional mismatch repair deficiency.

    Science.gov (United States)

    Wimmer, Katharina; Beilken, Andreas; Nustede, Rainer; Ripperger, Tim; Lamottke, Britta; Ure, Benno; Steinmann, Diana; Reineke-Plaass, Tanja; Lehmann, Ulrich; Zschocke, Johannes; Valle, Laura; Fauth, Christine; Kratz, Christian P

    2017-01-01

    In a 14-year-old boy with polyposis and rectosigmoid carcinoma, we identified a novel POLE germline mutation, p.(Val411Leu), previously found as recurrent somatic mutation in 'ultramutated' sporadic cancers. This is the youngest reported cancer patient with polymerase proofreading-associated polyposis indicating that POLE mutation p.(Val411Leu) may confer a more severe phenotype than previously reported POLE and POLD1 germline mutations. The patient had multiple café-au-lait macules and a pilomatricoma mimicking the clinical phenotype of constitutional mismatch repair deficiency. We hypothesize that these skin features may be common to different types of constitutional DNA repair defects associated with polyposis and early-onset cancer.

  17. The effects of MSH2 deficiency on spontaneous and radiation-induced mutation rates in the mouse germline

    International Nuclear Information System (INIS)

    Burr, Karen L-A.; Duyn-Goedhart, Annemarie van; Hickenbotham, Peter; Monger, Karen; Buul, Paul P.W. van; Dubrova, Yuri E.

    2007-01-01

    Mutation rates at two expanded simple tandem repeat (ESTR) loci were studied in the germline of mismatch repair deficient Msh2 knock-out mice. Spontaneous mutation rates in homozygous Msh2 -/- males were significantly higher than those in isogenic wild-type (Msh2 +/+ ) and heterozygous (Msh2 +/- ) mice. In contrast, the irradiated Msh2 -/- mice did not show any detectable increases in their mutation rate, whereas significant ESTR mutation induction was observed in the irradiated Msh2 +/+ and Msh2 +/- animals. Considering these data and the results of other publications, we propose that the Msh2-deficient mice possess a mutator phenotype in their germline and somatic tissues while the loss of a single Msh2 allele does not affect the stability of heterozygotes

  18. Mei-p26 cooperates with Bam, Bgcn and Sxl to promote early germline development in the Drosophila ovary.

    Directory of Open Access Journals (Sweden)

    Yun Li

    Full Text Available In the Drosophila female germline, spatially and temporally specific translation of mRNAs governs both stem cell maintenance and the differentiation of their progeny. However, the mechanisms that control and coordinate different modes of translational repression within this lineage remain incompletely understood. Here we present data showing that Mei-P26 associates with Bam, Bgcn and Sxl and nanos mRNA during early cyst development, suggesting that this protein helps to repress the translation of nanos mRNA. Together with recently published studies, these data suggest that Mei-P26 mediates both GSC self-renewal and germline differentiation through distinct modes of translational repression depending on the presence of Bam.

  19. Mei-P26 Cooperates with Bam, Bgcn and Sxl to Promote Early Germline Development in the Drosophila Ovary

    Science.gov (United States)

    Li, Yun; Zhang, Qiao; Carreira-Rosario, Arnaldo; Maines, Jean Z.; McKearin, Dennis M.; Buszczak, Michael

    2013-01-01

    In the Drosophila female germline, spatially and temporally specific translation of mRNAs governs both stem cell maintenance and the differentiation of their progeny. However, the mechanisms that control and coordinate different modes of translational repression within this lineage remain incompletely understood. Here we present data showing that Mei-P26 associates with Bam, Bgcn and Sxl and nanos mRNA during early cyst development, suggesting that this protein helps to repress the translation of nanos mRNA. Together with recently published studies, these data suggest that Mei-P26 mediates both GSC self-renewal and germline differentiation through distinct modes of translational repression depending on the presence of Bam. PMID:23526974

  20. Germ-line CAG repeat instability causes extreme CAG repeat expansion with infantile-onset spinocerebellar ataxia type 2

    DEFF Research Database (Denmark)

    Vinther-Jensen, Tua; Ek, Jakob; Duno, Morten

    2013-01-01

    The spinocerebellar ataxias (SCA) are a genetically and clinically heterogeneous group of diseases, characterized by dominant inheritance, progressive cerebellar ataxia and diverse extracerebellar symptoms. A subgroup of the ataxias is caused by unstable CAG-repeat expansions in their respective ...... of paternal germ-line repeat sequence instability of the expanded SCA2 locus.European Journal of Human Genetics advance online publication, 10 October 2012; doi:10.1038/ejhg.2012.231....

  1. NanoTIO2 (UV-Titan does not induce ESTR mutations in the germline of prenatally exposed female mice

    Directory of Open Access Journals (Sweden)

    Boisen Anne Mette

    2012-06-01

    Full Text Available Abstract Background Particulate air pollution has been linked to an increased risk of cardiovascular disease and cancer. Animal studies have shown that inhalation of air particulates induces mutations in the male germline. Expanded simple tandem repeat (ESTR loci in mice are sensitive markers of mutagenic effects on male germ cells resulting from environmental exposures; however, female germ cells have received little attention. Oocytes may be vulnerable during stages of active cell division (e.g., during fetal development. Accordingly, an increase in germline ESTR mutations in female mice prenatally exposed to radiation has previously been reported. Here we investigate the effects of nanoparticles on the female germline. Since pulmonary exposure to nanosized titanium dioxide (nanoTiO2 produces a long-lasting inflammatory response in mice, it was chosen for the present study. Findings Pregnant C57BL/6 mice were exposed by whole-body inhalation to the nanoTiO2 UV-Titan L181 (~42.4 mg UV-Titan/m3 or filtered clean air on gestation days (GD 8–18. Female C57BL/6 F1 offspring were raised to maturity and mated with unexposed CBA males. The F2 descendents were collected and ESTR germline mutation rates in this generation were estimated from full pedigrees (mother, father, offspring of F1 female mice (192 UV-Titan-exposed F2 offspring and 164 F2 controls. ESTR mutation rates of 0.029 (maternal allele and 0.047 (paternal allele in UV-Titan-exposed F2 offspring were not statistically different from those of F2 controls: 0.037 (maternal allele and 0.061 (paternal allele. Conclusions We found no evidence for increased ESTR mutation rates in F1 females exposed in utero to UV-Titan nanoparticles from GD8-18 relative to control females.

  2. Germline activating MTOR mutation arising through gonadal mosaicism in two brothers with megalencephaly and neurodevelopmental abnormalities.

    Science.gov (United States)

    Mroske, Cameron; Rasmussen, Kristen; Shinde, Deepali N; Huether, Robert; Powis, Zoe; Lu, Hsiao-Mei; Baxter, Ruth M; McPherson, Elizabeth; Tang, Sha

    2015-11-05

    In humans, Mammalian Target of Rapamycin (MTOR) encodes a 300 kDa serine/ threonine protein kinase that is ubiquitously expressed, particularly at high levels in brain. MTOR functions as an integrator of multiple cellular processes, and in so doing either directly or indirectly regulates the phosphorylation of at least 800 proteins. While somatic MTOR mutations have been recognized in tumors for many years, and more recently in hemimegalencephaly, germline MTOR mutations have rarely been described. We report the successful application of family-trio Diagnostic Exome Sequencing (DES) to identify the underlying molecular etiology in two brothers with multiple neurological and developmental lesions, and for whom previous testing was non-diagnostic. The affected brothers, who were 6 and 23 years of age at the time of DES, presented symptoms including but not limited to mild Autism Spectrum Disorder (ASD), megalencephaly, gross motor skill delay, cryptorchidism and bilateral iris coloboma. Importantly, we determined that each affected brother harbored the MTOR missense alteration p.E1799K (c.5395G>A). This exact variant has been previously identified in multiple independent human somatic cancer samples and has been shown to result in increased MTOR activation. Further, recent independent reports describe two unrelated families in whom p.E1799K co-segregated with megalencephaly and intellectual disability (ID); in both cases, p.E1799K was shown to have originated due to germline mosaicism. In the case of the family reported herein, the absence of p.E1799K in genomic DNA extracted from the blood of either parent suggests that this alteration most likely arose due to gonadal mosaicism. Further, the p.E1799K variant exerts its effect by a gain-of-function (GOF), autosomal dominant mechanism. Herein, we describe the use of DES to uncover an activating MTOR missense alteration of gonadal mosaic origin that is likely to be the causative mutation in two brothers who present

  3. Germline-specific MATH-BTB substrate adaptor MAB1 regulates spindle length and nuclei identity in maize.

    Science.gov (United States)

    Juranič, Martina; Srilunchang, Kanok-orn; Krohn, Nádia Graciele; Leljak-Levanic, Dunja; Sprunck, Stefanie; Dresselhaus, Thomas

    2012-12-01

    Germline and early embryo development constitute ideal model systems to study the establishment of polarity, cell identity, and asymmetric cell divisions (ACDs) in plants. We describe here the function of the MATH-BTB domain protein MAB1 that is exclusively expressed in the germ lineages and the zygote of maize (Zea mays). mab1 (RNA interference [RNAi]) mutant plants display chromosome segregation defects and short spindles during meiosis that cause insufficient separation and migration of nuclei. After the meiosis-to-mitosis transition, two attached nuclei of similar identity are formed in mab1 (RNAi) mutants leading to an arrest of further germline development. Transient expression studies of MAB1 in tobacco (Nicotiana tabacum) Bright Yellow-2 cells revealed a cell cycle-dependent nuclear localization pattern but no direct colocalization with the spindle apparatus. MAB1 is able to form homodimers and interacts with the E3 ubiquitin ligase component Cullin 3a (CUL3a) in the cytoplasm, likely as a substrate-specific adapter protein. The microtubule-severing subunit p60 of katanin was identified as a candidate substrate for MAB1, suggesting that MAB1 resembles the animal key ACD regulator Maternal Effect Lethal 26 (MEL-26). In summary, our findings provide further evidence for the importance of posttranslational regulation for asymmetric divisions and germline progression in plants and identified an unstable key protein that seems to be involved in regulating the stability of a spindle apparatus regulator(s).

  4. Germline-Specific MATH-BTB Substrate Adaptor MAB1 Regulates Spindle Length and Nuclei Identity in Maize[W

    Science.gov (United States)

    Juranić, Martina; Srilunchang, Kanok-orn; Krohn, Nádia Graciele; Leljak-Levanić, Dunja; Sprunck, Stefanie; Dresselhaus, Thomas

    2012-01-01

    Germline and early embryo development constitute ideal model systems to study the establishment of polarity, cell identity, and asymmetric cell divisions (ACDs) in plants. We describe here the function of the MATH-BTB domain protein MAB1 that is exclusively expressed in the germ lineages and the zygote of maize (Zea mays). mab1 (RNA interference [RNAi]) mutant plants display chromosome segregation defects and short spindles during meiosis that cause insufficient separation and migration of nuclei. After the meiosis-to-mitosis transition, two attached nuclei of similar identity are formed in mab1 (RNAi) mutants leading to an arrest of further germline development. Transient expression studies of MAB1 in tobacco (Nicotiana tabacum) Bright Yellow-2 cells revealed a cell cycle–dependent nuclear localization pattern but no direct colocalization with the spindle apparatus. MAB1 is able to form homodimers and interacts with the E3 ubiquitin ligase component Cullin 3a (CUL3a) in the cytoplasm, likely as a substrate-specific adapter protein. The microtubule-severing subunit p60 of katanin was identified as a candidate substrate for MAB1, suggesting that MAB1 resembles the animal key ACD regulator Maternal Effect Lethal 26 (MEL-26). In summary, our findings provide further evidence for the importance of posttranslational regulation for asymmetric divisions and germline progression in plants and identified an unstable key protein that seems to be involved in regulating the stability of a spindle apparatus regulator(s). PMID:23250449

  5. The effect of tributyltin chloride on Caenorhabditis elegans germline is mediated by a conserved DNA damage checkpoint pathway.

    Science.gov (United States)

    Cheng, Zhe; Tian, Huimin; Chu, Hongran; Wu, Jianjian; Li, Yingying; Wang, Yanhai

    2014-03-21

    Tributyltin (TBT), one of the environmental pollutants, has been shown to impact the reproduction of animals. However, due to the lack of appropriate animal model, analysis of the affected molecular pathways in germ cells is lagging and has been particularly challenging. In the present study, we investigated the effects of tributyltin chloride (TBTCL) on the nematode Caenorhabditis elegans germline. We show that exposure of C. elegans to TBTCL causes significantly elevated level of sterility and embryonic lethality. TBTCL exposure results in an increased number of meiotic DNA double-strand breaks in germ cells, subsequently leading to activated DNA damage checkpoint. Exposing C. elegans to TBTCL causes dose- and time-dependent germline apoptosis. This apoptotic response was blocked in loss-of-function mutants of hus-1 (op241), mrt-2 (e2663) and p53/cep-1 (gk138), indicating that checkpoints and p53 are essential for mediating TBTCL-induced germ cell apoptosis. Moreover, TBTCL exposure can inhibit germ cell proliferation, which is also mediated by the conserved checkpoint pathway. We thereby propose that TBT exhibits its effects on the germline by inducing DNA damage and impaired maintenance of genomic integrity. Copyright © 2014 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  6. Linking the environment, DAF-7/TGFβ signaling and LAG-2/DSL ligand expression in the germline stem cell niche.

    Science.gov (United States)

    Pekar, Olga; Ow, Maria C; Hui, Kailyn Y; Noyes, Marcus B; Hall, Sarah E; Hubbard, E Jane Albert

    2017-08-15

    The developmental accumulation of proliferative germ cells in the C. elegans hermaphrodite is sensitive to the organismal environment. Previously, we found that the TGFβ signaling pathway links the environment and proliferative germ cell accumulation. Neuronal DAF-7/TGFβ causes a DAF-1/TGFβR signaling cascade in the gonadal distal tip cell (DTC), the germline stem cell niche, where it negatively regulates a DAF-3 SMAD and DAF-5 Sno-Ski. LAG-2, a founding DSL ligand family member, is produced in the DTC and activates the GLP-1/Notch receptor on adjacent germ cells to maintain germline stem cell fate. Here, we show that DAF-7/TGFβ signaling promotes expression of lag-2 in the DTC in a daf-3- dependent manner. Using ChIP and one-hybrid assays, we find evidence for direct interaction between DAF-3 and the lag-2 promoter. We further identify a 25 bp DAF-3 binding element required for the DTC lag-2 reporter response to the environment and to DAF-7/TGFβ signaling. Our results implicate DAF-3 repressor complex activity as a key molecular mechanism whereby the environment influences DSL ligand expression in the niche to modulate developmental expansion of the germline stem cell pool. © 2017. Published by The Company of Biologists Ltd.

  7. Germline signaling mediates the synergistically prolonged longevity produced by double mutations in daf-2 and rsks-1 in C. elegans.

    Science.gov (United States)

    Chen, Di; Li, Patrick Wai-Lun; Goldstein, Benjamin A; Cai, Waijiao; Thomas, Emma Lynn; Chen, Fen; Hubbard, Alan E; Melov, Simon; Kapahi, Pankaj

    2013-12-26

    Inhibition of DAF-2 (insulin-like growth factor 1 [IGF-1] receptor) or RSKS-1 (S6K), key molecules in the insulin/IGF-1 signaling (IIS) and target of rapamycin (TOR) pathways, respectively, extend lifespan in Caenorhabditis elegans. However, it has not been clear how and in which tissues they interact with each other to modulate longevity. Here, we demonstrate that a combination of mutations in daf-2 and rsks-1 produces a nearly 5-fold increase in longevity that is much greater than the sum of single mutations. This synergistic lifespan extension requires positive feedback regulation of DAF-16 (FOXO) via the AMP-activated protein kinase (AMPK) complex. Furthermore, we identify germline as the key tissue for this synergistic longevity. Moreover, germline-specific inhibition of rsks-1 activates DAF-16 in the intestine. Together, our findings highlight the importance of the germline in the significantly increased longevity produced by daf-2 rsks-1, which has important implications for interactions between the two major conserved longevity pathways in more complex organisms. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  8. Mouse embryonic stem cells efficiently lipofected with nuclear localization peptide result in a high yield of chimeric mice and retain germline transmission potency.

    Science.gov (United States)

    Ma, Haiching; Liu, Qin; Diamond, Scott L; Pierce, Eric A

    2004-06-01

    Embryonic stem (ES) cells are an important tool in developmental biology, genomics, and transgenic methods, as well as in potential clinical applications such as gene therapy or tissue engineering. Electroporation is the standard transfection method for mouse ES (mES) cells because lipofection is quite inefficient. It is also unclear if mES cells treated with cationic lipids maintain pluripotency. We have developed a simple lipofection method for high efficiency transfection and stable transgene expression by employing the nonclassical nuclear localization signal M9 derived from the heterogeneous nuclear ribonucleoprotein A1. In contrast to using 20 microg DNA for 10 x 10(6) cells via electroporation which resulted in 10-20 positive cells/mm2, M9-assisted lipofection of 2 x 10(5) cells with 2 microg DNA resulted in > 150 positive cells/mm2. Electroporation produced only 0.16% EGFP positive cells with fluorescence intensity (FI) > 1000 by FACS assay, while M9-lipofection produced 36-fold more highly EGFP positive cells (5.75%) with FI > 1000. Using 2.5 x 10(6) ES cells and 6 microg linearized DNA followed by selection with G418, electroporation yielded 17 EGFP expressing colonies, while M9-assisted lipofection yielded 72 EGFP expressing colonies. The mES cells that stably expressed EGFP following M9-assisted lipofection yielded > 66% chimeric mice (8 of 12) and contributed efficiently to the germline. In an example of gene targeting, a knock-in mouse was produced from an ES clone screened from 200 G418-resistant colonies generated via M9-assisted lipofection. To our knowledge, this is the first report of generation of transgenic or knock-in mice obtained from lipofected mES cells and this method may facilitate large scale genomic studies of ES developmental biology or large scale generation of mouse models of human disease. Copyright 2003 Elsevier Inc.

  9. E-cadherin is required for centrosome and spindle orientation in Drosophila male germline stem cells.

    Directory of Open Access Journals (Sweden)

    Mayu Inaba

    2010-08-01

    Full Text Available Many adult stem cells reside in a special microenvironment known as the niche, where they receive essential signals that specify stem cell identity. Cell-cell adhesion mediated by cadherin and integrin plays a crucial role in maintaining stem cells within the niche. In Drosophila melanogaster, male germline stem cells (GSCs are attached to niche component cells (i.e., the hub via adherens junctions. The GSC centrosomes and spindle are oriented toward the hub-GSC junction, where E-cadherin-based adherens junctions are highly concentrated. For this reason, adherens junctions are thought to provide a polarity cue for GSCs to enable proper orientation of centrosomes and spindles, a critical step toward asymmetric stem cell division. However, understanding the role of E-cadherin in GSC polarity has been challenging, since GSCs carrying E-cadherin mutations are not maintained in the niche. Here, we tested whether E-cadherin is required for GSC polarity by expressing a dominant-negative form of E-cadherin. We found that E-cadherin is indeed required for polarizing GSCs toward the hub cells, an effect that may be mediated by Apc2. We also demonstrated that E-cadherin is required for the GSC centrosome orientation checkpoint, which prevents mitosis when centrosomes are not correctly oriented. We propose that E-cadherin orchestrates multiple aspects of stem cell behavior, including polarization of stem cells toward the stem cell-niche interface and adhesion of stem cells to the niche supporting cells.

  10. ALIX and ESCRT-III coordinately control cytokinetic abscission during germline stem cell division in vivo.

    Directory of Open Access Journals (Sweden)

    Åsmund H Eikenes

    2015-01-01

    Full Text Available Abscission is the final step of cytokinesis that involves the cleavage of the intercellular bridge connecting the two daughter cells. Recent studies have given novel insight into the spatiotemporal regulation and molecular mechanisms controlling abscission in cultured yeast and human cells. The mechanisms of abscission in living metazoan tissues are however not well understood. Here we show that ALIX and the ESCRT-III component Shrub are required for completion of abscission during Drosophila female germline stem cell (fGSC division. Loss of ALIX or Shrub function in fGSCs leads to delayed abscission and the consequent formation of stem cysts in which chains of daughter cells remain interconnected to the fGSC via midbody rings and fusome. We demonstrate that ALIX and Shrub interact and that they co-localize at midbody rings and midbodies during cytokinetic abscission in fGSCs. Mechanistically, we show that the direct interaction between ALIX and Shrub is required to ensure cytokinesis completion with normal kinetics in fGSCs. We conclude that ALIX and ESCRT-III coordinately control abscission in Drosophila fGSCs and that their complex formation is required for accurate abscission timing in GSCs in vivo.

  11. Structural polarity and dynamics of male germline stem cells in the milkweed bug (Oncopeltus fasciatus).

    Science.gov (United States)

    Schmidt, Esther D; Dorn, August

    2004-11-01

    The male germline stem cells (GSCs) of the milkweed bug present an extraordinary structural polarity that is, to our knowledge, unequalled by any other type of stem cells. They consist of a perikaryon and numerous projections arising from the cell pole directed toward the apical cells, the proposed niche of the GSCs. The projections can traverse a considerable distance until their terminals touch the apical cells. From hatching until death, the GSC projections undergo conspicuous changes, the sequence of which has been deduced from observations of all developmental stages. Projection formation starts from lobular cell protrusions showing trabecular ingrowths of the cell membrane. Finger-like projections result from a process of growth and "carving out". The newly formed projections contain mostly only free ribosomes other than a few mitochondria. A stereotyped degradation process commences in the projection terminals: profiles of circular, often concentric, cisternae of rough endoplasmic reticulum appear and turn into myelin bodies, whereas mitochondria become more numerous. The cytoplasm vesiculates, lysosomal bodies appear, and mitochondria become swollen. At the same time, the projection terminals are segregated by transverse ingrowths of the cell membrane. Finally, autophagic vacuoles and myelin bodies fill the segregated terminals, which then rupture. Simultaneously, new projections seem to sprout from the perikaryon of the GSCs. These dynamics, which are not synchronized among the GSCs, indicate that a novel type of signal exchange and transduction between the stem cells and their niche is involved in the regulation of asymmetric versus symmetric division of GSCs.

  12. Germline PRKACA amplification leads to Cushing syndrome caused by 3 adrenocortical pathologic phenotypes.

    Science.gov (United States)

    Carney, J Aidan; Lyssikatos, Charalampos; Lodish, Maya B; Stratakis, Constantine A

    2015-01-01

    We describe the pathology of 5 patients with germline PRKACA copy number gain and Cushing syndrome: 4 males and 1 female, aged 2 to 43 years, including a mother and son. Imaging showed normal or slightly enlarged adrenal glands in 4 patients and a unilateral mass in the fifth. Biochemically, the patients had corticotropin-independent hypercortisolism. Four underwent bilateral adrenalectomy; unilateral adrenalectomy was performed in the patient with the adrenal mass. Pathologically, 3 patients, including the 1 with the tumor (adenoma), had primary pigmented nodular adrenocortical disease with extranodular cortical atrophy and mild intracapsular and extracapsular extension of cortical cells. The other 2 patients had cortical hyperplasia and prominent capsular and extracapsular micronodular cortical hyperplasia. Immunoperoxidase staining revealed differences for synaptophysin, inhibin-A, and Ki-67 (nuclei) in the atrophic cortices (patients 1, 2, and 3) and hyperplastic cortices (patients 4 and 5) and for Ki-67 (nuclei) and vimentin in the extracortical nodules in the 2 groups of patients. β-Catenin stained the cell membrane, cytoplasm, and nuclei of the adenoma. The patients were well at follow-up (1-23 years); 24-hour urinary cortisol excretion was elevated in the patient who had unilateral adrenalectomy. Copyright © 2014 Elsevier Inc. All rights reserved.

  13. X-ray induced polyploidization in the male germline cells of Poekilocerus pictus (acrididoidea : orthopta)

    International Nuclear Information System (INIS)

    Gururaj, M.E.; Rajasekarasetty, M.R.

    1977-01-01

    After the irradiation of male germline cells of Poekilocerus pictus with 20r, 40r, 80r, 120r doses of X-rays, both first and second meiotic polyploid cells were recovered. While various degrees of polyploidy were encountered in first meiotic cells, second meiotic polyploid cells, second meitoic polyploid cells contained diploid number of half bivalents only. The former never progressed beyond leptotene and showed symptoms of degeneration. Among the latter, a few cells showed either emainingative tendencies like uncoiling and stickiness or failure of cellsted meiosis successfully. It has been shown that the dicentric bridges and/or laggards in anaphase-I interfere with the elongation and regression of the spindle, thereby giving rise to metaphase-II polyploid cells through restitution. The possible role of fragmentation of chromosomes in decreasing the incidence of metaphase-II polyploid cells at higher doses of irradiation and the causes for the differential fate of the first and second meiotic polyploid cells have been discussed. (author)

  14. Somatic insulin signaling regulates a germline starvation response in Drosophila egg chambers

    Science.gov (United States)

    Burn, K. Mahala; Shimada, Yuko; Ayers, Kathleen; Lu, Feiyue; Hudson, Andrew M.; Cooley, Lynn

    2014-01-01

    Egg chambers from starved Drosophila females contain large aggregates of processing (P) bodies and cortically enriched microtubules. As this response to starvation is rapidly reversed upon re-feeding females or culturing egg chambers with exogenous bovine insulin, we examined the role of endogenous insulin signaling in mediating the starvation response. We found that systemic Drosophila insulin-like peptides (dILPs) activate the insulin pathway in follicle cells, which then regulate both microtubule and P body organization in the underlying germline cells. This organization is modulated by the motor proteins Dynein and Kinesin. Dynein activity is required for microtubule and P body organization during starvation, while Kinesin activity is required during nutrient-rich conditions. Blocking the ability of egg chambers to form P body aggregates in response to starvation correlated with reduced progeny survival. These data suggest a potential mechanism to maximize fecundity even during periods of poor nutrient availability, by mounting a protective response in immature egg chambers. PMID:25481758

  15. Plasticity of male germline stem cells and their applications in reproductive and regenerative medicine

    Directory of Open Access Journals (Sweden)

    Zheng Chen

    2015-06-01

    Full Text Available Spermatogonial stem cells (SSCs, also known as male germline stem cells, are a small subpopulation of type A spermatogonia with the potential of self-renewal to maintain stem cell pool and differentiation into spermatids in mammalian testis. SSCs are previously regarded as the unipotent stem cells since they can only give rise to sperm within the seminiferous tubules. However, this concept has recently been challenged because numerous studies have demonstrated that SSCs cultured with growth factors can acquire pluripotency to become embryonic stem-like cells. The in vivo and in vitro studies from peers and us have clearly revealed that SSCs can directly transdifferentiate into morphologic, phenotypic, and functional cells of other lineages. Direct conversion to the cells of other tissues has important significance for regenerative medicine. SSCs from azoospermia patients could be induced to differentiate into spermatids with fertilization and developmental potentials. As such, SSCs could have significant applications in both reproductive and regenerative medicine due to their unique and great potentials. In this review, we address the important plasticity of SSCs, with focuses on their self-renewal, differentiation, dedifferentiation, transdifferentiation, and translational medicine studies.

  16. Germline Chromothripsis Driven by L1-Mediated Retrotransposition and Alu/Alu Homologous Recombination

    DEFF Research Database (Denmark)

    Nazaryan-Petersen, Lusine; Bertelsen, Birgitte; Bak, Mads

    2016-01-01

    Chromothripsis (CTH) is a phenomenon where multiple localized double-stranded DNA breaks result in complex genomic rearrangements. Although the DNA-repair mechanisms involved in CTH have been described, the mechanisms driving the localized "shattering" process remain unclear. High-throughput sequ......Chromothripsis (CTH) is a phenomenon where multiple localized double-stranded DNA breaks result in complex genomic rearrangements. Although the DNA-repair mechanisms involved in CTH have been described, the mechanisms driving the localized "shattering" process remain unclear. High......-throughput sequence analysis of a familial germline CTH revealed an inserted SVAE retrotransposon associated with a 110-kb deletion displaying hallmarks of L1-mediated retrotransposition. Our analysis suggests that the SVAE insertion did not occur prior to or after, but concurrent with the CTH event. We also observed...... L1-endonuclease potential target sites in other breakpoints. In addition, we found four Alu elements flanking the 110-kb deletion and associated with an inversion. We suggest that chromatin looping mediated by homologous Alu elements may have brought distal DNA regions into close proximity...

  17. Sexually Antagonistic Male Signals Manipulate Germline and Soma of C. elegans Hermaphrodites.

    Science.gov (United States)

    Aprison, Erin Z; Ruvinsky, Ilya

    2016-10-24

    Males and females pursue different reproductive strategies, which often bring them into conflict-many traits exist that benefit one sex at a cost to another [1]. Decreased female survival following mating dramatically demonstrates one aspect of this phenomenon [2-5]. Particularly intriguing is the evidence that secreted compounds can shorten lifespan of members of the opposite sex in Drosophila [6] and Caenorhabditid nematodes [7] even without copulation taking place. The purpose of such signals is not clear, however. While it is possible that they could limit subsequent mating with competitors or hasten post-reproductive demise, thus decreasing competition for resources, they are also likely to harm unmated individuals. Why would a system exist that reduces the vigor of potential mates prior to mating? Addressing this question could provide insights into mechanisms and evolution of sexual conflict and reveal sensory inputs that regulate aging. Here, we describe two distinct ways in which Caenorhabditis elegans males cause faster somatic aging of hermaphrodites but also manipulate different aspects of their reproductive physiology. The first, mediated by conserved ascaroside pheromones, delays the loss of germline progenitor cells. The second accelerates development, resulting in faster sexual maturation. These signals promote male reproductive strategy and the effects harmful to hermaphrodites appear to be collateral damage rather than the goal. Copyright © 2016 Elsevier Ltd. All rights reserved.

  18. Aubergine Controls Germline Stem Cell Self-Renewal and Progeny Differentiation via Distinct Mechanisms.

    Science.gov (United States)

    Ma, Xing; Zhu, Xiujuan; Han, Yingying; Story, Benjamin; Do, Trieu; Song, Xiaoqing; Wang, Su; Zhang, Ying; Blanchette, Marco; Gogol, Madelaine; Hall, Kate; Peak, Allison; Anoja, Perera; Xie, Ting

    2017-04-24

    Piwi family protein Aubergine (Aub) maintains genome integrity in late germ cells of the Drosophila ovary through Piwi-associated RNA-mediated repression of transposon activities. Although it is highly expressed in germline stem cells (GSCs) and early progeny, it remains unclear whether it plays any roles in early GSC lineage development. Here we report that Aub promotes GSC self-renewal and GSC progeny differentiation. RNA-iCLIP results show that Aub binds the mRNAs encoding self-renewal and differentiation factors in cultured GSCs. Aub controls GSC self-renewal by preventing DNA-damage-induced Chk2 activation and by translationally controlling the expression of self-renewal factors. It promotes GSC progeny differentiation by translationally controlling the expression of differentiation factors, including Bam. Therefore, this study reveals a function of Aub in GSCs and their progeny, which promotes translation of self-renewal and differentiation factors by directly binding to its target mRNAs and interacting with translational initiation factors. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Role of key-regulator genes in melanoma susceptibility and pathogenesis among patients from South Italy

    International Nuclear Information System (INIS)

    Casula, Milena; Sini, MariaCristina; Palomba, Grazia; The Italian Melanoma Intergroup; Palmieri, Giuseppe; Muggiano, Antonio; Cossu, Antonio; Budroni, Mario; Caracò, Corrado; Ascierto, Paolo A; Pagani, Elena; Stanganelli, Ignazio; Canzanella, Sergio

    2009-01-01

    Several genetic alterations have been demonstrated to contribute to the development and progression of melanoma. In this study, we further investigated the impact of key-regulator genes in susceptibility and pathogenesis of such a disease. A large series (N = 846) of sporadic and familial cases originating from South Italy was screened for germline mutations in p16 CDKN2A , BRCA2, and MC1R genes by DHPLC analysis and automated DNA sequencing. Paired primary melanomas and lymph node metastases from same patients (N = 35) as well as melanoma cell lines (N = 18) were analyzed for somatic mutations in NRAS, BRAF, and p16 CDKN2A genes. For melanoma susceptibility, investigations at germline level indicated that p16 CDKN2A was exclusively mutated in 16/545 (2.9%) non-Sardinian patients, whereas BRCA2 germline mutations were observed in 4/91 (4.4%) patients from North Sardinia only. Two MC1R germline variants, Arg151Cys and Asp294His, were significantly associated with melanoma in Sardinia. Regarding genetic events involved in melanoma pathogenesis at somatic level, mutually-exclusive mutations of NRAS and BRAF genes were observed at quite same rate (about two thirds) in cultured and in vivo melanomas (either primary or metastatic lesions). Conversely, p16 CDKN2A gene alterations were observed at increased rates moving from primary to metastatic melanomas and melanoma cell lines. Activation of the ERK gene product was demonstrated to be consistently induced by a combination of molecular alterations (NRAS/BRAF mutations and p16 CDKN2A silencing). Our findings further clarified that: a) mutation prevalence in melanoma susceptibility genes may vary within each specific geographical area; b) multiple molecular events are accumulating during melanomagenesis

  20. Preleukemic and second-hit mutational events in an acute myeloid leukemia patient with a novel germline RUNX1 mutation.

    Science.gov (United States)

    Ng, Isaac Ks; Lee, Joanne; Ng, Christopher; Kosmo, Bustamin; Chiu, Lily; Seah, Elaine; Mok, Michelle Meng Huang; Tan, Karen; Osato, Motomi; Chng, Wee-Joo; Yan, Benedict; Tan, Lip Kun

    2018-01-01

    Germline mutations in the RUNX1 transcription factor give rise to a rare autosomal dominant genetic condition classified under the entity: Familial Platelet Disorders with predisposition to Acute Myeloid Leukaemia (FPD/AML). While several studies have identified a myriad of germline RUNX1 mutations implicated in this disorder, second-hit mutational events are necessary for patients with hereditary thrombocytopenia to develop full-blown AML. The molecular picture behind this process remains unclear. We describe a patient of Malay descent with an unreported 7-bp germline RUNX1 frameshift deletion, who developed second-hit mutations that could have brought about the leukaemic transformation from a pre-leukaemic state. These mutations were charted through the course of the treatment and stem cell transplant, showing a clear correlation between her clinical presentation and the mutations present. The patient was a 27-year-old Malay woman who presented with AML on the background of hereditary thrombocytopenia affecting her father and 3 brothers. Initial molecular testing revealed the same novel RUNX1 mutation in all 5 individuals. The patient received standard induction, consolidation chemotherapy, and a haploidentical stem cell transplant from her mother with normal RUNX1 profile. Comprehensive genomic analyses were performed at diagnosis, post-chemotherapy and post-transplant. A total of 8 mutations ( RUNX1 , GATA2 , DNMT3A , BCORL1 , BCOR , 2 PHF6 and CDKN2A ) were identified in the pre-induction sample, of which 5 remained ( RUNX1 , DNMT3A , BCORL1 , BCOR and 1 out of 2 PHF6 ) in the post-treatment sample and none were present post-transplant. In brief, the 3 mutations which were lost along with the leukemic cells at complete morphological remission were most likely acquired leukemic driver mutations that were responsible for the AML transformation from a pre-leukemic germline RUNX1 -mutated state. On the contrary, the 5 mutations that persisted post

  1. Management of Gene Variants of Unknown Significance

    DEFF Research Database (Denmark)

    Alosi, Daniela; Bisgaard, Marie Luise; Hemmingsen, Sophie Nowak

    2017-01-01

    by germline mutations in the VHL gene, which predispose to the development of multiple tumors such as central nervous system hemangioblastomas and renal cell carcinoma (RCC). Objective: We propose a method for the evaluation of VUS pathogenicity through our experience with the VHL missense mutation c.241C...... (IHC); 3) Assessment of the variant’s impact on protein structure and function, using multiple databases, in silico algorithms, and reports of functional studies. Results: Only one family member had clinical signs of vHL with early-onset RCC. IHC analysis showed no VHL protein expressed in the tumor...

  2. Identification of genes expressed in the hermaphrodite germ line of C. elegans using SAGE

    Science.gov (United States)

    Wang, Xin; Zhao, Yongjun; Wong, Kim; Ehlers, Peter; Kohara, Yuji; Jones, Steven J; Marra, Marco A; Holt, Robert A; Moerman, Donald G; Hansen, Dave

    2009-01-01

    Background Germ cells must progress through elaborate developmental stages from an undifferentiated germ cell to a fully differentiated gamete. Some of these stages include exiting mitosis and entering meiosis, progressing through the various stages of meiotic prophase, adopting either a male (sperm) or female (oocyte) fate, and completing meiosis. Additionally, many of the factors needed to drive embryogenesis are synthesized in the germ line. To increase our understanding of the genes that might be necessary for the formation and function of the germ line, we have constructed a SAGE library from hand dissected C. elegans hermaphrodite gonads. Results We found that 4699 genes, roughly 21% of all known C. elegans genes, are expressed in the adult hermaphrodite germ line. Ribosomal genes are highly expressed in the germ line; roughly four fold above their expression levels in the soma. We further found that 1063 of the germline-expressed genes have enriched expression in the germ line as compared to the soma. A comparison of these 1063 germline-enriched genes with a similar list of genes prepared using microarrays revealed an overlap of 460 genes, mutually reinforcing the two lists. Additionally, we identified 603 germline-enriched genes, supported by in situ expression data, which were not previously identified. We also found >4 fold enrichment for RNA binding proteins in the germ line as compared to the soma. Conclusion Using multiple technological platforms provides a more complete picture of global gene expression patterns. Genes involved in RNA metabolism are expressed at a significantly higher level in the germ line than the soma, suggesting a stronger reliance on RNA metabolism for control of the expression of genes in the germ line. Additionally, the number and expression level of germ line expressed genes on the X chromosome is lower than expected based on a random distribution. PMID:19426519

  3. Identification of genes expressed in the hermaphrodite germ line of C. elegans using SAGE

    Directory of Open Access Journals (Sweden)

    Holt Robert A

    2009-05-01

    Full Text Available Abstract Background Germ cells must progress through elaborate developmental stages from an undifferentiated germ cell to a fully differentiated gamete. Some of these stages include exiting mitosis and entering meiosis, progressing through the various stages of meiotic prophase, adopting either a male (sperm or female (oocyte fate, and completing meiosis. Additionally, many of the factors needed to drive embryogenesis are synthesized in the germ line. To increase our understanding of the genes that might be necessary for the formation and function of the germ line, we have constructed a SAGE library from hand dissected C. elegans hermaphrodite gonads. Results We found that 4699 genes, roughly 21% of all known C. elegans genes, are expressed in the adult hermaphrodite germ line. Ribosomal genes are highly expressed in the germ line; roughly four fold above their expression levels in the soma. We further found that 1063 of the germline-expressed genes have enriched expression in the germ line as compared to the soma. A comparison of these 1063 germline-enriched genes with a similar list of genes prepared using microarrays revealed an overlap of 460 genes, mutually reinforcing the two lists. Additionally, we identified 603 germline-enriched genes, supported by in situ expression data, which were not previously identified. We also found >4 fold enrichment for RNA binding proteins in the germ line as compared to the soma. Conclusion Using multiple technological platforms provides a more complete picture of global gene expression patterns. Genes involved in RNA metabolism are expressed at a significantly higher level in the germ line than the soma, suggesting a stronger reliance on RNA metabolism for control of the expression of genes in the germ line. Additionally, the number and expression level of germ line expressed genes on the X chromosome is lower than expected based on a random distribution.

  4. Germline MLH1 Mutations Are Frequently Identified in Lynch Syndrome Patients With Colorectal and Endometrial Carcinoma Demonstrating Isolated Loss of PMS2 Immunohistochemical Expression.

    Science.gov (United States)

    Dudley, Beth; Brand, Randall E; Thull, Darcy; Bahary, Nathan; Nikiforova, Marina N; Pai, Reetesh K

    2015-08-01

    Current guidelines on germline mutation testing for patients suspected of having Lynch syndrome are not entirely clear in patients with tumors demonstrating isolated loss of PMS2 immunohistochemical expression. We analyzed the clinical and pathologic features of patients with tumors demonstrating isolated loss of PMS2 expression in an attempt to (1) determine the frequency of germline MLH1 and PMS2 mutations and (2) correlate mismatch-repair protein immunohistochemistry and tumor histology with germline mutation results. A total of 3213 consecutive colorectal carcinomas and 215 consecutive endometrial carcinomas were prospectively analyzed for DNA mismatch-repair protein expression by immunohistochemistry. In total, 32 tumors from 31 patients demonstrated isolated loss of PMS2 immunohistochemical expression, including 16 colorectal carcinomas and 16 endometrial carcinomas. Microsatellite instability (MSI) polymerase chain reaction was performed in 29 tumors from 28 patients with the following results: 28 tumors demonstrated high-level MSI, and 1 tumor demonstrated low-level MSI. Twenty of 31 (65%) patients in the study group had tumors demonstrating histopathology associated with high-level MSI. Seventeen patients underwent germline mutation analysis with the following results: 24% with MLH1 mutations, 35% with PMS2 mutations, 12% with PMS2 variants of undetermined significance, and 29% with no mutations in either MLH1 or PMS2. Three of the 4 patients with MLH1 germline mutations had a mutation that results in decreased stability and quantity of the MLH1 protein that compromises the MLH1-PMS2 protein complex, helping to explain the presence of immunogenic but functionally inactive MLH1 protein within the tumor. The high frequency of MLH1 germline mutations identified in our study has important implications for testing strategies in patients suspected of having Lynch syndrome and indicates that patients with tumors demonstrating isolated loss of PMS2 expression

  5. Frequency of CDH1 germline mutations in gastric carcinoma coming from high- and low-risk areas: metanalysis and systematic review of the literature

    International Nuclear Information System (INIS)

    Corso, Giovanni; Marrelli, Daniele; Pascale, Valeria; Vindigni, Carla; Roviello, Franco

    2012-01-01

    The frequency of E-cadherin germline mutations in countries with different incidence rates for gastric carcinoma has not been well established. The goal of this study was to assess the worldwide frequency of CDH1 germline mutations in gastric cancers coming from low- and high-risk areas. English articles using MEDLINE access (from 1998 to 2011). Search terms included CDH1, E-cadherin, germline mutation, gastric cancer, hereditary, familial and diffuse histotype. The study included all E-cadherin germline mutations identified in gastric cancer patients; somatic mutations and germline mutations reported in other tumors were excluded. The method of this study was scheduled in accordance with the 'PRISMA statement for reporting systematic reviews and meta-analyses'. Countries were classified as low- or middle/high risk-areas for gastric carcinoma incidence. Statistical analysis was performed to correlate the CDH1 mutation frequency with gastric cancer incidence areas. A total of 122 E-cadherin germline mutations have been identified; the majority (87.5%) occurred in gastric cancers coming from low-risk areas. In high-risk areas, we identified 16 mutations in which missense mutations were predominant. (68.8%). We verified a significant association between the mutation frequency and the gastric cancer risk area (p < 0.001: overall identified mutations in low- vs. middle/high-risk areas). E-cadherin genetic screenings performed in low-risk areas for gastric cancer identified a higher frequency of CDH1 germline mutations. This data could open new approaches in the gastric cancer prevention test; before proposing a proband candidate for the CDH1 genetic screening, geographic variability, alongside the family history should be considered

  6. Screening for Novel Germline Rare Mutations Associated with Aggressive Prostate Cancer

    Science.gov (United States)

    2015-12-01

    Tyrosine Kinase, Endothelial), CDH2 (Cadherin 2), and BEST2 (Bestrophin 2); while in the CAPS population, the gene showed significant association...Reduced proliferation of CD34(+) cells from patients with acute myeloid leukemia after gene transfer of INPP5D. Gene Ther 16: 570 -3. doi: 10.1038

  7. Association between common germline genetic variation in 94 candidate genes or regions and risks of invasive epithelial ovarian cancer

    DEFF Research Database (Denmark)

    Quaye, Lydia; Tyrer, Jonathan; Ramus, Susan J

    2009-01-01

    BACKGROUND: Recent studies have identified several single nucleotide polymorphisms (SNPs) in the population that are associated with variations in the risks of many different diseases including cancers such as breast, prostate and colorectal. For ovarian cancer, the known highly penetrant suscept...

  8. Germline mutations in people descendants occupationally exposed to ionizing radiation from Cesium 137

    International Nuclear Information System (INIS)

    Silva, Juliana Ferreira da

    2016-01-01

    significant differences between exposed and control groups using the Mann-Whitney U test. Thus, our data showed that CNVs are induced by IR exposure in a human population, while the losses were more frequent the gains in the exposed group. In addition, progeny from a population occupationally exposed to IR ∼ 1.15x showed CNV more new than healthy controls. Therefore, with the present study was possible to validate the use of a high resolution method to describe a mutagenic exposure by IR signature, thus legitimized the use of CNVs as a useful biomarker to assess germline mutation military occupationally exposed to RI. In addition to validating the use of this marker, the study also pioneered research germline mutation in humans exposed to RI. (author)

  9. Genes and Gene Therapy

    Science.gov (United States)

    ... correctly, a child can have a genetic disorder. Gene therapy is an experimental technique that uses genes to ... or prevent disease. The most common form of gene therapy involves inserting a normal gene to replace an ...

  10. Expanded simple tandem repeat (ESTR) mutation induction in the male germline: Lessons learned from lab mice

    Energy Technology Data Exchange (ETDEWEB)

    Somers, Christopher M. [University of Regina, Department of Biology, 3737 Wascana Parkway, Regina, SK, S4S 0A2 (Canada)]. E-mail: chris.somers@uregina.ca

    2006-06-25

    Expanded simple tandem repeat (ESTR) DNA loci that are unstable in the germline have provided the most sensitive tool ever developed for investigating low-dose heritable mutation induction in laboratory mice. Ionizing radiation exposures have shown that ESTR mutations occur mainly in pre-meiotic spermatogonia and stem cells. The average spermatogonial doubling dose is 0.62-0.69 Gy for low LET, and 0.18-0.34 Gy for high LET radiation. Chemical alkylating agents also cause significant ESTR mutation induction in pre-meiotic spermatogonia and stem cells, but are much less effective per unit dose than radiation. ESTR mutation induction efficiency is maximal at low doses of radiation or chemical mutagens, and may decrease at higher dose ranges. DNA repair deficient mice (SCID and PARP-1) with elevated levels of single and double-strand DNA breaks have spontaneously elevated ESTR mutation frequencies, and surprisingly do not show additional ESTR mutation induction following irradiation. In contrast, ESTR mutation induction in p53 knock-outs is indistinguishable from that of wild-type mice. Studies of sentinel mice exposed in situ to ambient air pollution showed elevated ESTR mutation frequencies in males exposed to high levels of particulate matter. These studies highlight the application of the ESTR assay for assessing environmental hazards under real-world conditions. All ESTR studies to date have shown untargeted mutations that occur at much higher frequencies than predicted. The mechanism of this untargeted mutation induction is unknown, and must be elucidated before we can fully understand the biological significance of ESTR mutations, or use these markers for formal risk assessment. Future studies should focus on the mechanism of ESTR mutation induction, refining dose responses, and developing ESTR markers for other animal species.

  11. Risk Profile of the RET A883F Germline Mutation: An International Collaborative Study.

    Science.gov (United States)

    Mathiesen, Jes Sloth; Habra, Mouhammed Amir; Bassett, John Howard Duncan; Choudhury, Sirazum Mubin; Balasubramanian, Sabapathy Prakash; Howlett, Trevor A; Robinson, Bruce G; Gimenez-Roqueplo, Anne-Paule; Castinetti, Frederic; Vestergaard, Peter; Frank-Raue, Karin

    2017-06-01

    The A883F germline mutation of the rearranged during transfection (RET) proto-oncogene causes multiple endocrine neoplasia 2B. In the revised American Thyroid Association (ATA) guidelines for the management of medullary thyroid carcinoma (MTC), the A883F mutation has been reclassified from the highest to the high-risk level, although no well-defined risk profile for this mutation exists. To create a risk profile for the A883F mutation for appropriate classification among the ATA risk levels. Retrospective analysis. International collaboration. Included were 13 A883F carriers. The intervention was thyroidectomy. Earliest age of MTC, regional lymph node metastases, distant metastases, age-related penetrance of MTC and pheochromocytoma (PHEO), overall and disease-specific survival, and biochemical cure rate. One and three carriers were diagnosed at age 7 to 9 years (median, 7.5 years) with a normal thyroid and C-cell hyperplasia, respectively. Nine carriers were diagnosed with MTC at age 10 to 39 years (median, 19 years). The earliest age of MTC, regional lymph node metastasis, and distant metastasis was 10, 20, and 20 years, respectively. Fifty percent penetrance of MTC and PHEO was achieved by age 19 and 34 years, respectively. Five- and 10-year survival rates (both overall and disease specific) were 88% and 88%, respectively. Biochemical cure for MTC at latest follow-up