Sample records for abl fusion gene


    Institute of Scientific and Technical Information of China (English)

    WANG Chun-hong; SUN Bing-zhong; YUAN Yue-chuan


    Objective: To investigate the effect of bcr-abl fusion gene on CML cell apoptosis. Methods: Apoptosis of exvivo cultured K562 cells were observed after exposure to synthetic 18 mer antisense oligodeoxynucleotide complementary to the bcr-abl junction (b3a2). Results: Apoptosis of K562 cells was significantly increased associated with inhibition of bcr-abl expression. Conclusion: bcr-abl fusion gene formation due to chromosome translocation may be the major mechanism of CML via inhibition of apoptosis.

  2. The Study on BCR/ABL Fusion Gene in Adult Acute Lymphoblastic Leukemia

    Institute of Scientific and Technical Information of China (English)


    In order to assess the significance of BCR/ABC fusion gene in adult acute lymphoblastic Leukemia (ALL), 28 patients who were diagnosed as ALL were enrolled to detect BCR/ABC gene using nested-RT PCR. The results showed that 9 cases (31.25%) were BCR/ABL positive ,and expressed P210 subtype. A mong them 7 cases were B-ALL, and one was T-ALL. The diagnosis was proved by monoclonal antibodies recognition by indirect immunofluorescence. Adult patients with BCR/ABL positive ALL were significantly older (p<0. 01) and had higher WBC count (p<0. 01) as compared with BCR/ABL-negative patients. There was no significant difference in sex, hemoglobin and splenomegaly between two group (p>0. 05). The induc tion failure rate was high in BCR/ABL positive patients and those who achieved complete remission usually relapsed earlier. In conclusion, adult ALL patients with BCR/ABL-positive have poorer prognosis.

  3. Biosensing of BCR/ABL fusion gene using an intensity-interrogation surface plasmon resonance imaging system (United States)

    Wu, Jiangling; Huang, Yu; Bian, Xintong; Li, DanDan; Cheng, Quan; Ding, Shijia


    In this work, a custom-made intensity-interrogation surface plasmon resonance imaging (SPRi) system has been developed to directly detect a specific sequence of BCR/ABL fusion gene in chronic myelogenous leukemia (CML). The variation in the reflected light intensity detected from the sensor chip composed of gold islands array is proportional to the change of refractive index due to the selective hybridization of surface-bound DNA probes with target ssDNA. SPRi measurements were performed with different concentrations of synthetic target DNA sequence. The calibration curve of synthetic target sequence shows a good relationship between the concentration of synthetic target and the change of reflected light intensity. The detection limit of this SPRi measurement could approach 10.29 nM. By comparing SPRi images, the target ssDNA and non-complementary DNA sequence are able to be distinguished. This SPRi system has been applied for assay of BCR/ABL fusion gene extracted from real samples. This nucleic acid-based SPRi biosensor therefore offers an alternative high-effective, high-throughput label-free tool for DNA detection in biomedical research and molecular diagnosis.

  4. Rapid detection of BCR-ABL fusion genes using a novel combined LUX primer, in-cell RT-PCR and flow cytometric method. (United States)

    Shi, Yan; Li, Li-Zhen; Sun, Jian-Zhi; Zhang, Ti; Peng, Jun; Xu, Cong-Gao


    Currently, quantitative and semiquantitative assays for minimal residual disease detection include fluorescence in situ hybridisation, multiparameter flow cytometric immunophenotyping and real-time quantitative polymerase chain reaction (RQ-PCR). We have developed a new approach to detect hybrid breakpoint cluster region and Abelson proto-oncogene (BCR-ABL) transcripts inside suspension cells using in situ RT-PCR and light upon extension (LUX) primer, followed by rapid quantitative analysis with flow cytometry. After cellular permeabilization and fixation of single cell suspension, the neoplastic mRNA was reverse transcribed and amplified by PCR with LUX primer. The results demonstrated that a strong positive yellow-green signal was observed in 99-100% cells of K562 cell line, only the red nucleus was detected in NB4 cell line and normal controls. The technique has been utilised to study 12 patients with chronic myeloid leukemia, and the results were compared with those of BCR-ABL fusion mRNA by RT-PCR and BCR-ABL fusion gene of the interphase cells by fluorescence in situ hybridization (FISH). In the five diagnosed patients, 90-98% cells were strongly positive. Four patients, including three patients treated with interferon-alpha and hydroxyurea and one patient treated with imatinib mesylate, had 26-82.5% positive cells. Three patients treated with imatinib mesylate were negative. The in situ RT-PCR results demonstrated complete concordance with the results of I-FISH and RT-PCR. A fluorescence signal was detectable at 1/10(4) cells and became negative below this threshold with flow cytometry. The results of the present study suggest that (1) LUX primers can be used to efficiently detect BCR-ABL fusion mRNA by in-cell RT-PCR; (2) the novel technique is a specific and sensitive way of detecting fusion gene with potential clinical usefulness.

  5. Characterization of leukemias with ETV6-ABL1 fusion (United States)

    Zaliova, Marketa; Moorman, Anthony V.; Cazzaniga, Giovanni; Stanulla, Martin; Harvey, Richard C.; Roberts, Kathryn G.; Heatley, Sue L.; Loh, Mignon L.; Konopleva, Marina; Chen, I-Ming; Zimmermannova, Olga; Schwab, Claire; Smith, Owen; Mozziconacci, Marie-Joelle; Chabannon, Christian; Kim, Myungshin; Frederik Falkenburg, J. H.; Norton, Alice; Marshall, Karen; Haas, Oskar A.; Starkova, Julia; Stuchly, Jan; Hunger, Stephen P.; White, Deborah; Mullighan, Charles G.; Willman, Cheryl L.; Stary, Jan; Trka, Jan; Zuna, Jan


    To characterize the incidence, clinical features and genetics of ETV6-ABL1 leukemias, representing targetable kinase-activating lesions, we analyzed 44 new and published cases of ETV6-ABL1-positive hematologic malignancies [22 cases of acute lymphoblastic leukemia (13 children, 9 adults) and 22 myeloid malignancies (18 myeloproliferative neoplasms, 4 acute myeloid leukemias)]. The presence of the ETV6-ABL1 fusion was ascertained by cytogenetics, fluorescence in-situ hybridization, reverse transcriptase-polymerase chain reaction and RNA sequencing. Genomic and gene expression profiling was performed by single nucleotide polymorphism and expression arrays. Systematic screening of more than 4,500 cases revealed that in acute lymphoblastic leukemia ETV6-ABL1 is rare in childhood (0.17% cases) and slightly more common in adults (0.38%). There is no systematic screening of myeloproliferative neoplasms; however, the number of ETV6-ABL1-positive cases and the relative incidence of acute lymphoblastic leukemia and myeloproliferative neoplasms suggest that in adulthood ETV6-ABL1 is more common in BCR-ABL1-negative chronic myeloid leukemia-like myeloproliferations than in acute lymphoblastic leukemia. The genomic profile of ETV6-ABL1 acute lymphoblastic leukemia resembled that of BCR-ABL1 and BCR-ABL1-like cases with 80% of patients having concurrent CDKN2A/B and IKZF1 deletions. In the gene expression profiling all the ETV6-ABL1-positive samples clustered in close vicinity to BCR-ABL1 cases. All but one of the cases of ETV6-ABL1 acute lymphoblastic leukemia were classified as BCR-ABL1-like by a standardized assay. Over 60% of patients died, irrespectively of the disease or age subgroup examined. In conclusion, ETV6-ABL1 fusion occurs in both lymphoid and myeloid leukemias; the genomic profile and clinical behavior resemble BCR-ABL1-positive malignancies, including the unfavorable prognosis, particularly of acute leukemias. The poor outcome suggests that treatment with

  6. Coupling a universal DNA circuit with graphene sheets/polyaniline/AuNPs nanocomposites for the detection of BCR/ABL fusion gene

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Xueping [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China); Wang, Li [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China); Department of Medical Laboratory, Chongqing Emergency Medical Center (Chongqing The Fourth Hospital), Chongqing, 400016 (China); Sheng, Shangchun [The No.2 Peoples' Hospital of Yibin, Sichuan, 644000 (China); Wang, Teng; Yang, Juan [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China); Xie, Guoming, E-mail: [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China); Feng, Wenli, E-mail: [Key Laboratory of Laboratory Medical Diagnostics of Education, Department of Laboratory Medicine, Chongqing Medical University, Chongqing, 400016 (China)


    This article described a novel method by coupling a universal DNA circuit with graphene sheets/polyaniline/AuNPs nanocomposites (GS/PANI/AuNPs) for highly sensitive and specific detection of BCR/ABL fusion gene (bcr/abl) in chronic myeloid leukemia (CML). DNA circuit known as catalyzed hairpin assembly (CHA) is enzyme-free and can be simply operated to achieve exponential amplification, which has been widely employed in biosensing. However, application of CHA has been hindered by the need of specially redesigned sequences for each single-stranded DNA input. Herein, a transducer hairpin (HP) was designed to obtain a universal DNA circuit with favorable signal-to-background ratio. To further improve signal amplification, GS/PANI/AuNPs with excellent conductivity and enlarged effective area were introduced into this DNA circuit. Consequently, by combining the advantages of CHA and GS/PANI/AuNPs, bcr/abl could be detected in a linear range from 10 pM to 20 nM with a detection limit of 1.05 pM. Moreover, this protocol showed excellent specificity, good stability and was successfully applied for the detection of real sample, which demonstrated its great potential in clinical application. - Highlights: • A transducer hairpin was designed to improve the versatility of DNA circuit. • GS/PANI/AuNPs were introduced to the DNA circuit for further signal amplification. • The established biosensor displayed high sensitivity and good specificity.

  7. Frequency of the ETV6-RUNX1, BCR-ABL1, TCF3-PBX1, and MLL-AFF1 fusion genes in Guatemalan pediatric acute lymphoblastic leukemia patients and their ethnic associations. (United States)

    Carranza, Claudia; Granados, Lilian; Morales, Oneida; Jo, Wendy; Villagran, Swuanny; Tinti, Damaris; Villegas, Mauricio; Antillón, Federico; Torselli, Silvana; Silva, Gabriel


    Fusion genes involved in acute lymphoblastic leukemia (ALL) occur mostly due to genetic and environmental factors, and only a limited number of studies have reported any ethnic influence. This study assesses whether an ethnic influence has an effect on the frequency of any of the four fusion genes: BCR-ABL1, ETV6-RUNX1, TCF3-PBX1, and MLL-AFF1 found in ALL. To study this ethnic influence, mononuclear cells were obtained from bone marrow samples from 143 patients with ALL. We performed RNA extraction and reverse transcription, then assessed the quality of the cDNA by amplifying the ABL1 control gene, and finally evaluated the presence of the four transcripts by multiplex polymerase chain reaction. We found 10 patients who had the BCR-ABL1 fusion gene (7%); 3 patients (2%) were TCF3-PBX1 positive; and 6 patients (4.5%) were ETV6-RUNX1 positive. The incidence of this last fusion gene is quite low when compared to the values reported in most countries. The low incidence of the ETV6-RUNX1 fusion gene found in Guatemala matches the incidence rates that have been reported in Spain and Indian Romani. Since it is known that an ethnic resemblance exists among these three populations, as shown by ancestral marker studies, the ALL data suggests an ethnic influence on the occurrence and frequency of this particular fusion gene.


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    Iuliu-Cristian Ivanov


    Full Text Available Acute lymphoblastic leukemia represents a heterogeneous group of hematological malignancies, defined by clonal proliferation of lymphoid cells. Immunophenotyping by flow cytometry and molecular analysis for the detection of genetic anomalies are clinical standard procedures for diagnosis, sub-classification and post-therapeutic evaluation. Samples from 105 patients diagnosed with acute lymphoblastic leukemia were immunophenotyped at diagnosis and were investigated by molecular analysis in order to identify the occurrence of four fusion genes: MLL-AF4, TEL-AML-1, BCR-ABL-p190, E2A-PBX-1. There were no associations found between the immunophenotype and the presence of any fusion genes evaluated. Both methods in combination remain a prerequisite for an improved subclassification of hematological malignancies, therapeutic decision, and evaluation of treatment response.

  9. Attomolar electrochemical detection of the BCR/ABL fusion gene based on an amplifying self-signal metal nanoparticle-conducting polymer hybrid composite. (United States)

    Avelino, Karen Y P S; Frias, Isaac A M; Lucena-Silva, Norma; Gomes, Renan G; de Melo, Celso P; Oliveira, Maria D L; Andrade, César A S


    In the last ten years, conjugated polymers started to be used in the immobilization of nucleic acids via non-covalent interactions. In the present study, we describe the construction and use of an electrochemical DNA biosensor based on a nanostructured polyaniline-gold composite, specifically developed for the detection of the BCR/ABL chimeric oncogene. This chromosome translocation is used as a biomarker to confirm the clinical diagnosis of both chronic myelogenous leukemia (CML) and acute lymphocytic leukemia (ALL). The working principle of the biosensor rests on measuring the conductivity resulting from the non-covalent interactions between the hybrid nanocomposite and the DNA probe. The nanostructured platform exhibits a large surface area that enhances the conductivity. Positive cases, which result from the hybridization between DNA probe and targeted gene, induce changes in the amperometric current and in the charge transfer resistance (RCT) responses. Atomic force microscopy (AFM) images showed changes in the genosensor surface after exposure to cDNA sample of patient with leukemia, evidencing the hybridization process. This new hybrid sensing-platform displayed high specificity and selectivity, and its detection limit is estimated to be as low as 69.4 aM. The biosensor showed excellent analytical performance for the detection of the BCR/ABL oncogene in clinical samples of patients with leukemia. Hence, this electrochemical sensor appears as a simple and attractive tool for the molecular diagnosis of the BCR/ABL oncogene even in early-stage cases of leukemia and for the monitoring of minimum levels of residual disease.

  10. Detection of BCR/ABL Fusion Gene by Fluorescence In Situ Hybridization and Its Clinical Application%荧光原位杂交技术检测BCR/ABL融合基因的临床应用

    Institute of Scientific and Technical Information of China (English)

    周瑞莲; 莫耀禧; 蓝梅; 林金盈


    This study was aimed to investigate the clinical value of detecting BCR/ABL fusion gene by fluorescence in situ hybridization (FISH). The conventional cytogenetic test and detection of BCR/ABL fusion gene by FISH for bone marrow of patients with newly diagnosed chronic myeloproliferative disease or myelodysplastic and myeloproliferative disorders, acute lymphocytic leukemia and chronic myelogeneous leukemia (CML) after allogeneic hematopoietic stem cell transplantation were carried out. The results showed that ( 1 ) out of 46 newly diagnosed as chronic myeloproliferative dsease or myelodysplastic and myeloproliferative disorders, 22 cases were diagnosed as CML, the FISH detection showed all positive (100% ), while cytogenetic test showed 86.4% (19/22) positive, in the other 24 patients who were diagnosed as other chronic myeloproliferative disease or myelodysplastic and myeloproliferative disorders, BCR/ABL fusion gene all were be detected as regative 100% by FISH, while the cytogenetic test of bone marrow in 3 cases supported the diagnosis of CML, and the diagnosis of myelodysplastic disorder in 1 case; (2) in 3 out of 7 acute lymphocytic leukemia cases the BCR/ABL fusion gene could not be detected by FISH; (3) the BCR/ABL fusion gene could be detected by FISH in 2 cases of CML received allogeneic hematopoietic stem cell transplantation, with abnormal threshold 6.5% and 1. 2% respectively. It is concluded that the detection of BCR/ABL fusion gene by FISH is sensitive and reliable, which is very important for the diagnosis and differential dignosis of chronic myeloproliferative disorders, myelodysplastic and myeloproliferative disease, as well as definite diagnosis of Ph* acute lymphoblastic leukemia. This method also has an important significance for monitor of minimal residual disease in CML patients received allogeneic hematopoietic stem cell transplantation.%本研究探讨应用荧光原位杂交技术( FISH)检测BCR/ABL融合基因在临床中的应用价

  11. Detection of p190BCR-ABL AND p210BCR-ABL fusion transcripts in patients with chronic myeloid leukemia (CML using qualitative RT-PCR

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    Aya Bonilla, Carlos Alberto


    Full Text Available Introduction: Chronic myelogenous leukemia (CML is characterized by the presence of the Philadelphia chromosome (Ph, resulting from the balanced reciprocal translocation t(9;22(q34;q11. This marker chromosome is found less frequently in patients suffering from acute lymphoblastic leukemia. Objective: To determine the frequency of BCR-ABL gene fusions encoding the p210BCR-ABL y p190 BCR-ABL transcripts in Colombian patients diagnosed with CML in different stages of the disease and/or its treatment. Materials and methods: Cross sectional, descriptive study of thirty one CML patients (aged 15-78. Analysis was carried out through qualitative nested PCR for the isoforms P210 BCR-ABL (b3a2 e b2a2 and P190 BCR-ABL (e1a2, and based on peripheral blood samples. Results: In 29 of the 31 patients (93.6% transcript p210BCR-ABL was detected; b2a2 and b3a2 gene fusions and the coexpression b3a2 y b2a2 were identified in 55.2% (16/29, 34.5% (10/29 and 10.3% (3/29 of the cases, respectively. Conclusion: b2a2 gene fusion was the most frequent in this CML population.

  12. PESV对K562细胞BCR/ABL融合基因及凋亡调控因子Bcl-2、Bad表达的影响%The Effects of PESV on the Expression of BCR/ABL Fusion Gene and Bcl-2, Bad of Apoptosis Regulators on the K562 Cells

    Institute of Scientific and Technical Information of China (English)

    于文俊; 杨文华; 杨向东; 史哲新; 王兴丽; 郝征; 张佳


    Objective: To investigate the PESV of K562 cells BCR / ABL fusion gene and apoptosis regulators bcl-2 and bad expression. Methods: K562 cells were cultured in vitro, by PESV for different times, the apoptosis rate by flow cytometry, fluorescence quantitative RT-PCR detection of BCR / ABL, Bcl-2, Bad mRNA level changes. Results: Compared with the control group, PESV treated K562 cells, apoptosis increased, BCR / ABL fusion gene reduced expression, anti-apoptotic gene Bcl-2 mRNA expression decreased, pro-apoptotic gene Bad mRNA expression. Conclusion: PESV reduced in K562 cells can reduce the BCR / ABL fusion gene, may regulate the expression of Bcl-2 and Bad, inhibit proliferation of K562 cells and promote their apoptosis.%目的:探讨PESV对K562细胞BCR/ABL融合基因及凋亡调控因子bcl-2和bad表达的影响.方法:将体外培养K562细胞,经PESV处理不同时间后,流式细胞术检测细胞凋亡率,荧光定量RT-PCR检测BCR/ABL、Bcl-2、Bad mRNA水平变化.结果:与对照组相比,PESV处理后K562细胞,凋亡率增加,BCR/ABL融合基因表达降低,抗凋亡相关基因Bcl-2 mRNA表达降低,促凋亡基因Bad mRNA表达增加.结论:PESV能降低降低K562细胞BCR/ABL融合基因的表达,可能通过调节Bcl-2和Bad表达,抑制K562细胞增殖,促进其凋亡.

  13. Detection of BCR-ABL Fusion mRNA Using Reverse Transcriptase Loop-mediated Isothermal Amplification

    Energy Technology Data Exchange (ETDEWEB)

    Dugan, L C; Hall, S; Kohlgruber, A; Urbin, S; Torres, C; Wilson, P


    RT-PCR is commonly used for the detection of Bcr-Abl fusion transcripts in patients diagnosed with chronic myelogenous leukemia, CML. Two fusion transcripts predominate in CML, Br-Abl e13a2 and e14a2. They have developed reverse transcriptase isothermal loop-mediated amplification (RT-LAMP) assays to detect these two fusion transcripts along with the normal Bcr transcript.

  14. DNA fusion gene vaccines

    DEFF Research Database (Denmark)

    Holst, Peter Johannes; Bassi, Maria Rosaria; Thomsen, Allan Randrup


    DNA vaccines are versatile and safe, but limited immunogenicity has prevented their use in the clinical setting. Experimentally, immunogenicity may be enhanced by the use of new delivery technologies, by coadministration of cytokines and pathogen-associated molecular patterns, or by fusion...... of antigens into molecular domains that enhance antigen presentation. More specifically, the immunogenicity of DNA vaccines may benefit from increased protein synthesis, increased T-cell help and MHC class I presentation, and the addition of a range of specific cytokines and pathogen-associated molecular...... with viral-vectored vaccines, various synergistic components may need to be incorporated into DNA vaccines. From the perspective of the future clinical use of DNA vaccines, it has been suggested that antigen presentation should be improved and cytokine coadministration attempted. However, even...

  15. 伴不典型BCR-ABL融合基因的慢性髓性白血病的临床和实验研究%A clinical and laboratory study of chronic myeloid leukemia with atypical BCR-ABL fusion gene subtypes

    Institute of Scientific and Technical Information of China (English)

    桂晓敏; 潘金兰; 仇惠英; 岑建农; 薛永权; 陈苏宁; 沈宏杰; 姚利; 张俊


    目的 探讨伴不典型BCR-ABL融合基因亚型el4a3和e19a2型慢性髓性白血病(CML)的临床和实验室特点.方法 对2004至2012年染色体核型分析有t(9;22) (q34;q11),荧光原位杂交(FISH)证实为BCR-ABL融合基因阳性,而常规实时定量PCR(RQ-PCR)检测常见BCR-ABL融合基因(b3a2、b2a2和e1a2)阴性的6例CML患者,重新设计引物进行PCR扩增,并将扩增产物测序,以明确不典型BCR-ABL融合基因类型.对BCR-ABL融合基因扩增产物进行突变检测.对患者的临床资料进行回顾性分析.结果 6例患者PCR扩增产物经测序分析,其中5例为e14a3型,1例为e19a2型.5例e14a3型CML患者中,男4例,女1例,中位年龄48岁,慢性期4例,加速期1例;1例e19a2型患者为女性,40岁,CML慢性期,PLT>1 000× 109/L.5例e14a3型CML患者中4例在羟基脲或IFN治疗无效后予以伊马替尼(IM)治疗,1例行造血干细胞移植(HSCT).前4例患者中1例因有E255K突变而对IM耐药,改用达沙替尼后获完全细胞遗传学反应(CCyR);1例在IM治疗获CCyR后3个月复发并急变,最终死亡;2例IM治疗后获CCyR,目前状态稳定,仍处于CCyR,尽管其中1例伴有I293T突变.行HSCT治疗患者目前处于CCyR.1例e19a2型CML患者羟基脲治疗后获得完全血液学反应,后改用IM治疗,很快获得CCyR.结论 伴不典型BCR-ABL融合基因的CML发病率极低,酪氨酸激酶抑制剂或HSCT都可以取得疗效,常规RQ-PCR可能漏检少见的不典型BCR-ABL融合基因亚型.%Objective To explore the clinical and laboratory features of chronic myeloid leukemia (CML) with atypical e14a3 and e19a2 BCR-ABL fusion gene subtypes.Methods We retrospectively analyzed a cohort of CML patients with Ph chromosome positive confirmed by cytogenetic and FISH but classical el3a3 (b2a2),e14a2 (b3a2)and ela2 fusion transcripts negative identified by conventional realtime quantification RT-PCR (RQ-PCR).Further RQ-PCR was done with the forward primer and reverse primer designed to

  16. Co-expression of the CBFβ-MYH11 and BCR-ABL fusion genes in chronic myeloid leukaemia / Coexistenţa genelor de fuziune CBFβ-MYH11 şi BCR-ABL în leucemia mieloidă cronică

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    Popescu Roxana


    Full Text Available Coexistenţa t(9;22 şi a inv(16 a fost descrisă într-un număr limitat de cazuri de LMC, LAM de novo sau LAM post chimioterapie. Raportăm un pacient cu LMC care a prezentat atât inversie de 16 cât şi cromozom Philadelphia şi care a evoluat spre criză blastică sub tratament cu Imatinib. Diagnosicul de laborator şi monitorizarea s-a realizat prin citometrie în flux, citogenetică convenţională şi tehnici de genetică moleculară. Inv(16, detectată prin cariotipare în clona Philadelphia pozitivă la momentul transformării blastice, a fost evaluată retrospective prin metoda real-time PCR, şi s-a dovedit a fi fost prezentă încă de la diagnostic. Biopsia de măduvă osoasă, efectuată în faza blastică a LMC, a confirmat prezenţa blaştilor aparţinând liniei mieloide, cu indicii de diferenţiere monocitoidă, frecvent asociată cu inv (16. De asemenea, cazul a asociat şi mutaţia F359V în domeniul kinazic al ABL, care determină rezistenţă intermediară la Imatinib şi Nilotinib, ceea ce a impus schimbarea terapiei cu Dasatinib. În cazul prezentat evoluţia a fost progresivă, urmată de deces ca urmare a lipsei de răspuns la inhibitorii de tirozin kinază, la 18 luni de la diagnosticare. Coexistenţa t(9; 22 şi inv(16 în LMC pare a fi asociată cu o evoluţie clinică agresivă şi rezistenţă la terapia cu inhibitori de tirozin kinază. Având în vedere numărul foarte mic de cazuri descrise în literatura de specialitate, deciziile terapeutice în cazul pacienţilor care prezintă aceste anomalii sunt încă dificile

  17. Cloning and expression of catalytic domain of Abl protein tyrosine kinase gene in E. coli

    Institute of Scientific and Technical Information of China (English)


    Protein tyrosine kinases (PTKs) regulate cell proliferation, differentiation and are involved in signal transduction. Uncontrolled signaling from receptor tyrosine kinases to intracellular tyrosine kinases can lead to inflamma tory responses and diseases such as cancer and atherosclerosis. Thus, inhibitors that block the activity of tyrosine kinases or the signaling pathways of PTKs activation could be assumed as the potential candidate for drug development. On this assumption, we cloned and expressed the Abl PTK gene in E. coli, and purified the PTK, which was used to screen the PTK inhibitors from the extracts of Chinese herbs. The catalytic domain sequence of PTK gene was amplified by PCR us ing the cDNA of abl from Abelson murine leukemia virus as template. The amplified fragment was then cloned into the GST-tagged expression vector pGEX2T. The recombinant plasmid was transformed into host cell E. coli DH5α and was induced to express PTK protein. The expression of the protein was detected using SDS-PAGE. The result showed that a specific protein was induced to express after 12 min induction, and reached peak level about 40% of the host total pro tein after 4 h induction. The molecular weight of the fusion protein was about 58 kD. The purified GST-PTK fusion pro tein presented higher activity for tyrosine phosphorylation.

  18. Evaluation of Morpholino Antisense Oligos’ Role on BCR-ABL Gene Silencing in the K562 Cell Line

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    Bahman Delalat


    Full Text Available Objective: Chronic myeloid leukemia (CML develops when a hematopoietic stem cellacquires the BCR/ABL fusion gene. This causes these transformed hematopoietic cellsto have a greater than normal proliferation rate. Scientists attempt to improve the CMLtreatment process by silencing the BCR/ABL oncogene. In this work, we used morpholinoantisense oligos to silence the BCR/ABL oncogene.Materials and Methods: In this study, the K562 was used as a BCR/ABL fusion-genepositive cell line and the Jurkat cell line as a control. We explored the inhibiting capacityof morpholino antisense oligos in the the expression of the BCR/ABL oncogene andstudied their p210 BCR/ABL suppression, inhibition of cell proliferation and stimulation ofapoptosis in the K562 cells after 24 and 48 hours. Endo-Porter was used for delivery ofmorpholino antisense oligos into cell cytosols. Meanwhile, flow cytometric analysis wasperformed in order to determine the appropriate concentration of morpholino antisenseoligos.Results: Prolonged exposure of the K562 cell line to the morpholino antisense oligostargeted against the BCR-ABL gene showed proliferation inhibition as its main feature.After western blotting, we found that complete silencing of BCR/ABL was achieved, butflow cytometric analysis showed no broad apoptosis.Conclusion: The results indicate that the Morpholino antisense oligo is able to inhibitp210 BCR/ABL; however, it cannot induce broad apoptosis due to co-silencing of BCR.

  19. BCR and its mutants, the reciprocal t(9;22-associated ABL/BCR fusion proteins, differentially regulate the cytoskeleton and cell motility

    Directory of Open Access Journals (Sweden)

    Puccetti Elena


    Full Text Available Abstract Background The reciprocal (9;22 translocation fuses the bcr (breakpoint cluster region gene on chromosome 22 to the abl (Abelson-leukemia-virus gene on chromosome 9. Depending on the breakpoint on chromosome 22 (the Philadelphia chromosome – Ph+ the derivative 9+ encodes either the p40(ABL/BCR fusion transcript, detectable in about 65% patients suffering from chronic myeloid leukemia, or the p96(ABL/BCR fusion transcript, detectable in 100% of Ph+ acute lymphatic leukemia patients. The ABL/BCRs are N-terminally truncated BCR mutants. The fact that BCR contains Rho-GEF and Rac-GAP functions strongly suggest an important role in cytoskeleton modeling by regulating the activity of Rho-like GTPases, such as Rho, Rac and cdc42. We, therefore, compared the function of the ABL/BCR proteins with that of wild-type BCR. Methods We investigated the effects of BCR and ABL/BCRs i. on the activation status of Rho, Rac and cdc42 in GTPase-activation assays; ii. on the actin cytoskeleton by direct immunofluorescence; and iii on cell motility by studying migration into a three-dimensional stroma spheroid model, adhesion on an endothelial cell layer under shear stress in a flow chamber model, and chemotaxis and endothelial transmigration in a transwell model with an SDF-1α gradient. Results Here we show that both ABL/BCRs lost fundamental functional features of BCR regarding the regulation of small Rho-like GTPases with negative consequences on cell motility, in particular on the capacity to adhere to endothelial cells. Conclusion Our data presented here describe for the first time an analysis of the biological function of the reciprocal t(9;22 ABL/BCR fusion proteins in comparison to their physiological counterpart BCR.

  20. 荧光原位杂交在慢性粒细胞白血病细胞BCR/ABL融合基因检测中的应用及其意义%The detection of BCR/ABL fusion gene by interphase fluorescence in situ hybridization in patients with chronic myeloid leukemia and its clinical significance

    Institute of Scientific and Technical Information of China (English)

    张济; 李君君; 颜家运; 邹礼衡; 刘静


    目的:探讨荧光原位杂交(FISH)技术检测慢性粒细胞白血病(CML)骨髓和外周血细胞的BCR /ABL融合基因的临床价值.方法:应用FISH 对正常对照组及CML患者骨髓和外周血细胞BCR/ ABL融合基因进行检测和分析.结果:慢性期CML 患者骨髓和外周血细胞该融合基因阳性细胞率分别为(61.9 ± 22.3)%和(68.4 ± 19.8)%,加速期为(77.2 ± 16.7)% 和(86.8 ± 12.1)%,急变期为(80.6 ± 17.5)%和(81.4 ± 18.0)%,两者细胞中BCR/ABL 基因的阳性率未见统计学差异(P>0.05),且骨髓和外周血细胞之间融合基因阳性细胞比率呈直线正相关.同时发现在完全临床缓解患者中,经伊马替尼治疗的CML 患者(71.4%)较经干扰素和羟基脲联合治疗者(10.0%)有更高的分子生物学缓解(P 0.05). A significant correlation was found between the BCR/ABL positive cells in peripheral blood and in bone marrow samples. Moreover, the higher rate of complete molecular remission was achieved by treating with imatinib than with interferon and hydroxyurea during complete remission phase (P < 0.05). Conclusions Detecting BCR/ABL fusion gene in peripheral blood and bone marrow samples by FISH are beneficial to diagnosis, treatment and minimal residual disease monitor in CML.

  1. Assessment of fusion gene status in sarcomas using a custom made fusion gene microarray.

    Directory of Open Access Journals (Sweden)

    Marthe Løvf

    Full Text Available Sarcomas are relatively rare malignancies and include a large number of histological subgroups. Based on morphology alone, the differential diagnoses of sarcoma subtypes can be challenging, but the identification of specific fusion genes aids correct diagnostication. The presence of individual fusion products are routinely investigated in Pathology labs. However, the methods used are time-consuming and based on prior knowledge about the expected fusion gene and often the most likely break-point. In this study, 16 sarcoma samples, representing seven different sarcoma subtypes with known fusion gene status from a diagnostic setting, were investigated using a fusion gene microarray. The microarray was designed to detect all possible exon-exon breakpoints between all known fusion genes in a single analysis. An automated scoring of the microarray data from the 38 known sarcoma-related fusion genes identified the correct fusion gene among the top-three hits in 11 of the samples. The analytical sensitivity may be further optimised, but we conclude that a sarcoma-fusion gene microarray is suitable as a time-saving screening tool to identify the majority of the correct fusion genes.

  2. ETS fusion genes in prostate cancer. (United States)

    Gasi Tandefelt, Delila; Boormans, Joost; Hermans, Karin; Trapman, Jan


    Prostate cancer is very common in elderly men in developed countries. Unravelling the molecular and biological processes that contribute to tumor development and progressive growth, including its heterogeneity, is a challenging task. The fusion of the genes ERG and TMPRSS2 is the most frequent genomic alteration in prostate cancer. ERG is an oncogene that encodes a member of the family of ETS transcription factors. At lower frequency, other members of this gene family are also rearranged and overexpressed in prostate cancer. TMPRSS2 is an androgen-regulated gene that is preferentially expressed in the prostate. Most of the less frequent ETS fusion partners are also androgen-regulated and prostate-specific. During the last few years, novel concepts of the process of gene fusion have emerged, and initial experimental results explaining the function of the ETS genes ERG and ETV1 in prostate cancer have been published. In this review, we focus on the most relevant ETS gene fusions and summarize the current knowledge of the role of ETS transcription factors in prostate cancer. Finally, we discuss the clinical relevance of TMRPSS2-ERG and other ETS gene fusions in prostate cancer.

  3. An Efficient Method for Identifying Gene Fusions by Targeted RNA Sequencing from Fresh Frozen and FFPE Samples.

    Directory of Open Access Journals (Sweden)

    Jonathan A Scolnick

    Full Text Available Fusion genes are known to be key drivers of tumor growth in several types of cancer. Traditionally, detecting fusion genes has been a difficult task based on fluorescent in situ hybridization to detect chromosomal abnormalities. More recently, RNA sequencing has enabled an increased pace of fusion gene identification. However, RNA-Seq is inefficient for the identification of fusion genes due to the high number of sequencing reads needed to detect the small number of fusion transcripts present in cells of interest. Here we describe a method, Single Primer Enrichment Technology (SPET, for targeted RNA sequencing that is customizable to any target genes, is simple to use, and efficiently detects gene fusions. Using SPET to target 5701 exons of 401 known cancer fusion genes for sequencing, we were able to identify known and previously unreported gene fusions from both fresh-frozen and formalin-fixed paraffin-embedded (FFPE tissue RNA in both normal tissue and cancer cells.

  4. An Efficient Method for Identifying Gene Fusions by Targeted RNA Sequencing from Fresh Frozen and FFPE Samples. (United States)

    Scolnick, Jonathan A; Dimon, Michelle; Wang, I-Ching; Huelga, Stephanie C; Amorese, Douglas A


    Fusion genes are known to be key drivers of tumor growth in several types of cancer. Traditionally, detecting fusion genes has been a difficult task based on fluorescent in situ hybridization to detect chromosomal abnormalities. More recently, RNA sequencing has enabled an increased pace of fusion gene identification. However, RNA-Seq is inefficient for the identification of fusion genes due to the high number of sequencing reads needed to detect the small number of fusion transcripts present in cells of interest. Here we describe a method, Single Primer Enrichment Technology (SPET), for targeted RNA sequencing that is customizable to any target genes, is simple to use, and efficiently detects gene fusions. Using SPET to target 5701 exons of 401 known cancer fusion genes for sequencing, we were able to identify known and previously unreported gene fusions from both fresh-frozen and formalin-fixed paraffin-embedded (FFPE) tissue RNA in both normal tissue and cancer cells.

  5. Molecular pathways: targeting ETS gene fusions in cancer. (United States)

    Feng, Felix Y; Brenner, J Chad; Hussain, Maha; Chinnaiyan, Arul M


    Rearrangements, or gene fusions, involving the ETS family of transcription factors are common driving events in both prostate cancer and Ewing sarcoma. These rearrangements result in pathogenic expression of the ETS genes and trigger activation of transcriptional programs enriched for invasion and other oncogenic features. Although ETS gene fusions represent intriguing therapeutic targets, transcription factors, such as those comprising the ETS family, have been notoriously difficult to target. Recently, preclinical studies have demonstrated an association between ETS gene fusions and components of the DNA damage response pathway, such as PARP1, the catalytic subunit of DNA protein kinase (DNAPK), and histone deactylase 1 (HDAC1), and have suggested that ETS fusions may confer sensitivity to inhibitors of these DNA repair proteins. In this review, we discuss the role of ETS fusions in cancer, the preclinical rationale for targeting ETS fusions with inhibitors of PARP1, DNAPK, and HDAC1, as well as ongoing clinical trials targeting ETS gene fusions.

  6. What we miss in order to be able to design and build a commercially viable fusion reactor

    Energy Technology Data Exchange (ETDEWEB)

    Andreani, R. [ENEA, Centro Ricerche Frascati, RM (Italy). Dipt. Energia


    The paper considers in a critical way the different areas in which work is required to provide sufficient information in view of designing a reliable and attractive fusion reactor. [Italian] Il rapporto considera in modo critico le differenti aree nelle quali si richiede ulteriore lavoro per fornire informazioni al fine di progettare un reattore a fusione affidabile ed economicamente competitivo.

  7. Emergence of FGFR family gene fusions as therapeutic targets in a wide spectrum of solid tumours. (United States)

    Parker, Brittany C; Engels, Manon; Annala, Matti; Zhang, Wei


    The emergence of fibroblast growth factor receptor (FGFR) family fusions across diverse cancers has brought attention to FGFR-derived cancer therapies. The discovery of the first recurrent FGFR fusion in glioblastoma was followed by discoveries of FGFR fusions in bladder, lung, breast, thyroid, oral, and prostate cancers. Drug targeting of FGFR fusions has shown promising results and should soon be translating into clinical trials. FGFR fusions form as a result of various mechanisms – predominantly deletion for FGFR1, translocation for FGFR2, and tandem duplication for FGFR3. The ability to exploit the unique targetability of FGFR fusions proves that FGFR-derived therapies could have a promising future in cancer therapeutics. Drug targeting of fusion genes has proven to be an extremely effective therapeutic approach for cancers such as the recurrent BCR–ABL1 fusion in chronic myeloid leukaemia. The recent discovery of recurrent FGFR family fusions in several cancer types has brought to attention the unique therapeutic potential for FGFR-positive patients. Understanding the diverse mechanisms of FGFR fusion formation and their oncogenic potential will shed light on the impact of FGFR-derived therapy in the future.

  8. Identification of targetable FGFR gene fusions in diverse cancers. (United States)

    Wu, Yi-Mi; Su, Fengyun; Kalyana-Sundaram, Shanker; Khazanov, Nickolay; Ateeq, Bushra; Cao, Xuhong; Lonigro, Robert J; Vats, Pankaj; Wang, Rui; Lin, Su-Fang; Cheng, Ann-Joy; Kunju, Lakshmi P; Siddiqui, Javed; Tomlins, Scott A; Wyngaard, Peter; Sadis, Seth; Roychowdhury, Sameek; Hussain, Maha H; Feng, Felix Y; Zalupski, Mark M; Talpaz, Moshe; Pienta, Kenneth J; Rhodes, Daniel R; Robinson, Dan R; Chinnaiyan, Arul M


    Through a prospective clinical sequencing program for advanced cancers, four index cases were identified which harbor gene rearrangements of FGFR2, including patients with cholangiocarcinoma, breast cancer, and prostate cancer. After extending our assessment of FGFR rearrangements across multiple tumor cohorts, we identified additional FGFR fusions with intact kinase domains in lung squamous cell cancer, bladder cancer, thyroid cancer, oral cancer, glioblastoma, and head and neck squamous cell cancer. All FGFR fusion partners tested exhibit oligomerization capability, suggesting a shared mode of kinase activation. Overexpression of FGFR fusion proteins induced cell proliferation. Two bladder cancer cell lines that harbor FGFR3 fusion proteins exhibited enhanced susceptibility to pharmacologic inhibition in vitro and in vivo. Because of the combinatorial possibilities of FGFR family fusion to a variety of oligomerization partners, clinical sequencing efforts, which incorporate transcriptome analysis for gene fusions, are poised to identify rare, targetable FGFR fusions across diverse cancer types.

  9. What we miss in order to be able to design and build a commercially viable fusion reactor

    Energy Technology Data Exchange (ETDEWEB)

    Andreani, R. [ENEA, Centro Ricerche Frascati, Rome (Italy). Dipt. Energia


    The paper considers in a critical way the different areas in which work is required to provide sufficient information in view of designing a reliable and attractive fusion reactor. Four main areas of activity are considered: physics, technology, engineering, safety. In physics the trend is positive towards a better understanding of suitable plasma regimes to be confirmed through further experimentation on the operating machines. Engineering has already proven itself by the design and construction of a number of experimental machines. In addition a large data base obtained from design and operation of fission reactors is available. Safety is reaching very satisfactory results in the analysis of the impact of fusion on man and the environment. Where it is still a large unsolved problem is concerning materials capable of standing the harsh fusion environment for an adequate number of years. An intense neutron source is needed in order to allow the necessary developments. [Italian] Il rapporto considera in modo critico le differenti aree nelle quali si richiede ulteriore lavoro per fornire informazini sufficienti al fine di progettare un reattore a fusione affidabile ed economicamente competitivo. Vengono considerate quattro aree principali di attivita': fisica, tecnologia, ingegneria, sicurezza. Nella fisica, vi e' una positiva tendenza verso una migliore comprensione di regimi di plasma favorevoli da confermare attraverso ulteriore sperimentazione sulle macchine funzionanti. L' ingegneria ha gia' dato dimostrazione di se' col progetto e la costruzione di un notevole numero di macchine sperimentali. In aggiunta e' disponibile un gran numero di dati ottenuti dalla progettazione, realizzazione e funzionamento dei reattori a fissione. La sicurezza sta raggiungendo risultati molto soddisfacenti nell'analisi dell'impatto della fusione sull'uomo e sull'ambiente. Un grosso problema tuttora irresoluto e' quello dei

  10. Detection of a rare BCR-ABL tyrosine kinase fusion protein in H929 multiple myeloma cells using immunoprecipitation (IP)-tandem mass spectrometry (MS/MS). (United States)

    Breitkopf, Susanne B; Yuan, Min; Pihan, German A; Asara, John M


    Hypothesis directed proteomics offers higher throughput over global analyses. We show that immunoprecipitation (IP)-tandem mass spectrometry (LC-MS/MS) in H929 multiple myeloma (MM) cancer cells led to the discovery of a rare and unexpected BCR-ABL fusion, informing a therapeutic intervention using imatinib (Gleevec). BCR-ABL is the driving mutation in chronic myeloid leukemia (CML) and is uncommon to other cancers. Three different IP-MS experiments central to cell signaling pathways were sufficient to discover a BCR-ABL fusion in H929 cells: phosphotyrosine (pY) peptide IP, p85 regulatory subunit of phosphoinositide-3-kinase (PI3K) IP, and the GRB2 adaptor IP. The pY peptides inform tyrosine kinase activity, p85 IP informs the activating adaptors and receptor tyrosine kinases (RTKs) involved in AKT activation and GRB2 IP identifies RTKs and adaptors leading to ERK activation. Integration of the bait-prey data from the three separate experiments identified the BCR-ABL protein complex, which was confirmed by biochemistry, cytogenetic methods, and DNA sequencing revealed the e14a2 fusion transcript. The tyrosine phosphatase SHP2 and the GAB2 adaptor protein, important for MAPK signaling, were common to all three IP-MS experiments. The comparative treatment of tyrosine kinase inhibitor (TKI) drugs revealed only imatinib, the standard of care in CML, was inhibitory to BCR-ABL leading to down-regulation of pERK and pS6K and inhibiting cell proliferation. These data suggest a model for directed proteomics from patient tumor samples for selecting the appropriate TKI drug(s) based on IP and LC-MS/MS. The data also suggest that MM patients, in addition to CML patients, may benefit from BCR-ABL diagnostic screening.

  11. FGFR-TACC gene fusions in human glioma. (United States)

    Lasorella, Anna; Sanson, Marc; Iavarone, Antonio


    Chromosomal translocations joining in-frame members of the fibroblast growth factor receptor-transforming acidic coiled-coil gene families (the FGFR-TACC gene fusions) were first discovered in human glioblastoma multiforme (GBM) and later in many other cancer types. Here, we review this rapidly expanding field of research and discuss the unique biological and clinical features conferred to isocitrate dehydrogenase wild-type glioma cells by FGFR-TACC fusions. FGFR-TACC fusions generate powerful oncogenes that combine growth-promoting effects with aneuploidy through the activation of as yet unclear intracellular signaling mechanisms. FGFR-TACC fusions appear to be clonal tumor-initiating events that confer strong sensitivity to FGFR tyrosine kinase inhibitors. Screening assays have recently been reported for the accurate identification of FGFR-TACC fusion variants in human cancer, and early clinical data have shown promising effects in cancer patients harboring FGFR-TACC fusions and treated with FGFR inhibitors. Thus, FGFR-TACC gene fusions provide a "low-hanging fruit" model for the validation of precision medicine paradigms in human GBM.

  12. [The progress of TMPRSS2-ETS gene fusions and their mechanism in prostate cancer]. (United States)

    Guo, Xiao-Qiang; Gui, Yao-Ting; Cai, Zhi-Ming


    The gene fusions between transmembrane protease serine 2 (TMPRSS2) and E26 (ETS) transcription factors are present in over 50% of patients with prostate cancer. TMPRSS2-ERG is the most common gene fusion type. The ERG overexpression induced by TMPRSS2-ERG gene fusion contributes to the development of prostate cancer. Both androgen receptor binding and genotoxic stress induce chromosomal proximity and TMPRSS2-ETS gene fusions. TMPRSS2-ERG gene fusion functions as a biomarker for prostate cancer, which can be easily detected in urine. This review focuses on the characteristics, oncogenic and rearranged mechanism, and clinical application of TMPRSS2-ETS gene fusions.

  13. ABI-like transcription factor gene TaABL1 from wheat improves multiple abiotic stress tolerances in transgenic plants. (United States)

    Xu, Dong-Bei; Gao, Shi-Qing; Ma, You-Zhi; Xu, Zhao-Shi; Zhao, Chang-Ping; Tang, Yi-Miao; Li, Xue-Yin; Li, Lian-Cheng; Chen, Yao-Feng; Chen, Ming


    The phytohormone abscisic acid (ABA) plays crucial roles in adaptive responses of plants to abiotic stresses. ABA-responsive element binding proteins (AREBs) are basic leucine zipper transcription factors that regulate the expression of downstream genes containing ABA-responsive elements (ABREs) in promoter regions. A novel ABI-like (ABA-insensitive) transcription factor gene, named TaABL1, containing a conserved basic leucine zipper (bZIP) domain was cloned from wheat. Southern blotting showed that three copies were present in the wheat genome. Phylogenetic analyses indicated that TaABL1 belonged to the AREB subfamily of the bZIP transcription factor family and was most closely related to ZmABI5 in maize and OsAREB2 in rice. Expression of TaABL1 was highly induced in wheat roots, stems, and leaves by ABA, drought, high salt, and low temperature stresses. TaABL1 was localized inside the nuclei of transformed wheat mesophyll protoplast. Overexpression of TaABL1 enhanced responses of transgenic plants to ABA and hastened stomatal closure under stress, thereby improving tolerance to multiple abiotic stresses. Furthermore, overexpression of TaABL1 upregulated or downregulated the expression of some stress-related genes controlling stomatal closure in transgenic plants under ABA and drought stress conditions, suggesting that TaABL1 might be a valuable genetic resource for transgenic molecular breeding.

  14. Two novel imatinib-responsive PDGFRA fusion genes in chronic eosinophilic leukaemia. (United States)

    Curtis, Claire E; Grand, Francis H; Musto, Pellegrino; Clark, Andrew; Murphy, John; Perla, Gianni; Minervini, Maria M; Stewart, Janet; Reiter, Andreas; Cross, Nicholas C P


    We identified two patients with a t(2;4)(p24;q12) and a t(4;12)(q2?3;p1?2), respectively, in association with BCR-ABL and FIP1L1-PDGFRA negative chronic eosinophilic leukaemia. Molecular analysis revealed a novel STRN-PDGFRA fusion for the t(2;4) and ETV6-PDGFRA for the t(4;12). The fusions were confirmed by specific amplification of the genomic breakpoints, reverse transcription polymerase chain reaction and fluorescence in situ hybridisation. Both patients were treated with imatinib and, following a rapid haematological response, achieved cytogenetic remission and a major molecular response. In conclusion, PDGFRA fuses to diverse partner genes in myeloid disorders. Identification of these fusions is important as they are particularly sensitive to imatinib.

  15. Immunologic evaluation of peptides derived from BCR/ABL-out-of-frame fusion protein in HLA A2.1 transgenic mice. (United States)

    Casnici, Claudia; Volpe, Gisella; Crotta, Katia; Lattuada, Donatella; Saglio, Giuseppe; Marelli, Ornella


    Philadelphia chromosome-positive chronic myelogenous leukemia and acute lymphocytic leukemia express, besides the main BCR/ABL transcripts, novel BCR/ABL transcripts derived from alternative splicing between BCR exons 1, 13, or 14 with ABL exons 4 and 5. Their translational products present at C-terminus an amino acid portion derived from out-of-frame (OOF) reading of the ABL gene. The presence of OOF-peptide-specific T cells in chronic myelogenous leukemia patients was demonstrated and a first study in in vivo model demonstrated that OOF ABL portion was immunogenic in human leukcocyte antigen (HLA)-A2.1 transgenic mice. Here we immunized HLA A2.1 mice with novel peptides designed on the ABL OOF sequence, containing epitopes with high affinity for HLA A2.1 molecule. The specific immune response, cellular and humoral, obtained ex vivo against HLA A2.1-positive human chronic myelogenous leukemia cells using peptide 22-53 and the cytotoxic activity induced by peptide 32mer confirm the possibility to use the ABL OOF portion as target to evoke a specific and multiple immune response in Philadelphia positive leukemic patients in cytogenetic remission.

  16. Multimodality imaging of reporter gene expression using a novel fusion vector in living cells and animals (United States)

    Gambhir; Sanjiv , Pritha; Ray


    Novel double and triple fusion reporter gene constructs harboring distinct imageable reporter genes are provided, as well as applications for the use of such double and triple fusion constructs in living cells and in living animals using distinct imaging technologies.

  17. Genome-Wide Overexpression Screen Identifies Genes Able to Bypass p16-Mediated Senescence in Melanoma. (United States)

    Lee, Won Jae; Škalamera, Dubravka; Dahmer-Heath, Mareike; Shakhbazov, Konstanin; Ranall, Max V; Fox, Carly; Lambie, Duncan; Stevenson, Alexander J; Yaswen, Paul; Gonda, Thomas J; Gabrielli, Brian


    Malignant melanomas often arise from nevi, which result from initial oncogene-induced hyperproliferation of melanocytes that are maintained in a CDKN2A/p16-mediated senescent state. Thus, genes that can bypass this senescence barrier are likely to contribute to melanoma development. We have performed a gain-of-function screen of 17,030 lentivirally expressed human open reading frames (ORFs) in a melanoma cell line containing an inducible p16 construct to identify such genes. Genes known to bypass p16-induced senescence arrest, including the human papilloma virus 18 E7 gene ( HPV18E7), and genes such as the p16-binding CDK6 with expected functions, as well as panel of novel genes, were identified, including high-mobility group box (HMGB) proteins. A number of these were further validated in two other models of p16-induced senescence. Tissue immunohistochemistry demonstrated higher levels of CDK6 in primary melanomas compared with normal skin and nevi. Reduction of CDK6 levels drove melanoma cells expressing functional p16 into senescence, demonstrating its contribution to bypass senescence.

  18. Gene Prioritization by Compressive Data Fusion and Chaining.

    Directory of Open Access Journals (Sweden)

    Marinka Žitnik


    Full Text Available Data integration procedures combine heterogeneous data sets into predictive models, but they are limited to data explicitly related to the target object type, such as genes. Collage is a new data fusion approach to gene prioritization. It considers data sets of various association levels with the prediction task, utilizes collective matrix factorization to compress the data, and chaining to relate different object types contained in a data compendium. Collage prioritizes genes based on their similarity to several seed genes. We tested Collage by prioritizing bacterial response genes in Dictyostelium as a novel model system for prokaryote-eukaryote interactions. Using 4 seed genes and 14 data sets, only one of which was directly related to the bacterial response, Collage proposed 8 candidate genes that were readily validated as necessary for the response of Dictyostelium to Gram-negative bacteria. These findings establish Collage as a method for inferring biological knowledge from the integration of heterogeneous and coarsely related data sets.

  19. Analysis of API2-MALT1 fusion, trisomies, and immunoglobulin VH genes in pulmonary mucosa-associated lymphoid tissue lymphoma. (United States)

    Xia, Hongjing; Nakayama, Takahisa; Sakuma, Hidenori; Yamada, Seiji; Sato, Fumihiko; Takino, Hisashi; Okabe, Mitsukuni; Fujiyoshi, Yukio; Hattori, Hideo; Inagaki, Hiroshi


    Pulmonary mucosa-associated lymphoid tissue lymphoma is unique in that chronic inflammation is rare and that API2-MALT1 fusion, resulting from t(11;18)(q21;q21), occurs frequently. In this study, we examined 20 cases for API2-MALT1 fusion using the multiplex reverse-transcription polymerase chain reaction and looked for trisomy 3, trisomy 18, and abnormalities of MALT1 and IGH genes using fluorescence in situ hybridization. In addition, we analyzed VH genes by subcloning of the monoclonal polymerase chain reaction products. Of 20 cases studied, we detected gene abnormalities in 16: API2-MALT1 fusion in 9, trisomy 3 in 5, trisomy 18 in 4, MALT1 abnormality in 13, and IGH abnormality in 1. MALT1 gene abnormalities were concordant with API2-MALT1 fusion or trisomy 18. One case showed API2-MALT1 fusion and trisomy 3. On detection of API2-MALT1 fusion and trisomies, we were able to divide our cases into 3 groups, API2-MALT1 positive, trisomy positive, and no detectable gene abnormality, suggesting that tumor development had processed along different genetic pathways. All 20 cases were analyzed for VH genes. Most of the VH genes selected by the lymphomas belonged to the VH3 family, but there was no restriction to any particular VH fragment. Of interest, VH genes were unmutated in 7 cases, suggesting that T-cell-independent extrafollicular B-cell maturation may be important in the development of this lymphoma. In addition, both mutated and unmutated tumor cases were found to carry the API2-MALT1 fusion and trisomy 3. This observation suggests that these gene abnormalities may occur in microenvironments found before or outside of follicular germinal centers.

  20. ETS Gene Fusions as Predictive Biomarkers of Resistance to Radiation Therapy for Prostate Cancer (United States)


    Award Number: W81XWH-10-1-0582 TITLE: ETS Gene Fusions as Predictive Biomarkers of Resistance to Radiation Therapy for Prostate Cancer PRINCIPAL...ETS gene fusion status associated with clinical outcomes following radiation therapy , by analyzing both the collected biomarker and clinical data...denotes absence of an ERG fusion). ETS gene fusions status did not predict outcomes following radiation therapy , as demonstrated by Kaplan Meier

  1. Rooting the eukaryote tree by using a derived gene fusion. (United States)

    Stechmann, Alexandra; Cavalier-Smith, Thomas


    Single-gene trees have failed to locate the root of the eukaryote tree because of systematic biases in sequence evolution. Structural genetic data should yield more reliable insights into deep phylogenetic relationships. We searched major protist groups for the presence or absence of a gene fusion in order to locate the root of the eukaryote tree. In striking contrast to previous molecular studies, we show that all eukaryote groups ancestrally with two cilia (bikonts) are evolutionarily derived. The root lies between bikonts and opisthokonts (animals, Fungi, Choanozoa). Amoebozoa either diverged even earlier or are sister of bikonts or (less likely) opisthokonts.

  2. Application of next generation sequencing to human gene fusion detection: computational tools, features and perspectives. (United States)

    Wang, Qingguo; Xia, Junfeng; Jia, Peilin; Pao, William; Zhao, Zhongming


    Gene fusions are important genomic events in human cancer because their fusion gene products can drive the development of cancer and thus are potential prognostic tools or therapeutic targets in anti-cancer treatment. Major advancements have been made in computational approaches for fusion gene discovery over the past 3 years due to improvements and widespread applications of high-throughput next generation sequencing (NGS) technologies. To identify fusions from NGS data, existing methods typically leverage the strengths of both sequencing technologies and computational strategies. In this article, we review the NGS and computational features of existing methods for fusion gene detection and suggest directions for future development.

  3. Repeated evolution of chimeric fusion genes in the β-globin gene family of laurasiatherian mammals. (United States)

    Gaudry, Michael J; Storz, Jay F; Butts, Gary Tyler; Campbell, Kevin L; Hoffmann, Federico G


    The evolutionary fate of chimeric fusion genes may be strongly influenced by their recombinational mode of origin and the nature of functional divergence between the parental genes. In the β-globin gene family of placental mammals, the two postnatally expressed δ- and β-globin genes (HBD and HBB, respectively) have a propensity for recombinational exchange via gene conversion and unequal crossing-over. In the latter case, there are good reasons to expect differences in retention rates for the reciprocal HBB/HBD and HBD/HBB fusion genes due to thalassemia pathologies associated with the HBD/HBB "Lepore" deletion mutant in humans. Here, we report a comparative genomic analysis of the mammalian β-globin gene cluster, which revealed that chimeric HBB/HBD fusion genes originated independently in four separate lineages of laurasiatherian mammals: Eulipotyphlans (shrews, moles, and hedgehogs), carnivores, microchiropteran bats, and cetaceans. In cases where an independently derived "anti-Lepore" duplication mutant has become fixed, the parental HBD and/or HBB genes have typically been inactivated or deleted, so that the newly created HBB/HBD fusion gene is primarily responsible for synthesizing the β-type subunits of adult and fetal hemoglobin (Hb). Contrary to conventional wisdom that the HBD gene is a vestigial relict that is typically inactivated or expressed at negligible levels, we show that HBD-like genes often encode a substantial fraction (20-100%) of β-chain Hbs in laurasiatherian taxa. Our results indicate that the ascendancy or resuscitation of genes with HBD-like coding sequence requires the secondary acquisition of HBB-like promoter sequence via unequal crossing-over or interparalog gene conversion.

  4. Establishment and characterization of A novel Philadelphia-chromosome positive chronic myeloid leukemia cell line, TCC-S, expressing P210 and P190 BCR/ABL transcripts but missing normal ABL gene. (United States)

    Van, Phan Nguyen Thanh; Xinh, Phan Thi; Kano, Yasuhiko; Tokunaga, Katsushi; Sato, Yuko


    A novel Philadelphia-chromosome positive (Ph+) cell line, TCC-S, has been established from a patient with Ph+ chronic myeloid leukemia (CML) in the blastic crisis. TCC-S cells were shown to express both P210 and P190 BCR/ABL transcripts by reverse transcriptase-polymerase chain reaction (PCR), although quantitative-PCR revealed that TCC-S cells mainly expressed P210 BCR/ABL transcript. Karyotype analysis revealed several triploid clones which constantly harbored two der(9)del(9) (p12)t(9;22) (q34;qll)s and two del(9) (q21)s. The der(9)del(9) (p12)t(9;22) (q34;q11) is rarely found in other CML cell lines. Moreover, to the best of our knowledge, del(9) (q21) resulting in missing of a restrict region including normal ABL gene has not been found among CML cell lines previously described. Thus, TCC-S cells with only BCR/ABL gene and no normal ABL gene may be a useful tool for functional study of ABL in Ph+ CML.

  5. Transforming fusions of FGFR and TACC genes in human glioblastoma. (United States)

    Singh, Devendra; Chan, Joseph Minhow; Zoppoli, Pietro; Niola, Francesco; Sullivan, Ryan; Castano, Angelica; Liu, Eric Minwei; Reichel, Jonathan; Porrati, Paola; Pellegatta, Serena; Qiu, Kunlong; Gao, Zhibo; Ceccarelli, Michele; Riccardi, Riccardo; Brat, Daniel J; Guha, Abhijit; Aldape, Ken; Golfinos, John G; Zagzag, David; Mikkelsen, Tom; Finocchiaro, Gaetano; Lasorella, Anna; Rabadan, Raul; Iavarone, Antonio


    The brain tumor glioblastoma multiforme (GBM) is among the most lethal forms of human cancer. Here, we report that a small subset of GBMs (3.1%; 3 of 97 tumors examined) harbors oncogenic chromosomal translocations that fuse in-frame the tyrosine kinase coding domains of fibroblast growth factor receptor (FGFR) genes (FGFR1 or FGFR3) to the transforming acidic coiled-coil (TACC) coding domains of TACC1 or TACC3, respectively. The FGFR-TACC fusion protein displays oncogenic activity when introduced into astrocytes or stereotactically transduced in the mouse brain. The fusion protein, which localizes to mitotic spindle poles, has constitutive kinase activity and induces mitotic and chromosomal segregation defects and triggers aneuploidy. Inhibition of FGFR kinase corrects the aneuploidy, and oral administration of an FGFR inhibitor prolongs survival of mice harboring intracranial FGFR3-TACC3-initiated glioma. FGFR-TACC fusions could potentially identify a subset of GBM patients who would benefit from targeted FGFR kinase inhibition.

  6. Fusion genes in solid tumors:an emerging target for cancer diagnosis and treatment

    Institute of Scientific and Technical Information of China (English)

    Brittany C. Parker; Wei Zhang


    Studies over the past decades have uncovered fusion genes, a class of oncogenes that provide immense diagnostic and therapeutic advantages because of their tumor-specific expression. Originally associated with hemotologic cancers, fusion genes have recently been discovered in a wide array of solid tumors, including sarcomas, carcinomas, and tumors of the central nervous system. Fusion genes are attractive as both therapeutic targets and diagnostic tools due to their inherent expression in tumor tissue alone. Therefore, the discovery and elucidation of fusion genes in various cancer types may provide more effective therapies in the future for cancer patients.

  7. Fusion

    CERN Document Server

    Mahaffey, James A


    As energy problems of the world grow, work toward fusion power continues at a greater pace than ever before. The topic of fusion is one that is often met with the most recognition and interest in the nuclear power arena. Written in clear and jargon-free prose, Fusion explores the big bang of creation to the blackout death of worn-out stars. A brief history of fusion research, beginning with the first tentative theories in the early 20th century, is also discussed, as well as the race for fusion power. This brand-new, full-color resource examines the various programs currently being funded or p

  8. Cadmium-regulated gene fusions in Pseudomonas fluorescens. (United States)

    Rossbach, S; Kukuk, M L; Wilson, T L; Feng, S F; Pearson, M M; Fisher, M A


    To study the mechanisms soil bacteria use to cope with elevated concentrations of heavy metals in the environment, a mutagenesis with the lacZ-based reporter gene transposon Tn5B20 was performed. Random gene fusions in the genome of the common soil bacterium Pseudomonas fluorescens strain ATCC 13525 were used to create a bank of 5,000 P. fluorescens mutants. This mutant bank was screened for differential gene expression in the presence of the toxic metal cadmium. Fourteen mutants were identified that responded with increased or reduced gene expression to the presence of cadmium. The mutants were characterized with respect to their metal-dependent gene expression and their metal tolerance. Half the identified mutants reacted with differential gene expression specifically to the metal cadmium, whereas some of the other mutants also responded to elevated concentrations of copper and zinc ions. One of the mutants, strain C8, also showed increased gene expression in the presence of the solvent ethanol, but otherwise no overlap between cadmium-induced gene expression and general stress response was detected. Molecular analysis of the corresponding genetic loci was performed using arbitrary polymerase chain reaction (PCR), DNA sequencing and comparison of the deduced protein products with sequences deposited in genetic databases. Some of the genetic loci targeted by the transposon did not show any similarities to any known genes; thus, they may represent 'novel' loci. The hypothesis that genes that are differentially expressed in the presence of heavy metals play a role in metal tolerance was verified for one of the mutants. This mutant, strain C11, was hypersensitive to cadmium and zinc ions. In mutant C11, the transposon had inserted into a genetic region displaying similarity to genes encoding the sensor/regulator protein pairs of two-component systems that regulate gene expression in metal-resistant bacteria, including czcRS of Ralstonia eutropha, czrRS of Pseudomonas

  9. ETS gene fusions in prostate cancer: from discovery to daily clinical practice.

    NARCIS (Netherlands)

    Tomlins, S.A.; Bjartell, A.; Chinnaiyan, A.M.; Jenster, G.; Nam, R.K.; Rubin, M.A.; Schalken, J.A.


    CONTEXT: In 2005, fusions between the androgen-regulated transmembrane protease serine 2 gene, TMPRSS2, and E twenty-six (ETS) transcription factors were discovered in prostate cancer. OBJECTIVE: To review advances in our understanding of ETS gene fusions, focusing on challenges affecting translatio

  10. Construction of hpaA gene from a clinical isolate of Helicobacter pyloriand identification of fusion protein

    Institute of Scientific and Technical Information of China (English)

    Ya-Fei Mao; Jie Yan; Li-Wei Li; Shu-Ping Li


    AIM: To clone hpaA gene from a clinical strain of Helicobacter pylori and to construct the expression vector of the gene and to identify immunity of the fusion protein.METHODS: The hpaA gene from a clinical isolate Y06 of H.pylori was amplified by high fidelity PCR. The nucleotide sequence of the target DNA amplification fragment was sequenced after T-A cloning. The recombinant expression vector inserted with hpaA gene was constructed. The expression of HpaA fusion protein in E. coli BL21DE3 induced by IPTG at different dosages was examined by SDS-PAGE.Western blot with commercial antibody against whole cell of H. pylorias well as immunodiffusion assay with selfprepared rabbit antiserum against HpaA fusion protein were applied to determine immunity of the fusion protein. ELISA was used to detect the antibody against HpaA in sera of 125 patients infected with H. pylori and to examine HpaA expression of 109 clinical isolates of H. pylori.RESULTS: In comparison with the reported corresponding sequences, the homologies of nucleotide and putative amino acid sequences of the cloned hpaA gene were from 94.25-97.32 % and 95.38-98.46 %, respectively. The output of HpaA fusion protein in its expression system of pET32ahpaA-BL21DE3 was approximately 40 % of the total bacterial proteins. HpaA fusion protein was able to combine with the commercial antibody against whole cell of H. pyloriand to induce rabbit producing specific antiserum with 1:4immunodiffusion titer after the animal was immunized with the fusion protein. 81.6 % of the serum samples from 125patients infected with H.pylori(102/125) were positive for HpaA antibody and all of the tested isolates of H.pylori(109/109) were detectable for HpaA.CONCLUSION: A prokaryotic expression system with high efficiency of H.pylorihpaA gene was successfully established.The HpaA expressing fusion protein showed satisfactory immunoreactivity and antigenicity. High frequencies of HpaA expression in different H. pyloriclinicalstrains

  11. DNA immunization with fusion genes containing HCV core region and HBV core region

    Institute of Scientific and Technical Information of China (English)

    杨莉; 刘晶; 孔玉英; 汪垣; 李光地


    The eucaryotic expression plasmids were constructed to express the complete (HCc191) or the truncated (HCc69 and HCc40) HCV core genes, solely or fused with the HBV core gene (HBc144). These constructions were transiently expressed in COS cells under the control of the CMV promoter. The antigenicity of HBc and HCc could be detected in the expression products by ELISA and Western blot. The mice immunized with these expression plasmids efficiently produced the anti-HCc antibodies, and also anti-HBc antibodies when the plasmids contained the fusion genes. In addition, the antibodies induced by the fusion genes were more persistent than those induced by the non-fusion HCV core genes. These indicate that the fusion of HCc genes to HBc gene is in favor of the immunogenicity of HCc, while the immunogenicity of HBc is not affected.

  12. Origin and ascendancy of a chimeric fusion gene: the beta/delta-globin gene of paenungulate mammals. (United States)

    Opazo, Juan C; Sloan, Angela M; Campbell, Kevin L; Storz, Jay F


    The delta-globin gene (HBD) of eutherian mammals exhibits a propensity for recombinational exchange with the closely linked beta-globin gene (HBB) and has been independently converted by the HBB gene in multiple lineages. Here we report the presence of a chimeric beta/delta fusion gene in the African elephant (Loxodonta africana) that was created by unequal crossing-over between misaligned HBD and HBB paralogs. The recombinant chromosome that harbors the beta/delta fusion gene in elephants is structurally similar to the "anti-Lepore" duplication mutant of humans (the reciprocal exchange product of the hemoglobin Lepore deletion mutant). However, the situation in the African elephant is unique in that the chimeric beta/delta fusion gene supplanted the parental HBB gene and is therefore solely responsible for synthesizing the beta-chain subunits of adult hemoglobin. A phylogenetic survey of beta-like globin genes in afrotherian and xenarthran mammals revealed that the origin of the chimeric beta/delta fusion gene and the concomitant inactivation of the HBB gene predated the radiation of "Paenungulata," a clade of afrotherian mammals that includes three orders: Proboscidea (elephants), Sirenia (dugongs and manatees), and Hyracoidea (hyraxes). The reduced fitness of the human Hb Lepore deletion mutant helps to explain why independently derived beta/delta fusion genes (which occur on an anti-Lepore chromosome) have been fixed in a number of mammalian lineages, whereas the reciprocal delta/beta fusion gene (which occurs on a Lepore chromosome) has yet to be documented in any nonhuman mammal. This illustrates how the evolutionary fates of chimeric fusion genes can be strongly influenced by their recombinational mode of origin.

  13. The Investigation of EWS-FLI-1 Fusion Gene in the Ewing Family of Tumors

    Institute of Scientific and Technical Information of China (English)

    GangFeng; ZhongquanZhao; DonglinWang


    There is evidence that 95% of the Ewing family of tumors (EFT) have a EWS-FLI-1 fusion gene. EWS-FL1-1 is a transcription factor with a pivotal function and it is known to bind to a special DNA sequence. Research has demonstrated that the EWS-FLI-1 fusion gene occurrence is related to the EFT, and it has been used to diagnose, treat and serve as a basis for EFT prognosis. We have briefly summarized the progress of the EWS-FLI-1 fusion gene in basic and clinical investigation within the past several years.

  14. Construction and characterization of calreticulin-HBsAg fusion gene recombinant adenovirus expression vector

    Institute of Scientific and Technical Information of China (English)


    AIM: To generate recombinant adenoviral vector con-taining calreticulin (CRT)-hepatitis B surface antigen (HBsAg) fusion gene for developing a safe, effective and HBsAg-specific therapeutic vaccine.METHODS: CRT and HBsAg gene were fused using polymerase chain reaction (PCR), endonuclease diges-tion and ligation methods. The fusion gene was cloned into pENTR/D-TOPO transfer vector after the base pairs of DNA (CACC) sequence was added to the 5′ end. Adenoviral expression vector containing CRT-HBsAg fusion gen...

  15. Novel fusion genes and chimeric transcripts in ependymal tumors

    DEFF Research Database (Denmark)

    Olsen, Thale Kristin; Panagopoulos, Ioannis; Gorunova, Ludmila


    We have previously identified two ALK rearrangements in a subset of ependymal tumors using a combination of cytogenetic data and RNA sequencing. The aim of this study was to perform an unbiased search for fusion transcripts in our entire series of ependymal tumors. Fusion analysis was performed...... using the FusionCatcher algorithm on 12 RNA-sequenced ependymal tumors. Candidate transcripts were prioritized based on the software's filtering and manual visualization using the BLAST (Basic Local Alignment Search Tool) and BLAT (BLAST-like alignment tool) tools. Genomic and reverse transcriptase PCR...... with subsequent Sanger sequencing was used to validate the potential fusions. Fluorescent in situ hybridization (FISH) using locus-specific probes was also performed. A total of 841 candidate chimeric transcripts were identified in the 12 tumors, with an average of 49 unique candidate fusions per tumor. After...

  16. LINE FUSION GENES: a database of LINE expression in human genes

    Directory of Open Access Journals (Sweden)

    Park Hong-Seog


    Full Text Available Abstract Background Long Interspersed Nuclear Elements (LINEs are the most abundant retrotransposons in humans. About 79% of human genes are estimated to contain at least one segment of LINE per transcription unit. Recent studies have shown that LINE elements can affect protein sequences, splicing patterns and expression of human genes. Description We have developed a database, LINE FUSION GENES, for elucidating LINE expression throughout the human gene database. We searched the 28,171 genes listed in the NCBI database for LINE elements and analyzed their structures and expression patterns. The results show that the mRNA sequences of 1,329 genes were affected by LINE expression. The LINE expression types were classified on the basis of LINEs in the 5' UTR, exon or 3' UTR sequences of the mRNAs. Our database provides further information, such as the tissue distribution and chromosomal location of the genes, and the domain structure that is changed by LINE integration. We have linked all the accession numbers to the NCBI data bank to provide mRNA sequences for subsequent users. Conclusion We believe that our work will interest genome scientists and might help them to gain insight into the implications of LINE expression for human evolution and disease. Availability

  17. NUP98 gene fusions and hematopoietic malignancies: common themes and new biologic insights. (United States)

    Gough, Sheryl M; Slape, Christopher I; Aplan, Peter D


    Structural chromosomal rearrangements of the Nucleoporin 98 gene (NUP98), primarily balanced translocations and inversions, are associated with a wide array of hematopoietic malignancies. NUP98 is known to be fused to at least 28 different partner genes in patients with hematopoietic malignancies, including acute myeloid leukemia, chronic myeloid leukemia in blast crisis, myelodysplastic syndrome, acute lymphoblastic leukemia, and bilineage/biphenotypic leukemia. NUP98 gene fusions typically encode a fusion protein that retains the amino terminus of NUP98; in this context, it is important to note that several recent studies have demonstrated that the amino-terminal portion of NUP98 exhibits transcription activation potential. Approximately half of the NUP98 fusion partners encode homeodomain proteins, and at least 5 NUP98 fusions involve known histone-modifying genes. Several of the NUP98 fusions, including NUP98-homeobox (HOX)A9, NUP98-HOXD13, and NUP98-JARID1A, have been used to generate animal models of both lymphoid and myeloid malignancy; these models typically up-regulate HOXA cluster genes, including HOXA5, HOXA7, HOXA9, and HOXA10. In addition, several of the NUP98 fusion proteins have been shown to inhibit differentiation of hematopoietic precursors and to increase self-renewal of hematopoietic stem or progenitor cells, providing a potential mechanism for malignant transformation.

  18. The tumorigenic FGFR3-TACC3 gene fusion escapes miR-99a regulation in glioblastoma (United States)

    Parker, Brittany C.; Annala, Matti J.; Cogdell, David E.; Granberg, Kirsi J.; Sun, Yan; Ji, Ping; Li, Xia; Gumin, Joy; Zheng, Hong; Hu, Limei; Yli-Harja, Olli; Haapasalo, Hannu; Visakorpi, Tapio; Liu, Xiuping; Liu, Chang-gong; Sawaya, Raymond; Fuller, Gregory N.; Chen, Kexin; Lang, Frederick F.; Nykter, Matti; Zhang, Wei


    Fusion genes are chromosomal aberrations that are found in many cancers and can be used as prognostic markers and drug targets in clinical practice. Fusions can lead to production of oncogenic fusion proteins or to enhanced expression of oncogenes. Several recent studies have reported that some fusion genes can escape microRNA regulation via 3′–untranslated region (3′-UTR) deletion. We performed whole transcriptome sequencing to identify fusion genes in glioma and discovered FGFR3-TACC3 fusions in 4 of 48 glioblastoma samples from patients both of mixed European and of Asian descent, but not in any of 43 low-grade glioma samples tested. The fusion, caused by tandem duplication on 4p16.3, led to the loss of the 3′-UTR of FGFR3, blocking gene regulation of miR-99a and enhancing expression of the fusion gene. The fusion gene was mutually exclusive with EGFR, PDGFR, or MET amplification. Using cultured glioblastoma cells and a mouse xenograft model, we found that fusion protein expression promoted cell proliferation and tumor progression, while WT FGFR3 protein was not tumorigenic, even under forced overexpression. These results demonstrated that the FGFR3-TACC3 gene fusion is expressed in human cancer and generates an oncogenic protein that promotes tumorigenesis in glioblastoma. PMID:23298836

  19. Recurrent Fusion Genes in Gastric Cancer: CLDN18-ARHGAP26 Induces Loss of Epithelial Integrity

    Directory of Open Access Journals (Sweden)

    Fei Yao


    Full Text Available Genome rearrangements, a hallmark of cancer, can result in gene fusions with oncogenic properties. Using DNA paired-end-tag (DNA-PET whole-genome sequencing, we analyzed 15 gastric cancers (GCs from Southeast Asians. Rearrangements were enriched in open chromatin and shaped by chromatin structure. We identified seven rearrangement hot spots and 136 gene fusions. In three out of 100 GC cases, we found recurrent fusions between CLDN18, a tight junction gene, and ARHGAP26, a gene encoding a RHOA inhibitor. Epithelial cell lines expressing CLDN18-ARHGAP26 displayed a dramatic loss of epithelial phenotype and long protrusions indicative of epithelial-mesenchymal transition (EMT. Fusion-positive cell lines showed impaired barrier properties, reduced cell-cell and cell-extracellular matrix adhesion, retarded wound healing, and inhibition of RHOA. Gain of invasion was seen in cancer cell lines expressing the fusion. Thus, CLDN18-ARHGAP26 mediates epithelial disintegration, possibly leading to stomach H+ leakage, and the fusion might contribute to invasiveness once a cell is transformed.

  20. Low frequency of ESRRA-C11orf20 fusion gene in ovarian carcinomas. (United States)

    Micci, Francesca; Panagopoulos, Ioannis; Thorsen, Jim; Davidson, Ben; Tropé, Claes Gøran; Heim, Sverre


    The identification of recurrent gene fusions in common epithelial cancers--for example, TMPRSS2/ERG in prostate cancer and EML4/ALK in nonsmall cell lung carcinomas--has raised the question of whether fusion genes are pathogenetically important also in ovarian carcinomas. The first recurrent fusion transcript in serous ovarian carcinomas was reported by Salzman et al. in 2011, who used deep paired-end sequencing to detect the fusion gene ESRRA-C11orf20 in 10 out of 67 (15%) serous ovarian carcinomas examined, a finding that holds great promise for our understanding of ovarian tumorigenesis as well as, potentially, for new treatment strategies. We wanted to test how frequent the ESRRA/C11orf20 fusion is in ovarian carcinomas of all subtypes, and therefore examined a series of 230 ovarian carcinomas of which 197 were of the serous subtype and 163 of the 197 were of stages III and IV--that is, the very same carcinoma subset where the fusion transcript had been found. We performed PCR and high-throughput sequencing analyses in search of the fusion transcript. We used the same primers described previously for the detection of the fusion and the same primer combination, but found no ESRRA/C11orf20 fusion in our series. A synthetic DNA plasmid containing the reported ESRRA/C11orf20 fusion was included as a positive control for our PCR experiments. Data from high-throughput sequencing of 23 ovarian carcinomas were screened in search of alternative partner(s) for the ESRRA and/or C11orf20 gene, but none was found. We conclude that the frequency of the ESRRA/C11orf20 gene fusion in serous ovarian carcinomas of stages III and IV must be considerable less than that reported previously (0/163 in our experience compared with 10/67 in the previous study). At the very least, it seems clear that the said fusion cannot be a common pathogenetic event in this tumor type.

  1. Low frequency of ESRRA-C11orf20 fusion gene in ovarian carcinomas.

    Directory of Open Access Journals (Sweden)

    Francesca Micci


    Full Text Available The identification of recurrent gene fusions in common epithelial cancers--for example, TMPRSS2/ERG in prostate cancer and EML4/ALK in nonsmall cell lung carcinomas--has raised the question of whether fusion genes are pathogenetically important also in ovarian carcinomas. The first recurrent fusion transcript in serous ovarian carcinomas was reported by Salzman et al. in 2011, who used deep paired-end sequencing to detect the fusion gene ESRRA-C11orf20 in 10 out of 67 (15% serous ovarian carcinomas examined, a finding that holds great promise for our understanding of ovarian tumorigenesis as well as, potentially, for new treatment strategies. We wanted to test how frequent the ESRRA/C11orf20 fusion is in ovarian carcinomas of all subtypes, and therefore examined a series of 230 ovarian carcinomas of which 197 were of the serous subtype and 163 of the 197 were of stages III and IV--that is, the very same carcinoma subset where the fusion transcript had been found. We performed PCR and high-throughput sequencing analyses in search of the fusion transcript. We used the same primers described previously for the detection of the fusion and the same primer combination, but found no ESRRA/C11orf20 fusion in our series. A synthetic DNA plasmid containing the reported ESRRA/C11orf20 fusion was included as a positive control for our PCR experiments. Data from high-throughput sequencing of 23 ovarian carcinomas were screened in search of alternative partner(s for the ESRRA and/or C11orf20 gene, but none was found. We conclude that the frequency of the ESRRA/C11orf20 gene fusion in serous ovarian carcinomas of stages III and IV must be considerable less than that reported previously (0/163 in our experience compared with 10/67 in the previous study. At the very least, it seems clear that the said fusion cannot be a common pathogenetic event in this tumor type.

  2. Stable Expression of Hantavirus H8205 Strain G1/IL-2 Gene and Immune Protection of the Fusion Gene

    Institute of Scientific and Technical Information of China (English)

    XIONG Ying; YUAN Yuan; JIA Min; YU Bing; HUANG Hanju


    To explore the feasibility of stable expression of Hantavirus H8205 strain G1 segment and human IL-2 fusion gene in Vero cells, and to examine the immune protection effects on mice vaccinated with this recombinant eukaryotic expression vector containing Hantavirus G1 gene and IL-2 gene. With the help of lipofectamine, the Vero cells were transfected with pcDNA3.1/HisB-IL-2-G1 and the positive cells were selected by G418. IFAT and SDS-PAGE electrophoresis were used to determine the stable transfection and expression of recombinant protein.Each mouse was inoculated with plasmids intramuscularly (i.m.) three times, 2 boosts were given at 2-week intervals, serum anti-hantavirus antibodies were detected by ELISA and neutralizing antibodies (NAb) were detected by Plaque Reduction Neutralization Test. The fusion protein expressed in Vero cells was 78 kD, corresponding to the estimated molecular size. The neutralizing antibody titers of mice with pcDNA3.1/HisB-IL-2-G1 were 1:20-1:80. IL-2/G1 fusion gene could be transferred in Vero cells and stably express the fusion protein. Specific humeral immune responses in mice can be induced with the recombinant eukaryotic expression vector containing the fusion gene, which lays the foundation for further development of therapeutic HTNV vaccine.

  3. ESRRA-C11orf20 is a recurrent gene fusion in serous ovarian carcinoma.

    Directory of Open Access Journals (Sweden)

    Julia Salzman


    Full Text Available Every year, ovarian cancer kills approximately 14,000 women in the United States and more than 140,000 women worldwide. Most of these deaths are caused by tumors of the serous histological type, which is rarely diagnosed before it has disseminated. By deep paired-end sequencing of mRNA from serous ovarian cancers, followed by deep sequencing of the corresponding genomic region, we identified a recurrent fusion transcript. The fusion transcript joins the 5' exons of ESRRA, encoding a ligand-independent member of the nuclear-hormone receptor superfamily, to the 3' exons of C11orf20, a conserved but uncharacterized gene located immediately upstream of ESRRA in the reference genome. To estimate the prevalence of the fusion, we tested 67 cases of serous ovarian cancer by RT-PCR and sequencing and confirmed its presence in 10 of these. Targeted resequencing of the corresponding genomic region from two fusion-positive tumor samples identified a nearly clonal chromosomal rearrangement positioning ESRRA upstream of C11orf20 in one tumor, and evidence of local copy number variation in the ESRRA locus in the second tumor. We hypothesize that the recurrent novel fusion transcript may play a role in pathogenesis of a substantial fraction of serous ovarian cancers and could provide a molecular marker for detection of the cancer. Gene fusions involving adjacent or nearby genes can readily escape detection but may play important roles in the development and progression of cancer.

  4. ESRRA-C11orf20 is a recurrent gene fusion in serous ovarian carcinoma. (United States)

    Salzman, Julia; Marinelli, Robert J; Wang, Peter L; Green, Ann E; Nielsen, Julie S; Nelson, Brad H; Drescher, Charles W; Brown, Patrick O


    Every year, ovarian cancer kills approximately 14,000 women in the United States and more than 140,000 women worldwide. Most of these deaths are caused by tumors of the serous histological type, which is rarely diagnosed before it has disseminated. By deep paired-end sequencing of mRNA from serous ovarian cancers, followed by deep sequencing of the corresponding genomic region, we identified a recurrent fusion transcript. The fusion transcript joins the 5' exons of ESRRA, encoding a ligand-independent member of the nuclear-hormone receptor superfamily, to the 3' exons of C11orf20, a conserved but uncharacterized gene located immediately upstream of ESRRA in the reference genome. To estimate the prevalence of the fusion, we tested 67 cases of serous ovarian cancer by RT-PCR and sequencing and confirmed its presence in 10 of these. Targeted resequencing of the corresponding genomic region from two fusion-positive tumor samples identified a nearly clonal chromosomal rearrangement positioning ESRRA upstream of C11orf20 in one tumor, and evidence of local copy number variation in the ESRRA locus in the second tumor. We hypothesize that the recurrent novel fusion transcript may play a role in pathogenesis of a substantial fraction of serous ovarian cancers and could provide a molecular marker for detection of the cancer. Gene fusions involving adjacent or nearby genes can readily escape detection but may play important roles in the development and progression of cancer.

  5. A simple, flexible and efficient PCR-fusion/Gateway cloning procedure for gene fusion, site-directed mutagenesis, short sequence insertion and domain deletions and swaps

    Directory of Open Access Journals (Sweden)

    Etchells J Peter


    Full Text Available Abstract Background The progress and completion of various plant genome sequencing projects has paved the way for diverse functional genomic studies that involve cloning, modification and subsequent expression of target genes. This requires flexible and efficient procedures for generating binary vectors containing: gene fusions, variants from site-directed mutagenesis, addition of protein tags together with domain swaps and deletions. Furthermore, efficient cloning procedures, ideally high throughput, are essential for pyramiding of multiple gene constructs. Results Here, we present a simple, flexible and efficient PCR-fusion/Gateway cloning procedure for construction of binary vectors for a range of gene fusions or variants with single or multiple nucleotide substitutions, short sequence insertions, domain deletions and swaps. Results from selected applications of the procedure which include ORF fusion, introduction of Cys>Ser mutations, insertion of StrepII tag sequence and domain swaps for Arabidopsis secondary cell wall AtCesA genes are demonstrated. Conclusion The PCR-fusion/Gateway cloning procedure described provides an elegant, simple and efficient solution for a wide range of diverse and complicated cloning tasks. Through streamlined cloning of sets of gene fusions and modification variants into binary vectors for systematic functional studies of gene families, our method allows for efficient utilization of the growing sequence and expression data.

  6. Dual-therapeutic reporter genes fusion for enhanced cancer gene therapy and imaging. (United States)

    Sekar, T V; Foygel, K; Willmann, J K; Paulmurugan, R


    Two of the successful gene-directed enzyme prodrug therapies include herpes simplex virus-thymidine kinase (HSV1-TK) enzyme-ganciclovir prodrug and the Escherichia coli nitroreductase (NTR) enzyme-CB1954 prodrug strategies; these enzyme-prodrug combinations produce activated cytotoxic metabolites of the prodrugs capable of tumor cell death by inhibiting DNA synthesis and killing quiescent cells, respectively. Both these strategies also affect significant bystander cell killing of neighboring tumor cells that do not express these enzymes. We have developed a dual-combination gene strategy, where we identified HSV1-TK and NTR fused in a particular orientation can effectively kill tumor cells when the tumor cells are treated with a fusion HSV1-TK-NTR gene- along with a prodrug combination of GCV and CB1954. In order to determine whether the dual-system demonstrate superior therapeutic efficacy than either HSV1-TK or NTR systems alone, we conducted both in vitro and in vivo tumor xenograft studies using triple negative SUM159 breast cancer cells, by evaluating the efficacy of cell death by apoptosis and necrosis upon treatment with the dual HSV1-TK genes-GCV-CB1954 prodrugs system, and compared the efficiency to HSV1-TK-GCV and NTR-CB1954. Our cell-based studies, tumor regression studies in xenograft mice, histological analyses of treated tumors and bystander studies indicate that the dual HSV1-TK-NTR-prodrug system is two times more efficient even with half the doses of both prodrugs than the respective single gene-prodrug system, as evidenced by enhanced apoptosis and necrosis of tumor cells in vitro in culture and xenograft of tumor tissues in animals.

  7. Nuclear fusion occurs during mating in Candida albicans and is dependent on the KAR3 gene. (United States)

    Bennett, Richard J; Miller, Mathew G; Chua, Penelope R; Maxon, Mary E; Johnson, Alexander D


    It is now well established that mating can occur between diploid a and alpha cells of Candida albicans. There is, however, controversy over when, and with what efficiency, nuclear fusion follows cell fusion to create stable tetraploid a/alpha cells. In this study, we have analysed the mating process between C. albicans strains using both cytological and genetic approaches. Using strains derived from SC5314, we used a number of techniques, including time-lapse microscopy, to demonstrate that efficient nuclear fusion occurs in the zygote before formation of the first daughter cell. Consistent with these observations, zygotes micromanipulated from mating mixes gave rise to mononuclear tetraploid cells, even when no selection for successful mating was applied to them. Mating between different clinical isolates of C. albicans revealed that while all isolates could undergo nuclear fusion, the efficiency of nuclear fusion varied in different crosses. We also show that nuclear fusion in C. albicans requires the Kar3 microtubule motor protein. Deletion of the CaKAR3 gene from both mating partners had little or no effect on zygote formation but reduced the formation of stable tetraploids more than 600-fold, as determined by quantitative mating assays. These findings demonstrate that nuclear fusion is an active process that can occur in C. albicans at high frequency to produce stable, mononucleate mating products.

  8. Construction of expression vector for NT4-ADNF-9 fusion gene

    Institute of Scientific and Technical Information of China (English)


    Objective To construct the prokaryotic expression vector bearing fusion gene NT4-ADNF-9 and lay foundation for further study on genetic therapy of neurosensory deafness. Methods By means of asymmetrical primer/ template,double stranded cDNA of activity dependent neurotrophic factor-9 (ADNF-9) was obtained,which included restriction enzymes sites on the two extremities. ADNF-9 cDNA was ligated to the signal and leader peptides of neurotrophin 4 (NT4),and the fusion gene was named NT4-ADNF-9. Then it was subc...

  9. CRTC1-MAML2 gene fusion in mucoepidermoid carcinoma of the lacrimal gland

    DEFF Research Database (Denmark)

    von Holstein, Sarah Linea; Fehr, André; Heegaard, Steffen


    -grade MEC of the lacrimal gland. There were no signs of recurrence or metastases during a five-year follow-up. Using RT-PCR and FISH we demonstrated that the tumor was positive for the CRTC1-MAML2 gene fusion previously shown to be associated with in particular low-grade salivary MECs with favorable...... prognosis. By immunohistochemistry we showed that the majority of tumor cells, including epidermoid, intermediate and mucous producing cells, expressed the CRTC1-MAML2 fusion protein. In contrast, 15 non-MEC lacrimal neoplasm were fusion-negative. Our findings show that lacrimal MEC is not only clinically...... anatomical sites and organs. Moreover, our findings indicate that the CRTC1-MAML2 fusion may be a useful diagnostic and prognostic biomarker for lacrimal MEC....

  10. ChimerDB 3.0: an enhanced database for fusion genes from cancer transcriptome and literature data mining (United States)

    Lee, Myunggyo; Lee, Kyubum; Yu, Namhee; Jang, Insu; Choi, Ikjung; Kim, Pora; Jang, Ye Eun; Kim, Byounggun; Kim, Sunkyu; Lee, Byungwook; Kang, Jaewoo; Lee, Sanghyuk


    Fusion gene is an important class of therapeutic targets and prognostic markers in cancer. ChimerDB is a comprehensive database of fusion genes encompassing analysis of deep sequencing data and manual curations. In this update, the database coverage was enhanced considerably by adding two new modules of The Cancer Genome Atlas (TCGA) RNA-Seq analysis and PubMed abstract mining. ChimerDB 3.0 is composed of three modules of ChimerKB, ChimerPub and ChimerSeq. ChimerKB represents a knowledgebase including 1066 fusion genes with manual curation that were compiled from public resources of fusion genes with experimental evidences. ChimerPub includes 2767 fusion genes obtained from text mining of PubMed abstracts. ChimerSeq module is designed to archive the fusion candidates from deep sequencing data. Importantly, we have analyzed RNA-Seq data of the TCGA project covering 4569 patients in 23 cancer types using two reliable programs of FusionScan and TopHat-Fusion. The new user interface supports diverse search options and graphic representation of fusion gene structure. ChimerDB 3.0 is available at PMID:27899563

  11. Drug-efflux and target-site gene expression patterns in Haemonchus contortus larvae able to survive increasing concentrations of levamisole in vitro (United States)

    Sarai, Ranbir S.; Kopp, Steven R.; Coleman, Glen T.; Kotze, Andrew C.


    While there is some evidence that changes in nicotinic acetylcholine receptor (nAChR) subunits confer resistance to levamisole in gastrointestinal helminth parasites, the exact nature of the resistance mechanism(s) is unclear. We utilised the presence of a resistant fraction within the Wallangra 2003 isolate of Haemonchus contortus larvae in order to subdivide the population into three subpopulations of larvae able to survive increasing concentrations of the drug. We then measured gene expression levels in the subpopulations and the larval population as a whole, focusing on genes encoding the subunit components of levamisole-sensitive receptors, genes encoding ancillary proteins involved in receptor assembly, and P-glycoprotein (P-gp) genes. The subpopulation surviving the lowest levamisole concentration showed increases of 1.5- to 3-fold in a number of P-gp genes (Hco-pgp-3, -4, -10, and -14) alongside unchanged receptor genes, compared to the whole Wallangra larval population. On the other hand, the subpopulation surviving the intermediate levamisole concentration showed an increase in only a single P-gp (Hco-pgp-14), alongside decreases in some receptor subunit (Hco-unc-63a) and ancillary protein genes (Hco-unc-50, Hco-ric-3.1 and 3.1). The subpopulation surviving the highest levamisole concentration showed further decreases in receptor subunit genes (Hco-unc-63a and Hco-unc-29 paralogs) as well as genes involved in receptor assembly (Hco-unc-74, Hco-unc-50, Hco-ric-3.1 and 3.1), alongside no increased P-gp gene levels. This suggests a biphasic pattern of drug resistance in the larvae of this worm isolate, in which a non-specific P-gp-mediated mechanism confers low levels of resistance, while higher level resistance is due to altered receptor subunit composition as a result of changes in both subunit composition and in the levels of proteins involved in receptor assembly. PMID:25057457

  12. Promoting lumbar spinal fusion by adenovirus-mediated bone morphogenetic protein-4 gene therapy

    Institute of Scientific and Technical Information of China (English)

    ZHAO Jian; ZHAO Dun-yan; SHEN Ai-guo; LIU Fan; ZHANG Feng; SUN Yu; WU Hong-fu; LU Chun-feng; SHI Hong-guang


    Objective: To determine whether an adenoviral construct containing bone morphogenetic protein-4 (BMP-4) gene can be used for lumbar spinal fusion. Methods: Twelve New Zealand white rabbits were randomly divided into two groups, 8 in the experimental group and 4 in the control group. Recombinant, replication-defective type 5 adenovirus with the cytomegalovirus (CMV) promoter and BMP-4 gene (Ad-BMP-4) was used. Another adenovirus constructed with the CMV promoter and β-galactosidase gene (Ad-β-gal) was used as control. Using collagen sponge as a carrier, Ad-BMP-4 (2.9×108 pfu/ml ) was directly implanted on the surface of L5-L6 lamina in the experimental group, while Ad-β-gal was implanted simultaneously in the control group. X-ray was obtained at 3, 6, and 12 weeks postoperatively to observe new bone formation. When new bone formation was identified, CT scans and three-dimensional reconstruction were obtained. After that, the animals were killed and underwent histological inspection.Results: In 12 weeks after operation, new bone formation and fusion were observed on CT scans in the experimental group, without the evidence of ectopic calcification in the canal. Negative results were found in the control group. Histological analysis demonstrated endochondral bone formation at the operative site and fusion at early stage was testified.Conclusions: In vivo gene therapy using Ad-BMP-4 for lumbar posterolateral spinal fusion is practicable and effective.

  13. IGF1 is a common target gene of Ewing's sarcoma fusion proteins in mesenchymal progenitor cells.

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    Luisa Cironi

    Full Text Available BACKGROUND: The EWS-FLI-1 fusion protein is associated with 85-90% of Ewing's sarcoma family tumors (ESFT, the remaining 10-15% of cases expressing chimeric genes encoding EWS or FUS fused to one of several ets transcription factor family members, including ERG-1, FEV, ETV1 and ETV6. ESFT are dependent on insulin-like growth factor-1 (IGF-1 for growth and survival and recent evidence suggests that mesenchymal progenitor/stem cells constitute a candidate ESFT origin. METHODOLOGY/PRINCIPAL FINDINGS: To address the functional relatedness between ESFT-associated fusion proteins, we compared mouse progenitor cell (MPC permissiveness for EWS-FLI-1, EWS-ERG and FUS-ERG expression and assessed the corresponding expression profile changes. Whereas all MPC isolates tested could stably express EWS-FLI-1, only some sustained stable EWS-ERG expression and none could express FUS-ERG for more than 3-5 days. Only 14% and 4% of the total number of genes that were respectively induced and repressed in MPCs by the three fusion proteins were shared. However, all three fusion proteins, but neither FLI-1 nor ERG-1 alone, activated the IGF1 promoter and induced IGF1 expression. CONCLUSION/SIGNIFICANCE: Whereas expression of different ESFT-associated fusion proteins may require distinct cellular microenvironments and induce transcriptome changes of limited similarity, IGF1 induction may provide one common mechanism for their implication in ESFT pathogenesis.

  14. Optimization of protoplast fusion conditions of bacteria able to de-grade bensulfuron-methyl and butachlor%苄嘧磺隆和丁草胺降解菌原生质体融合条件优化

    Institute of Scientific and Technical Information of China (English)

    李春艳; 吴志洋; 冯丽萍; 熊明华; 成小松


    In this study, functional fusants with dual functions to simultaneously degrade bensulfuron-methyl and butachlor were constructed through protoplast fusion of Rhodococcus sp. BX2 and Acinetobacter sp.LYC-1. The protoplast fusion condition was optimized and finally the fusion frequency reached to 2.67 × 10-7 under the following condition:40%of PEG4000, 10 min, 35℃, 500μL of freshly-prepared calcium phosphate, pH 7.5. The fusant F1 was obtained with the use of penicillin G and fosfomycin as selection markers and determined to be able to pass more than eight generations stablely on the plates containing bensulfuron-methyl and butachlor. The fusant F1 had respective high degradation rate of 65.35%BSM and 62.41%butachlor in mineral medium supplemented by BSM 100 mg·L-1 and butachlor 100 mg·L-1.%研究以苄嘧磺隆降解菌Rhodococcus sp. BX2与丁草胺降解菌Acinetobacter sp. LYC-1为亲本,优化用于构建可同时降解苄嘧磺隆与丁草胺的融合子的原生质体融合条件。通过单因素试验和正交试验,确定原生质体融合的最佳条件:PEG400040%、35℃、10 min,500μL新生磷酸钙溶液,pH 7.5,此条件下融合频率达到2.67×10-7。以青霉素和磷霉素作为筛选的遗传标记,最终获得在含有上述两种除草剂的无机盐培养基中可稳定继代培养8代以上的融合子F1。该融合子在含有100 mg·mL-1苄嘧磺隆和100 mg·mL-1丁草胺的无机盐培养基中,F1的降解率分别为65.35%和62.41%。

  15. Developing transgenic maize (Zea mays L.) with insect resistance and glyphosate tolerance by fusion gene transformation

    Institute of Scientific and Technical Information of China (English)

    SUN He; LANG Zhi-hong; LU Wei; ZHANG Jie; HE Kang-lai; ZHU Li; LIN Min; HUANG Da-fang


    Using linker peptide LP4/2A for multiple gene transformation is considered to be an effective method to stack or pyramid several traits in plants. Bacil us thuringiensis (Bt) cry gene and epsps (5-enolpyruvylshikimate-3-phosphate synthase) gene are two important genes for culturing pest-resistant and glyphosate-tolerant crops. We used linker peptide LP4/2A to connect the Bt cry1Ah gene with the 2mG2-epsps gene and combined the wide-used manA gene as a selective marker to construct one coordinated expression vector cal ed p2EPUHLAGN. The expression vector was transferred into maize by Agrobacterium tumefaciens-mediated transformation, and 60 plants were obtained, 40%of which were positive transformants. Molecular detection demonstrated that the two genes in the fusion vector were expressed simultaneously and spliced correctly in translation processing;meanwhile bioassay detection proved the transgenic maize had preferable pest resistance and glyphosate tolerance. Therefore, linker peptide LP4/2A provided a simple and reliable strategy for producing gene stacking in maize and the result showed that the fusion gene transformation system of LP4/2A was feasible in monocot plants.

  16. Targeted DNA demethylation and activation of endogenous genes using programmable TALE-TET1 fusion proteins. (United States)

    Maeder, Morgan L; Angstman, James F; Richardson, Marcy E; Linder, Samantha J; Cascio, Vincent M; Tsai, Shengdar Q; Ho, Quan H; Sander, Jeffry D; Reyon, Deepak; Bernstein, Bradley E; Costello, Joseph F; Wilkinson, Miles F; Joung, J Keith


    Genome-wide studies have defined cell type-specific patterns of DNA methylation that are important for regulating gene expression in both normal development and disease. However, determining the functional significance of specific methylation events remains challenging, owing to the lack of methods for removing such modifications in a targeted manner. Here we describe an approach for efficient targeted demethylation of specific CpGs in human cells using fusions of engineered transcription activator-like effector (TALE) repeat arrays and the TET1 hydroxylase catalytic domain. Using these TALE-TET1 fusions, we demonstrate that modification of critical methylated promoter CpG positions can lead to substantial increases in the expression of endogenous human genes. Our results delineate a strategy for understanding the functional significance of specific CpG methylation marks in the context of endogenous gene loci and validate programmable DNA demethylation reagents with potential utility for research and therapeutic applications.

  17. Recurrent BCAM-AKT2 fusion gene leads to a constitutively activated AKT2 fusion kinase in high-grade serous ovarian carcinoma (United States)

    Kannan, Kalpana; Coarfa, Cristian; Chao, Pei-Wen; Luo, Liming; Wang, Yan; Brinegar, Amy E.; Hawkins, Shannon M.; Milosavljevic, Aleksandar; Matzuk, Martin M.; Yen, Laising


    High-grade serous ovarian cancer (HGSC) is among the most lethal forms of cancer in women. Excessive genomic rearrangements, which are expected to create fusion oncogenes, are the hallmark of this cancer. Here we report a cancer-specific gene fusion between BCAM, a membrane adhesion molecule, and AKT2, a key kinase in the PI3K signaling pathway. This fusion is present in 7% of the 60 patient cancers tested, a significant frequency considering the highly heterogeneous nature of this malignancy. Further, we provide direct evidence that BCAM-AKT2 is translated into an in-frame fusion protein in the patient’s tumor. The resulting AKT2 fusion kinase is membrane-associated, constitutively phosphorylated, and activated as a functional kinase in cells. Unlike endogenous AKT2, whose activity is tightly regulated by external stimuli, BCAM-AKT2 escapes the regulation from external stimuli. Moreover, a BCAM-AKT2 fusion gene generated via chromosomal translocation using the CRISPR/Cas9 system leads to focus formation in both OVCAR8 and HEK-293T cell lines, suggesting that BCAM-AKT2 is oncogenic. Together, the results indicate that BCAM-AKT2 expression is a new mechanism of AKT2 kinase activation in HGSC. BCAM-AKT2 is the only fusion gene in HGSC that is proven to translate an aberrant yet functional kinase fusion protein with oncogenic properties. This recurrent genomic alteration is a potential therapeutic target and marker of a clinically relevant subtype for tailored therapy of HGSC. PMID:25733895

  18. [Standardization of quantitative detection of BCR-ABL gene expression by RQ-PCR in patients with chronic myeloid leukemia in cooperation with European Leukemia Net]. (United States)

    Sacha, Tomasz; Zawada, Magdalena; Czekalska, Sylwia; Florek, Izabela; Mueller, Martin; Gniot, Michał; Jaźwiec, Bozena; Kyrcz-Krzemień, Sławomira; Leszczyńska, Aleksandra; Lewandowski, Krzysztof; Matiakowska, Karolina; Solarska, Iwona; Stokłosa, Tomasz; Skotnicki, Aleksander B


    Monitoring of chronic myeloid leukemia treatment efficacy requires very sensitive methods of BCR-ABL gene detection based on polymerase chain reaction (PCR). The lack of comparability of BCR-ABL mRNA quantification results generated by various methodologies in different laboratories was the cause of an international multicenter trial initiation with the participation of 133 laboratories in 24 European countries cooperating within the "EUTOS for CML" project. Pracownia Diagnostyki Molekularnej Kliniki Hematologii is taking part in standardisation rounds organised since 2005. The compatibility of methodology used in Pracownia with European Leukemia Net (ELN) standards was confirmed, and correction factor for the expression of RQ-PCR results in an international scale was calculated. Pracownia was charge by ELN with a task of conducting the standardisation in polish molecular biology laboratories. Test probes were prepared and sent to eight cooperating laboratories. The results obtained in six laboratories were concordant with results from laboratory in Krakow after conversion to international scale, therefore it was possible to calculate individual correction factors. The participation of polish laboratories in international standardization process created the opportunity for unification of BCR-ABL quantification methodologies with recommendations of international experts, and showed that the quality of analyses performed in majority of them was satisfactory enough to calculate correction factor and to express the RQ-PCR results in widely accepted international scale.


    Directory of Open Access Journals (Sweden)

    M. Bocchia


    Full Text Available Chronic Myeloid Leukemia (CML is a myeloproliferative pluripotent stem cell disorder characterized by the presence of a cytogenetic hallmark, the Philadelphia (Ph chromosome, and accounts for 15% of adult leukemias. The disease progresses from a chronic phase through an accelerated phase to a blast phase and its natural course accounts for a median 4 years survival1. The Ph chromosome is derived by a reciprocal translocation termed t(9;22 in which the c-abl oncogene has moved from chromosome 9 into the breakpoint cluster region (bcr, within the bcr gene on chromosome 22, resulting in a chimeric bcr-abl fusion gene that encodes a 210 KD protein (p210 with constitutive tyrosine kinase activity. Two major alternative chimeric p210 can result from this fusion gene: p210-b2a2 where the junction occurs between bcr exon 2 (b2 and abl exon 2 (a2 and p210-b3a2 where the the junction occurs between bcr exon 3 (b3 and abl exon 2 (a2. About 40% of CML patients harbor the p210-b2a2 and about 60% of them show the p210-b3a2.

  20. Gene expression, single nucleotide variant and fusion transcript discovery in archival material from breast tumors.

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    Nadine Norton

    Full Text Available Advantages of RNA-Seq over array based platforms are quantitative gene expression and discovery of expressed single nucleotide variants (eSNVs and fusion transcripts from a single platform, but the sensitivity for each of these characteristics is unknown. We measured gene expression in a set of manually degraded RNAs, nine pairs of matched fresh-frozen, and FFPE RNA isolated from breast tumor with the hybridization based, NanoString nCounter (226 gene panel and with whole transcriptome RNA-Seq using RiboZeroGold ScriptSeq V2 library preparation kits. We performed correlation analyses of gene expression between samples and across platforms. We then specifically assessed whole transcriptome expression of lincRNA and discovery of eSNVs and fusion transcripts in the FFPE RNA-Seq data. For gene expression in the manually degraded samples, we observed Pearson correlations of >0.94 and >0.80 with NanoString and ScriptSeq protocols, respectively. Gene expression data for matched fresh-frozen and FFPE samples yielded mean Pearson correlations of 0.874 and 0.783 for NanoString (226 genes and ScriptSeq whole transcriptome protocols respectively, p<2x10(-16. Specifically for lincRNAs, we observed superb Pearson correlation (0.988 between matched fresh-frozen and FFPE pairs. FFPE samples across NanoString and RNA-Seq platforms gave a mean Pearson correlation of 0.838. In FFPE libraries, we detected 53.4% of high confidence SNVs and 24% of high confidence fusion transcripts. Sensitivity of fusion transcript detection was not overcome by an increase in depth of sequencing up to 3-fold (increase from ~56 to ~159 million reads. Both NanoString and ScriptSeq RNA-Seq technologies yield reliable gene expression data for degraded and FFPE material. The high degree of correlation between NanoString and RNA-Seq platforms suggests discovery based whole transcriptome studies from FFPE material will produce reliable expression data. The RiboZeroGold ScriptSeq protocol

  1. Gene fusion analysis in the battle against the African endemic sleeping sickness.

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    Philip Trimpalis

    Full Text Available The protozoan Trypanosoma brucei causes African Trypanosomiasis or sleeping sickness in humans, which can be lethal if untreated. Most available pharmacological treatments for the disease have severe side-effects. The purpose of this analysis was to detect novel protein-protein interactions (PPIs, vital for the parasite, which could lead to the development of drugs against this disease to block the specific interactions. In this work, the Domain Fusion Analysis (Rosetta Stone method was used to identify novel PPIs, by comparing T. brucei to 19 organisms covering all major lineages of the tree of life. Overall, 49 possible protein-protein interactions were detected, and classified based on (a statistical significance (BLAST e-value, domain length etc., (b their involvement in crucial metabolic pathways, and (c their evolutionary history, particularly focusing on whether a protein pair is split in T. brucei and fused in the human host. We also evaluated fusion events including hypothetical proteins, and suggest a possible molecular function or involvement in a certain biological process. This work has produced valuable results which could be further studied through structural biology or other experimental approaches so as to validate the protein-protein interactions proposed here. The evolutionary analysis of the proteins involved showed that, gene fusion or gene fission events can happen in all organisms, while some protein domains are more prone to fusion and fission events and present complex evolutionary patterns.

  2. A Search for Gene Fusions/Translocations in Breast Cancer (United States)


    scoring meta-outliers across 3 highly contributory datasets: Miller et al. (26), Hess et al. (27), and Wang et al. (28). Genes are ranked by their MetaCOPA...effects, and patient survival. Proc Natl Acad Sci USA 102:13550–13555. 27. Hess KR, et al. (2006) Pharmacogenomic predictor of sensitivity to...Papillary Thyroid Carcinoma* 5’ Partner 3’ Partner Prevalence References t(17;19)(q21;q13) KLK2 ETV4 ə% Hermans et al. Cancer Res 2008 inv(17;17)(q22;q25

  3. Identification of target genes of synovial sarcoma-associated fusion oncoprotein using human pluripotent stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Hayakawa, Kazuo [Department of Tissue Regeneration, Institute for Frontier Medical Sciences, Kyoto University, Kyoto (Japan); Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Department of Orthopaedic Surgery, Graduate School of Medical Sciences, Nagoya City University, Nagoya (Japan); Ikeya, Makoto [Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Fukuta, Makoto [Department of Tissue Regeneration, Institute for Frontier Medical Sciences, Kyoto University, Kyoto (Japan); Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Department of Orthopaedic Surgery, Graduate School of Medical Sciences, Nagoya City University, Nagoya (Japan); Woltjen, Knut [Department of Reprogramming Sciences, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Tamaki, Sakura; Takahara, Naoko; Kato, Tomohisa; Sato, Shingo [Department of Tissue Regeneration, Institute for Frontier Medical Sciences, Kyoto University, Kyoto (Japan); Otsuka, Takanobu [Department of Orthopaedic Surgery, Graduate School of Medical Sciences, Nagoya City University, Nagoya (Japan); Toguchida, Junya, E-mail: [Department of Tissue Regeneration, Institute for Frontier Medical Sciences, Kyoto University, Kyoto (Japan); Department of Cell Growth and Differentiation, Center for iPS Cell Research and Application, Kyoto University, Kyoto (Japan); Department of Orthopaedic Surgery, Graduate School of Medicine, Kyoto University, Kyoto (Japan)


    Highlights: ► We tried to identify targets of synovial sarcoma (SS)-associated SYT–SSX fusion gene. ► We established pluripotent stem cell (PSC) lines with inducible SYT–SSX gene. ► SYT–SSX responsive genes were identified by the induction of SYT–SSX in PSC. ► SS-related genes were selected from database by in silico analyses. ► 51 genes were finally identified among SS-related genes as targets of SYT–SSX in PSC. -- Abstract: Synovial sarcoma (SS) is a malignant soft tissue tumor harboring chromosomal translocation t(X; 18)(p11.2; q11.2), which produces SS-specific fusion gene, SYT–SSX. Although precise function of SYT–SSX remains to be investigated, accumulating evidences suggest its role in gene regulation via epigenetic mechanisms, and the product of SYT–SSX target genes may serve as biomarkers of SS. Lack of knowledge about the cell-of-origin of SS, however, has placed obstacle in the way of target identification. Here we report a novel approach to identify SYT–SSX2 target genes using human pluripotent stem cells (hPSCs) containing a doxycycline-inducible SYT–SSX2 gene. SYT–SSX2 was efficiently induced both at mRNA and protein levels within three hours after doxycycline administration, while no morphological change of hPSCs was observed until 24 h. Serial microarray analyses identified genes of which the expression level changed more than twofold within 24 h. Surprisingly, the majority (297/312, 95.2%) were up-regulated genes and a result inconsistent with the current concept of SYT–SSX as a transcriptional repressor. Comparing these genes with SS-related genes which were selected by a series of in silico analyses, 49 and 2 genes were finally identified as candidates of up- and down-regulated target of SYT–SSX, respectively. Association of these genes with SYT–SSX in SS cells was confirmed by knockdown experiments. Expression profiles of SS-related genes in hPSCs and human mesenchymal stem cells (hMSCs) were strikingly

  4. Construction of expression vector for NT4-ADNF-9 fusion gene

    Institute of Scientific and Technical Information of China (English)

    Guo-xi Zheng; Kang Zhu; Yang Jing; Jun-rong Wei; Hong-liang Zhu


    Objective To construct the prokaryotic expression vector bearing fusion gene NT4-ADNF-9 and lay foundation for further study on genetic therapy of neuraseusory deafness. Methods By means of asymmetrical prince/ template, double stranded eDNA of activity dependent neurotrophic factor-9 (ADNF-9) was obtained, which included restriction enzymes sites on the two extremities. ADNF-9 eDNA was ligated to the signal and leader peptides of nenrotrophin 4 (NT4), and the fusion gene was named NT4-ADNF-9. Then it was suheluned into prokaryotic expression vector pBV220, and called pBV220/ NT4-ADNF-9. Results Evidences of DNA sequence analysis and restrtction enzymes digestion showed that we recombined ADNF-9 eDNA to the 3'terminal of the signal and leader peptides of NT4, and the fusion gene was subcluned into pBV220 successfully. Bioactivity of the products was proved that it could support the cell survival and neurite growth in the primary cultures of dorsal root ganglia (DRG) of embryonic day-8 cbicken neurons as compared to the control. Conclusion Prokaryotic expression vector pBV220/NT4-ADNF-9 can be constructed successfully and the bioactivtty is satisfactory.

  5. Expanding the molecular toolbox for Lactococcus lactis: construction of an inducible thioredoxin gene fusion expression system

    LENUS (Irish Health Repository)

    Douillard, Francois P


    Abstract Background The development of the Nisin Inducible Controlled Expression (NICE) system in the food-grade bacterium Lactococcus lactis subsp. cremoris represents a cornerstone in the use of Gram-positive bacterial expression systems for biotechnological purposes. However, proteins that are subjected to such over-expression in L. lactis may suffer from improper folding, inclusion body formation and\\/or protein degradation, thereby significantly reducing the yield of soluble target protein. Although such drawbacks are not specific to L. lactis, no molecular tools have been developed to prevent or circumvent these recurrent problems of protein expression in L. lactis. Results Mimicking thioredoxin gene fusion systems available for E. coli, two nisin-inducible expression vectors were constructed to over-produce various proteins in L. lactis as thioredoxin fusion proteins. In this study, we demonstrate that our novel L. lactis fusion partner expression vectors allow high-level expression of soluble heterologous proteins Tuc2009 ORF40, Bbr_0140 and Tuc2009 BppU\\/BppL that were previously insoluble or not expressed using existing L. lactis expression vectors. Over-expressed proteins were subsequently purified by Ni-TED affinity chromatography. Intact heterologous proteins were detected by immunoblotting analyses. We also show that the thioredoxin moiety of the purified fusion protein was specifically and efficiently cleaved off by enterokinase treatment. Conclusions This study is the first description of a thioredoxin gene fusion expression system, purposely developed to circumvent problems associated with protein over-expression in L. lactis. It was shown to prevent protein insolubility and degradation, allowing sufficient production of soluble proteins for further structural and functional characterization.

  6. Detection of EMI4-ALK fusion gene in non-small cell lung cancer and its clinicopathologic correlation

    Institute of Scientific and Technical Information of China (English)



    Objective To investigate the frequency of EML4-ALK fusion gene in non small-cell lung cancer NSCLC patients,and its correlation with clinicopathologic features.Methods Real-time PCR was used to detect

  7. Two different point mutations in ABL gene ATP-binding domain conferring Primary Imatinib resistance in a Chronic Myeloid Leukemia (CML patient: A case report

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    Iqbal Zafar


    Full Text Available Imatinib (Gleevec is the effective therapy for BCR-ABL positive CML patients. Point mutations have been detected in ATP-binding domain of ABL gene which disturbs the binding of Gleevec to this target leading to resistance. Detection of mutations is helpful in clinical management of imatinib resistance. We established a very sensitive (ASO PCR to detect mutations in an imatinib-resistant CML patient. Mutations C944T and T1052C were detected which cause complete partial imatinib resistance, respectively. This is the first report of multiple point mutations conferring primary imatinib resistance in same patient at the same time. Understanding the biological reasons of primary imatinib resistance is one of the emerging issues of pharmacogenomics and will be helpful in understanding primary resistance of molecularly-targeted cancer therapies. It will also be of great utilization in clinical management of imatinib resistance. Moreover, this ASO-PCR assay is very effective in detecting mutations related to imatinib resistance.

  8. Knockout of the dhfr-ts gene in Trypanosoma cruzi generates attenuated parasites able to confer protection against a virulent challenge.

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    Cecilia Perez Brandan


    Full Text Available BACKGROUND: Trypanosoma cruzi is a protozoan parasite that causes severe disease in millions of habitants of developing countries. Currently there is no vaccine to prevent this disease and the available drugs have the consequences of side effects. Live vaccines are likely to be more effective in inducing protection than recombinant proteins or DNA vaccines; however, safety problems associated to their use have been pointed out. In recent years, increasing knowledge on the molecular genetics of Trypanosomes has allowed the identification and elimination of genes that may be necessary for parasite infectivity and survival. In this sense, targeted deletion or disruption of specific genes in the parasite genome may protect against such reversion to virulent genotypes. METHODS AND FINDINGS: By targeted gene disruption we generated monoallelic mutant parasites for the dhfr-ts gene in a T. cruzi strain that has been shown to be naturally attenuated. In comparison to T. cruzi wild type epimastigotes, impairment in growth of dhfr-ts(+/- mutant parasites was observed and mutant clones displayed decreased virulence in mice. Also, a lower number of T. cruzi-specific CD8(+ T cells, in comparison to those induced by wild type parasites, was detected in mice infected with mutant parasites. However, no remarkable differences in the protective effect of TCC wild type versus TCC mutant parasites were observed. Mice challenged with virulent parasites a year after the original infection with the mutant parasites still displayed a significant control over the secondary infection. CONCLUSION: This study indicates that it is possible to generate genetically attenuated T. cruzi parasites able to confer protection against further T. cruzi infections.

  9. Biological impact of hepatitis B virus X-hepatitis C virus core fusion gene on

    Institute of Scientific and Technical Information of China (English)

    Ma Zhen; Qin-Hai Shen; Guo-Min Chen; Da-Zhi Zhang


    AIM: To investigate the biological impact of hepatitis B virus X- hepatitis C virus core (HBV X-HCV C) fusion gene on hepatoma cells.METHODS: The recombinant adenoviruses AdXC,Ad-X and Ad-C expressing HBV X-HCV C fusion gene,HBVX gene and HCV C gene were constructed,respectively.Hepatoma cells were infected with different recombinant adenoviruses.MTT,colonyforming experiment,FCM,TUNEL assay were performed to observe the biological impact of the HBV X-HCV C fusion gene on liver cells.RESULTS: MTT showed that the Ad-XC group cells grew faster than the other group cells.Colony-forming experiment showed that the colony-forming rate for the Ad-XC group cells was significantly higher than that for the other group cells.FCM analysis showed that Ad-XC/Ad-X/Ad-C infection enhanced the progression of GIS phase in the HepG2 cell cycle.The apoptosis index of the Ad-XC,Ad-X,Ad-C group cells was significantly lower than that of the AdO and control group cells.Semi-quantitative RT-PCR showed that the expression level of c-myc was the highest in AdXC infected cells.Tumor formation was found at the injected site of mice inoculated with Ad-XC-infected LO2 cells,but not in control mice.CONCLUSION: Ad-XC,Ad-X and Ad-C facilitate the proliferation activity of HepG2 cells and inhibit their apoptosis in vitro.The effect of Ad-XC is significantly stronger than that of Ad-X and Ad-C.Up-regulation of c-myc may be one of the mechanisms underlying the synergism of HBVX and HCV C genes on hepatocarcinogenesis in athymic nude mice.

  10. Discovery of CTCF-sensitive Cis-spliced fusion RNAs between adjacent genes in human prostate cells.

    Directory of Open Access Journals (Sweden)

    Fujun Qin


    Full Text Available Genes or their encoded products are not expected to mingle with each other unless in some disease situations. In cancer, a frequent mechanism that can produce gene fusions is chromosomal rearrangement. However, recent discoveries of RNA trans-splicing and cis-splicing between adjacent genes (cis-SAGe support for other mechanisms in generating fusion RNAs. In our transcriptome analyses of 28 prostate normal and cancer samples, 30% fusion RNAs on average are the transcripts that contain exons belonging to same-strand neighboring genes. These fusion RNAs may be the products of cis-SAGe, which was previously thought to be rare. To validate this finding and to better understand the phenomenon, we used LNCaP, a prostate cell line as a model, and identified 16 additional cis-SAGe events by silencing transcription factor CTCF and paired-end RNA sequencing. About half of the fusions are expressed at a significant level compared to their parental genes. Silencing one of the in-frame fusions resulted in reduced cell motility. Most out-of-frame fusions are likely to function as non-coding RNAs. The majority of the 16 fusions are also detected in other prostate cell lines, as well as in the 14 clinical prostate normal and cancer pairs. By studying the features associated with these fusions, we developed a set of rules: 1 the parental genes are same-strand-neighboring genes; 2 the distance between the genes is within 30kb; 3 the 5' genes are actively transcribing; and 4 the chimeras tend to have the second-to-last exon in the 5' genes joined to the second exon in the 3' genes. We then randomly selected 20 neighboring genes in the genome, and detected four fusion events using these rules in prostate cancer and non-cancerous cells. These results suggest that splicing between neighboring gene transcripts is a rather frequent phenomenon, and it is not a feature unique to cancer cells.

  11. Kinase impact assessment in the landscape of fusion genes that retain kinase domains: a pan-cancer study. (United States)

    Kim, Pora; Jia, Peilin; Zhao, Zhongming


    Assessing the impact of kinase in gene fusion is essential for both identifying driver fusion genes (FGs) and developing molecular targeted therapies. Kinase domain retention is a crucial factor in kinase fusion genes (KFGs), but such a systematic investigation has not been done yet. To this end, we analyzed kinase domain retention (KDR) status in chimeric protein sequences of 914 KFGs covering 312 kinases across 13 major cancer types. Based on 171 kinase domain-retained KFGs including 101 kinases, we studied their recurrence, kinase groups, fusion partners, exon-based expression depth, short DNA motifs around the break points and networks. Our results, such as more KDR than 5'-kinase fusion genes, combinatorial effects between 3'-KDR kinases and their 5'-partners and a signal transduction-specific DNA sequence motif in the break point intronic sequences, supported positive selection on 3'-kinase fusion genes in cancer. We introduced a degree-of-frequency (DoF) score to measure the possible number of KFGs of a kinase. Interestingly, kinases with high DoF scores tended to undergo strong gene expression alteration at the break points. Furthermore, our KDR gene fusion network analysis revealed six of the seven kinases with the highest DoF scores (ALK, BRAF, MET, NTRK1, NTRK3 and RET) were all observed in thyroid carcinoma. Finally, we summarized common features of 'effective' (highly recurrent) kinases in gene fusions such as expression alteration at break point, redundant usage in multiple cancer types and 3'-location tendency. Collectively, our findings are useful for prioritizing driver kinases and FGs and provided insights into KFGs' clinical implications.

  12. Doxorubicin Differentially Induces Apoptosis, Expression of Mitochondrial Apoptosis-Related Genes, and Mitochondrial Potential in BCR-ABL1-Expressing Cells Sensitive and Resistant to Imatinib

    Directory of Open Access Journals (Sweden)

    Ewelina Synowiec


    Full Text Available Imatinib resistance is an emerging problem in the therapy of chronic myeloid leukemia (CML. Because imatinib induces apoptosis, which may be coupled with mitochondria and DNA damage is a prototype apoptosis-inducing factor, we hypothesized that imatinib-sensitive and -resistant CML cells might differentially express apoptosis-related mitochondrially encoded genes in response to genotoxic stress. We investigated the effect of doxorubicin (DOX, a DNA-damaging anticancer drug, on apoptosis and the expression of the mitochondrial NADH dehydrogenase 3 (MT-ND3 and cytochrome b (MT-CYB in model CML cells showing imatinib resistance caused by Y253H mutation in the BCR-ABL1 gene (253 or culturing imatinib-sensitive (S cells in increasing concentrations of imatinib (AR. The imatinib-resistant 253 cells displayed higher sensitivity to apoptosis induced by 1 μM DOX and this was confirmed by an increased activity of executioner caspases 3 and 7 in those cells. Native mitochondrial potential was lower in imatinib-resistant cells than in their sensitive counterparts and DOX lowered it. MT-CYB mRNA expression in 253 cells was lower than that in S cells and 0.1 μM DOX kept this relationship. In conclusion, imatinib resistance may be associated with altered mitochondrial response to genotoxic stress, which may be further exploited in CML therapy in patients with imatinib resistance.

  13. Construction of a fusion gene comprising the Taka-amylase A promoter and the Escherichia coli beta-glucuronidase gene and analysis of its expression in Aspergillus oryzae. (United States)

    Tada, S; Gomi, K; Kitamoto, K; Takahashi, K; Tamura, G; Hara, S


    Northern blot analysis of glucose-grown and starch-grown mycelia of Aspergillus oryzae RIB40 was conducted using the cloned Taka-amylase A (TAA) gene as a probe. The amount of mRNA homologous to the TAA gene was increased when this fungus was grown with starch as a sole carbon source. In order to analyze the induction mechanism, we inserted the Escherichia coli uidA gene encoding beta-glucuronidase (GUS) down-stream of the TAA promoter and introduced the resultant fusion gene into the A. oryzae genome. Production of a functional GUS protein was induced by starch, but not by glucose. When the effects of various sugars on expression of the fusion gene were examined, the results suggested that the expression of the fusion gene was under control of the TAA gene promoter.

  14. Origin of the plant Tm-1-like gene via two independent horizontal transfer events and one gene fusion event. (United States)

    Yang, Zefeng; Liu, Li; Fang, Huimin; Li, Pengcheng; Xu, Shuhui; Cao, Wei; Xu, Chenwu; Huang, Jinling; Zhou, Yong


    The Tomato mosaic virus (ToMV) resistance gene Tm-1 encodes a direct inhibitor of ToMV RNA replication to protect tomato from infection. The plant Tm-1-like (Tm-1L) protein is predicted to contain an uncharacterized N-terminal UPF0261 domain and a C-terminal TIM-barrel signal transduction (TBST) domain. Homologous searches revealed that proteins containing both of these two domains are mainly present in charophyte green algae and land plants but absent from glaucophytes, red algae and chlorophyte green algae. Although Tm-1 homologs are widely present in bacteria, archaea and fungi, UPF0261- and TBST-domain-containing proteins are generally encoded by different genes in these linages. A co-evolution analysis also suggested a putative interaction between UPF0261- and TBST-domain-containing proteins. Phylogenetic analyses based on homologs of these two domains revealed that plants have acquired UPF0261- and TBST-domain-encoding genes through two independent horizontal gene transfer (HGT) events before the origin of land plants from charophytes. Subsequently, gene fusion occurred between these two horizontally acquired genes and resulted in the origin of the Tm-1L gene in streptophytes. Our results demonstrate a novel evolutionary mechanism through which the recipient organism may acquire genes with functional interaction through two different HGT events and further fuse them into one functional gene.

  15. Analysis of EML4-ALK Gene Fusion Mutation in Patients 
with Non-small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Xuzhou WANG


    Full Text Available Background and objective Non-small cell lung cancer (NSCLC is the main type of lung cancer, and the related locus mutation detection research has become a hot direction of molecular targeted therapy, studying on gene mutation status of echinodem microtubule associated protein like 4-Anaplastic lymphoma kinase (EML4-ALK and epidermal growth factor receptor (EGFR, detecting the sensitivity of EML4-ALK gene fusion and gene mutation of EGFR. Methods EML4-ALK gene fusion in 85 cases of paraffin embedded tumor tissue and adjacent lung tissue was detected with the application of immunohistochemistry (IHC, Scorpions amplification refractory mutation system (Scorpions ARMS fluorescence quantitative PCR and fluorescence in situ hybridization (FISH technology, and EGFR gene in 18, 19, 20 and 21 exon mutation status was detected with the application of ARMS method. Results In 115 cases of NSCLC, IHC showed 32 cases with ALK (D5F3 expression, the expression rate was 27.8%; ARMS showed 27 cases with EML4-ALK fusion gene mutation, the mutation detection rate was 23.5%; 53 cases were detected with EGFR mutation, the mutation rate was 46%. While FISH showed 23 cases with EML4-ALK fusion gene mutation, the detection rate was 20%, slightly lower than the ARMS detection results, suggesting that ARMS more sensitive. Conclusion The application of IHC, ARMS fluorescence quantitative PCR and FISH technology can make a rapid and accurate evaluation of EML4-ALK gene fusion.

  16. Membrane fusion inducers, chloroquine and spermidine increase lipoplex-mediated gene transfection

    Energy Technology Data Exchange (ETDEWEB)

    Wong-Baeza, Carlos; Bustos, Israel; Serna, Manuel; Tescucano, Alonso; Alcantara-Farfan, Veronica; Ibanez, Miguel [Biochemistry Department, National Polytechnic Institute (IPN), Mexico City 11340 (Mexico); Montanez, Cecilia [Department of Genetics and Molecular Biology, Centre for Research and Advanced Studies (CINVESTAV), IPN, Mexico City 07360 (Mexico); Wong, Carlos [Biochemistry Department, National Polytechnic Institute (IPN), Mexico City 11340 (Mexico); Baeza, Isabel, E-mail: [Biochemistry Department, National Polytechnic Institute (IPN), Mexico City 11340 (Mexico)


    Gene transfection into mammalian cells can be achieved with viral and non-viral vectors. Non-viral vectors, such as cationic lipids that form lipoplexes with DNA, are safer and more stable than viral vectors, but their transfection efficiencies are lower. Here we describe that the simultaneous treatment with a membrane fusion inducer (chlorpromazine or procainamide) plus the lysosomotropic agent chloroquine increases lipoplex-mediated gene transfection in human (HEK293 and C-33 A) and rat (PC12) cell lines (up to 9.2-fold), as well as in situ in BALB/c mice spleens and livers (up to 6-fold); and that the polyamine spermidine increases lipoplex-mediated gene transfection and expression in cell cultures. The use of these four drugs provides a novel, safe and relatively inexpensive way to considerably increase lipoplex-mediated gene transfection efficiency.

  17. Deep RNA sequencing analysis of readthrough gene fusions in human prostate adenocarcinoma and reference samples

    Directory of Open Access Journals (Sweden)

    Nacu Serban


    Full Text Available Abstract Background Readthrough fusions across adjacent genes in the genome, or transcription-induced chimeras (TICs, have been estimated using expressed sequence tag (EST libraries to involve 4-6% of all genes. Deep transcriptional sequencing (RNA-Seq now makes it possible to study the occurrence and expression levels of TICs in individual samples across the genome. Methods We performed single-end RNA-Seq on three human prostate adenocarcinoma samples and their corresponding normal tissues, as well as brain and universal reference samples. We developed two bioinformatics methods to specifically identify TIC events: a targeted alignment method using artificial exon-exon junctions within 200,000 bp from adjacent genes, and genomic alignment allowing splicing within individual reads. We performed further experimental verification and characterization of selected TIC and fusion events using quantitative RT-PCR and comparative genomic hybridization microarrays. Results Targeted alignment against artificial exon-exon junctions yielded 339 distinct TIC events, including 32 gene pairs with multiple isoforms. The false discovery rate was estimated to be 1.5%. Spliced alignment to the genome was less sensitive, finding only 18% of those found by targeted alignment in 33-nt reads and 59% of those in 50-nt reads. However, spliced alignment revealed 30 cases of TICs with intervening exons, in addition to distant inversions, scrambled genes, and translocations. Our findings increase the catalog of observed TIC gene pairs by 66%. We verified 6 of 6 predicted TICs in all prostate samples, and 2 of 5 predicted novel distant gene fusions, both private events among 54 prostate tumor samples tested. Expression of TICs correlates with that of the upstream gene, which can explain the prostate-specific pattern of some TIC events and the restriction of the SLC45A3-ELK4 e4-e2 TIC to ERG-negative prostate samples, as confirmed in 20 matched prostate tumor and normal

  18. FN1-EGF gene fusions are recurrent in calcifying aponeurotic fibroma. (United States)

    Puls, Florian; Hofvander, Jakob; Magnusson, Linda; Nilsson, Jenny; Haywood, Elaine; Sumathi, Vaiyapuri P; Mangham, D Chas; Kindblom, Lars-Gunnar; Mertens, Fredrik


    Calcifying aponeurotic fibroma (CAF) is a soft tissue neoplasm with a predilection for the hands and feet in children and adolescents. Its molecular basis is unknown. We used chromosome banding analysis, fluorescence in situ hybridization (FISH), mRNA sequencing (RNA-seq), RT-PCR, and immunohistochemistry to characterize a series of CAFs. An insertion ins(2;4)(q35;q25q?) was identified in the index case. Fusion of the FN1 and EGF genes, mapping to the breakpoint regions on chromosomes 2 and 4, respectively, was detected by RNA-seq and confirmed by RT-PCR in the index case and two additional cases. FISH on five additional tumours identified FN1-EGF fusions in all cases. CAFs analysed by RT-PCR showed that FN1 exon 23, 27 or 42 was fused to EGF exon 17 or 19. High-level expression of the entire FN1 gene in CAF suggests that strong FN1 promoter activity drives inappropriate expression of the biologically active portion of EGF, which was detected immunohistochemically in 8/9 cases. The FN1-EGF fusion, which has not been observed in any other neoplasm, appears to be the main driver mutation in CAF. Although further functional studies are required to understand the exact pathogenesis of CAF, the composition of the chimera suggests an autocrine/paracrine mechanism of transformation. Copyright © 2015 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

  19. Construction and Expression of GST and Carbon Skeleton of Amatoxins Fusion Gene

    Institute of Scientific and Technical Information of China (English)

    Zhao Jian(赵建); Cao Mei; Zhang Jie; Hou Ruotong; Chen Yaofeng; Yang Zhirong


    The primer, AT For1, AT For2 and AT Back, are designed and synthesized for adding-PCR that is used to construct the fusion GST-AT gene. Depending on two-time adding-PCR amplification and the insertion of coding sequence of octapeptide carbon skeleton into the 3 terminus of GST gene, the authors get the masculine recon, pGAT-1, confirmed by sequence determination and analysis, whose ORF is read-through. After IPTG induction and partial purification, SDS-PAGE electrophoresis is employed to detect the gene expression. The expressions of total bacterial proteins are about 21.3%, 22.5%, 19.32%, 21.73% in E. coli BL21, DH5α, JM109 and Top10 strains, respectively. Considering the quantity of induced total bacteria, the E. coli BL21 is the best one among the four recipient strains. This article supplies an academic foundation for producing biological active amatoxins.

  20. Molecular Characterization of Inflammatory Myofibroblastic Tumors With Frequent ALK and ROS1 Gene Fusions and Rare Novel RET Rearrangement

    NARCIS (Netherlands)

    Antonescu, Cristina R.; Suurmeijer, Albert J. H.; Zhang, Lei; Sung, Yun-Shao; Jungbluth, Achim A.; Travis, William D.; Al-Ahmadie, Hikmat; Fletcher, Christopher D. M.; Alaggio, Rita


    Approximately 50% of conventional inflammatory myofibroblastic tumors (IMTs) harbor ALK gene rearrangement and overexpress ALK. Recently, gene fusions involving other kinases have been implicated in the pathogenesis of IMT, including ROS1 and in 1 patient PDGFRB. However, it remains uncertain whethe

  1. Regulation of RAD54- and RAD52-lacZ gene fusions in Saccharomyces cerevisiae in response to DNA damage

    Energy Technology Data Exchange (ETDEWEB)

    Cole, G.M.; Schild, D.; Lovett, S.T.; Mortimer, R.K.


    The RAD52 and RAD54 genes in the yeast Saccharomyces cerevisiae are involved in both DNA repair and DNA recombination. RAD54 has recently been shown to be inducible by X-rays, while RAD52 is not. To further investigate the regulation of these genes, we constructed gene fusions using 5' regions upstream of the RAD52 and RAD54 genes and a 3'-terminal fragment of the Escherichia coli beta-galactosidase gene. Yeast transformants with either an integrated or an autonomously replicating plasmid containing these fusions expressed beta-galactosidase activity constitutively. In addition, the RAD54 gene fusion was inducible in both haploid and diploid cells in response to the DNA-damaging agents X-rays, UV light, and methyl methanesulfonate, but not in response to heat shock. The RAD52-lacZ gene fusion showed little or no induction in response to X-ray or UV radiation nor methyl methanesulfonate. Typical induction levels for RAD54 in cells exposed to such agents were from 3- to 12-fold, in good agreement with previous mRNA analyses. When MATa cells were arrested in G1 with alpha-factor, RAD54 was still inducible after DNA damage, indicating that the observed induction is independent of the cell cycle. Using a yeast vector containing the EcoRI structural gene fused to the GAL1 promoter, we showed that double-strand breaks alone are sufficient in vivo for induction of RAD54.

  2. Decoding of exon splicing patterns in the human RUNX1-RUNX1T1 fusion gene. (United States)

    Grinev, Vasily V; Migas, Alexandr A; Kirsanava, Aksana D; Mishkova, Olga A; Siomava, Natalia; Ramanouskaya, Tatiana V; Vaitsiankova, Alina V; Ilyushonak, Ilia M; Nazarov, Petr V; Vallar, Laurent; Aleinikova, Olga V


    The t(8;21) translocation is the most widespread genetic defect found in human acute myeloid leukemia. This translocation results in the RUNX1-RUNX1T1 fusion gene that produces a wide variety of alternative transcripts and influences the course of the disease. The rules of combinatorics and splicing of exons in the RUNX1-RUNX1T1 transcripts are not known. To address this issue, we developed an exon graph model of the fusion gene organization and evaluated its local exon combinatorics by the exon combinatorial index (ECI). Here we show that the local exon combinatorics of the RUNX1-RUNX1T1 gene follows a power-law behavior and (i) the vast majority of exons has a low ECI, (ii) only a small part is represented by "exons-hubs" of splicing with very high ECI values, and (iii) it is scale-free and very sensitive to targeted skipping of "exons-hubs". Stochasticity of the splicing machinery and preferred usage of exons in alternative splicing can explain such behavior of the system. Stochasticity may explain up to 12% of the ECI variance and results in a number of non-coding and unproductive transcripts that can be considered as a noise. Half-life of these transcripts is increased due to the deregulation of some key genes of the nonsense-mediated decay system in leukemia cells. On the other hand, preferred usage of exons may explain up to 75% of the ECI variability. Our analysis revealed a set of splicing-related cis-regulatory motifs that can explain "attractiveness" of exons in alternative splicing but only when they are considered together. Cis-regulatory motifs are guides for splicing trans-factors and we observed a leukemia-specific profile of expression of the splicing genes in t(8;21)-positive blasts. Altogether, our results show that alternative splicing of the RUNX1-RUNX1T1 transcripts follows strict rules and that the power-law component of the fusion gene organization confers a high flexibility to this process.

  3. High-speed biosensing strategy for non-invasive profiling of multiple cancer fusion genes in urine. (United States)

    Koo, Kevin M; Wee, Eugene J H; Trau, Matt


    Aberrant chromosal rearrangements, such as the multiple variants of TMPRSS2:ERG fusion gene mutations in prostate cancer (PCa), are promising diagnostic and prognostic biomarkers due to their specific expression in cancerous tissue only. Additionally, TMPRSS2:ERG variants are detectable in urine to provide non-invasive PCa diagnostic sampling as an attractive surrogate for needle biopsies. Therefore, rapid and simplistic assays for identifying multiple urinary TMPRSS2:ERG variants are potentially useful to aid in early cancer detection, immediate patient risk stratification, and prompt personalized treatment. However, current strategies for simultaneous detection of multiple gene fusions are limited by tedious and prolonged experimental protocols, thus limiting their use as rapid clinical screening tools. Herein, we report a simple and rapid gene fusion strategy which expliots the specificity of DNA ligase and the speed of isothermal amplification to simultaneously detect multiple fusion gene RNAs within a short sample-to-answer timeframe of 60min. The method has a low detection limit of 2 amol (1000 copies), and was successfully applied for non-invasive fusion gene profiling in patient urine samples with subsequent validation by a PCR-based gold standard approach.

  4. SPR Detection and Discrimination of the Oligonucleotides Related to the Normal and the Hybrid bcr-abl Genes by Two Stringency Control Strategies (United States)

    Matsishin, M. J.; Ushenin, Iu. V.; Rachkov, A. E.; Solatkin, A. P.


    In this study, we applied two stringency control strategies for surface plasmon resonance (SPR) detection of DNA hybridization and discrimination of completely and partially complementary 24-mer sequences. These sequences are specific to the human normal bcr and the hybrid bcr-abl genes, protein products of which are responsible for some leukemia. SPR sensors based on resonance phenomena in nanoscale gold films are well suited for label-free, real-time investigations of the macromolecule interactions. Thermodynamic parameters obtained using the web server DINAMelt allowed supposing the possibility for realization (a) stringency control based on the ionic strength of the hybridization buffer and (b) stringency control based on the temperature elevation. The first one resulted in that the discrimination index of completely complementary and partially complementary oligonucleotides depending on the target concentration varied from 1.3 to 1.8 in 2 × SSC and from 2.0 to 2.9 in 0.5 × SSC. For implementation of the second stringency control strategy, SPR spectrometer measuring flow cell with built-in high-precision temperature control and regulation as well as corresponding software was created. It is shown that the duplexes formed by the immobilized probes mod-Ph and completely complementary oligonucleotides P1 remained without significant changes until ~50 °C, while the duplexes formed with partially complementary oligonucleotide Bcrex14 almost entirely disrupted at 40 °C. Thus, the absolutely effective thermodiscrimination of this pair of oligonucleotides was achieved in this temperature range (40-50 °C).

  5. Spatial and temporal analysis of gene expression during growth and fusion of the mouse facial prominences.

    Directory of Open Access Journals (Sweden)

    Weiguo Feng

    Full Text Available Orofacial malformations resulting from genetic and/or environmental causes are frequent human birth defects yet their etiology is often unclear because of insufficient information concerning the molecular, cellular and morphogenetic processes responsible for normal facial development. We have, therefore, derived a comprehensive expression dataset for mouse orofacial development, interrogating three distinct regions - the mandibular, maxillary and frontonasal prominences. To capture the dynamic changes in the transcriptome during face formation, we sampled five time points between E10.5-E12.5, spanning the developmental period from establishment of the prominences to their fusion to form the mature facial platform. Seven independent biological replicates were used for each sample ensuring robustness and quality of the dataset. Here, we provide a general overview of the dataset, characterizing aspects of gene expression changes at both the spatial and temporal level. Considerable coordinate regulation occurs across the three prominences during this period of facial growth and morphogenesis, with a switch from expression of genes involved in cell proliferation to those associated with differentiation. An accompanying shift in the expression of polycomb and trithorax genes presumably maintains appropriate patterns of gene expression in precursor or differentiated cells, respectively. Superimposed on the many coordinated changes are prominence-specific differences in the expression of genes encoding transcription factors, extracellular matrix components, and signaling molecules. Thus, the elaboration of each prominence will be driven by particular combinations of transcription factors coupled with specific cell:cell and cell:matrix interactions. The dataset also reveals several prominence-specific genes not previously associated with orofacial development, a subset of which we externally validate. Several of these latter genes are components of

  6. Spatial and temporal analysis of gene expression during growth and fusion of the mouse facial prominences. (United States)

    Feng, Weiguo; Leach, Sonia M; Tipney, Hannah; Phang, Tzulip; Geraci, Mark; Spritz, Richard A; Hunter, Lawrence E; Williams, Trevor


    Orofacial malformations resulting from genetic and/or environmental causes are frequent human birth defects yet their etiology is often unclear because of insufficient information concerning the molecular, cellular and morphogenetic processes responsible for normal facial development. We have, therefore, derived a comprehensive expression dataset for mouse orofacial development, interrogating three distinct regions - the mandibular, maxillary and frontonasal prominences. To capture the dynamic changes in the transcriptome during face formation, we sampled five time points between E10.5-E12.5, spanning the developmental period from establishment of the prominences to their fusion to form the mature facial platform. Seven independent biological replicates were used for each sample ensuring robustness and quality of the dataset. Here, we provide a general overview of the dataset, characterizing aspects of gene expression changes at both the spatial and temporal level. Considerable coordinate regulation occurs across the three prominences during this period of facial growth and morphogenesis, with a switch from expression of genes involved in cell proliferation to those associated with differentiation. An accompanying shift in the expression of polycomb and trithorax genes presumably maintains appropriate patterns of gene expression in precursor or differentiated cells, respectively. Superimposed on the many coordinated changes are prominence-specific differences in the expression of genes encoding transcription factors, extracellular matrix components, and signaling molecules. Thus, the elaboration of each prominence will be driven by particular combinations of transcription factors coupled with specific cell:cell and cell:matrix interactions. The dataset also reveals several prominence-specific genes not previously associated with orofacial development, a subset of which we externally validate. Several of these latter genes are components of bidirectional

  7. On the origin of protein synthesis factors: a gene duplication/fusion model. (United States)

    Cousineau, B; Leclerc, F; Cedergren, R


    Sequence similarity has given rise to the proposal that IF-2, EF-G, and EF-Tu are related through a common ancestor. We evaluate this proposition and whether the relationship can be extended to other factors of protein synthesis. Analysis of amino acid sequence similarity gives statistical support for an evolutionary affiliation among IF-1, IF-2, IF-3, EF-Tu, EF-Ts, and EF-G and suggests further that this association is a result of gene duplication/fusion events. In support of this mechanism, the three-dimensional structures of IF-3, EF-Tu, and EF-G display a predictable domain structure and overall conformational similarity. The model that we propose consists of three consecutives duplication/fusion events which would have taken place before the divergence of the three superkingdoms: eubacteria, archaea, and eukaryotes. The root of this protein superfamily tree would be an ancestor of the modern IF-1 gene sequence. The repeated fundamental motif of this protein superfamily is a small RNA binding domain composed of two alpha-helices packed along side of an antiparallel beta-sheet.

  8. Expression pattern of the RAR alpha-PML fusion gene in acute promyelocytic leukemia. (United States)

    Alcalay, M; Zangrilli, D; Fagioli, M; Pandolfi, P P; Mencarelli, A; Lo Coco, F; Biondi, A; Grignani, F; Pelicci, P G


    Two chimeric genes, PML-RAR alpha and RAR alpha-PML, are formed as a consequence of the acute promyelocytic leukemia (APL)-specific reciprocal translocation of chromosomes 15 and 17 [t(15;17)]. PML-RAR alpha is expressed as a fusion protein. We investigated the organization and expression pattern of the RAR alpha-PML gene in a series of APL patients representative of the molecular heterogeneity of the t(15;17) and found (i) two types of RAR alpha-PML mRNA junctions (RAR alpha exon 2/PML exon 4 or RAR alpha exon 2/PML exon 7) that maintain the RAR alpha and PML longest open reading frames aligned and are the result of chromosome 15 breaking at two different sites; and (ii) 10 different RAR alpha-PML fusion transcripts that differ for the assembly of their PML coding exons. A RAR alpha-PML transcript was present in most, but not all, APL patients. Images PMID:1317574

  9. PL1 fusion gene: a novel visual selectable marker gene that confers tolerance to multiple abiotic stresses in transgenic tomato. (United States)

    Jin, Feng; Li, Shu; Dang, Lijie; Chai, Wenting; Li, Pengli; Wang, Ning Ning


    Visual selectable markers, including the purple color caused by the accumulation of anthocyanins, have been proposed for use as antibiotic-free alternatives. However, the excessive accumulation of anthocyanins seriously inhibits the growth and development of transgenic plants. In our study, the AtDWF4 promoter from Arabidopsis and the tomato LeANT1 gene, encoding a MYB transcription factor, were used to construct the PL1 fusion gene to test whether it could be used as a visual selectable marker gene for tomato transformation. All the PL1 transgenic shoots exhibited intense purple color on shoot induction medium. In the transgenic tomato plants, PL1 was highly expressed in the cotyledons, but expressed only slightly in the true leaves and other organs. The expression of PL1 had no significantly adverse effects on the growth or development of the transgenic tomato plants, and conferred tolerance to multiple abiotic stresses in them. With the “cut off green shoots” method, multiple independent 35S::GFP transgenic tomato lines were successfully obtained using PL1 as the selectable marker gene. These results suggest that PL1 has potential application of visual selectable marker gene for tomato transformation.

  10. Horizontal gene transfers and cell fusions in microbiology, immunology and oncology (Review). (United States)

    Sinkovics, Joseph G


    Evolving young genomes of archaea, prokaryota and unicellular eukaryota were wide open for the acceptance of alien genomic sequences, which they often preserved and vertically transferred to their descendants throughout three billion years of evolution. Established complex large genomes, although seeded with ancestral retroelements, have come to regulate strictly their integrity. However, intruding retroelements, especially the descendents of Ty3/Gypsy, the chromoviruses, continue to find their ways into even the most established genomes. The simian and hominoid-Homo genomes preserved and accommodated a large number of endogenous retroviral genomic segments. These retroelements may mature into exogenous retroviruses, or into functional new genes. Phages and viruses have been instrumental in incorporating and transferring host cell genes. These events profoundly influenced and altered the course of evolution. Horizontal (lateral) gene transfers (HGT) overwhelmed the genomes of the ancient protocells and the evolving unicellular microorganisms, actually leading to their Cambrian explosion. While the rigidly organized genomes of multicellular organisms increasingly resist H/LGT, de-differentiated cells assuming the metabolism of their onto- or phylogenetic ancestors, open up widely to the practice of H/LGT by direct transfer, or to transfers mediated by viruses, or by cell fusions. This activity is intensified in malignantly transformed cells, thus rendering these subjects receptive to therapy with oncolytic viruses and with viral vectors of tumor-suppressive or immunogenic genetic materials. Naturally formed hybrids of dendritic and tumor cells are often tolerogenic, whereas laboratory products of these unisons may be immunogenic in the hosts of origin. As human breast cancer stem cells are induced by a treacherous class of CD8+ T cells to undergo epithelial to mesenchymal (ETM) transition and to yield to malignant transformation by the omnipresent proto

  11. NAN fusions: a synthetic sialidase reporter gene as a sensitive and versatile partner for GUS. (United States)

    Kirby, J; Kavanagh, T A


    GUS continues to be the reporter of choice for many gene fusion applications, due to the unparalleled sensitivity of the encoded enzyme and the ease with which it can be quantified in cell-free extracts and visualized histochemically in cells and tissues. A compatible and functionally equivalent reporter gene would facilitate dual promoter studies and internal standardization of expression analyses in the same plant. A search for a candidate enzyme activity not found in plants, which might form the basis of a novel GUS-compatible reporter system, led us to investigate nanH, a Clostridium perfringens gene which encodes the so-called 'small' cytoplasmic sialidase. Expression of the native, AT-rich nanH gene in transgenic plants did not, however, result in detectable sialidase activity. For this reason, a codon-optimized derivative, NAN, was synthesized which possesses a GC content similar to that found in highly expressed plant genes. NAN enzyme activity was expressed at high levels in both stably and transiently transformed cells, possessed kinetic and stability properties similar to those of GUS, and showed optimal activity in GUS buffer. Moreover, NAN and GUS activity could be visualized simultaneously in polyacrylamide gels using the corresponding methylumbelliferone-based substrates, and in whole seedlings and tissue sections using the histochemical substrates 5-bromo-4-chloro-3-indolyl alpha-d-N-acetylneuraminic acid (X-NeuNAc) and 5-bromo-6-chloro-3-indolyl beta-d-glucuronide (X-GlucM), respectively.

  12. Dysregulated Glycoprotein B-Mediated Cell-Cell Fusion Disrupts Varicella-Zoster Virus and Host Gene Transcription during Infection. (United States)

    Oliver, Stefan L; Yang, Edward; Arvin, Ann M


    The highly conserved herpesvirus glycoprotein complex gB/gH-gL mediates membrane fusion during virion entry and cell-cell fusion. Varicella-zoster virus (VZV) characteristically forms multinucleated cells, or syncytia, during the infection of human tissues, but little is known about this process. The cytoplasmic domain of VZV gB (gBcyt) has been implicated in cell-cell fusion regulation because a gB[Y881F] substitution causes hyperfusion. gBcyt regulation is necessary for VZV pathogenesis, as the hyperfusogenic mutant gB[Y881F] is severely attenuated in human skin xenografts. In this study, gBcyt-regulated fusion was investigated by comparing melanoma cells infected with wild-type-like VZV or hyperfusogenic mutants. The gB[Y881F] mutant exhibited dramatically accelerated syncytium formation in melanoma cells caused by fusion of infected cells with many uninfected cells, increased cytoskeleton reorganization, and rapid displacement of nuclei to dense central structures compared to pOka using live-cell confocal microscopy. VZV and human transcriptomes were concurrently investigated using whole transcriptome sequencing (RNA-seq) to identify viral and cellular responses induced when gBcyt regulation was disrupted by the gB[Y881F] substitution. The expression of four vital VZV genes, ORF61 and the genes for glycoproteins gC, gE, and gI, was significantly reduced at 36 h postinfection for the hyperfusogenic mutants. Importantly, hierarchical clustering demonstrated an association of differential gene expression with dysregulated gBcyt-mediated fusion. A subset of Ras GTPase genes linked to membrane remodeling were upregulated in cells infected with the hyperfusogenic mutants. These data implicate gBcyt in the regulation of gB fusion function that, if unmodulated, triggers cellular processes leading to hyperfusion that attenuates VZV infection.

  13. Evolution by Pervasive Gene Fusion in Antibiotic Resistance and Antibiotic Synthesizing Genes

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    Orla Coleman


    Full Text Available Phylogenetic (tree-based approaches to understanding evolutionary history are unable to incorporate convergent evolutionary events where two genes merge into one. In this study, as exemplars of what can be achieved when a tree is not assumed a priori, we have analysed the evolutionary histories of polyketide synthase genes and antibiotic resistance genes and have shown that their history is replete with convergent events as well as divergent events. We demonstrate that the overall histories of these genes more closely resembles the remodelling that might be seen with the children’s toy Lego, than the standard model of the phylogenetic tree. This work demonstrates further that genes can act as public goods, available for re-use and incorporation into other genetic goods.

  14. Acquiring transgenic tobacco plants with insect resistance and glyphosate tolerance by fusion gene transformation. (United States)

    Sun, He; Lang, Zhihong; Zhu, Li; Huang, Dafang


    The advantages of gene 'stacking' or 'pyramiding' are obvious in genetically modified (GM) crops, and several different multi-transgene-stacking methods are available. Using linker peptides for multiple gene transformation is considered to be a good method to meet a variety of needs. In our experiment, the Bt cry1Ah gene, which encodes the insect-resistance protein, and the mG ( 2 ) -epsps gene, which encodes the glyphosate-tolerance protein, were connected by a 2A or LP4/2A linker. Linker 2A is a peptide from the foot-and-mouth disease virus (FMDV) that has self-cleavage activity. LP4 is a peptide from Raphanus sativus seeds that has a recognition site and is cleaved by a protease. LP4/2A is a hybrid peptide that contains the first 9 amino acids of LP4 and 20 amino acids from 2A. We used the linker peptide to construct four coordinated expression vectors: pHAG, pHLAG, pGAH and pGLAH. Two single gene expression vectors, pSAh and pSmG(2), were used as controls. The six expression vectors and the pCAMBIA2301 vector were transferred into tobacco by Agrobacterium tumefaciens-mediated transformation, and 529 transformants were obtained. Molecular detection and bioassay detection data demonstrated that the transgenic tobaccos possessed good pest resistance and glyphosate tolerance. The two genes in the fusion vector were expressed simultaneously. The plants with the genes linked by the LP4/2A peptide showed better pest resistance and glyphosate tolerance than the plants with the genes linked by 2A. The expression level of the two genes linked by LP4/2A was not significantly different from the single gene vector. Key message The expression level of the two genes linked by LP4/2A was higher than those linked by 2A and was not significantly different from the single gene vector.

  15. Anti-colorectal cancer effect of interleukin-2 and interferon-β fusion gene driven by carcinoembryonic antigen promoter

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    Wang Y


    Full Text Available Yan Wang, Mengchun Wang, Yan LiDepartment of Gastroenterology, Shengjing Hospital of China Medical University, Shenyang, People’s Republic of ChinaAbstract: This study was designed to investigate the antitumor effects of combined interleukin-2/interferon-β-based gene therapy in colorectal cancer. Transfection of the fusion gene expression plasmid induced significant apoptosis of Lovo cells. Additionally, the fusion gene exhibited strong inhibitory activity against tumor growth and apoptosis when being injected into the nude mice implanted with human colon cancer cells. Furthermore, the tail-vein injection showed a more notable effect than direct injection into tumor. These results suggest that the combined interleukin-2/interferon-β-based gene therapy with the carcinoembryonic antigen promoter might be an effective antitumor strategy.Keywords: apoptosis, interferon-β, interleukin-2, antitumor, combined gene therapy

  16. Rapid and sensitive lentivirus vector-based conditional gene expression assay to monitor and quantify cell fusion activity.

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    Manuel A F V Gonçalves

    Full Text Available Cell-to-cell fusion is involved in multiple fundamental biological processes. Prominent examples include osteoclast and giant cell formation, fertilization and skeletal myogenesis which involve macrophage, sperm-egg and myoblast fusion, respectively. Indeed, the importance of cell fusion is underscored by the wide range of homeostatic as well as pathologic processes in which it plays a key role. Therefore, rapid and sensitive systems to trace and measure cell fusion events in various experimental systems are in demand. Here, we introduce a bipartite cell fusion monitoring system based on a genetic switch responsive to the site-specific recombinase FLP. To allow flexible deployment in both dividing as well as non-dividing cell populations, inducer and reporter modules were incorporated in lentivirus vector particles. Moreover, the recombinase-inducible transcription units were designed in such a way as to minimize basal activity and chromosomal position effects in the "off" and "on" states, respectively. The lentivirus vector-based conditional gene expression assay was validated in primary human mesenchymal stem cells and in a differentiation model based on muscle progenitor cells from a Duchenne muscular dystrophy patient using reporter genes compatible with live- and single-cell imaging and with whole population measurements. Using the skeletal muscle cell differentiation model, we showed that the new assay displays low background activity, a 2-log dynamic range, high sensitivity and is amenable to the investigation of cell fusion kinetics. The utility of the bipartite cell fusion monitoring system was underscored by a study on the impact of drug- and RNAi-mediated p38 MAPK inhibition on human myocyte differentiation. Finally, building on the capacity of lentivirus vectors to readily generate transgenic animals the present FLP-inducible system should be adaptable, alone or together with Cre/loxP-based assays, to cell lineage tracing and

  17. Fusion gene and splice variant analyses in liquid biopsies of lung cancer patients (United States)

    Giménez-Capitán, Ana; Karachaliou, Niki; Pérez-Rosado, Ana; Viteri, Santiago; Morales-Espinosa, Daniela; Rosell, Rafael


    Obtaining a biopsy of solid tumors requires invasive procedures that strongly limit patient compliance. In contrast, a blood extraction is safe, can be performed at many time points during the course disease and encourages appropriate therapy modifications, potentially improving the patient’s clinical outcome and quality of life. Fusion of the tyrosine kinase genes anaplastic lymphoma kinase (ALK), C-ROS oncogen 1 (ROS 1), rearranged during transfection (RET) and neurotrophic tyrosine kinase 1 (NTRK1) occur in 1–5% of lung adenocarcinomas and constitute therapeutic targets for tyrosine kinase inhibitors. In addition, a MET splicing variant of exon 14, has been reported in 2–4% of lung adenocarcinoma and recent studies suggests that targeted therapies inhibiting MET signaling would be beneficial for patients with this alteration. In this review, we will summarize the new techniques recently developed to detect ALK, RET, ROS and NTRK1 fusions and MET exon 14 splicing variant in liquid biopsy using plasma, serum, circulating tumor cells (CTCs), platelets and exosomes as starting material. PMID:27826534

  18. Discovery and characterization of a novel CCND1/MRCK gene fusion in mantle cell lymphoma

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    Chioniso Patience Masamha


    Full Text Available Abstract The t(11;14 translocation resulting in constitutive cyclin D1 expression is an early event in mantle cell lymphoma (MCL transformation. Patients with a highly proliferative phenotype produce cyclin D1 transcripts with truncated 3′UTRs that evade miRNA regulation. Here, we report the recurrence of a novel gene fusion in MCL cell lines and MCL patient isolates that consists of the full protein coding region of cyclin D1 (CCND1 and a 3′UTR consisting of sequences from both the CCND1 3′UTR and myotonic dystrophy kinase-related Cdc42-binding kinase's (MRCK intron one. The resulting CCND1/MRCK mRNA is resistant to CCND1-targeted miRNA regulation, and targeting the MRCK region of the chimeric 3′UTR with siRNA results in decreased CCND1 levels.

  19. Inducible expression of a fusion gene encoding two proteinase inhibitors leads to insect and pathogen resistance in transgenic rice. (United States)

    Quilis, Jordi; López-García, Belén; Meynard, Donaldo; Guiderdoni, Emmanuel; San Segundo, Blanca


    Plant proteinase inhibitors (PIs) are considered as candidates for increased insect resistance in transgenic plants. Insect adaptation to PI ingestion might, however, compromise the benefits received by transgenic expression of PIs. In this study, the maize proteinase inhibitor (MPI), an inhibitor of insect serine proteinases, and the potato carboxypeptidase inhibitor (PCI) were fused into a single open reading frame and introduced into rice plants. The two PIs were linked using either the processing site of the Bacillus thuringiensis Cry1B precursor protein or the 2A sequence from the foot-and-mouth disease virus (FMDV). Expression of each fusion gene was driven by the wound- and pathogen-inducible mpi promoter. The mpi-pci fusion gene was stably inherited for at least three generations with no penalty on plant phenotype. An important reduction in larval weight of Chilo suppressalis fed on mpi-pci rice, compared with larvae fed on wild-type plants, was observed. Expression of the mpi-pci fusion gene confers resistance to C. suppressalis (striped stem borer), one of the most important insect pest of rice. The mpi-pci expression systems described may represent a suitable strategy for insect pest control, better than strategies based on the use of single PI genes, by preventing insect adaptive responses. The rice plants expressing the mpi-pci fusion gene also showed enhanced resistance to infection by the fungus Magnaporthe oryzae, the causal agent of the rice blast disease. Our results illustrate the usefulness of the inducible expression of the mpi-pci fusion gene for dual resistance against insects and pathogens in rice plants.

  20. Species based synonymous codon usage in fusion protein gene of Newcastle disease virus.

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    Chandra Shekhar Kumar

    Full Text Available Newcastle disease is highly pathogenic to poultry and many other avian species. However, the Newcastle disease virus (NDV has also been reported from many non-avian species. The NDV fusion protein (F is a major determinant of its pathogenicity and virulence. The functionalities of F gene have been explored for the development of vaccine and diagnostics against NDV. Although the F protein is well studied but the codon usage and its nucleotide composition from NDV isolated from different species have not yet been explored. In present study, we have analyzed the factors responsible for the determination of codon usage in NDV isolated from four major avian host species. The F gene of NDV is analyzed for its base composition and its correlation with the bias in codon usage. Our result showed that random mutational pressure is responsible for codon usage bias in F protein of NDV isolates. Aromaticity, GC3s, and aliphatic index were not found responsible for species based synonymous codon usage bias in F gene of NDV. Moreover, the low amount of codon usage bias and expression level was further confirmed by a low CAI value. The phylogenetic analysis of isolates was found in corroboration with the relatedness of species based on codon usage bias. The relationship between the host species and the NDV isolates from the host does not represent a significant correlation in our study. The present study provides a basic understanding of the mechanism involved in codon usage among species.

  1. Analysis of Magnaporthe oryzae genome reveals a fungal effector, which is able to induce resistance response in transgenic rice line containing resistance gene, Pi54

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    Soham Ray


    Full Text Available Rice blast caused by Magnaporthe oryzae is one of the most important diseases of rice. Pi54, a rice gene that imparts resistance to M. oryzae isolates prevalent in India, was already cloned but its avirulent counterpart in the pathogen was not known.. After decoding the whole genome of an avirulent isolate of M. oryzae, we predicted 11440 protein coding genes and then identified four candidate effector proteins which are exclusively expressed in the infectious structure, appresoria. In silico protein modeling followed by interaction analysis between Pi54 protein model and selected four candidate effector proteins models revealed that Mo-01947_9 protein model encoded by a gene located at chromosome 4 of M. oryzae, interacted best at the Leucine Rich Repeat domain of Pi54 protein model. Yeast-two-hybrid analysis showed that Mo-01947_9 protein physically interacts with Pi54 protein. Nicotiana benthamiana leaf infiltration assay confirmed induction of hypersensitive response in the presence of Pi54 gene in a heterologous system. Genetic complementation test also proved that Mo-01947_9 protein induces avirulence response in the pathogen in the presence of Pi54 gene. Here, we report identification and cloning of a new fungal effector gene which interacts with resistance gene Pi54 gene in rice.

  2. Drosophila Erect wing (Ewg) controls mitochondrial fusion during muscle growth and maintenance by regulation of the Opa1-like gene. (United States)

    Rai, Mamta; Katti, Prasanna; Nongthomba, Upendra


    Mitochondrial biogenesis and morphological changes are associated with tissue-specific functional demand, but the factors and pathways that regulate these processes have not been completely identified. A lack of mitochondrial fusion has been implicated in various developmental and pathological defects. The spatiotemporal regulation of mitochondrial fusion in a tissue such as muscle is not well understood. Here, we show in Drosophila indirect flight muscles (IFMs) that the nuclear-encoded mitochondrial inner membrane fusion gene, Opa1-like, is regulated in a spatiotemporal fashion by the transcription factor/co-activator Erect wing (Ewg). In IFMs null for Ewg, mitochondria undergo mitophagy and/or autophagy accompanied by reduced mitochondrial functioning and muscle degeneration. By following the dynamics of mitochondrial growth and shape in IFMs, we found that mitochondria grow extensively and fuse during late pupal development to form the large tubular mitochondria. Our evidence shows that Ewg expression during early IFM development is sufficient to upregulate Opa1-like, which itself is a requisite for both late pupal mitochondrial fusion and muscle maintenance. Concomitantly, by knocking down Opa1-like during early muscle development, we show that it is important for mitochondrial fusion, muscle differentiation and muscle organization. However, knocking down Opa1-like, after the expression window of Ewg did not cause mitochondrial or muscle defects. This study identifies a mechanism by which mitochondrial fusion is regulated spatiotemporally by Ewg through Opa1-like during IFM differentiation and growth.

  3. Evolutionary Origins of the Eukaryotic Shikimate Pathway: Gene Fusions, Horizontal Gene Transfer, and Endosymbiotic Replacements†



    Currently the shikimate pathway is reported as a metabolic feature of prokaryotes, ascomycete fungi, apicomplexans, and plants. The plant shikimate pathway enzymes have similarities to prokaryote homologues and are largely active in chloroplasts, suggesting ancestry from the plastid progenitor genome. Toxoplasma gondii, which also possesses an alga-derived plastid organelle, encodes a shikimate pathway with similarities to ascomycete genes, including a five-enzyme pentafunctional arom. These ...

  4. FISH技术在儿童ALL常见融合基因检测中的应用价值%Value of fluorescence in situ hybridization in Identification of fusion gene in pediatric cases with acute lymphoblastic leukemia

    Institute of Scientific and Technical Information of China (English)

    徐雪琴; 林小玲; 谢番妮; 郑昭科; 唐少华


    Objective:To discuss the role of interphase fluorescence in situ hybridization (I-FISH) in TELAML1 and BCR/ABL fusion gene detection in pediatric cases with acute lymphoblastic leukemia.Methods:Dual color I-FISH and conventional cytogenetic analysis (CCA) were combined to detect t(12;21) TEL/AML1 and t(9;22) BCR/ABL fusion gene in bone marrow mononuclear cells from 35 newly and 4 oddly pediatric ALL patients.Results:Twelve cases were found the TEL/AML1 fusion gene by FISH (30.8%).But no dubious t (12; 21) gene was detected by CCA.Four cases were found the BCR/ABL fusion gene by FISH (10.3%).Only 1 case was found dubious t (9;22) gene by CCA,the incidence was 2.5%.TEL/AML1 fusion gene was more than BCR/ABL fusion gene in pediatric ALL patients and showed lower peripheral tumor load in diagnosis.Conclusion:Dual color IFISH is more sensitive and specific than conventional,cytogenetic analysis (CCA)in the identification of TELAML1 and BCP/ABL fusion gene.So it is playing a significant role in diagnosis and therapy of pediatric cases with acute lymphoblastic leukemia.%目的:探讨双色间期荧光原位杂交(I-FISH)技术在儿童急性淋巴细胞白血病TEL/AML1和BCR/ABL融合基因检测中的应用价值.方法:联合双色间期荧光原位杂交(I-FISH)技术和常规染色体核型分析技术(CCA)对35例儿童初治ALL和4例儿童复发ALL的骨髓有核细胞进行t(12;21) TEL/AML1和t(9;22)BCR/ABL融合基因进行检测.结果:12例患几经FISH检测发现TEL/AML1融合基因,占总病例的30.8%;而CCA检测均未发现有可疑t(12;21).4例患儿经FISH检测发现BCR/ABL融合基因,占总病例的10.3%;而CCA检测发现1例可疑t(9;22),占总病例的2.5%.TEL/AML1在儿童ALL中阳性率较BCR/ABL高,该融合基因阳性的患儿病情较轻.结论:FISH技术较常规染色体核型分析技术特异性强、敏感度高,可以有效检测出TEL/AML1和BCR/ABL融合基因,从而为儿童ALL的诊断和个体化治疗方案提供重要的依据.

  5. Gene fusion detection in formalin-fixed paraffin-embedded benign fibrous histiocytomas using fluorescence in situ hybridization and RNA sequencing. (United States)

    Walther, Charles; Hofvander, Jakob; Nilsson, Jenny; Magnusson, Linda; Domanski, Henryk A; Gisselsson, David; Tayebwa, Johnbosco; Doyle, Leona A; Fletcher, Christopher D M; Mertens, Fredrik


    Benign fibrous histiocytomas (FH) can be subdivided into several morphological and clinical subgroups. Recently, gene fusions involving either one of two protein kinase C genes (PRKCB and PRKCD) or the ALK gene were described in FH. We here wanted to evaluate the frequency of PRKCB and PRKCD gene fusions in FH. Using interphase fluorescence in situ hybridization on sections from formalin-fixed paraffin-embedded (FFPE) tumors, 36 cases could be analyzed. PRKCB or PRKCD rearrangements were seen in five tumors: 1/7 regular, 0/3 aneurysmal, 0/6 cellular, 2/7 epithelioid, 0/1 atypical, 2/10 deep, and 0/2 metastatic lesions. We also evaluated the status of the ALK gene in selected cases, finding rearrangements in 3/7 epithelioid and 0/1 atypical lesions. To assess the gene fusion status of FH further, deep sequencing of RNA (RNA-Seq) was performed on FFPE tissue from eight cases with unknown gene fusion status, as well as on two FH and six soft tissue sarcomas with known gene fusions; of the latter eight positive controls, the expected fusion transcript was found in all but one, while 2/8 FH with unknown genetic status showed fusion transcripts, including a novel KIRREL/PRKCA chimera. Thus, also a third member of the PRKC family is involved in FH tumorigenesis. We conclude that gene fusions involving PRKC genes occur in several morphological (regular, cellular, aneurysmal, epithelioid) and clinical (cutaneous, deep) subsets of FH, but they seem to account for only a minority of the cases. In epithelioid lesions, however, rearrangements of PRKC or ALK were seen, as mutually exclusive events, in the majority (5/7) of cases. Finally, the study also shows that RNA-Seq is a promising tool for identifying gene fusions in FFPE tissues.

  6. Hh/Gli antagonist in acute myeloid leukemia with CBFA2T3-GLIS2 fusion gene

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    Riccardo Masetti


    Full Text Available Abstract Background CBFA2T3-GLIS2 is a fusion gene found in 17% of non-Down syndrome acute megakaryoblastic leukemia (non-DS AMKL, FAB M7 and in 8% of pediatric cytogenetically normal acute myeloid leukemia (CN-AML, in association with several French-American-British (FAB subtypes. Children with AML harboring this aberration have a poor outcome, regardless of the FAB subtype. This fusion gene drives a peculiar expression pattern and leads to overexpression of some of Hedgehog-related genes. GLI-similar protein 2 (GLIS2 is closely related to the GLI family, the final effectors of classic Hedgehog pathway. These observations lend compelling support to the application of GLI inhibitors in the treatment of AML with the aberration CBFA2T3-GLIS2. GANT61 is, nowadays, the most potent inhibitor of GLI family proteins. Methods We exposed to GANT61 AML cell lines and primary cells positive and negative for CBFA2T3-GLIS2 and analyzed the effect on cellular viability, induction of apoptosis, cell cycle, and expression profile. Results As compared to AML cells without GLIS2 fusion, GANT61 exposure resulted in higher sensitivity of both cell lines and primary AML cells carrying CBFA2T3-GLIS2 to undergo apoptosis and G1 cell cycle arrest. Remarkably, gene expression studies demonstrated downregulation of GLIS2-specific signature genes in both treated cell lines and primary cells, in comparison with untreated cells. Moreover, chromatin immunoprecipitation analysis revealed direct regulation by GLIS2 chimeric protein of DNMT1 and DNMT3B, two genes implicated in important epigenetic functions. Conclusions Our findings indicate that the GLI inhibitor GANT61 may be used to specifically target the CBFA2T3-GLIS2 fusion gene in pediatric AML.

  7. VviAPRT3 and VviFSEX: Two Genes Involved in Sex Specification Able to Distinguish Different Flower Types in Vitis (United States)

    Coito, João L.; Ramos, Miguel J. N.; Cunha, Jorge; Silva, Helena G.; Amâncio, Sara; Costa, Maria M. R.; Rocheta, Margarida


    Vitis vinifera vinifera is a hermaphrodite subspecies, while its ancestor, Vitis vinifera sylvestris, is dioecious. We have identified two genes that together allow the discrimination between male, female and hermaphrodite Vitis plants. The sex locus region on chromosome 2 was screened resulting in the discovery of a new gene, VviFSEX. The same screening revealed another gene, VviAPRT3, located in the sex region, that be used as a sex marker. Both genes are good candidates to be involved in flower sex differentiation in grapevine. To assess their role in sex specification, spatial and temporal expression analysis was performed. The expression of VviFSEX is detected in petals, stamens and carpel primordia of all flower types, making its putative function unclear; however, female plants display a single allele for this gene, while male and hermaphrodites display two alleles. On the other hand, the specific expression of VviAPRT3 in the carpel primordial of male plants suggests a possible role in the abortion of pistil structures. We propose a model to explain the carpel abortion in male flowers and the absence of stamen viability in female flowers. In addition, this work reinforces the presence of a sex locus on Vitis chromosome 2. PMID:28197167

  8. Establishment of cells to monitor Microprocessor through fusion genes of microRNA and GFP. (United States)

    Tsutsui, Motomu; Hasegawa, Hitoki; Adachi, Koichi; Miyata, Maiko; Huang, Peng; Ishiguro, Naoki; Hamaguchi, Michinari; Iwamoto, Takashi


    Microprocessor, the complex of Drosha and DGCR8, promotes the processing of primary microRNA to precursor microRNA, which is a crucial step for microRNA maturation. So far, no convenient assay systems have been developed for observing this step in vivo. Here we report the establishment of highly sensitive cellular systems where we can visually monitor the function of Microprocessor. During a series of screening of transfectants with fusion genes of the EGFP cDNA and primary microRNA genes, we have obtained certain cell lines where introduction of siRNA against DGCR8 or Drosha strikingly augments GFP signals. In contrast, these cells have not responded to Dicer siRNA; thus they have a unique character that GFP signals should be negatively and specifically correlated to the action of Microprocessor among biogenesis of microRNA. These cell lines can be useful tools for real-time analysis of Microprocessor action in vivo and identifying its novel modulators.

  9. Comprehensive genomic analysis of malignant pleural mesothelioma identifies recurrent mutations, gene fusions and splicing alterations. (United States)

    Bueno, Raphael; Stawiski, Eric W; Goldstein, Leonard D; Durinck, Steffen; De Rienzo, Assunta; Modrusan, Zora; Gnad, Florian; Nguyen, Thong T; Jaiswal, Bijay S; Chirieac, Lucian R; Sciaranghella, Daniele; Dao, Nhien; Gustafson, Corinne E; Munir, Kiara J; Hackney, Jason A; Chaudhuri, Amitabha; Gupta, Ravi; Guillory, Joseph; Toy, Karen; Ha, Connie; Chen, Ying-Jiun; Stinson, Jeremy; Chaudhuri, Subhra; Zhang, Na; Wu, Thomas D; Sugarbaker, David J; de Sauvage, Frederic J; Richards, William G; Seshagiri, Somasekar


    We analyzed transcriptomes (n = 211), whole exomes (n = 99) and targeted exomes (n = 103) from 216 malignant pleural mesothelioma (MPM) tumors. Using RNA-seq data, we identified four distinct molecular subtypes: sarcomatoid, epithelioid, biphasic-epithelioid (biphasic-E) and biphasic-sarcomatoid (biphasic-S). Through exome analysis, we found BAP1, NF2, TP53, SETD2, DDX3X, ULK2, RYR2, CFAP45, SETDB1 and DDX51 to be significantly mutated (q-score ≥ 0.8) in MPMs. We identified recurrent mutations in several genes, including SF3B1 (∼2%; 4/216) and TRAF7 (∼2%; 5/216). SF3B1-mutant samples showed a splicing profile distinct from that of wild-type tumors. TRAF7 alterations occurred primarily in the WD40 domain and were, except in one case, mutually exclusive with NF2 alterations. We found recurrent gene fusions and splice alterations to be frequent mechanisms for inactivation of NF2, BAP1 and SETD2. Through integrated analyses, we identified alterations in Hippo, mTOR, histone methylation, RNA helicase and p53 signaling pathways in MPMs.

  10. Therapeutic efficacy of a tuberculosis DNA vaccine encoding heat shock protein 65 of Mycobacterium tuberculosis and the human interleukin 2 fusion gene. (United States)

    Changhong, Shi; Hai, Zhang; Limei, Wang; Jiaze, An; Li, Xi; Tingfen, Zhang; Zhikai, Xu; Yong, Zhao


    Use of therapeutic DNA vaccines is a promising strategy against tuberculosis (TB), however, their immunogenicity still needs to be improved. In this study, a plasmid DNA vaccine expressing heat shock protein 65 (HSP65) and the human interleukin 2 (IL-2) fusion gene was constructed. Immune responses induced by the vaccine in the mice and protection against Mycobacterium tuberculosis (MTB) were investigated, along with the therapeutic effect of the DNA vaccine on tuberculosis in mice. Administration of the HSP65-IL-2-DNA vaccine enhanced Th1-type cellular responses by producing greater amounts of interferon-gamma (IFN-gamma) and IL-2 with a higher titer of antigen-specific anti-Hsp65 IgG2a. Compared with the Bacille Calmette-Guérin (BCG) vaccine, the DNA vaccine was able to evoke both CD4 and CD8 T-cell responses, with an especially high percentage of CD8 T-cells. The DNA vaccine was also able to induce high antigen-specific cytotoxicity activity against target cells. When the mice were challenged with virulent MTB H37Rv, a dramatic decrease in the numbers of MTB colony forming units in the spleen and lungs was observed in the mice immunized with HSP65-IL-2-DNA (P<0.05). Meanwhile, the bacterial numbers in TB infected mice treated with the DNA vaccine were also significantly reduced. The protective and therapeutic effects of the HSP65-IL-2-DNA vaccine in the spleen and lungs were superior to that of the HSP65-DNA vaccine (P<0.05). These results suggest that the DNA vaccine expression of IL-2 and the HSP65 fusion gene enhances the immunogenicity and protective as well as therapeutic effects of the HSP65-DNA vaccine against TB in mice by improving the Th1-type response.

  11. Limited inter- and intra-patient sequence diversity of the genetic lineage A human metapneumovirus fusion gene

    DEFF Research Database (Denmark)

    Winther, Thilde Nordmann; Madsen, Chris D; Pedersen, Anders;


    Human metapneumovirus (hMPV) is associated with respiratory tract illness especially in young children. Two hMPV genetic lineages, A and B, and four sublineages A1, A2 and B1, B2 have been defined. Infection with hMPV occurs through membrane fusion mediated by the hMPV fusion (F) protein. In this...... diversity observed lay emphasis on the hMPV F gene as a putative target for future vaccine development....... been infected with at least two viruses. Several independent viruses contained premature stop codons in exactly identical positions resulting in truncated fusion proteins. Possibly this is a mechanism for immune system evasion. The F protein is a major antigenic determinant, and the limited sequence...

  12. Limited inter- and intra-patient sequence diversity of the genetic lineage a human metapneumovirus fusion gene

    DEFF Research Database (Denmark)

    Winther, T.N.; Madsen, C.D.; Pedersen, Anders Gorm;


    Human metapneumovirus (hMPV) is associated with respiratory tract illness especially in young children. Two hMPV genetic lineages, A and B, and four sublineages A1, A2 and B1, B2 have been defined. Infection with hMPV occurs through membrane fusion mediated by the hMPV fusion (F) protein. In this...... diversity observed lay emphasis on the hMPV F gene as a putative target for future vaccine development....... been infected with at least two viruses. Several independent viruses contained premature stop codons in exactly identical positions resulting in truncated fusion proteins. Possibly this is a mechanism for immune system evasion. The F protein is a major antigenic determinant, and the limited sequence...

  13. Development of RNA-FISH Assay for Detection of Oncogenic FGFR3-TACC3 Fusion Genes in FFPE Samples (United States)

    Kojima, Takahiro; Nishimura, Kouichi; Kandori, Shuya; Kawahara, Takashi; Yoshino, Takayuki; Ueno, Satoshi; Iizumi, Yuichi; Mitsuzuka, Koji; Arai, Yoichi; Tsuruta, Hiroshi; Habuchi, Tomonori; Kobayashi, Takashi; Matsui, Yoshiyuki; Ogawa, Osamu; Sugimoto, Mikio; Kakehi, Yoshiyuki; Nagumo, Yoshiyuki; Tsutsumi, Masakazu; Oikawa, Takehiro; Kikuchi, Koji; Nishiyama, Hiroyuki


    Introduction and Objectives Oncogenic FGFR3-TACC3 fusions and FGFR3 mutations are target candidates for small molecule inhibitors in bladder cancer (BC). Because FGFR3 and TACC3 genes are located very closely on chromosome 4p16.3, detection of the fusion by DNA-FISH (fluorescent in situ hybridization) is not a feasible option. In this study, we developed a novel RNA-FISH assay using branched DNA probe to detect FGFR3-TACC3 fusions in formaldehyde-fixed paraffin-embedded (FFPE) human BC samples. Materials and Methods The RNA-FISH assay was developed and validated using a mouse xenograft model with human BC cell lines. Next, we assessed the consistency of the RNA-FISH assay using 104 human BC samples. In this study, primary BC tissues were stored as frozen and FFPE tissues. FGFR3-TACC3 fusions were independently detected in FFPE sections by the RNA-FISH assay and in frozen tissues by RT-PCR. We also analyzed the presence of FGFR3 mutations by targeted sequencing of genomic DNA extracted from deparaffinized FFPE sections. Results FGFR3-TACC3 fusion transcripts were identified by RNA-FISH and RT-PCR in mouse xenograft FFPE tissues using the human BC cell lines RT112 and RT4. These cell lines have been reported to be fusion-positive. Signals for FGFR3-TACC3 fusions by RNA-FISH were positive in 2/60 (3%) of non-muscle-invasive BC (NMIBC) and 2/44 (5%) muscle-invasive BC (MIBC) patients. The results of RT-PCR of all 104 patients were identical to those of RNA-FISH. FGFR3 mutations were detected in 27/60 (45%) NMIBC and 8/44 (18%) MIBC patients. Except for one NMIBC patient, FGFR3 mutation and FGFR3-TACC3 fusion were mutually exclusive. Conclusions We developed an RNA-FISH assay for detection of the FGFR3-TACC3 fusion in FFPE samples of human BC tissues. Screening for not only FGFR3 mutations, but also for FGFR3-TACC3 fusion transcripts has the potential to identify additional patients that can be treated with FGFR inhibitors. PMID:27930669

  14. Construction and Expression of Methionine-rich and Lysine-rich Fusion Gene inBacillus natto

    Institute of Scientific and Technical Information of China (English)

    Zhang Shuang; Luo Chao-chao; Wu Cai-xia; Gao Xue-jun


    Methionine and lysine are restrictive essential amino acids of livestock, they are also the most attentive indexes in the feed production to carry out the quality control and quality evaluation. Their contents in feed directly affect livestock protein synthesis. Bacillus natto has excellent probiotic properties. In this experiment, we used the genetic engineering method, fusion PCR technique, to connect methionine-rich gene (zein) from maize endosperm protein with lysine-rich gene (Cflr) from the pepper anther, then the fusion gene was inserted into the expression vector pHT43, and the recombinant plasmid pHT43/zein-Cflr was constructed. The recombinant plasmid was transferred intoBacillus natto, and induced by IPTG for the expression of the fusion gene. We found an apparent band at 40 ku site for the recombinant strain by SDS-PAGE. The contents of methionine and lysine were individually detected with HPLC, the quantities of methionine and lysine in the recombinant strain increased by 18.37% and 24.68% than the wild one, respectively. We also verified the stability of the recombinant bacterium during passaging, and found the stability was 100%. This study provided research-basis for the application of the recombinedBacillus nattoas feed additive.

  15. Gene fusions and gene duplications: relevance to genomic annotation and functional analysis

    Directory of Open Access Journals (Sweden)

    Riley Monica


    Full Text Available Abstract Background Escherichia coli a model organism provides information for annotation of other genomes. Our analysis of its genome has shown that proteins encoded by fused genes need special attention. Such composite (multimodular proteins consist of two or more components (modules encoding distinct functions. Multimodular proteins have been found to complicate both annotation and generation of sequence similar groups. Previous work overstated the number of multimodular proteins in E. coli. This work corrects the identification of modules by including sequence information from proteins in 50 sequenced microbial genomes. Results Multimodular E. coli K-12 proteins were identified from sequence similarities between their component modules and non-fused proteins in 50 genomes and from the literature. We found 109 multimodular proteins in E. coli containing either two or three modules. Most modules had standalone sequence relatives in other genomes. The separated modules together with all the single (un-fused proteins constitute the sum of all unimodular proteins of E. coli. Pairwise sequence relationships among all E. coli unimodular proteins generated 490 sequence similar, paralogous groups. Groups ranged in size from 92 to 2 members and had varying degrees of relatedness among their members. Some E. coli enzyme groups were compared to homologs in other bacterial genomes. Conclusion The deleterious effects of multimodular proteins on annotation and on the formation of groups of paralogs are emphasized. To improve annotation results, all multimodular proteins in an organism should be detected and when known each function should be connected with its location in the sequence of the protein. When transferring functions by sequence similarity, alignment locations must be noted, particularly when alignments cover only part of the sequences, in order to enable transfer of the correct function. Separating multimodular proteins into module units makes

  16. Fusion of ZMYND8 and RELA genes in acute erythroid leukemia

    DEFF Research Database (Denmark)

    Panagopoulos, Ioannis; Micci, Francesca; Thorsen, Jim


    Acute erythroid leukemia was diagnosed in a 4-month-old boy. Cytogenetic analysis of bone marrow (BM) cells showed a t(11;20)(p11;q11) translocation. RNA extracted from the BM was sequenced and analyzed for fusion transcripts using the software FusionMap. A ZMYND8-RELA fusion was ranked first. RT...... the translocation. The putative ZMYND8-RELA fusion protein contains the Zinc-PHD finger domain, a bromodomain, a PWWP domain, a MYND type of zinc finger of ZMYND8, and the entire RELA protein, indicating that it might act leukemogenically by influencing several cellular processes including the NF-kappa-B pathway....

  17. ZNF384-related fusion genes define a subgroup of childhood B-cell precursor acute lymphoblastic leukemia with a characteristic immunotype (United States)

    Hirabayashi, Shinsuke; Ohki, Kentaro; Nakabayashi, Kazuhiko; Ichikawa, Hitoshi; Momozawa, Yukihide; Okamura, Kohji; Yaguchi, Akinori; Terada, Kazuki; Saito, Yuya; Yoshimi, Ai; Ogata-Kawata, Hiroko; Sakamoto, Hiromi; Kato, Motohiro; Fujimura, Junya; Hino, Moeko; Kinoshita, Akitoshi; Kakuda, Harumi; Kurosawa, Hidemitsu; Kato, Keisuke; Kajiwara, Ryosuke; Moriwaki, Koichi; Morimoto, Tsuyoshi; Nakamura, Kozue; Noguchi, Yasushi; Osumi, Tomoo; Sakashita, Kazuo; Takita, Junko; Yuza, Yuki; Matsuda, Koich; Yoshida, Teruhiko; Matsumoto, Kenji; Hata, Kenichiro; Kubo, Michiaki; Matsubara, Yoichi; Fukushima, Takashi; Koh, Katsuyoshi; Manabe, Atsushi; Ohara, Akira; Kiyokawa, Nobutaka


    Fusion genes involving ZNF384 have recently been identified in B-cell precursor acute lymphoblastic leukemia, and 7 fusion partners have been reported. We further characterized this type of fusion gene by whole transcriptome sequencing and/or polymerase chain reaction. In addition to previously reported genes, we identified BMP2K as a novel fusion partner for ZNF384. Including the EP300-ZNF384 that we reported recently, the total frequency of ZNF384-related fusion genes was 4.1% in 291 B-cell precursor acute lymphoblastic leukemia patients enrolled in a single clinical trial, and TCF3-ZNF384 was the most recurrent, with a frequency of 2.4%. The characteristic immunophenotype of weak CD10 and aberrant CD13 and/or CD33 expression was revealed to be a common feature of the leukemic cells harboring ZNF384-related fusion genes. The signature gene expression profile in TCF3-ZNF384-positive patients was enriched in hematopoietic stem cell features and related to that of EP300-ZNF384-positive patients, but was significantly distinct from that of TCF3-PBX1-positive and ZNF384-fusion-negative patients. However, clinical features of TCF3-ZNF384-positive patients are markedly different from those of EP300-ZNF384-positive patients, exhibiting higher cell counts and a younger age at presentation. TCF3-ZNF384-positive patients revealed a significantly poorer steroid response and a higher frequency of relapse, and the additional activating mutations in RAS signaling pathway genes were detected by whole exome analysis in some of the cases. Our observations indicate that ZNF384-related fusion genes consist of a distinct subgroup of B-cell precursor acute lymphoblastic leukemia with a characteristic immunophenotype, while the clinical features depend on the functional properties of individual fusion partners. PMID:27634205

  18. Analysis of KIAA1549-BRAF fusion gene expression and IDH1/IDH2 mutations in low grade pediatric astrocytomas. (United States)

    Cruz, Gabriela Rampazzo; Dias Oliveira, Indhira; Moraes, Laís; Del Giudice Paniago, Mário; de Seixas Alves, Maria Teresa; Capellano, Andrea Maria; Saba-Silva, Nasjla; Cavalheiro, Sérgio; Cerutti, Janete Maria; Toledo, Silvia Regina Caminada


    Low-grade astrocytomas comprise about 30 % of the central nervous system tumors in children. Several investigations have searched a correlation between the BRAF gene fusions alterations and mutations at IDH1 and IDH2 genes in low grade pediatric astrocytomas. This study identified the expression of KIAA1549-BRAF fusion gene and BRAF V600E mutation, mutations at exon 4 of the IDH1 and IDH2 genes in samples of pilocytic astrocytomas (PA) and grade-II astrocytomas (A-II) pediatric patients. The correlation between these alterations and the clinical profile of the patients was also evaluated. Eighty-two samples of low-grade astrocytomas (65 PA and 17 A-II) were analyzed by PCR and sequencing for each of the targets identified. We identified the KIAA1549-BRAF fusion transcript in 45 % of the samples. BRAF V600E and BRAFins598T mutations were detected in 7 and 1 % of the samples, respectively. Mutations in the R132/R172 residues of the IDH1/IDH2 genes were detected in only two samples, and the G105G polymorphism (rs11554137:C>T) was identified in ten patients. Additionally, we observed two mutations out of the usual hotspots at IDH1 and IDH2 genes. We observed a smaller frequency of mutations in IDHs genes than previously described, but since the prior studies were composed of adult or mixed (adults and children) samples, we believe that our results represent a relevant contribution to the growing knowledge in low grade childhood astrocytomas.

  19. Cloned s-Lap Gene Coding Area, Expression and Localizationof s-Lap/GFP Fusion Protein in Mammal Cells

    Institute of Scientific and Technical Information of China (English)

    SONG Yi-shu; SONG Zhi-yu; LI Hong-jun; Wu Yin; BAO Yong-li; TAN Da-peng; LI Yu-xin


    s-Lap is a new gene sequence from pig retinal pigment epithelial(RPE) cells, which was found and cloned in the early period of apoptosis of RPE cells damaged with visible light. We cloned the coding area sequence of the novel gene of s-Lap and constructed its recombinant eukaryotic plasmid pcDNA3.1-GFP/s-lap with the recombinant DNA technique. The expression and localization of s-lap/GFP fusion protein in CHO and B16 cell lines were studied with the instantaneously transfected pcDNA3.1-GFP/s-lap recombinant plasmid. s-Lap/GFP fusion protein can be expressed in CHO and B16 cells with a high rate expression in the nuclei.

  20. A BCR-ABL1 cutoff of 1.5% at 3 months, determined by the GeneXpert system, predicts an optimal response in patients with chronic myeloid leukemia (United States)

    García-Gutiérrez, Valentín; Gómez-Casares, María T.; Puerta, José M.; Alonso-Domínguez, Juan M.; Osorio, Santiago; Hernández-Boluda, Juan C.; Collado, Rosa; Ramírez, María J.; Ibáñez, Fátima; Martín, María L.; Rodríguez-Gambarte, Juan D.; Martínez-Laperche, Carolina; Gómez, Montse; Fiallo, Dolly V.; Redondo, Sara; Rodríguez, Alicia; Ruiz-Nuño, Concepción; Steegmann, Juan L.


    In chronic myeloid leukemia (CML) patients, 3-month BCR-ABL1 levels have consistently been correlated with further outcomes. Monitoring molecular responses in CML using the GeneXpert (Cepheid) platform has shown an optimal correlation with standardized RQ-PCR (IS) when measuring BCR-ABL1 levels lower than 10%, as it is not accurate for values over 10%. The aim of the present study was to determine the predictive molecular value at three months on different outcome variables using the Xpert BCR-ABL1 MonitorTM assay (Xpert BCR-ABL1). We monitored 125 newly diagnosed consecutive CML patients in the chronic phase (CML-CP) using an automated method: Xpert BCR-ABL1. Only 5% of patients did not achieve an optimal response at 3 months, and the 10% BCR-ABL1 cutoff defined by RQ-PCR (IS) methods was unable to identify significant differences in the probabilities of achieving a complete cytogenetic response (CCyR) (50% vs. 87%, p = 0.1) or a major molecular response (MMR) (60% vs. 80%, p = 0.29) by 12 months. In contrast, a cutoff of 1.5% more accurately identified differences in the probabilities of achieving CCyR (98% vs. 54%, p<0.001) and MMR (88% vs. 56%, p<0.001) by 12 months, as well as probabilities of treatment changes (p = 0.005). Therefore, when using the Xpert BCR-ABL1 assay, a cutoff of 1.5% at 3 months could with high probability identify patients able to achieve an optimal response at 12 months. PMID:28278193

  1. Evaluation of reverse transcription-PCR protocols based on the fusion gene for diagnosis of bovine respiratory syncytial virus infections


    Selim A.; Gaede W.


    Bovine respiratory syncytial virus (BRSV) is a pneumovirus in the family paramyxoviridae, is an important cause of acute respiratory disease in postweaning calves and feedlot cattle. The real-time reverse transcriptase PCR protocols were developed to detect BRSV infection in infected animals. The sensitivity of RT-PCR protocols based on fusion gene were evaluated using different Mastermixes such as Qiagen One Step RT-PCR (Qiagen) for conventional RT-PCR, Su...

  2. A human ESC model for MLL-AF4 leukemic fusion gene reveals an impaired early hematopoietic-endothelial specification

    Institute of Scientific and Technical Information of China (English)

    Clara Bueno; Agustin F Femández; Mario F Fraga; Inmaculada Moreno-Gimeno; Deborah Burks; Maria del Carmen Plaza-Calonge; Juan C Rodríguez-Manzaneque; Pablo Menendez; Rosa Montes; Gustavo J Melen; Verónica Ramos-Mejia; Pedro J Real; Verónica Ayllón; Laura Sanchez; Gertrudis Ligero; Iván Gutierrez-Aranda


    The MLL-AF4 fusion gene is a hallmark genomic aberration in high-risk acute lymphoblastic leukemia in inants.Although it is well established that MLL-AF4 arises prenatally during human development,its effects on hematopoieric development in utero remain unexplored.We have created a human-specific cellular system to study early hemato-endothelial development in MLL-AF4-expressing human embryonic stem cells (hESCs).Functional studies,clonal analysis and gene expression profiling reveal that expression of MLL-AF4 in hESCs has a phenotypic,functional and gene expression impact.MLL-AF4 acts as a global transcriptional activator and a positive regulator of homeobox gene expression in hESCs.Functionally,MLL-AF4 enhances the specification of hemogenic precursors from hESCs but strongly impairs further hematopoietic commitment in favor of an endothelial cell fate.MLL-AF4 hESCs are transcriptionally primed to differentiate towards hemogenic precursors prone to endothelial maturation,as reflected by the marked upregulation of master genes associated to vascular-endothelial functions and early hematopoiesis.Furthermore,we report that MLL-AF4 expression is not sufficient to transform hESC-derived hematopoietic cells.This work illustrates how hESCs may provide unique insights into human development and further our understanding of how leukemic fusion genes,known to arise prenatally,regulate human embryonic hematopoietic specification.

  3. [Expression of SET-NUP214 fusion gene in patients with T-cell acute lymphoblastic leukemia and its clinical significance]. (United States)

    Dai, Hai-Ping; Wang, Qian; Wu, Li-Li; Ping, Na-Na; Wu, Chun-Xiao; Xie, Jun-Dan; Pan, Jin-Lan; Xue, Yong-Quan; Wu, De-Pei; Chen, Su-Ning


    This study was aimed to investigate the occurrence and clinical significance of the SET-NUP214 fusion gene in patients with T-cell acute lymphoblastic leukemia (T-ALL), analyse clinical and biological characteristics in this disease. RT-PCR was used to detect the expression of SET-NUP214 fusion gene in 58 T-ALL cases. Interphase FISH and Array-CGH were used to detect the deletion of 9q34. Direct sequencing was applied to detect mutations of PHF6 and NOTCH1. The results showed that 6 out of 58 T-ALL cases (10.3%) were detected to have the SET-NUP214 fusion gene by RT-PCR. Besides T-lineage antigens, expression of CD13 and(or) CD33 were detected in all the 6 cases. Deletions of 9q34 were detected in 4 out of the 6 patients by FISH. Array-CGH results of 3 SET-NUP214 positive T-ALL patients confirmed that this fusion gene was resulted from a cryptic deletion of 9q34.11q34.13. PHF6 and NOTCH1 gene mutations were found in 4 and 5 out of 6 SET-NUP214 positive T-ALL patients, respectively. It is concluded that SET-NUP214 fusion gene is often resulted from del(9)(q34). PHF6 and NOTCH1 mutations may be potential leukemogenic event in SET-NUP214 fusion gene.

  4. Clinicopathological differences between variants of the NAB2-STAT6 fusion gene in solitary fibrous tumors of the meninges and extra-central nervous system. (United States)

    Nakada, Satoko; Minato, Hiroshi; Nojima, Takayuki


    Investigations on the NAB2-STAT6 fusion gene in solitary fibrous tumors (SFTs) and hemangiopericytomas (HPCs) have increased since its discovery in 2013. Although several SFTs reported without NAB2-STAT6 fusion gene analysis, we reviewed 546 SFTs/HPCs with NAB2-STAT6 fusion gene analysis in this study and investigated differences between the gene variants. In total, 452 cases tested positive for the NAB2-STAT6 fusion gene, with more than 40 variants being detected. The most frequent of these were NAB2 exon 6-STAT6 exon 16/17/18 and NAB2 exon 4-STAT6 exon 2/3, with the former occurring most frequently in SFTs in meninges, soft tissues, and head and neck; the latter predominated in SFTs in the pleura and lung. There was no difference between the histology of SFTs and fusion gene variants. A follow-up analysis of SFTs showed that 51 of 202 cases had a recurrence, with 18 of 53 meningeal SFTs having a local recurrence and/or metastasis within 0-19 years. In meninges and soft tissue, SFTs with the NAB2 exon 6-STAT6 exon 16/17/18 tended to recur more frequently than SFTs with the NAB2 exon 4-STAT6 exon 2/3. Clinicopathological data, including yearly follow-ups, are required for meningeal SFTs/HPCs to define the correlation of variants of NAB2-STAT6 fusion gene.

  5. Systematic mapping of occluded genes by cell fusion reveals prevalence and stability of cis-mediated silencing in somatic cells (United States)

    Looney, Timothy J.; Zhang, Li; Chen, Chih-Hsin; Lee, Jae Hyun; Chari, Sheila; Mao, Frank Fuxiang; Pelizzola, Mattia; Zhang, Lu; Lister, Ryan; Baker, Samuel W.; Fernandes, Croydon J.; Gaetz, Jedidiah; Foshay, Kara M.; Clift, Kayla L.; Zhang, Zhenyu; Li, Wei-Qiang; Vallender, Eric J.; Wagner, Ulrich; Qin, Jane Yuxia; Michelini, Katelyn J.; Bugarija, Branimir; Park, Donghyun; Aryee, Emmanuel; Stricker, Thomas; Zhou, Jie; White, Kevin P.; Ren, Bing; Schroth, Gary P.; Ecker, Joseph R.; Xiang, Andy Peng; Lahn, Bruce T.


    Both diffusible factors acting in trans and chromatin components acting in cis are implicated in gene regulation, but the extent to which either process causally determines a cell's transcriptional identity is unclear. We recently used cell fusion to define a class of silent genes termed “cis-silenced” (or “occluded”) genes, which remain silent even in the presence of trans-acting transcriptional activators. We further showed that occlusion of lineage-inappropriate genes plays a critical role in maintaining the transcriptional identities of somatic cells. Here, we present, for the first time, a comprehensive map of occluded genes in somatic cells. Specifically, we mapped occluded genes in mouse fibroblasts via fusion to a dozen different rat cell types followed by whole-transcriptome profiling. We found that occluded genes are highly prevalent and stable in somatic cells, representing a sizeable fraction of silent genes. Occluded genes are also highly enriched for important developmental regulators of alternative lineages, consistent with the role of occlusion in safeguarding cell identities. Alongside this map, we also present whole-genome maps of DNA methylation and eight other chromatin marks. These maps uncover a complex relationship between chromatin state and occlusion. Furthermore, we found that DNA methylation functions as the memory of occlusion in a subset of occluded genes, while histone deacetylation contributes to the implementation but not memory of occlusion. Our data suggest that the identities of individual cell types are defined largely by the occlusion status of their genomes. The comprehensive reference maps reported here provide the foundation for future studies aimed at understanding the role of occlusion in development and disease. PMID:24310002

  6. 融合基因构建方法的建立%Establishment of fusion gene construction

    Institute of Scientific and Technical Information of China (English)

    张丽丽; 李振军; 李景富; 王傲雪


    Overlap extension PCR(OE-PCR) is frequently used for gene fragments fusion based on polymerase chain reaction. PCR, restriction enzyme digestion and ligation are primarily applied to the construction of recombinant vectors. These two techniques are in widespread application in the fields of gene engineering and molecular biology. LhCBF gene (Lhirsutum CRT-binding factor 1), EGFP gene and promoter RD29A sequence were taken as example and successfully obtained LhCBF1-EGFP andRD29A..EGFP fusion genes by using one step and two step OE-PCR, PCR-digestion-ligation method and optimized PCR protocol and system. This study provides significant theoretical basis for OE-PCR, PCR-digestion-ligation and fusion gene construction techniques.%重叠PCR技术是以聚合酶链式反应为基础,将多条DNA片段进行融合;PCR-醇切连接主要应用于基因片段连接及载体构建,这两种方法在基因工程与分子生物学领域已得到广泛应用.以多毛番茄抗冷转录因子LhCBF1基因、EGFP基因及RD29A启动子为例,分别采用一步法重叠PCR、二步法重叠PCR及PCR-酶切连接技术,优化PCR体系及反应条件,成功构建LhCBF1-EGFP及RD29A::EGFP融合基因.研究为重叠PCR及PCR-酶切连接技术的改进及构建融合基因方法的选择提供了有益参考.

  7. TBL1XR1/TP63: a novel recurrent gene fusion in B-cell non-Hodgkin lymphoma | Office of Cancer Genomics (United States)

    Recently, the landscape of single base mutations in diffuse large B-cell lymphoma (DLBCL) was described. Here we report the discovery of a gene fusion between TBL1XR1 and TP63, the only recurrent somatic novel gene fusion identified in our analysis of transcriptome data from 96 DLBCL cases. Based on this cohort and a further 157 DLBCL cases analyzed by FISH, the incidence in de novo germinal center B cell-like (GCB) DLBCL is 5% (6 of 115).

  8. The role of FLI-1-EWS, a fusion gene reciprocal to EWS-FLI-1, in Ewing sarcoma


    Elzi, David J.; Song, Meihua; Houghton, Peter J.; Chen, Yidong; Shiio, Yuzuru


    Ewing sarcoma is a cancer of bone and soft tissue in children that is characterized by a chromosomal translocation involving EWS and an Ets family transcription factor, most commonly FLI-1. The EWS-FLI-1 fusion oncogene is widely believed to play a central role in Ewing sarcoma. The EWS-FLI-1 gene product regulates the expression of a number of genes important for cancer progression, can transform mouse cells such as NIH3T3 and C3H10T1/2, and is necessary for proliferation and tumorigenicity ...

  9. Antitumor effects and radiosensitization of cytosine deaminase and thymidine kinase fusion suicide gene on colorectal carcinoma cells

    Institute of Scientific and Technical Information of China (English)

    De-Hua Wu; Li Liu; Long-Hua Chen


    AIM: To investigate the killing effect and radiosensitization of double suicide gene mediated by adenovirus on colorectal carcinoma cells.METHODS: Colorectal carcinoma cell line SW480 was transfected with adenovirus expression vector containing cytosine deaminase (CD) and thymidine kinase (Tk) fusion gene. The expression of CD-TK fusion gene was detected by reverse transcriptase-polymerase chain reaction. The toxic effect of ganciclovir (GCV) and 5-fiuorocytosine (5FC) on infected cells was determined by MTT assay. The radiosensitization of double suicide gene was evaluated by clonogenic assay.RESULTS: After prodrugs were used, the survival rate of colorectal carcinoma cells was markedly decreased. When GCV and 5-FC were used in combination, the cytotoxicity and bystandereffect were markedly superior to a single prodrug (x2 = 30.371, P<0.01). Both GCV and 5-FC could sensitize colorectal carcinoma cells to the toxic effect of radiation, and greater radiosensitization was achieved when both prodrug were used in combination. CONCLUSION: CD-TK double suicide gene can kill and radiosensitize colorectal carcinoma cells.

  10. The anti-tumour effect of a DNA vaccine carrying a fusion gene of human VEGFR2 and IL-12

    Directory of Open Access Journals (Sweden)

    Sha Wen


    Full Text Available Because of tumour dependence on angiogenesis, anti-angiogenic therapy has become the most attractive area of basic and clinical study in the field of cancer research. In order to create a synergistic effect on angiogenesis and immune regulation, we designed and constructed a new type of DNA vaccine that can express VEGFR2 (vascular endothelial growth factor receptor 2 and the prostate cancer antigen IL-12 (interleukin 12 in the same reading frame. The aim of this study was to investigate the anti-tumour activity of a eukaryotic expression plasmid carrying a fusion gene of human VEGFR2 and IL-12. According to the gene sequences in GenBank, we synthesized the human VEGFR2 and IL-12 genes. VEGFR2 and IL-12 were joined by a sequence encoding a Furin recognition site and a 2A cleavage site, and the resulting fusion gene was cloned into the eukaryotic expression vector pVAX1 to construct the expression plasmid pVAX1-VEGFR2-F2A-IL-12. The expression of VEGFR2 and IL-12 could be detected in 293T cells transfected with pVAX1-VEGFR2-F2A-IL-12 by enzyme-linked immunosorbent assay. Each of these proteins, and in particular co-expression of both proteins, can result in humoral and cellular immune responses in C57BL/6 mice. After injection into the tumour-bearing mouse model, the plasmid showed stronger inhibition of tumour growth than a plasmid expressing VEGFR2 alone. Our results demonstrate that a DNA vaccine carrying a fusion gene of human VEGFR2 and IL-12 could represent a promising approach for tumour immunotherapy.

  11. [Construction of eukaryotic recombinant vector and expression in COS7 cell of LipL32-HlyX fusion gene from Leptospira serovar Lai]. (United States)

    Huang, Bi; Bao, Lang; Zhong, Qi; Zhang, Huidong; Zhang, Ying


    This study was conducted to construct eukaryotic recombinant vector of LipL32-HlyX fusion gene from Leptospira serovar Lai and express it in mammalian cell. Both of LipL32 gene and HlyX gene were amplified from Leptospira strain O17 genomic DNA by PCR. Then with the two genes as template, LipL32-HlyX fusion gene was obtained by SOE PCR (gene splicing by overlap extension PCR). The fusion gene was then cloned into pcDNA3.1 by restriction nuclease digestion. Having been transformed into E. coli DH5alpha, the recombiant plasmid was identified by restriction nuclease digestion, PCR analysis and sequencing. The recombinant plasmid was then transfected into COS7 cell whose expression was detected by RT-PCR and Western blotting analysis. RT-PCR amplified a fragment about 2000 bp and Western blotting analysis found a specific band about 75 KD which was consistent with the expected fusion protein size. In conclusion, the successful construction of eukaryotic recombinant vector containing LipL32-HlyX fusion gene and the effective expression in mammalian have laid a foundation for the application of Leptospira DNA vaccine.

  12. Characterization of foot-and-mouth disease virus gene products with antisera against bacterially synthesized fusion proteins

    Energy Technology Data Exchange (ETDEWEB)

    Strebel, K.; Beck, E.; Strohmaier, K.; Schaller, H.


    Defined segments of the cloned foot-and-mouth disease virus genome corresponding to all parts of the coding region were expressed in Escherichia coli as fusions to the N-terminal part of the MS2-polymerase gene under the control of the inducible lambdaPL promoter. All constructs yielded large amounts of proteins, which were purified and used to raise sequence-specific antisera in rabbits. These antisera were used to identify the corresponding viral gene products in /sup 35/S-labeled extracts from foot-and-mouth disease virus-infected BHK cells. This allowed us to locate unequivocally all mature foot-and-mouth disease virus gene products in the nucleotide sequence, to identify precursor-product relationships, and to detect several foot-and mouth disease virus gene products not previously identified in vivo or in vitro.

  13. TMPRSS2-ERG Gene Fusion Causing ERG Overexpression Precedes Chromosome Copy Number Changes in Prostate Carcinomas, Paired HGPIN Lesions

    Directory of Open Access Journals (Sweden)

    Nuno Cerveira


    Full Text Available TMPRSS2-ETS gene fusions have been found recurrently in prostate carcinomas, but not in the presumed precursor lesion, high-grade prostatic intraepithelial neoplasia (HGPIN. However, HGPIN lesions may share chromosomal changes with prostate cancer. To determine the relative order of genetic events in prostate carcinogenesis, we have analyzed 34 prostate carcinomas, 19 paired HGPIN lesions, 14 benign prostate hyperplasias, 11 morphologically normal prostatic tissues for TMPRSS2-ERG, TMPRSS2-ETV1 rearrangements, genomic imbalances. TMPRSS2 exon 1 was fused in-frame with ERG exon 4 in 17 of 34 (50% prostate carcinomas, in 4 of 19 (21% HGPIN lesions, but in none of controls. The findings were further validated by sequencing analysis, by the real-time polymerase chain reaction quantification of TMPRSS2-ERG fusion transcript, the ERG exons 5/6:exons 1/2 expression ratio. Chromosome copy number changes were detected by comparative genomic hybridization in 42% of clinically confined carcinomas, in none of the 16 HGPIN lesions analyzed. We demonstrate for the first time that the TMPRSS2-ERG fusion gene can be detected in a proportion of HGPIN lesions, that this molecular rearrangement is an early event that may precede chromosome-level alterations in prostate carcinogenesis.

  14. Analysis of NAB2-STAT6 Gene Fusion in 17 Cases of Meningeal Solitary Fibrous Tumor/Hemangiopericytoma: Review of the Literature. (United States)

    Yuzawa, Sayaka; Nishihara, Hiroshi; Wang, Lei; Tsuda, Masumi; Kimura, Taichi; Tanino, Mishie; Tanaka, Shinya


    Solitary fibrous tumor/hemangiopericytoma (SFT/HPC) is a mesenchymal tumor that can affect virtually any region of the body. SFT/HPC of the thoracic cavity and soft tissue has been histologically considered a single biological entity termed SFT; in fact, NAB2-STAT6 gene fusion was recently identified in both diseases. In contrast, meningeal SFT and HPC still need to be investigated in detail with regard to gene fusion variants. The aim of this study was to verify the frequency of NAB2-STAT6 fusion and the relationship between fusion variants and clinicopathologic findings of SFT/HPC, especially meningeal SFT/HPC. We examined the NAB2-STAT6 fusion by reverse transcription polymerase chain reaction with 4 cases of meningeal SFT and 13 cases of meningeal HPC. NAB2-STAT6 fusion transcripts were identified in 12 of 17 cases, including NAB2ex6-STAT6ex17 (4/17, 24%), NAB2ex6-STAT6ex16 and NAB2ex4-STAT6ex2 (3/17, 18%, respectively), and NAB2ex5-STAT6ex16 (2/17, 12%). Three cases showed a pseudopapillary pattern, and 2 of them carried NAB2ex6-STAT6ex17. In addition, our meta-analysis revealed that the major fusion variant in meningeal SFT/HPC was NAB2ex6-STAT6ex16/17 (29/54, 54%), which was also common in soft tissue and intraperitoneum/retroperitoneum but rare in thoracic SFT. Fusion variant significantly correlated with age and histologic diagnosis in meningeal SFT/HPC but not with prognosis. Our results represented that meningeal SFT and HPC were in a single biological spectrum with NAB2-STAT6 gene fusion as was nonmeningeal SFT and further confirmed the organ-specific tumorigenic process and morphologic differences on the basis of fusion variants in meningeal SFT/HPC.

  15. The EWSR1/NR4A3 fusion protein of extraskeletal myxoid chondrosarcoma activates the PPARG nuclear receptor gene. (United States)

    Filion, C; Motoi, T; Olshen, A B; Laé, M; Emnett, R J; Gutmann, D H; Perry, A; Ladanyi, M; Labelle, Y


    The NR4A3 nuclear receptor is implicated in the development of extraskeletal myxoid chondrosarcoma (EMC), primitive sarcoma unrelated to conventional chondrosarcomas, through a specific fusion with EWSR1 resulting in an aberrant fusion protein that is thought to disrupt the transcriptional regulation of specific target genes. We performed an expression microarray analysis of EMC tumours expressing the EWSR1/NR4A3 fusion protein, comparing their expression profiles to those of other sarcoma types. We thereby identified a set of genes significantly overexpressed in EMC relative to other sarcomas, including PPARG and NDRG2. Western blot or immunohistochemical analyses confirm that PPARG and NDRG2 are expressed in tumours positive for EWSR1/NR4A3. Bioinformatic analysis identified a DNA response element for EWSR1/NR4A3 in the PPARG promoter, and band-shift experiments and transient transfections indicate that EWSR1/NR4A3 can activate transcription through this element. Western blots further show that an isoform of the native NR4A3 receptor lacking the C-terminal domain is very highly expressed in tumours positive for EWSR1/NR4A3, and co-transfections of this isoform along with EWSR1/NR4A3 indicate that it may negatively regulate the activity of the fusion protein on the PPARG promoter. These results suggest that the overall expression of PPARG in EMC may be regulated in part by the balance between EWSR1/NR4A3 and NR4A3, and that PPARG may play a crucial role in the development of these tumours. The specific up-regulation of PPARG by EWSR1/NR4A3 may also have potential therapeutic implications.

  16. Bifunctional chimeric SuperCD suicide gene -YCD: YUPRT fusion is highly effective in a rat hepatoma model

    Institute of Scientific and Technical Information of China (English)

    Florian Graepler; Ulrike A Lauer; Reinhard Vonthein; Michael Gregor; Sorin Armeanu; Michael Bitzer; Ulrich M. Lauer; Marie-Luise Lemken; Wolfgang A Wybranietz; Ulrike Schmidt; Irina Smirnow; Christine D Groβ; Martin Spiegel; Andrea Schenk; Hansj(o)rg Graf


    AIM: To investigate the effects of catalytically superior gene-directed enzyme prodrug therapy systems on a rat hepatoma model.METHODS: To increase hepatoma cell chemosensitivity for the prodrug 5-fluorocytosine (5-FC), we generated a chimeric bifunctional SuperCD suicide gene, a fusion of the yeast cytosine deaminase (YCD) and the yeast uracil phosphoribosyltransferase (YUPRT) gene.RESULTS: In vitro stably transduced Morris rat hepatoma cells (MH) expressing the bifunctional SuperCD suicide gene (MH SuperCD) showed a clearly marked enhancement in cell killing when incubated with 5-FC as compared with MH ceils stably expressing YCD solely (MH YCD) or the cytosine deaminase gene of bacterial origin(MH BCD), respectively. In vivo, MH SuperCD tumors implanted both subcutaneously as well as orthotopically into the livers of syngeneic ACI rats demonstrated significant tumor regressions (P<0.01) under both high dose as well as low dose systemic 5-FC application,whereas MH tumors without transgene expression (MH naive) showed rapid progression. For the first time, an order of in vivo suicide gene effectiveness (SuperCD>>YCD > > BCD > > > negative control) was defi ned as a result of a directin vivo comparison of all three suicide genes.CONCLUSION: Bifunctional SuperCD suicide gene expression is highly effective in a rat hepatoma model,thereby significantly improving both the therapeutic index and the efficacy of hepatocellular carcinoma killing by fluorocytosine.

  17. Expression of Chlamydomonas actin-gfp fusion gene in to-bacco suspension cell and polymerization of the actin-gfp protein in vitro

    Institute of Scientific and Technical Information of China (English)


    The fusion gene of actin (cDNA of Chlamydo- monas reinhardtii) and green fluorescence protein (gfp) had been constructed into two expression vectors which could be expressed in E. coli and tobacco suspension cells BY2. The correct expression was observed in E. coli and BY2 with a fluorescence microscopy. The fusion protein, which took part in the membrane skeleton, was mainly located peripherally along the membrane, specially the fusion protein was dis-tributed around nucleus and cell plate, while the fusion pro-tein also forms F-actin in the cell. The fusion protein was purified from Bl21plus by ammonium sulfate fractionation, ion exchange chromatography and hydrophobic interaction chromatography. The purified production could polymerize into F-actin when the actin polymerizing buffer was added. It was demonstrated that the characteristics and function of actin in Chlamydomonas was similar with those of animals and higher plants.

  18. FGFR3–TACC3: A novel gene fusion in cervical cancer

    Directory of Open Access Journals (Sweden)

    Benedito A. Carneiro


    Full Text Available Cervical cancer epitomizes the success of cancer prevention through the human papillomavirus (HPV vaccine, but significant challenges remain in the treatment of advanced disease. We report the first three cases of cervical carcinoma harboring an FGFR3–TACC3 fusion, which serves as a novel therapeutic target. The fusion, identified by comprehensive genomic profiling, activates the FGFR pathway that has been implicated in HPV-driven carcinogenesis. One of the patients whose tumor contained the FGFR3–TACC3 fusion was treated with an investigational FGFR tyrosine kinase inhibitor. Concomitant molecular alterations involving the PI3K/AKT/mTOR and RAF/MEK pathways were also identified and suggest other treatment strategies that deserve investigation. This case series highlights the role of comprehensive genomic profiling in the identification of new therapeutic targets and in targeted therapy selection for patients with cervical cancer.

  19. Retrovirus-mediated transfer of the fusion gene encoding EGFP-BMP2 in mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)

    Zhang Yingang; Guo Xiong; Liu Zheng; Wang Shijie


    Objective To develop retrovirus-mediated transfer of the fusion gene encoding EGFP-BMP2 in mesenchymal stem cells. Methods Mesenchymal stem cells from New Zealand white rabbits were transduced with retroviral pLEGFP-BMP2 vector by the optimized retroviral transduction protocol. Fluorescent microscopy's examination was to evaluate the results of the transduction, flow cytometer's analysis was to evaluate the transduction efficiency and the Fluorescence-activated cell sorting method was to sort the transduced cells. Bioactivity test from C2C12K4 cells was to show the expression and bio-activity of the fusion gene. Results Fluorescent microscopy showed the success of the transduction. By flow cytometer's analysis, the mean efficiency of the transduction with EGFP was (42.8±6.1)% SD. Transduced cells were sorted efficiently by the fluorescence-activated cell sorting method and after sorting, almost of those showed the expression of BMP2. Fluorescently and strongly bioactivity test for C2C12K4 cells demonstrated that fluorescent materials were located the surface of cells and the activity of luciferase increased compared with the control. Analysis of long-term expression showed there was no difference between 2 week-time point and 3 month-time point of culture post-sorting. Conclusion Mesenchymal stem cells can be transduced efficiently by retrovirus-mediated transfer of the fusion gene encoding EGFP-BMP2, the highly pure transduced cells are obtained by the fluorescence-activated cell sorting technique, the expressed chimeric protein embraced the double bioactivity of EGFP and BMP2, and moreover, the expression had not attenuated over time.

  20. A system for the measurement of gene targeting efficiency in human cell lines using an antibiotic resistance-GFP fusion gene. (United States)

    Konishi, Yuko; Karnan, Sivasundaram; Takahashi, Miyuki; Ota, Akinobu; Damdindorj, Lkhagvasuren; Hosokawa, Yoshitaka; Konishi, Hiroyuki


    Gene targeting in a broad range of human somatic cell lines has been hampered by inefficient homologous recombination. To improve this technology and facilitate its widespread application, it is critical to first have a robust and efficient research system for measuring gene targeting efficiency. Here, using a fusion gene consisting of hygromycin B phosphotransferase and 3'-truncated enhanced GFP (HygR-5' EGFP) as a reporter gene, we created a molecular system monitoring the ratio of homologous to random integration (H/R ratio) of targeting vectors into the genome. Cell clones transduced with a reporter vector containing HygR-5' EGFP were efficiently established from two human somatic cell lines. Established HygR-5' EGFP reporter clones retained their capacity to monitor gene targeting efficiency for a longer duration than a conventional reporter system using an unfused 5' EGFP gene. With the HygR-5' EGFP reporter system, we reproduced previous findings of gene targeting frequency being up-regulated by the use of an adeno-associated viral (AAV) backbone, a promoter-trap system, or a longer homology arm in a targeting vector, suggesting that this system accurately monitors H/R ratio. Thus, our HygR-5' EGFP reporter system will assist in the development of an efficient AAV-based gene targeting technology.

  1. Construction of prokaryotic expression system of TGF-β1 epitope gene and identification of recombinant fusion protein immunity

    Institute of Scientific and Technical Information of China (English)

    Yong-Hong Guo; Zhi-Ming Hao; Jin-Yan Luo; Jun-Hong Wang


    AIM: To insert the constructed TGF-β1the el loop of C-terminus of truncated hepatitis B core antigen to increase TGF-β1expression system and to identify immunity of the expressed recombinant protein in order to exploit the possibility for obtaining anti- TGF-β1METHODS: The TGF-β1mature TGF-β1TGF-32) was amplified by polymerase chain reaction from the recombinant pGEM-7z/TGF-β1HBcAg gene fragments (encoding HBcAg from 1-71 and 89-144 amino acid residues) were amplified from PYTA1-HBcAg vector. The recombinant vector pGEMEX-1 was used to insert HBcAg1-71, TGF-β1into restrictive endonuclease enzyme and ligated with T4ligase. The fusion gene fragments HBcAg1-71-TGF-β1HBcAg89-144 were recloned to pET28a(+) and the DNA sequence was confirmed by the dideoxy chain termination method. The recombinant vector pET28a (+)/CTC was transformed and expressed in E.. Coli BL21 (DE3)under induction of IPTG. After purification with Ni+2-NTA agarose resins, the antigenicity of purified protein was detected by ELISA and Western blot and visualized under electron microscope.RESULTS: Enzyme digestion analysis and sequencing showed that TGF-β1loop of C-terminus of truncated hepatitis B core antigen.SDS-PAGE analysis showed that relative molecular mass(Mr) of the expressed product by pET28a (+)/CTC was Mr 24 600.The output of the target recombinant protein was approximately 34.8% of the total bacterial protein,mainly presented in the form of inclusion body. Western blotting and ELISA demonstrated that the fusion protein could combine with anti-TGF-β1not with anti-HBcAg. The purity of protein was about 90% and the protein was in the form of self-assembling particles visualized under electron microscope. This fusion protein had good anti-TGF-β1could be used as anti-TGF-β1CONCLUSION: A recombinant prokaryotic expression system with high expression efficiency of the target TGF- epitope gene was successfully established.The fusion protein is in the form of self-assembling particles

  2. Fusion of the homeobox gene HLXB9 and the ETV6 gene in infant acute myeloid leukemias with the t(7;12)(q36;p13). (United States)

    Beverloo, H B; Panagopoulos, I; Isaksson, M; van Wering, E; van Drunen, E; de Klein, A; Johansson, B; Slater, R


    Recently, we and others reported a recurrent t(7;12)(q36;p13) found in myeloid malignancies in children < or =18 months of age and associated with a poor prognosis. Fluorescence in situ hybridization studies mapped the 12p13 breakpoint to the first intron of ETV6 and narrowed down the region of 7q36 involved. By using the sequences made public recently by the Human Genome Project, two candidate genes in 7q36 were identified: the homeobox gene HLXB9 and c7orf3, a gene with unknown function. Reverse transcription-PCR of two cases with t(7;12), using primers for c7orf3 and ETV6, was negative. However, reverse transcription-PCR for HLXB9-ETV6 demonstrated alternative splicing; the two major bands corresponded to fusion of exon 1 of HLXB9 to exons 2 and 3, respectively, of ETV6. The reciprocal ETV6-HLXB9 transcript was not detected. It remains to be elucidated if the leukemic phenotype is attributable to the formation of the HLXB9-ETV6 fusion protein, which includes the helix-loop-helix and E26 transformation-specific DNA binding domains of ETV6 or to the disruption of the normal ETV6 protein.

  3. State-of-the-Art Fusion-Finder Algorithms Sensitivity and Specificity

    Directory of Open Access Journals (Sweden)

    Matteo Carrara


    Full Text Available Background. Gene fusions arising from chromosomal translocations have been implicated in cancer. RNA-seq has the potential to discover such rearrangements generating functional proteins (chimera/fusion. Recently, many methods for chimeras detection have been published. However, specificity and sensitivity of those tools were not extensively investigated in a comparative way. Results. We tested eight fusion-detection tools (FusionHunter, FusionMap, FusionFinder, MapSplice, deFuse, Bellerophontes, ChimeraScan, and TopHat-fusion to detect fusion events using synthetic and real datasets encompassing chimeras. The comparison analysis run only on synthetic data could generate misleading results since we found no counterpart on real dataset. Furthermore, most tools report a very high number of false positive chimeras. In particular, the most sensitive tool, ChimeraScan, reports a large number of false positives that we were able to significantly reduce by devising and applying two filters to remove fusions not supported by fusion junction-spanning reads or encompassing large intronic regions. Conclusions. The discordant results obtained using synthetic and real datasets suggest that synthetic datasets encompassing fusion events may not fully catch the complexity of RNA-seq experiment. Moreover, fusion detection tools are still limited in sensitivity or specificity; thus, there is space for further improvement in the fusion-finder algorithms.

  4. Imaging of dihydrofolate reductase fusion gene expression in xenografts of human liver metastases of colorectal cancer in living rats

    Energy Technology Data Exchange (ETDEWEB)

    Mayer-Kuckuk, Philipp; Bertino, Joseph R.; Banerjee, Debabrata [Molecular Pharmacology and Therapeutics Program, Memorial Sloan-Kettering Cancer Center, New York, NY (United States); The Cancer Institute of New Jersey, Robert Wood Johnson Medical School/UMDNJ, 195 Little Albany Street, NJ 08903, New Brunswick (United States); Doubrovin, Mikhail; Blasberg, Ronald; Tjuvajev, Juri Gelovani [Department of Neurooncology, Memorial Sloan-Kettering Cancer Center, New York, NY (United States); Gusani, Niraj J.; Fong, Yuman [Department of Surgery, Memorial Sloan-Kettering Cancer Center, New York, NY (United States); Gade, Terence; Koutcher, Jason A. [Department of Medical Physics, Memorial Sloan-Kettering Cancer Center, New York, NY (United States); Balatoni, Julius; Finn, Ronald [Radiochemistry/Cyclotron Core Facility, Memorial Sloan-Kettering Cancer Center, New York, NY (United States); Akhurst, Tim; Larson, Steven [Nuclear Medicine Service, Memorial Sloan-Kettering Cancer Center, New York, NY (United States)


    Radionuclide imaging has been demonstrated to be feasible to monitor transgene expression in vivo. We hypothesized that a potential application of this technique is to non-invasively detect in deep tissue, such as cancer cells metastatic to the liver, a specific molecular response following systemic drug treatment. Utilizing human colon adenocarcinoma cells derived from a patient's liver lesion we first developed a nude rat xenograft model for colorectal cancer metastatic to the liver. Expression of a dihydrofolate reductase-herpes simplex virus 1 thymidine kinase fusion (DHFR-HSV1 TK) transgene in the hepatic tumors was monitored in individual animals using the tracer [{sup 124}I]2'-fluoro-2'-deoxy-5-iodouracil-{beta}-d-arabinofuranoside (FIAU) and a small animal micro positron emission tomograph (microPET), while groups of rats were imaged using the tracer [{sup 131}I]FIAU and a clinical gamma camera. Growth of the human metastatic colorectal cancer cells in the rat liver was detected using magnetic resonance imaging and confirmed by surgical inspection. Single as well as multiple lesions of different sizes and sites were observed in the liver of the animals. Next, using a subset of rats bearing hepatic tumors, which were retrovirally bulk transduced to express the DHFR-HSV1 TK transgene, we imaged the fusion protein expression in the hepatic tumor of living rats using the tracer [{sup 124}I]FIAU and a microPET. The observed deep tissue signals were highly specific for the tumors expressing the DHFR-HSV1 TK fusion protein compared with parental untransduced tumors and other tissues as determined by gamma counting of tissue samples. A subsequent study used the tracer [{sup 131}I]FIAU and a gamma camera to monitor two groups of transduced hepatic tumor-bearing rats. Prior to imaging, one group was treated with trimetrexate to exploit DHFR-mediated upregulation of the fusion gene product. Imaging in the living animal as well as subsequent gamma

  5. EnABLing microprocessor for apoptosis. (United States)

    Tu, Chi-Chiang; Wang, Jean Y J

    The Microprocessor complex consisting of DROSHA (a type III ribonuclease) and DGCR8 (DiGeorge syndrome critical region gene 8-encoded RNA binding protein) recognizes and cleaves the precursor microRNA hairpin (pre-miRNA) from the primary microRNA transcript (pri-miRNA). The Abelson tyrosine kinase 1 (ABL) phosphorylates DGCR8 to stimulate the cleavage of a subset of pro-apoptotic pri-miRNAs, thus expanding the nuclear functions of ABL to include regulation of RNA processing.

  6. The zinc binuclear cluster activator AlcR is able to bind to single sites but requires multiple repeated sites for synergistic activation of the alcA gene in Aspergillus nidulans. (United States)

    Panozzo, C; Capuano, V; Fillinger, S; Felenbok, B


    The alcA gene which is part of the recently identified ethanol regulon, is one of the most strongly inducible genes in Aspergillus nidulans. Its transcriptional activation is mediated by the AlcR transactivator which contains a DNA-binding domain belonging to the C6 zinc binuclear cluster family. AlcR differs from the other members of this family by several features, the most striking characteristic being its binding to both symmetric and asymmetric DNA sites with the same apparent affinity. However, AlcR is also able to bind to a single site with high affinity, suggesting that unlike the other C6 proteins, AlcR binds as a monomer. In this report, we show that AlcR targets, to be functional in vivo, have to be organized as inverted or direct repeats. In addition, we show a strong synergistic activation of alcA transcription in which the number and the position of the AlcR-binding sites are crucial. The fact that the AlcR unit for in vitro binding is a single site whereas the in vivo functional unit is a repeat opens the question of the mechanism of the strong alcA transactivation. These results show that AlcR displays both in vitro and in vivo a new range of binding specificity and provides a novel example in the C6 zinc cluster protein family.

  7. Teaching mathematics to able children

    CERN Document Server

    Koshy, Valsa


    This book enables teachers to effectively meet the needs of their most able mathematicians. Using a tried and tested set of principles developed and used by The Able Children's Education Unit at Brunel University, the author demonstrates how to: identify high mathematical ability in a pupil, plan suitably challenging activities and teach them most effectively within the existing National Numeracy framework, make the most of the classroom resources available, including ICT and external agencies, implement strategies for differentiation, illustrated with real-life classroom examples. Ac

  8. TALE-PvuII fusion proteins--novel tools for gene targeting.

    Directory of Open Access Journals (Sweden)

    Mert Yanik

    Full Text Available Zinc finger nucleases (ZFNs consist of zinc fingers as DNA-binding module and the non-specific DNA-cleavage domain of the restriction endonuclease FokI as DNA-cleavage module. This architecture is also used by TALE nucleases (TALENs, in which the DNA-binding modules of the ZFNs have been replaced by DNA-binding domains based on transcription activator like effector (TALE proteins. Both TALENs and ZFNs are programmable nucleases which rely on the dimerization of FokI to induce double-strand DNA cleavage at the target site after recognition of the target DNA by the respective DNA-binding module. TALENs seem to have an advantage over ZFNs, as the assembly of TALE proteins is easier than that of ZFNs. Here, we present evidence that variant TALENs can be produced by replacing the catalytic domain of FokI with the restriction endonuclease PvuII. These fusion proteins recognize only the composite recognition site consisting of the target site of the TALE protein and the PvuII recognition sequence (addressed site, but not isolated TALE or PvuII recognition sites (unaddressed sites, even at high excess of protein over DNA and long incubation times. In vitro, their preference for an addressed over an unaddressed site is > 34,000-fold. Moreover, TALE-PvuII fusion proteins are active in cellula with minimal cytotoxicity.

  9. TALE-PvuII fusion proteins--novel tools for gene targeting. (United States)

    Yanik, Mert; Alzubi, Jamal; Lahaye, Thomas; Cathomen, Toni; Pingoud, Alfred; Wende, Wolfgang


    Zinc finger nucleases (ZFNs) consist of zinc fingers as DNA-binding module and the non-specific DNA-cleavage domain of the restriction endonuclease FokI as DNA-cleavage module. This architecture is also used by TALE nucleases (TALENs), in which the DNA-binding modules of the ZFNs have been replaced by DNA-binding domains based on transcription activator like effector (TALE) proteins. Both TALENs and ZFNs are programmable nucleases which rely on the dimerization of FokI to induce double-strand DNA cleavage at the target site after recognition of the target DNA by the respective DNA-binding module. TALENs seem to have an advantage over ZFNs, as the assembly of TALE proteins is easier than that of ZFNs. Here, we present evidence that variant TALENs can be produced by replacing the catalytic domain of FokI with the restriction endonuclease PvuII. These fusion proteins recognize only the composite recognition site consisting of the target site of the TALE protein and the PvuII recognition sequence (addressed site), but not isolated TALE or PvuII recognition sites (unaddressed sites), even at high excess of protein over DNA and long incubation times. In vitro, their preference for an addressed over an unaddressed site is > 34,000-fold. Moreover, TALE-PvuII fusion proteins are active in cellula with minimal cytotoxicity.

  10. Cysteine-rich secretory protein-3 (CRISP3 is strongly up-regulated in prostate carcinomas with the TMPRSS2-ERG fusion gene.

    Directory of Open Access Journals (Sweden)

    Franclim R Ribeiro

    Full Text Available A large percentage of prostate cancers harbor TMPRSS2-ERG gene fusions, leading to aberrant overexpression of the transcription factor ERG. The target genes deregulated by this rearrangement, however, remain mostly unknown. To address this subject we performed genome-wide mRNA expression analysis on 6 non-malignant prostate samples and 24 prostate carcinomas with (n = 16 and without (n = 8 TMPRSS2-ERG fusion as determined by FISH. The top-most differentially expressed genes and their associations with ERG over-expression were technically validated by quantitative real-time PCR and biologically validated in an independent series of 200 prostate carcinomas. Several genes encoding metabolic enzymes or extracellular/transmembrane proteins involved in cell adhesion, matrix remodeling and signal transduction pathways were found to be co-expressed with ERG. Within those significantly over-expressed in fusion-positive carcinomas, CRISP3 showed more than a 50-fold increase when compared to fusion-negative carcinomas, whose expression levels were in turn similar to that of non-malignant samples. In the independent validation series, ERG and CRISP3 mRNA levels were strongly correlated (r(s = 0.65, p<0.001 and both were associated with pT3 disease staging. Furthermore, immunohistochemistry results showed CRISP3 protein overexpression in 63% of the carcinomas and chromatin immunoprecipitation with an anti-ERG antibody showed that CRISP3 is a direct target of the transcription factor ERG. We conclude that ERG rearrangement is associated with significant expression alterations in genes involved in critical cellular pathways that define a subset of locally advanced PCa. In particular, we show that CRISP3 is a direct target of ERG that is strongly overexpressed in PCa with the TMPRSS2-ERG fusion gene.

  11. pBaSysBioll : an integrative plasmid generating gfp transcriptional fusions for high-throughput analysis of gene expression in Bacillus subtilis

    NARCIS (Netherlands)

    Botella, Eric; Fogg, Mark; Jules, Matthieu; Piersma, Sjouke; Doherty, Geoff; Hansen, Annette; Denham, Emma. L.; Le Chat, Ludovic; Veiga, Patrick; Bailey, Kirra; Lewis, Peter J.; van Dijl, Jan Maarten; Aymerich, Stephane; Wilkinson, Anthony J.; Devine, Kevin M.


    Plasmid pBaSysBioll was constructed for high-throughput analysis of gene expression in Bacillus subtilis. It is an integrative plasmid with a ligation-independent cloning (LIC) site, allowing the generation of transcriptional gfpmut3 fusions with desired promoters. Integration is by a Campbell-type

  12. Fusion protein gene nucleotide sequence similarities, shared antigenic sites and phylogenetic analysis suggest that phocid distemper virus 2 and canine distemper virus belong to the same virus entity.

    NARCIS (Netherlands)

    I.K.G. Visser (Ilona); R.W.J. van der Heijden (Roger); M.W.G. van de Bildt (Marco); M.J.H. Kenter (Marcel); C. Örvell; A.D.M.E. Osterhaus (Albert)


    textabstractNucleotide sequencing of the fusion protein (F) gene of phocid distemper virus-2 (PDV-2), recently isolated from Baikal seals (Phoca sibirica), revealed an open reading frame (nucleotides 84 to 2075) with two potential in-frame ATG translation initiation codons. We suggest that the secon

  13. Mammary Analogue Secretory Carcinoma of Salivary Glands: Molecular Analysis of 25 ETV6 Gene Rearranged Tumors With Lack of Detection of Classical ETV6-NTRK3 Fusion Transcript by Standard RT-PCR: Report of 4 Cases Harboring ETV6-X Gene Fusion. (United States)

    Skálová, Alena; Vanecek, Tomas; Simpson, Roderick H W; Laco, Jan; Majewska, Hanna; Baneckova, Martina; Steiner, Petr; Michal, Michal


    ETV6 gene abnormalities are well described in tumor pathology. Many fusion partners of ETV6 have been reported in a variety of epithelial and hematological malignancies. In salivary gland tumor pathology, however, the ETV6-NTRK3 translocation is specific for mammary analogue secretory carcinoma (MASC), and has not been documented in any other salivary tumor type. The present study comprised a clinical and molecular analysis of 25 cases morphologically and immunohistochemically typical of MASC. They all also displayed the ETV6 rearrangement as visualized by fluorescent in situ hybridization but lacked the classical ETV6-NTRK3 fusion transcript by standard reverse-transcriptase-polymerase chain reaction. In 4 cases, the classical fusion transcript was found by more sensitive, nested reverse-transcription-polymerase chain reaction. Five other cases harbored atypical fusion transcripts as detected by both standard and nested reverse-transcription-polymerase chain reaction. In addition, fluorescent in situ hybridization with an NTRK3 break-apart probe was also performed; rearrangement of NTRK3 gene was detected in 16 of 25 cases. In 3 other cases, the tissue was not analyzable, and in 2 further cases analysis could not be performed because of a lack of appropriate tissue material. Finally, in the 4 remaining cases whose profile was NTRK3 split-negative and ETV6 split-positive, unknown (non-NTRK) genes appeared to fuse with ETV6 (ETV6-X fusion). In looking for possible fusion partners, analysis of rearrangement of other kinase genes known to fuse with ETV6 was also performed, but without positive results. Although numbers were small, correlating the clinico-pathologic features of the 4 ETV6-X fusion tumors and 5 MASC cases with atypical fusion transcripts raises the possibility of that they may behave more aggressively.

  14. In-vitro activation of cytotoxic T lymphocytes by fusion of mouse hepatocellular carcinoma cells and lymphotactin gene-modified dendritic cells

    Institute of Scientific and Technical Information of China (English)


    AIM: To investigate the in-vitro activation of cytotoxic T lymphocytes (CTLs) by fusion of mouse hepatocellular carcinoma (HCC) cells and lymphotactin gene-modified dendritic cells (DCs).METHODS: Lymphotactin gene modified DCs (DCLptn) were prepared by lymphotactin recombinant adenovirus transduction of mature DCs which differentiated from mouse bone marrow cells by stimulation with granulocyte/macrophage colony-stimulating factor (GM-CSF), interleukin-4 (IL-4) and tumor necrosis factor alpha (TNF-α). DCLptn and H22 fusion was prepared using 50% PEG. Lymphotactin gene and protein expression levels were measured by RT-PCR and ELISA, respectively. Lymphotactin chemotactic responses were examined by in-vitro chemotaxis assay. In-vitro activation of CTLs by DCLptn/H22 fusion was measured by detecting CD25 expression and cytokine production after autologous T cell stimulation. Cytotoxic function of activated T lymphocytes stimulated with DCLptn/H22 cells was determined by LDH cytotoxicity assay.RESULTS: Lymphotactin gene could be efficiently transduced to DCs by adenovirus vector and showed an effective biological activity. After fusion, the hybrid DCLptn/H22 cells acquired the phenotypes of both DCLptn and H22 cells. In T cell proliferation assay, flow cytometry showed a very high CD25 expression, and cytokine release assay showed a significantly higher concentration of IFN-γ and IL-2 in DCLptn/H22 group than in DCLptn, DCLptn+H22, DC/H22 or H22 groups. Cytotoxicity assay revealed that T cells derived from DCLptn/H22 group had much higher anti-tumor activity than those derived from DCLptn, H22, DCLptn + H22, DC/H22 groups.CONCLUSION: Lymphotactin gene-modified dendritoma induces T-cell proliferation and strong CTL reaction against allogenic HCC cells. Immunization-engineered fusion hybrid vaccine is an attractive strategy in prevention and treatment of HCC metastases.

  15. Fusion of the Dhfr/Mtx and IR/MAR gene amplification methods produces a rapid and efficient method for stable recombinant protein production. (United States)

    Noguchi, Chiemi; Araki, Yoshio; Miki, Daisuke; Shimizu, Noriaki


    Amplification of the dihydrofolate reductase gene (Dhfr) by methotrexate (Mtx) exposure is commonly used for recombinant protein expression in Chinese hamster ovary (CHO) cells. However, this method is both time- and labor-intensive, and the high-producing cells that are generated are frequently unstable in culture. Another gene amplification method is based on using a plasmid bearing a mammalian replication initiation region (IR) and a matrix attachment region (MAR), which result in the spontaneous initiation of gene amplification in transfected cells. The IR/MAR and Dhfr/Mtx methods of gene amplification are based on entirely different principles. In this study, we combine these two methods to yield a novel method, termed the IR/MAR-Dhfr fusion method, which was used to express three proteins, the Fc receptor, GFP, and recombinant antibody. The fusion method resulted in a dramatic increase in expression of all three proteins in two CHO sub-lines, DXB-11, and DG44. The IR/MAR-Dhfr fusion amplified the genes rapidly and efficiently, and produced larger amounts of antibody than the Dhfr/Mtx or IR/MAR methods alone. While the amplified structure produced by the Dhfr/Mtx method was highly unstable, and the antibody production rate rapidly decreased with the culture time of the cells, the IR/MAR-Dhfr fusion method resulted in stable amplification and generated clonal cells that produced large amounts of antibody protein over a long period of time. In summary, the novel IR/MAR-Dhfr fusion method enables isolation of stable cells that produce larger amounts of a target recombinant protein more rapidly and easily than either the Dhfr/Mtx or IR/MAR methods alone.

  16. Studies on virus-induced cell fusion. Progress report, August 1, 1977--June 30, 1978

    Energy Technology Data Exchange (ETDEWEB)

    Person, S.


    We have previously postulated that wild-type Herpes Simplex Virus type I (HSV-1) infections are characterized by the presence of a fusion factor and a fusion inhibitor activity. The fusion inhibitor presumably is dominant so that a small fraction of cells fuse in a typical wild-type infection. Furthermore, the syn mutants isolated in our laboratory are thought to cause extensive cell fusion because the production of active fusion inhibitor in cell membranes is delayed. If mutations existed that altered both the fusion factor and fusion inhibitor activity then separate viruses containing these two mutations might be able to complement each other, each supplying the defective gene product missing in the other virus. This would produce a wild type and not a syncytial mutant response. Complementation tests between two viruses, tsB5 and syn 20, which are thought to contain defects in the production of active fusion factor and fusion inhibitor activity, respectively, were done. A wild-type response was observed indicating that the mutations affecting fusion were in two separate genes.

  17. NAB2-STAT6 gene fusion and STAT6 immunoexpression in extrathoracic solitary fibrous tumors: the association between fusion variants and locations. (United States)

    Chuang, I-Chieh; Liao, Kuan-Cho; Huang, Hsuan-Ying; Kao, Yu-Chien; Li, Chien-Feng; Huang, Shih-Chiang; Tsai, Jen-Wei; Chen, Ko-Chin; Lan, Jui; Lin, Po-Chun


    Solitary fibrous tumor (SFT) is a rare mesenchymal neoplasm harboring NAB2-STAT6 fusion, which drives STAT6 nuclear relocation. For extrathoracic SFTs, the clinical relevance of this molecular hallmark remains obscure. We assessed STAT6 immunoexpression for 61 extrathoracic SFTs exclusive of the meninges and head and neck, and 25 had analyzable RNAs to distinguish fusion variants by RT-PCR. The immunohistochemical and molecular findings were correlated with clincopathological features and disease-free survival (DFS). Twenty-eight males and 33 females had SFTs in the body cavities (n = 31), extremities (n = 17), and trunk (n = 13), categorized into 53 non-malignant and 8 malignant tumors. The vast majority (n = 57, 93%) exhibited distinctive STAT6 nuclear expression, including malignant ones. The common fusion variants were NAB2ex6-STAT6ex16/17 in 13 SFTs and NAB2ex4-STAT6ex2 in 8, while miscellaneous variants were detected only in 4 SFTs in the limbs and trunk but not in any body cavity-based cases (P = 0.026). The worse DFS was univariately associated with malignant histology (P = 0.04) but unrelated to tumor size, location, or fusion variant. Conclusively, extrathoracic SFTs mostly harbor NAB2ex6-STAT6ex16/17, followed by NAB2ex4-STAT6ex2. Miscellaneous variants are significantly rare in SFTs within the body cavities. The clinical aggressiveness of extrathoraic SFTs is associated with malignant histology but unrelated to the NAB2-STAT6 fusion variants.

  18. RNA-seq analysis of prostate cancer in the Chinese population identifies recurrent gene fusions,cancer-associated long noncoding RNAs and aberrant alternative splicings

    Institute of Scientific and Technical Information of China (English)

    Shancheng Ren; Weidong Xu; Chao Chen; Fubo Wang; Xinwu Guo; Ji Lu; Jun Yang; Min Wei; Zhijian Tian; Yinghui Guan; Liang Tang; Zhiyu Peng; Chuanliang Xu; Linhui Wang; Xu Gao; Wei Tian; Jian Wang; Huanming Yang; Jun Wang; Yinghao Sun; Jian-Hua Mao; Yongwei Yu; Changjun Yin; Xin Gao; Zilian Cui; Jibin Zhang; Kang Yi


    There are remarkable disparities among patients of different races with prostate cancer; however,the mechanism underlying this difference remains unclear.Here,we present a comprehensive landscape of the transcriptome profiles of 14 primary prostate cancers and their paired normal counterparts from the Chinese population using RNA-seq,revealing tremendous diversity across prostate cancer transcriptomes with respect to gene fusions,long noneoding RNAs (long ncRNA),alternative splicing and somatic mutations.Three of the 14 tumors (21.4%) harbored a TMPRSS2-ERG fusion,and the low prevalence of this fusion in Chinese patients was further confirmed in an additional tumor set (10/54=18.5%).Notably,two novel gene fusions,CTAGE5-KHDRBS3 (20/54=37%) and USP9Y-TTTY15(19/54=35.2%),occurred frequently in our patient cohort.Further systematic transcriptional profiling identified numerous long ncRNAs that were differentially expressed in the tumors.An analysis of the correlation between expression of long ncRNA and genes suggested that long ncRNAs may have functions beyond transcriptional regulation.This study yielded new insights into the pathogenesis of prostate cancer in the Chinese population.

  19. Plant expansins in bacteria and fungi: evolution by horizontal gene transfer and independent domain fusion. (United States)

    Nikolaidis, Nikolas; Doran, Nicole; Cosgrove, Daniel J


    Horizontal gene transfer (HGT) has been described as a common mechanism of transferring genetic material between prokaryotes, whereas genetic transfers from eukaryotes to prokaryotes have been rarely documented. Here we report a rare case of HGT in which plant expansin genes that code for plant cell-wall loosening proteins were transferred from plants to bacteria, fungi, and amoebozoa. In several cases, the species in which the expansin gene was found is either in intimate association with plants or is a known plant pathogen. Our analyses suggest that at least two independent genetic transfers occurred from plants to bacteria and fungi. These events were followed by multiple HGT events within bacteria and fungi. We have also observed that in bacteria expansin genes have been independently fused to DNA fragments that code for an endoglucanase domain or for a carbohydrate binding module, pointing to functional convergence at the molecular level. Furthermore, the functional similarities between microbial expansins and their plant xenologs suggest that these proteins mediate microbial-plant interactions by altering the plant cell wall and therefore may provide adaptive advantages to these species. The evolution of these nonplant expansins represents a unique case in which bacteria and fungi have found innovative and adaptive ways to interact with and infect plants by acquiring genes from their host. This evolutionary paradigm suggests that despite their low frequency such HGT events may have significantly contributed to the evolution of prokaryotic and eukaryotic species.

  20. Secretory breast carcinomas with ETV6-NTRK3 fusion gene belong to the basal-like carcinoma spectrum. (United States)

    Laé, Marick; Fréneaux, Paul; Sastre-Garau, Xavier; Chouchane, Olfa; Sigal-Zafrani, Brigitte; Vincent-Salomon, Anne


    Secretory breast carcinomas (carcinoma. A series of six secretory breast carcinomas were identified in our files. The ETV6 rearrangement was confirmed in all cases by fluorescence in situ hybridization. Immunophenotype was assessed with anti-ER, PR, ERBB2, KIT, EGFR, E-cadherin, vimentin, PS100, smooth muscle actin, basal (CK5/6 and 14), luminal cytokeratins (CK8/18) and p63 antibodies. In situ and invasive components shared the same immunoprofile and were ER, PR, ERBB2 negative with expression of basal cytokeratins. ETV6 gene alterations were present in both in situ and invasive components, highlighting their genetic similarities. The immunoprofile data (triple-negative with expression of basal markers) showed that secretory breast carcinomas with ETV6-NTRK3 fusion gene belong to the phenotypic basal-like spectrum of breast carcinomas. These results support the hypothesis that secretory breast carcinomas have immunohistochemical and genetic features that distinguish them from other basal-like tumors of the breast.

  1. Able to Adapt and Conquer (United States)


    penalty for failing to comply with a collection of information if it does not display a currently valid OMB control number. 1. REPORT DATE 10 MAR...2009 2. REPORT TYPE 3. DATES COVERED 00-00-2009 to 00-00-2009 4. TITLE AND SUBTITLE Able to Adapt and Conquer 5a. CONTRACT NUMBER 5b. GRANT...Afghanistan. This tells of how our unit moved in-between Regional Commands to deter and destroy the enemy, set up multiple Combat Out Posts (COP), trained

  2. Bioinformatic analysis of patient-derived ASPS gene expressions and ASPL-TFE3 fusion transcript levels identify potential therapeutic targets.

    Directory of Open Access Journals (Sweden)

    David G Covell

    Full Text Available Gene expression data, collected from ASPS tumors of seven different patients and from one immortalized ASPS cell line (ASPS-1, was analyzed jointly with patient ASPL-TFE3 (t(X;17(p11;q25 fusion transcript data to identify disease-specific pathways and their component genes. Data analysis of the pooled patient and ASPS-1 gene expression data, using conventional clustering methods, revealed a relatively small set of pathways and genes characterizing the biology of ASPS. These results could be largely recapitulated using only the gene expression data collected from patient tumor samples. The concordance between expression measures derived from ASPS-1 and both pooled and individual patient tumor data provided a rationale for extending the analysis to include patient ASPL-TFE3 fusion transcript data. A novel linear model was exploited to link gene expressions to fusion transcript data and used to identify a small set of ASPS-specific pathways and their gene expression. Cellular pathways that appear aberrantly regulated in response to the t(X;17(p11;q25 translocation include the cell cycle and cell adhesion. The identification of pathways and gene subsets characteristic of ASPS support current therapeutic strategies that target the FLT1 and MET, while also proposing additional targeting of genes found in pathways involved in the cell cycle (CHK1, cell adhesion (ARHGD1A, cell division (CDC6, control of meiosis (RAD51L3 and mitosis (BIRC5, and chemokine-related protein tyrosine kinase activity (CCL4.

  3. Enforced effect of tk-MCP-1 fusion gene in ovarian cancer

    Directory of Open Access Journals (Sweden)

    Hong Shuhui


    Full Text Available Abstract Objective The efficiency of HSV-tk/GCV system is not high because of insufficient gene transfer and incompletely initiative of host antineoplastic potency. The present study was designed to assess the antitumor efficacy of tk-MCP-1 on ovarian cancer in vitro and vivo. Methods A novel bicistronic expression system can help to improve the expression level of a gene in a stable manner. pLXSN/tk-MCP-1 co-expressing tk and MCP-1 genes was constructed using a pLXSN retroviral vector and an internal ribosome entry site sequence by restriction enzyme. Western blot was performed to determine tk and MCP-1 expression in the infected SKOV3. The GCV-sensitively tumoricidal activities of SKOV3/tk-MCP-1 with or without monocytes were compared to those of SKOV3 expressing HSV-tk or MCP-1. We investigated the growth of subcutaneous tumors in SCID mice immuno-reconstituted, and evaluated the antitumor effect of MCP-1 in conjunction with suicide gene. Results The significant GCV-sensitively tumoricidal activity of pLXSN/tk-MCP-1 was observed when compared with those of pLXSN/tk, pLXSN/MCP-1 and pLXSN/neo, especially when monocytes were added. The growth of subcutaneous tumors in SCID mice immuno-reconstituted was markedly suppressed by co-delivery of HSV-tk and MCP-1 genes, and the enhanced antitumor effect was associated with the recruitment of monocytes. Conclusion These results demonstrated pLXSN/tk-MCP-1 presented an enhanced antitumor effects on ovarian cancer by orchestration of immune responses.

  4. Fusion of the TBL1XR1 and HMGA1 genes in splenic hemangioma with t(3;6)(q26;p21). (United States)

    Panagopoulos, Ioannis; Gorunova, Ludmila; Bjerkehagen, Bodil; Lobmaier, Ingvild; Heim, Sverre


    RNA-sequencing of a splenic hemangioma with the karyotype 45~47,XX,t(3;6)(q26;p21) showed that this translocation generated a chimeric TBL1XR1-HMGA1 gene. This is the first time that this tumor has been subjected to genetic analysis, but the finding of an acquired clonal chromosome abnormality in cells cultured from the lesion and the presence of the TBL1XR1-HMGA1 fusion in them strongly favor the conclusion that splenic hemangiomas are of a neoplastic nature. Genomic PCR confirmed the presence of the TBL1XR1-HMGA1 fusion gene, and RT-PCR together with Sanger sequencing verified the presence of the fusion transcripts. The molecular consequences of the t(3;6) would be substantial. The cells carrying the translocation would retain only one functional copy of the wild-type TBL1XR1 gene while the other, rearranged allele could produce a putative truncated form of TBL1XR1 protein containing the LiSH and F-box-like domains. In the TBL1XR1-HMGA1 fusion transcript, furthermore, untranslated exons of HMGA1 are replaced by the first 5 exons of the TBL1XR1 gene. The result is that the entire coding region of HMGA1 comes under the control of the TBL1XR1 promoter, bringing about dysregulation of HMGA1. This is reminiscent of similar pathogenetic mechanisms involving high mobility genes in benign connective tissue tumors such as lipomas and leiomyomas.

  5. ETS Gene Fusions as Predictive Biomarkers of Resistance to Radiation Therapy for Prostate Cancer (United States)


    facilitates DNA end processing and rejoin- ing in a multistep procedure that requires the XRCC4/ DNA Ligase IV complex. In fact, XRCC4 and DNA Ligase IV are...Targeted disruption of the gene encoding DNA ligase IV leads to lethality in embryonic mice. Curr. Biol. 8, 1395–1398. Bennett, E.J., and Harper, J.W. (2008...1998). Late embryonic lethality and impaired V(D)J recombination in mice lacking DNA ligase IV. Nature 396, 173–177. Gallagher, D.J., Gaudet, M.M

  6. Role of the three-dimensional distribution of abl and bcr genes in the formation of bcr/abl fusion gene in interphase neucleus%间期细胞核中abl和bcr基因的三维空间分布在bcr/abl融合基因形成中的作用

    Institute of Scientific and Technical Information of China (English)

    张青; 周淑芸; 刘晓力; 牛超; 徐岚; 陈赛娟



  7. Mutations in the Drosophila pushover gene confer increased neuronal excitability and spontaneous synaptic vesicle fusion

    Energy Technology Data Exchange (ETDEWEB)

    Richards, S.; Hillman, T.; Stern, M. [Rice Univ., Houston, TX (United States)


    We describe the identification of a gene called pushover (push), which affects both behavior and synaptic transmission at the neuromuscular junction. Adults carrying either of two mutations in push exhibit sluggishness, uncoordination, a defective escape response, and male sterility. Larvae defective in push exhibit increased release of transmitter at the neuromuscular junction. In particular, the frequency of spontaneous transmitter release and the amount of transmitter release evoked by nerve stimulation are each increased two- to threefold in push mutants at the lowest external [(Ca{sup 2+})] tested (0.15 mM). Furthermore, these mutants are more sensitive than wild type to application of the potassium channel-blocking drug quinidine: following quinidine application, push mutants, but not wild-type, display repetitive firing of the motor axon, leading to repetitive muscle postsynaptic potentials. The push gene thus might affect both neuronal excitability and the transmitter release process. Complementation tests and recombinational mapping suggest that the push mutations are allelic to a previously identified P-element-induced mutation, which also causes behavorial abnormalities and male sterility. 43 refs., 5 figs., 1 tab.

  8. Using a GFP-gene fusion technique to study the cell cycle-dependent distribution of calmodulin in living cells

    Institute of Scientific and Technical Information of China (English)

    李朝军; 吕品; 张东才


    In this study, a green fluorescent protein (GFP)-calmodulin (CaM) fusion gene method was used to examine the distribution of calmodulin during various stages of cell cycle. First, it was found that the distribution of CaM in living cells changes with the cell cycle. CaM was found mainly in the cytoplasm during G1 phase. It began to move into the nucleus when the cell entered S phase. At G2 phase, CaM became more concentrated in the nucleus than in cytoplasm. Second, the accumulation of CaM in the nucleus during G2 phase appeared to be related to the onset of mitosis, since inhibiting the activation of CaM at this stage resulted in blocking the nuclear membrane breakdown and chromatin condensation. Finally, after the cell entered mitosis, a high concentration of CaM was found at the polar regions of the mitotic spindle. At this time, inhibiting the activity of CaM would cause a disruption of the spindle structure. The relationship between the stage-specific distribution of CaM and its function in regulat

  9. The molecular pathology examination analysis of BRAF gene mutation and RET fusion gene in thyroid cancer%甲状腺癌中BARF基因突变和RET融合基因的分子病理检测分析

    Institute of Scientific and Technical Information of China (English)

    吴永芳; 许春伟; 宋业颖; 班怡; 张博; 李晓兵


    Objective: To investigate the mutations of BRAF gene and RET fusion gene in thyroid cancer. Methods:Taqman-ARMS was used to test in 106 cases of thyroid cancer with BRAF gene mutation and RET fusion gene. Results:hTe total mutation rate of BRAF gene was 66.98%(71/106) in thyroid cancer, and they were all V600E mutation. BRAF gene mutation with age (0.05). Conclusion:BRAF gene mutation has a higher mutation rate in thyroid cancer, especially in young, capsule invasion or metastasis, andⅠorⅡstage. Fusion partner CCDC6-or NCOA4-is common RET fusion gene in thyroid cancer.%目的:探讨甲状腺癌中BRAF基因突变和RET融合基因各亚型情况。方法:应用Taqman-ARMS方法检测106例甲状腺癌中BRAF基因突变和RET融合基因情况。结果:甲状腺癌中BRAF基因总突变率为66.98%(71/106),均为V600E突变,BRAF基因突变在年龄(0.05)。结论:甲状腺癌患者中BRAF基因存在较高的突变率,低龄、被膜未浸润或转移及Ⅰ或Ⅱ期甲状腺癌多见,RET融合基因中融合伙伴CCDC6-或NCOA4-较为常见。

  10. The Neurospora Transcription Factor ADV-1 Transduces Light Signals and Temporal Information to Control Rhythmic Expression of Genes Involved in Cell Fusion

    Directory of Open Access Journals (Sweden)

    Rigzin Dekhang


    Full Text Available Light and the circadian clock have a profound effect on the biology of organisms through the regulation of large sets of genes. Toward understanding how light and the circadian clock regulate gene expression, we used genome-wide approaches to identify the direct and indirect targets of the light-responsive and clock-controlled transcription factor ADV-1 in Neurospora crassa. A large proportion of ADV-1 targets were found to be light- and/or clock-controlled, and enriched for genes involved in development, metabolism, cell growth, and cell fusion. We show that ADV-1 is necessary for transducing light and/or temporal information to its immediate downstream targets, including controlling rhythms in genes critical to somatic cell fusion. However, while ADV-1 targets are altered in predictable ways in Δadv-1 cells in response to light, this is not always the case for rhythmic target gene expression. These data suggest that a complex regulatory network downstream of ADV-1 functions to generate distinct temporal dynamics of target gene expression relative to the central clock mechanism.

  11. Genes encoding Cher-TPR fusion proteins are predominantly found in gene clusters encoding chemosensory pathways with alternative cellular functions. (United States)

    Muñoz-Martínez, Francisco; García-Fontana, Cristina; Rico-Jiménez, Miriam; Alfonso, Carlos; Krell, Tino


    Chemosensory pathways correspond to major signal transduction mechanisms and can be classified into the functional families flagellum-mediated taxis, type four pili-mediated taxis or pathways with alternative cellular functions (ACF). CheR methyltransferases are core enzymes in all of these families. CheR proteins fused to tetratricopeptide repeat (TPR) domains have been reported and we present an analysis of this uncharacterized family. We show that CheR-TPRs are widely distributed in GRAM-negative but almost absent from GRAM-positive bacteria. Most strains contain a single CheR-TPR and its abundance does not correlate with the number of chemoreceptors. The TPR domain fused to CheR is comparatively short and frequently composed of 2 repeats. The majority of CheR-TPR genes were found in gene clusters that harbor multidomain response regulators in which the REC domain is fused to different output domains like HK, GGDEF, EAL, HPT, AAA, PAS, GAF, additional REC, HTH, phosphatase or combinations thereof. The response regulator architectures coincide with those reported for the ACF family of pathways. Since the presence of multidomain response regulators is a distinctive feature of this pathway family, we conclude that CheR-TPR proteins form part of ACF type pathways. The diversity of response regulator output domains suggests that the ACF pathways form a superfamily which regroups many different regulatory mechanisms, in which all CheR-TPR proteins appear to participate. In the second part we characterize WspC of Pseudomonas putida, a representative example of CheR-TPR. The affinities of WspC-Pp for S-adenosylmethionine and S-adenosylhomocysteine were comparable to those of prototypal CheR, indicating that WspC-Pp activity is in analogy to prototypal CheRs controlled by product feed-back inhibition. The removal of the TPR domain did not impact significantly on the binding constants and consequently not on the product feed-back inhibition. WspC-Pp was found to be

  12. Intraparenchymal mesenchymal chondrosarcoma of the frontal lobe--a case report and molecular detection of specific gene fusions from archival FFPE sample. (United States)

    Sajjad, Emir Ahmed; Sikora, Katarzyna; Paciejewski, Tomasz; Garbicz, Filip; Paskal, Wiktor; Szacht, Milena; Grajkowska, Wieslawa; Włodarski, Pawel Krzysztof


    Mesenchymal chondrosarcoma is a rare tumor of cartilaginous origin characterized by its bimorphic pattern composed of highly undifferentiated small round cells separated by islands of well-differentiated hyaline cartilage. It exhibits higher malignancy and earlier occurrence in comparison to classic chondrosarcomas. Recently identified HEY1-NCOA2 and IRF2BP2-CDX1 gene fusions confirm their distinct molecular origin and pose a promising diagnostic marker. The majority of cases arise from craniofacial bones. In this study, we present a rare case of mesenchymal chondrosarcoma encompassed within the brain parenchyma of the frontal lobe without any dural or bone attachment. We demonstrate histopathological findings and confirm the HEY1-NCOA2 gene fusion in a formalin-fixed paraffin-embedded archival sample using simple reverse transcription polymerase chain reaction (RT-PCR) method. IRF2BP2-CDX1 gene fusion was absent in the analyzed sample. The clinical follow-up is also presented with a review of treatment modalities for this entity.

  13. Atrophic dermatofibrosarcoma protuberans with the fusion gene COL1A1-PDGFB detected by RT-PCR using only a single primer pair. (United States)

    Xu, Wen-Jun; Wang, Ju-Sheng


    Dermatofibrosarcoma protuberans (DFSPs) is an uncommon dermal tumor of intermediate to low-grade malignancy. A few patients have clinically persistent plaques that might be atrophic, and they are difficult to be diagnosed clinically. With the development of cytogenetic and molecular biology techniques, the detection of fusion transcripts of the collagen type 1a1 (COL1A1) and platelet-derived growth factor-BB (PDGFB) genes has been recognized as a reliable and valuable molecular tool for the diagnosis of DFSPs. We reported a 24-year-old woman who had a 2 years history of atrophic DFSPs, and detected the gene fusion between COL1A1 to PDGFB by one-step method of RT-PCR using only a single primer pair. The gene fusion detected by this rapid and efficient one-step method in our patient appears to be the first report of atrophic DFSPs, and we detected a novel COL1A1 breakpoint between exon 2 and exon 3.

  14. Genetic interaction between Tmprss2-ERG gene fusion and Nkx3.1-loss does not enhance prostate tumorigenesis in mouse models. (United States)

    Linn, Douglas E; Bronson, Roderick T; Li, Zhe


    Gene fusions involving ETS family transcription factors (mainly TMPRSS2-ERG and TMPRSS2-ETV1 fusions) have been found in ~50% of human prostate cancer cases. Although expression of TMPRSS2-ERG or TMPRSS2-ETV1 fusion alone is insufficient to initiate prostate tumorigenesis, they appear to sensitize prostate epithelial cells for cooperation with additional oncogenic mutations to drive frank prostate adenocarcinoma. To search for such ETS-cooperating oncogenic events, we focused on a well-studied prostate tumor suppressor NKX3.1, as loss of NKX3.1 is another common genetic alteration in human prostate cancer. Previous studies have shown that deletions at 8p21 (harboring NKX3.1) and 21q22 (resulting in TMPRSS2-ERG fusion) were both present in a subtype of prostate cancer cases, and that ERG can lead to epigenetic silencing of NKX3.1 in prostate cancer cells, whereas NKX3.1 can in turn negatively regulate TMPRSS2-ERG fusion expression via suppression of the TMPRSS2 promoter activity. We recently generated knockin mouse models for TMPRSS2-ERG and TMPRSS2-ETV1 fusions, utilizing the endogenous Tmprss2 promoter. We crossed these knockin models to an Nkx3.1 knockout mouse model. In Tmprss2-ERG;Nkx3.1+/- (or -/-) male mice, although we observed a slight but significant upregulation of Tmprss2-ERG fusion expression upon Nkx3.1 loss, we did not detect any significant cooperation between these two genetic events to enhance prostate tumorigenesis in vivo. Furthermore, retrospective analysis of a previously published human prostate cancer dataset revealed that within ERG-overexpressing prostate cancer cases, NKX3.1 loss or deletion did not predict biochemical relapse after radical prostatectomy. Collectively, these data suggest that although TMPRSS2-ERG fusion and loss of NKX3.1 are among the most common mutational events found in prostate cancer, and although each of them can sensitize prostate epithelial cells for cooperating with other oncogenic events, these two events

  15. Comparative study of DNA damage, cell cycle and apoptosis in human K562 and CCRF-CEM leukemia cells: role of BCR/ABL in therapeutic resistance. (United States)

    Pytel, Dariusz; Wysocki, Tomasz; Majsterek, Ireneusz


    The Philadelphia translocation t(9;22) resulting in the bcr/abl fusion gene is the pathogenic principle of almost 95% of human chronic myelogenous leukemia (CML). Imatinib mesylate (STI571) is a specific inhibitor of the BCR/ABL fusion tyrosine kinase that exhibits potent antileukemic effects in CML. BCR/ABL-positive K562 and -negative CCRF-CEM human leukemia cells were investigated. MTT survival assay and clonogenic test of the cell proliferation ability were used to estimate resistance against idarubicin. DNA damage after cell treatment with the drug at the concentrations from 0.001 to 3 microM with or without STI571 pre-treatment were examined by the alkaline comet assay. We found that the level of DNA damages was lower in K562 cells after STI571 pre-treatment. It is suggested that BCR/ABL activity may promote genomic instability, moreover K562 cells were found to be resistant to the drug treatment. Further, we provided evidence of apoptosis inhibition in BCR/ABL-positive cells using caspase-3 activity colorimetric assay and DAPI nuclear staining for chromatin condensation. We suggest that these processes associated with cell cycle arrest in G2/M checkpoint detected in K562 BCR/ABL-positive compared to CCRF-CEM cells without BCR/ABL expression might promote clone selection resistance to drug treatment.

  16. Pandemic H1N1 influenza A directly induces a robust and acute inflammatory gene signature in primary human bronchial epithelial cells downstream of membrane fusion

    Energy Technology Data Exchange (ETDEWEB)

    Paquette, Stéphane G. [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Institute of Medical Science, Faculty of Medicine, University of Toronto, Toronto, Ontario (Canada); Banner, David [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Chi, Le Thi Bao [Department of Microbiology, Hue University of Medicine and Pharmacy, Thua Thien Hue (Viet Nam); Carlo Urbani Centre, Hue University of Medicine and Pharmacy, Thua Thien Hue (Viet Nam); Leon, Alberto J. [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); International Institute of Infection and Immunity, Shantou University Medical College, Shantou, Guangdong (China); Xu, Luoling; Ran, Longsi [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Huang, Stephen S.H. [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); Department of Immunology, Faculty of Medicine, University of Toronto, Toronto, Ontario (Canada); Farooqui, Amber [Division of Experimental Therapeutics, Toronto General Hospital Research Institute, University Health Network, Toronto, Ontario (Canada); International Institute of Infection and Immunity, Shantou University Medical College, Shantou, Guangdong (China); and others


    Pandemic H1N1 influenza A (H1N1pdm) elicits stronger pulmonary inflammation than previously circulating seasonal H1N1 influenza A (sH1N1), yet mechanisms of inflammatory activation in respiratory epithelial cells during H1N1pdm infection are unclear. We investigated host responses to H1N1pdm/sH1N1 infection and virus entry mechanisms in primary human bronchial epithelial cells in vitro. H1N1pdm infection rapidly initiated a robust inflammatory gene signature (3 h post-infection) not elicited by sH1N1 infection. Protein secretion inhibition had no effect on gene induction. Infection with membrane fusion deficient H1N1pdm failed to induce robust inflammatory gene expression which was rescued with restoration of fusion ability, suggesting H1N1pdm directly triggered the inflammatory signature downstream of membrane fusion. Investigation of intra-virion components revealed H1N1pdm viral RNA (vRNA) triggered a stronger inflammatory phenotype than sH1N1 vRNA. Thus, our study is first to report H1N1pdm induces greater inflammatory gene expression than sH1N1 in vitro due to direct virus–epithelial cell interaction. - Highlights: • We investigated H1N1pdm/sH1N1 infection in primary epithelial cells. • H1N1pdm directly initiated a robust inflammatory gene signature, sH1N1 did not. • H1N1pdm viral RNA triggered a stronger response than sH1N1. • H1N1pdm induces greater response due to direct virus–cell interaction. • These results have potential to impact vaccine and therapeutic development.

  17. Two novel alkane hydroxylase-rubredoxin fusion genes isolated from a Dietzia bacterium and the functions of fused rubredoxin domains in long-chain n-alkane degradation. (United States)

    Nie, Yong; Liang, Jieliang; Fang, Hui; Tang, Yue-Qin; Wu, Xiao-Lei


    Two alkane hydroxylase-rubredoxin fusion gene homologs (alkW1 and alkW2) were cloned from a Dietzia strain, designated DQ12-45-1b, which can grow on crude oil and n-alkanes ranging in length from 6 to 40 carbon atoms as sole carbon sources. Both AlkW1 and AlkW2 have an integral-membrane alkane monooxygenase (AlkB) conserved domain and a rubredoxin (Rd) conserved domain which are fused together. Phylogenetic analysis showed that these two AlkB-fused Rd domains formed a novel third cluster with all the Rds from the alkane hydroxylase-rubredoxin fusion gene clusters in Gram-positive bacteria and that this third cluster was distant from the known AlkG1- and AlkG2-type Rds. Expression of the alkW1 gene in DQ12-45-1b was induced when cells were grown on C(8) to C(32) n-alkanes as sole carbon sources, but expression of the alkW2 gene was not detected. Functional heterologous expression in an alkB deletion mutant of Pseudomonas fluorescens KOB2Δ1 suggested the alkW1 could restore the growth of KOB2Δ1 on C(14) and C(16) n-alkanes and induce faster growth on C(18) to C(32) n-alkanes than alkW1ΔRd, the Rd domain deletion mutant gene of alkW1, which also caused faster growth than KOB2Δ1 itself. In addition, the artificial fusion of AlkB from the Gram-negative P. fluorescens CHA0 and the Rds from both Gram-negative P. fluorescens CHA0 and Gram-positive Dietzia sp. DQ12-45-1b significantly increased the degradation of C(32) alkane compared to that seen with AlkB itself. In conclusion, the alkW1 gene cloned from Dietzia species encoded an alkane hydroxylase which increased growth on and degradation of n-alkanes up to C(32) in length, with its fused rubredoxin domain being necessary to maintain the functions. In addition, the fusion of alkane hydroxylase and rubredoxin genes from both Gram-positive and -negative bacteria can increase the degradation of long-chain n-alkanes (such as C(32)) in the Gram-negative bacterium.

  18. Biodegradable charged polyester-based vectors (BCPVs) as an efficient non-viral transfection nanoagent for gene knockdown of the BCR-ABL hybrid oncogene in a human chronic myeloid leukemia cell line (United States)

    Yang, Chengbin; Panwar, Nishtha; Wang, Yucheng; Zhang, Butian; Liu, Maixian; Toh, Huiting; Yoon, Ho Sup; Tjin, Swee Chuan; Chong, Peter Han Joo; Law, Wing-Cheung; Chen, Chih-Kuang; Yong, Ken-Tye


    First-line therapy of chronic myelogenous leukemia (CML) has always involved the use of BCR-ABL tyrosine-kinase inhibitors which is associated with an abnormal chromosome called Philadelphia chromosome. Although the overall survival rate has been improved by the current therapeutic regime, the presence of resistance has resulted in limited efficacy. In this study, an RNA interference (RNAi)-based therapeutic regime is proposed with the aim to knockdown the BCR-ABL hybrid oncogene using small interfering RNA (siRNA). The siRNA transfection rates have usually been limited due to the declining contact probability among polyplexes and the non-adherent nature of leukemic cells. Our work aims at addressing this limitation by using a biodegradable charged polyester-based vector (BCPV) as a nanocarrier for the delivery of BCR-ABL-specific siRNA to the suspension culture of a K562 CML cell line. BCR-ABL siRNAs were encapsulated in the BCPVs by electrostatic force. Cell internalization was facilitated by the BCPV and assessed by confocal microscopy and flow cytometry. The regulation of the BCR-ABL level in K562 cells as a result of RNAi was analyzed by real-time polymerase chain reaction (RT-PCR). We observed that BCPV was able to form stable nanoplexes with siRNA molecules, even in the presence of fetal bovine serum (FBS), and successfully assisted in vitro siRNA transfection in the non-adherent K562 cells. As a consequence of downregulation of BCR-ABL, BCPV-siRNA nanoplexes inhibited cell proliferation and promoted cell apoptosis. All results were compared with a commercial transfection reagent, Lipofectamine2000™, which served as a positive control. More importantly, this class of non-viral vector exhibits biodegradable features and negligible cytotoxicity, thus providing a versatile platform to deliver siRNA to non-adherent leukemia cells with high transfection efficiency by effectively overcoming extra- and intra-cellular barriers. Due to the excellent in vitro

  19. Preliminary comparing the toxicities of the hybrid cry1Acs fused with different heterogenous genes provided guidance for the fusion expression of Cry proteins. (United States)

    Tang, Ying; Tong, Jinying; Zhang, Yunlei; Wang, Lei; Hu, Shengbiao; Li, Wenping; Lv, Yuan


    In order to provide guidance for selecting suitable heterogenous gene that can efficiently enhance toxicity or broaden insecticidal spectrum of Cry1Ac through fusion expression, two hybrid cry1Acs fused with chitinase-encoding gene tchiB and neurotoxin gene hwtx-1 respectively were constructed and their toxicities were compared. A Bacillus thuringiensis strain harboring the cry1Ac gene in vector pHT315 was used as control. Bioassay revealed that LC(50) (after 72 h) of Cry1Ac protoxin was 41.01 μg mL(-1), while the hybrid cry1Acs fused with tchiB and hwtx-1 were 4.89 and 23.14 μg mL(-1), which were 8.23- and 1.77-fold higher than Cry1Ac protoxin in terms of relative toxicity respectively. Both fusion crystals had a higher toxicity than the original Cry1Ac protein and the toxicity of hybrid cry1Acs fused with hwtx-1 experienced a more significant increase than that fused with tchiB.

  20. Horizontal transmission of malignancy: in-vivo fusion of human lymphomas with hamster stroma produces tumors retaining human genes and lymphoid pathology.

    Directory of Open Access Journals (Sweden)

    David M Goldenberg

    Full Text Available We report the in-vivo fusion of two Hodgkin lymphomas with golden hamster cheek pouch cells, resulting in serially-transplanted (over 5-6 years GW-532 and GW-584 heterosynkaryon tumor cells displaying both human and hamster DNA (by FISH, lymphoma-like morphology, aggressive metastasis, and retention of 7 human genes (CD74, CXCR4, CD19, CD20, CD71, CD79b, and VIM out of 24 tested by PCR. The prevalence of B-cell restricted genes (CD19, CD20, and CD79b suggests that this uniform population may be the clonal initiating (malignant cells of Hodgkin lymphoma, despite their not showing translation to their respective proteins by immunohistochemical analysis. This is believed to be the first report of in-vivo cell-cell fusion of human lymphoma and rodent host cells, and may be a method to disclose genes regulating both organoid and metastasis signatures, suggesting that the horizontal transfer of tumor DNA to adjacent stromal cells may be implicated in tumor heterogeneity and progression. The B-cell gene signature of the hybrid xenografts suggests that Hodgkin lymphoma, or its initiating cells, is a B-cell malignancy.

  1. Detection of BCR/ABL Fusion Gene in Interphase Cens with Bi-labelling Hybridization in Situ%双标记原位杂交法检测间期细胞BCR/ABL融合基因

    Institute of Scientific and Technical Information of China (English)

    袁长吉; 王建伟; 李舜华; 易永林



  2. Acute promyelocytic leukaemia in patients originating in Latin America is associated with an increased frequency of the bcr1 subtype of the PML/RARalpha fusion gene. (United States)

    Douer, Dan; Santillana, Sergio; Ramezani, Laleh; Samanez, Cesar; Slovak, Marilyn L; Lee, Ming S; Watkins, Kristy; Williams, Tony; Vallejos, Carlos


    The PML/RARalpha fusion gene in acute promyelocytic leukaemia (APL) has three subtypes based on the breakpoint site of the PML gene: long (bcr1), short (bcr3) and variable (bcr2) subtypes. The PML/RARalpha fusion protein is involved in the pathogenesis of APL and the breakpoint site of the PML gene might be associated with aetiological factor(s). Because APL is over-represented in patients that originate in Latin America (Latinos), we evaluated whether the distribution of the PML/RARalpha fusion mRNA in this population is different to that reported in non-Latinos. Among 52 APL patients (28 from Mexico and Central America diagnosed in Los Angeles and 24 from Peru, South America), bcr1, bcr2 and bcr3 expression was 75%, 10% and 15% respectively. However, bcr1 breakpoints were significantly higher compared with non-Latino patients (340/654, 52%) reported in four studies. Often bcr1 and bcr2 are reported together; 862 (60%) of 1429 non-Latino APL patients reported in nine studies were either bcr1 or bcr2, compared with 44 (85%) in our 52 Latino patients. This difference was also statistically significant when our patients were compared to each of the individual studies from USA and Europe, but not for a small series from China and Japan. These results suggest that the overrepresentation of APL among Latin American patients can be accounted for by an increase of a single subtype--bcr1, and the breakage sites in the PML gene may not be random but possibly influenced by genetic and/or environmental factor(s).

  3. Detection of Echinoderm Microtubule Associated Protein Like 4-Anaplastic Lymphoma Kinase Fusion Genes in Non-small Cell Lung Cancer Clinical Samples by a Real-time Quantitative Reverse Transcription Polymerase Chain Reaction Method. (United States)

    Zhao, Jing; Zhao, Jin-Yin; Chen, Zhi-Xia; Zhong, Wei; Li, Long-Yun; Liu, Li-Cheng; Hu, Xiao-Xu; Chen, Wei-Jun; Wang, Meng-Zhao


    Objective To establish a real-time quantitative reverse transcription polymerase chain reaction assay (qRT-PCR) for the rapid, sensitive, and specific detection of echinoderm microtubule associated protein like 4-anaplastic lymphoma kinase (EML4-ALK) fusion genes in non-small cell lung cancer. Methods The specific primers for the four variants of EML4-ALK fusion genes (V1, V2, V3a, and V3b) and Taqman fluorescence probes for the detection of the target sequences were carefully designed by the Primer Premier 5.0 software. Then, using pseudovirus containing EML4-ALK fusion genes variants (V1, V2, V3a, and V3b) as the study objects, we further analyzed the lower limit, sensitivity, and specificity of this method. Finally, 50 clinical samples, including 3 ALK-fluorescence in situ hybridization (FISH) positive specimens, were collected and used to detect EML4-ALK fusion genes using this method. Results The lower limit of this method for the detection of EML4-ALK fusion genes was 10 copies/μl if no interference of background RNA existed. Regarding the method's sensitivity, the detection resolution was as high as 1% and 0.5% in the background of 500 and 5000 copies/μl wild-type ALK gene, respectively. Regarding the method's specificity, no non-specific amplification was found when it was used to detect EML4-ALK fusion genes in leukocyte and plasma RNA samples from healthy volunteers. Among the 50 clinical samples, 47 ALK-FISH negative samples were also negative. Among 3 ALK-FISH positive samples, 2 cases were detected positive using this method, but another was not detected because of the failure of RNA extraction. Conclusion The proposed qRT-PCR assay for the detection of EML4-ALK fusion genes is rapid, simple, sensitive, and specific, which is deserved to be validated and widely used in clinical settings.

  4. Differential transactivation by orphan nuclear receptor NOR1 and its fusion gene product EWS/NOR1: possible involvement of poly(ADP-ribose) polymerase I, PARP-1. (United States)

    Ohkura, Naganari; Nagamura, Yuko; Tsukada, Toshihiko


    In extraskeletal myxoid chondrosarcoma, a chromosomal translocation creates a gene fusion between EWS and an orphan nuclear receptor, NOR1. The resulting fusion protein EWS/NOR1 has been believed to lead to malignant transformation by functioning as a transactivator for NOR1-target genes. By comparing the gene expression profiles of NOR1- and EWS/NOR1-overexpressing cells, we found that they largely shared up-regulated genes, but no significant correlation was observed with respect to the transactivation levels of each gene. In addition, the proteins associated with NOR1 and EWS/NOR1 were mostly the same in these cells. The results suggest that these proteins differentially transactivate overlapping target genes through a similar transcriptional machinery. To clarify the mechanisms underlying the transcriptional divergence between NOR1 and EWS/NOR1, we searched for alternatively associated proteins, and identified poly(ADP-ribose) polymerase I (PARP-1) as an NOR1-specific binding protein. Consistent with its binding properties, PARP-1 acted as a transcriptional repressor of NOR1, but not EWS/NOR1, in a luciferase reporter assay employing PARP-1(-/-) fibroblasts. Interestingly, suppressive activity of PARP-1 was observed in a DNA response element-specific manner, and in a subtype-specific manner toward the NR4A family (Nur77, Nurr1, and NOR1), suggesting that PARP-1 plays a role in the diversity of transcriptional regulation mediated by the NR4A family in normal cells. Altogether, our findings suggest that NOR1 and EWS/NOR1 regulate overlapping target genes differently by utilizing associated proteins, including PARP-1; and that EWS/NOR1 may acquire oncogenic activities by avoiding (or gaining) transcription factor-specific modulation by the associated proteins.

  5. Osteoclast Fusion

    DEFF Research Database (Denmark)

    Marie Julie Møller, Anaïs; Delaissé, Jean-Marie; Søe, Kent


    suggesting that fusion partners may specifically select each other and that heterogeneity between the partners seems to play a role. Therefore, we set out to directly test the hypothesis that fusion factors have a heterogenic involvement at different stages of nuclearity. Therefore, we have analyzed...... on the nuclearity of fusion partners. While CD47 promotes cell fusions involving mono-nucleated pre-osteoclasts, syncytin-1 promotes fusion of two multi-nucleated osteoclasts, but also reduces the number of fusions between mono-nucleated pre-osteoclasts. Furthermore, CD47 seems to mediate fusion mostly through......Investigations addressing the molecular keys of osteoclast fusion are primarily based on end-point analyses. No matter if investigations are performed in vivo or in vitro the impact of a given factor is predominantly analyzed by counting the number of multi-nucleated cells, the number of nuclei per...

  6. CDKN2D-WDFY2 is a cancer-specific fusion gene recurrent in high-grade serous ovarian carcinoma.

    Directory of Open Access Journals (Sweden)

    Kalpana Kannan


    Full Text Available Ovarian cancer is the fifth leading cause of cancer death in women. Almost 70% of ovarian cancer deaths are due to the high-grade serous subtype, which is typically detected only after it has metastasized. Characterization of high-grade serous cancer is further complicated by the significant heterogeneity and genome instability displayed by this cancer. Other than mutations in TP53, which is common to many cancers, highly recurrent recombinant events specific to this cancer have yet to be identified. Using high-throughput transcriptome sequencing of seven patient samples combined with experimental validation at DNA, RNA and protein levels, we identified a cancer-specific and inter-chromosomal fusion gene CDKN2D-WDFY2 that occurs at a frequency of 20% among sixty high-grade serous cancer samples but is absent in non-cancerous ovary and fallopian tube samples. This is the most frequent recombinant event identified so far in high-grade serous cancer implying a major cellular lineage in this highly heterogeneous cancer. In addition, the same fusion transcript was also detected in OV-90, an established high-grade serous type cell line. The genomic breakpoint was identified in intron 1 of CDKN2D and intron 2 of WDFY2 in patient tumor, providing direct evidence that this is a fusion gene. The parental gene, CDKN2D, is a cell-cycle modulator that is also involved in DNA repair, while WDFY2 is known to modulate AKT interactions with its substrates. Transfection of cloned fusion construct led to loss of wildtype CDKN2D and wildtype WDFY2 protein expression, and a gain of a short WDFY2 protein isoform that is presumably under the control of the CDKN2D promoter. The expression of short WDFY2 protein in transfected cells appears to alter the PI3K/AKT pathway that is known to play a role in oncogenesis. CDKN2D-WDFY2 fusion could be an important molecular signature for understanding and classifying sub-lineages among heterogeneous high-grade serous ovarian

  7. CDKN2D-WDFY2 Is a Cancer-Specific Fusion Gene Recurrent in High-Grade Serous Ovarian Carcinoma (United States)

    Rajapakshe, Kimal; Hawkins, Shannon M.; Matzuk, Martin M.; Milosavljevic, Aleksandar; Yen, Laising


    Ovarian cancer is the fifth leading cause of cancer death in women. Almost 70% of ovarian cancer deaths are due to the high-grade serous subtype, which is typically detected only after it has metastasized. Characterization of high-grade serous cancer is further complicated by the significant heterogeneity and genome instability displayed by this cancer. Other than mutations in TP53, which is common to many cancers, highly recurrent recombinant events specific to this cancer have yet to be identified. Using high-throughput transcriptome sequencing of seven patient samples combined with experimental validation at DNA, RNA and protein levels, we identified a cancer-specific and inter-chromosomal fusion gene CDKN2D-WDFY2 that occurs at a frequency of 20% among sixty high-grade serous cancer samples but is absent in non-cancerous ovary and fallopian tube samples. This is the most frequent recombinant event identified so far in high-grade serous cancer implying a major cellular lineage in this highly heterogeneous cancer. In addition, the same fusion transcript was also detected in OV-90, an established high-grade serous type cell line. The genomic breakpoint was identified in intron 1 of CDKN2D and intron 2 of WDFY2 in patient tumor, providing direct evidence that this is a fusion gene. The parental gene, CDKN2D, is a cell-cycle modulator that is also involved in DNA repair, while WDFY2 is known to modulate AKT interactions with its substrates. Transfection of cloned fusion construct led to loss of wildtype CDKN2D and wildtype WDFY2 protein expression, and a gain of a short WDFY2 protein isoform that is presumably under the control of the CDKN2D promoter. The expression of short WDFY2 protein in transfected cells appears to alter the PI3K/AKT pathway that is known to play a role in oncogenesis. CDKN2D-WDFY2 fusion could be an important molecular signature for understanding and classifying sub-lineages among heterogeneous high-grade serous ovarian carcinomas. PMID

  8. Membrane fusion

    DEFF Research Database (Denmark)

    Bendix, Pól Martin


    At Stanford University, Boxer lab, I worked on membrane fusion of small unilamellar lipid vesicles to flat membranes tethered to glass surfaces. This geometry closely resembles biological systems in which liposomes fuse to plasma membranes. The fusion mechanism was studied using DNA zippering...... between complementary strands linked to the two apposing membranes closely mimicking the zippering mechanism of SNARE fusion complexes....

  9. Estandarización de la TR-PCR para la detección de las fusiones génicas BCR-ABL en pacientes con leucemia Mieloide Crónica (LMC y Linfoide Aguda (LLA provenientes de HUSVP y Clíncia León XIII

    Directory of Open Access Journals (Sweden)

    Gonzálo Vásquez Palacio


    Full Text Available La translocación recíproca t(9:22(q34;q11 involucra el proto-oncogen ABL y el gen BCR, originando un gen de fusión BCR-ABL, que codifica una proteína con elevada actividad tirosina quinasa, implicada en la leucemogénesis.

  10. The evolution of the histone methyltransferase gene Su(var3-9 in metazoans includes a fusion with and a re-fission from a functionally unrelated gene

    Directory of Open Access Journals (Sweden)

    Patties Ina


    Full Text Available Abstract Background In eukaryotes, histone H3 lysine 9 (H3K9 methylation is a common mechanism involved in gene silencing and the establishment of heterochromatin. The loci of the major heterochromatic H3K9 methyltransferase Su(var3-9 and the functionally unrelated γ subunit of the translation initiation factor eIF2 are fused in Drosophila melanogaster. Here we examined the phylogenetic distribution of this unusual gene fusion and the molecular evolution of the H3K9 HMTase Su(var3-9. Results We show that the gene fusion had taken place in the ancestral line of winged insects and silverfishs (Dicondylia about 400 million years ago. We cloned Su(var3-9 genes from a collembolan and a spider where both genes ancestrally exist as independent transcription units. In contrast, we found a Su(var3-9-specific exon inside the conserved intron position 81-1 of the eIF2γ gene structure in species of eight different insect orders. Intriguinly, in the pea aphid Acyrthosiphon pisum, we detected only sequence remains of this Su(var3-9 exon in the eIF2γ intron, along with an eIF2γ-independent Su(var3-9 gene. This reveals an evolutionary re-fission of both genes in aphids. Su(var3-9 chromo domains are similar to HP1 chromo domains, which points to a potential binding activity to methylated K9 of histone H3. SET domain comparisons suggest a weaker methyltransferase activity of Su(var3-9 in comparison to other H3K9 HMTases. Astonishingly, 11 of 19 previously described, deleterious amino acid substitutions found in Drosophila Su(var3-9 are seemingly compensable through accompanying substitutions during evolution. Conclusion Examination of the Su(var3-9 evolution revealed strong evidence for the establishment of the Su(var3-9/eIF2γ gene fusion in an ancestor of dicondylic insects and a re-fission of this fusion during the evolution of aphids. Our comparison of 65 selected chromo domains and 93 selected SET domains from Su(var3-9 and related proteins offers

  11. Temsirolimus in the treatment of renal cell carcinoma associated with Xp11.2 translocation/TFE gene fusion proteins: a case report and review of literature

    Directory of Open Access Journals (Sweden)

    James Brown


    Full Text Available Xp11.2 translocation renal cell carcinomas (TRCCs are a rare family of tumors newly recognized by the World Health Organization (WHO in 2004. These tumors result in the fusion of partner genes to the TFE3 gene located on Xp11.2. They are most common in the pediatric population, but have been recently implicated in adult renal cell carcinoma (RCC presenting at an early age. TFE3-mediated direct transcriptional upregulation of the Met tyrosine kinase receptor triggers dramatic activation of downstream signaling pathways including the protein kinase B (Akt/phosphatidylinositol-3 kinase (PI3K and mammalian target of rapamycin (mTOR pathways. Temsirolimus is an inhibitor of mammalian target of rapamycin (mTOR kinase, a component of intracellular signaling pathways involved in the growth and proliferation of malignant cells. Here we present a case of a 22-year old female who has been treated with temsirolimus for her Xp11.2/TFE3 gene fusion RCC.

  12. Construction of prokaryotic expression system of ItB-ureB fusion gene and identification of the recombinant protein immunity and adjuvanticity

    Institute of Scientific and Technical Information of China (English)

    Jie Yan; Yuan Wang; Shi-He Shao; Ya-Fei Mao; Hua-Wen Li; Yi-Hui Luo


    AIM: To construct ItB-ureB fusion gene and its prokaryotic expression system and identify immunity and adjuvanticity of the expressed recombinant protein.METHODS: The ureB gene from a clinical Helicobacterpylori(Hpylori) strain Y06 and the ItB gene from Escherichiacoli(E coli) strain 44851 were linked into ItB-ureB fusiongene by PCR. The fusion gene sequence was analyzedafter T-A cloning. A prokaryotic recombinant expressionvector pET32a inserted with ItB-ureB fusion gene (pET32aItB-ureB) was constructed. Expression of the recombinantLTB-UreB protein (rLTB-UreB)in E. coliBL21DE3 inducedby isopropylthio-β-D-galactoside (IPTG) at differentconcentrations was detected by SDS-PAGE. Western blot assays were used to examine the immunoreaction of rLTBUreB by a commercial antibody against whole cell of H pylori and a self-prepared rabbit anti-rUreB serum, respectively, and determine the antigenicity of the recombinant proteinon inducing specific antibody in rabbits. GM1-ELISA wasused to demonstrate the adjuvanticity of rLTB-UreB. Immunoreaction of rLTB-UreB to the UreB antibody positivesera from 125 gastric patients was determined by using ELISA. RESULTS: In comparison with the corresponding sequences of original genes, the nucleotide sequence homologies of the cloned ItB-ureB fusion gene were 100%. IPTG withdifferent dosages of 0.1-1.0 mmol/L could efficiently inducepET32a-ItB-ureB-E.coli BL21DE3 to express the rLTB-UreB. The output of the target recombinant protein expressed by pET32a-ureB-E. coli BL21DE3 was approximately 35%of the total bacterial proteins. rLTB-UreB mainly presented in the form of inclusion body. Western blotting results demonstrated that rLTB-UreB could combine with the commercial antibody against whole cell of H pylori andanti-rUreB serum as well as induce rabbit to produce specific antibody. The strong ability of rLTB-UreB bindingbovine GM1 indicated the existence of adjuvanticity of the recombinant protein. All the UreB antibody positive sera from

  13. Checkpoint genes and Exo1 regulate nearby inverted repeat fusions that form dicentric chromosomes in Saccharomyces cerevisiae. (United States)

    Kaochar, Salma; Shanks, Lisa; Weinert, Ted


    Genomic rearrangements are common, occur by largely unknown mechanisms, and can lead to human diseases. We previously demonstrated that some genome rearrangements occur in budding yeast through the fusion of two DNA sequences that contain limited sequence homology, lie in inverted orientation, and are within 5 kb of one another. This inverted repeat fusion reaction forms dicentric chromosomes, which are well-known intermediates to additional rearrangements. We have previously provided evidence indicating that an error of stalled or disrupted DNA replication forks can cause inverted repeat fusion. Here we analyze how checkpoint protein regulatory pathways known to stabilize stalled forks affect this form of instability. We find that two checkpoint pathways suppress inverted repeat fusion, and that their activities are distinguishable by their interactions with exonuclease 1 (Exo1). The checkpoint kinase Rad53 (Chk2) and recombination protein complex MRX(MRN) inhibit Exo1 in one pathway, whereas in a second pathway the ATR-like kinases Mec1 and Tel1, adaptor protein Rad9, and effector kinases Chk1 and Dun1 act independently of Exo1 to prevent inverted repeat fusion. We provide a model that indicates how in Rad53 or MRX mutants, an inappropriately active Exo1 may facilitate faulty template switching between nearby inverted repeats to form dicentric chromosomes. We further investigate the role of Rad53, using hypomorphic alleles of Rad53 and null mutations in Rad9 and Mrc1, and provide evidence that only local, as opposed to global, activity of Rad53 is sufficient to prevent inverted repeat fusion.

  14. Molecular Characterization of TMPRSS2-ERG Gene Fusion in the NCI-H660 Prostate Cancer Cell Line: A New Perspective for an Old Model

    Directory of Open Access Journals (Sweden)

    Kirsten D. Mertz


    Full Text Available Recent studies have established that a significant fraction of prostate cancers harbor a signature gene fusion between the 5' region of androgen-regulated TMPRSS2 and an ETS family transcription factor, most commonly ERG. Studies on the molecular mechanisms and functional consequences of this important chromosomal rearrangement are currently limited to the VCaP cell line derived from a vertebral bone metastasis of a hormone-refractory prostate tumor. Here we report on the NCI-H660 cell line, derived from a metastasic site of an extrapulmonary small cell carcinoma arising from the prostate. NCI-H660 harbors TMPRSS2-ERG fusion with a homozygous intronic deletion between TMPRSS2 and ERG. We demonstrate this by real-time quantitative polymerase chain reaction, a two-stage dual-color interphase fluorescence in situ hybridization (FISH assay testing for TMPRSS2 and ERG break-aparts, and single-nucleotide polymorphism oligonucleotide arrays. The deletion is consistent with the common intronic deletion found on chromosome 21q22.2-3 in human prostate cancer samples. We demonstrate the physical juxtaposition of TMPRSS2 and ERG on the DNA level by fiber FISH. The androgen receptor-negative NCI-H660 cell line expresses ERG in an androgen-independent fashion. This in vitro model system has the potential to provide important pathobiologic insights into TMPRSS2-ERG fusion prostate cancer.

  15. Fusion rings and fusion ideals

    DEFF Research Database (Denmark)

    Andersen, Troels Bak

    by the so-called fusion ideals. The fusion rings of Wess-Zumino-Witten models have been widely studied and are well understood in terms of precise combinatorial descriptions and explicit generating sets of the fusion ideals. They also appear in another, more general, setting via tilting modules for quantum...

  16. Lack of a synergistic effect of a non-viral ALS gene therapy based on BDNF and a TTC fusion molecule

    Directory of Open Access Journals (Sweden)

    Navarro Xavier


    Full Text Available Abstract Background Amyotrophic lateral sclerosis (ALS is one of the most devastating neurodegenerative diseases. Neurotrophic factors have been widely tested to counteract neurodegenerative conditions, despite their unspecific neuronal access. The non-toxic C-terminal fragment of the tetanus toxin (TTC heavy chain has been studied not only as a carrier molecule to the CNS but also as a neuroprotective agent. Because the neurotrophic effects of BDNF have been demonstrated in vitro and in vivo, the question addressed in this work is whether a fusion molecule of BDNF-TTC may have a synergistic effect and enhance the neuroprotective properties of TTC alone in a mouse model of ALS. Methods Recombinant plasmid constructs (pCMV-TTC and pCMV-BDNF-TTC were injected into the quadriceps femoris and triceps brachialis muscles of SOD1G93A transgenic mice at 8 weeks of age. The hanging wire and rotarod tests were performed to assess motor coordination, strength and balance. Electrophysiological tests, morphological assays of spinal cord sections of L2 and L4 segments, and gene and protein expression analyses were performed. The Kaplan-Meier survival analysis test was used for comparisons of survival. Multiple comparisons of data were analyzed using a one-way analysis of variance (ANOVA. Results Treatment with the fusion-molecule BDNF-TTC and with TTC alone significantly delayed the onset of symptoms and functional deficits of SOD1G93A mice. Muscle innervation was partially preserved with these treatments, and the number of surviving motoneurons in L2 spinal cord segment was increased particularly by the fusion protein induction. Inhibition of pro-apoptotic protein targets (caspase-3 and Bax and significant phosphorylation of Akt and ERK were also found in the spinal cord of treated mice. Conclusions Significant improvements in behavioral and electrophysiological results, motoneuron survival and anti-apoptotic/survival-activated pathways were observed with

  17. Transformation of Arabidopsis by Rice OsWRKY78:: GFP Fusion Gene and Subcellular Localization of OsWRKY78 Protein

    Institute of Scientific and Technical Information of China (English)

    Shunzhi LIU; Mei ZHANG; Xin TANG; Xiaolan WANG


    [Objective] The study was to understand the subcellular localization of OsWRKY78 protein in plants. [Method] Primers specific for OsWRKY78 gene were designed according to the OsWRKY78 full length sequence in Genbank. The gene was cloned by RT-PCR method. The gene was then recombined into a plasmid ex- pression vector carrying green fluorescent protein (GFP) gene, pBinGFP. The re- combinant was confirmed by PCR and enzyme digestion. The recombinant plasmid pBinGFP-OsWRKY was transformed into Arabidopsis through Agrobacterium tume- faciens strain GV3101 and transgenic plants were obtained. [Result] Measured by fluorescence microscopy, the expression of OsWRKY78 and GFP fusion protein in root tip cells was localized in the nucleus. [Conclusion] This study laid the foundation for further investigating the function of OsWRKY78 gene and its role in related sig- nal transduction and provided theoretical basis for exploring the relation between OsWRKY78 gene and brown planthoppers.

  18. Translocation t(1;11(p32;q23 with MLL-EPS15 fusion gene formation in acute leukemias: a review and 6 new case reports. Approaches to minimal residual disease monitoring

    Directory of Open Access Journals (Sweden)

    G. A. Tsaur


    Full Text Available We performed clinical and laboratory characterization of patients with rare translocation t(1;11(p32;q23 leading to MLL-EPS15 fusion gene formation. Study cohort consisted of 33 primary acute leukemia (AL cases including 6 newly diagnosed and 27 patients previously described in literature. Among study group patients t(1;11(p32;q23 was found most frequently in infant AL cases (median age 8 months. In acute lymphoblastic leukemia (ALL male/female ratio was 1:3, in acute myeloid leukemia (AML it was 1:1. Additional cytogenetic aberrations in 38 % of patients were revealed. The most frequent breakpoint position in EPS15 gene was intron 1. Four different types of MLLEPS15 fusion gene transcripts were detected. Primers-probe-plasmid combination for MLL-EPS15 fusion gene transcript monitoring by realtime quantitative polymerase chain reaction (RQ-PCR was developed and successfully applied. In 3 patients RQ-PCR was done on genomic DNA for absolute quantification of MLL-EPS15 fusion gene. High qualitative concordance rate (92 % was noted between minimal residual disease data obtained in cDNA and genomic DNA for MLL-EPS15 fusion detection.

  19. Translocation t(1;11(p32;q23 with MLL-EPS15 fusion gene formation in acute leukemias: a review and 6 new case reports. Approaches to minimal residual disease monitoring

    Directory of Open Access Journals (Sweden)

    G. A. Tsaur


    Full Text Available We performed clinical and laboratory characterization of patients with rare translocation t(1;11(p32;q23 leading to MLL-EPS15 fusion gene formation. Study cohort consisted of 33 primary acute leukemia (AL cases including 6 newly diagnosed and 27 patients previously described in literature. Among study group patients t(1;11(p32;q23 was found most frequently in infant AL cases (median age 8 months. In acute lymphoblastic leukemia (ALL male/female ratio was 1:3, in acute myeloid leukemia (AML it was 1:1. Additional cytogenetic aberrations in 38 % of patients were revealed. The most frequent breakpoint position in EPS15 gene was intron 1. Four different types of MLLEPS15 fusion gene transcripts were detected. Primers-probe-plasmid combination for MLL-EPS15 fusion gene transcript monitoring by realtime quantitative polymerase chain reaction (RQ-PCR was developed and successfully applied. In 3 patients RQ-PCR was done on genomic DNA for absolute quantification of MLL-EPS15 fusion gene. High qualitative concordance rate (92 % was noted between minimal residual disease data obtained in cDNA and genomic DNA for MLL-EPS15 fusion detection.

  20. ABL Tyrosine Kinase Stimulates PUMA Protein Expression


    Oon, Chet K


    ABL is an ubiquitously expressed non-receptor tyrosine kinase involved in multiple cellular functions including programmed cell death. Upon DNA damage, ABL has been shown to upregulate PUMA, p53 upregulated modulator of apoptosis, and causes downstream mitochondrial intrinsic apoptotic events. However, the mechanism by which ABL regulates PUMA expression remains unknown. We have shown that ABL does not change PUMA protein subcellular localization through immunofluorescence. Through protein an...

  1. BCR-ABL1 kinase inhibits uracil DNA glycosylase UNG2 to enhance oxidative DNA damage and stimulate genomic instability (United States)

    Slupianek, Artur; Falinski, Rafal; Znojek, Pawel; Stoklosa, Tomasz; Flis, Sylwia; Doneddu, Valentina; Pytel, Dariusz; Synowiec, Ewelina; Blasiak, Janusz; Bellacosa, Alfonso; Skorski, Tomasz


    Tyrosine kinase inhibitors (TKIs) revolutionized the treatment of CML-CP. Unfortunately, 25% of TKI-naive patients and 50–90% of TKI-responding patients carry CML clones expressing TKI resistant BCR-ABL1 kinase mutants. We reported that CML-CP leukemia stem and progenitor cell populations accumulate high amounts of reactive oxygen species (ROS), which may result in accumulation of uracil derivatives in genomic DNA. Unfaithful and/or inefficient repair of these lesions generates TKI resistant point mutations in BCR-ABL1 kinase. Using an array of specific substrates and inhibitors/blocking antibodies we found that uracil-DNA glycosylase UNG2 were inhibited in BCR-ABL1 –transformed cell lines and CD34+ CML cells. The inhibitory effect was not accompanied by downregulation of nuclear expression and/or chromatin association of UNG2. The effect was BCR-ABL1 kinase-specific because several other fusion tyrosine kinases did not reduce UNG2 activity. Using UNG2-specific inhibitor UGI we found that reduction of UNG2 activity increased the number of uracil derivatives in genomic DNA detected by modified comet assay and facilitated accumulation of ouabain-resistant point mutations in reporter gene Na+/K+ATPase. In conclusion, we postulate that BCR-ABL1 kinase-mediated inhibition of UNG2 contributes to accumulation of point mutations responsible for TKI-resistance causing the disease relapse, and perhaps also other point mutations facilitating malignant progression of CML. PMID:23047475

  2. Genetic transformation of peanut (Arachis hypogaea L.) using cotyledonary node as explant and a promoterless gus::nptII fusion gene based vector

    Indian Academy of Sciences (India)

    T Swathi Anuradha; S K Jami; R S Datla; P B Kirti


    We have generated putative promoter tagged transgenic lines in Arachis hypogaea cv JL-24 using cotyledonary node (CN) as an explant and a promoterless gus::nptII bifunctional fusion gene mediated by Agrobacterium transformation. MS medium fortified with 6-benzylaminopurine (BAP) at 4 mg/l in combination with 0.1 mg/l -napthaleneacetic acid (NAA) was the most effective out of the various BAP and NAA combinations tested in multiple shoot bud formation. Parameters enhancing genetic transformation viz. seedling age, Agrobacterium genetic background and co-cultivation periods were studied by using the binary vector p35SGUSINT. Genetic transformation with CN explants from 6-dayold seedlings co-cultivated with Agrobacterium GV2260 strain for 3 days resulted in high kanamycin resistant shoot induction percentage (45%); approximately 31% transformation frequency was achieved with p35S GUSINT in -glucuronidase (GUS) assays. Among the in vivo GUS fusions studied with promoterless gus::nptII construct, GUS-positive sectors occupied 38% of the total transient GUS percentage. We have generated over 141 putative T0 plants by using the promoterless construct and transferred them to the field. Among these, 82 plants survived well in the green house and 5 plants corresponding to 3.54% showed stable integration of the fusion gene as evidenced by GUS, polymerase chain reaction (PCR) and Southern blot analyses. Twenty-four plants were positive for GUS showing either tissue-specific expression or blue spots in at least one plant part. The progeny of 15 T0 plants indicated Mendelian inheritance pattern of segregation for single-copy integration. The tissue-specific GUS expression patterns were more or less similar in both T0 and corresponding T1 progeny plants. We present the differential patterns of GUS expression identified in the putative promoter-tagged transgenic lines in the present communication.

  3. High Level Information Fusion (HLIF) with nested fusion loops (United States)

    Woodley, Robert; Gosnell, Michael; Fischer, Amber


    Situation modeling and threat prediction require higher levels of data fusion in order to provide actionable information. Beyond the sensor data and sources the analyst has access to, the use of out-sourced and re-sourced data is becoming common. Through the years, some common frameworks have emerged for dealing with information fusion—perhaps the most ubiquitous being the JDL Data Fusion Group and their initial 4-level data fusion model. Since these initial developments, numerous models of information fusion have emerged, hoping to better capture the human-centric process of data analyses within a machine-centric framework. 21st Century Systems, Inc. has developed Fusion with Uncertainty Reasoning using Nested Assessment Characterizer Elements (FURNACE) to address challenges of high level information fusion and handle bias, ambiguity, and uncertainty (BAU) for Situation Modeling, Threat Modeling, and Threat Prediction. It combines JDL fusion levels with nested fusion loops and state-of-the-art data reasoning. Initial research has shown that FURNACE is able to reduce BAU and improve the fusion process by allowing high level information fusion (HLIF) to affect lower levels without the double counting of information or other biasing issues. The initial FURNACE project was focused on the underlying algorithms to produce a fusion system able to handle BAU and repurposed data in a cohesive manner. FURNACE supports analyst's efforts to develop situation models, threat models, and threat predictions to increase situational awareness of the battlespace. FURNACE will not only revolutionize the military intelligence realm, but also benefit the larger homeland defense, law enforcement, and business intelligence markets.

  4. [Construction of fusion gene vaccine of WT1 multi-epitope fused with stimulating epitope of mycobacterium tuberculosis heat shock protein 70 and its expression and immunogenicity]. (United States)

    Tian, Wei-Wei; Qiao, Zhen-Hua; Yang, Lin-Hua; Wang, Hong-Wei; Tang, Yan-Hong; Bian, Si-Cheng


    This study was purposed to construct a fusion DNA vaccine containing WT1 multi-epitope and stimulating epitope of mycobacterium tuberculosis heat shock protein 70 and to detect its expression and immunogenicity. On the basis of published data, a multi-epitope gene (Multi-WT1) containing three HLA *0201-restricted CTL epitopes: one HLA*2402-restricted CTL epitope, two Th epitopes and one universal Th Pan-DR epitope (PADRE) was constructed. DNA-coding sequence was modified by Computer-Aided Design (CAD) to optimize proteasome-mediated epitope processing through the introduction of different amino acid spacer sequences. The synthetic nucleotide sequence was then inserted into an eukaryotic vector to construct the plasmid pcDNA3.1-WT1.For enhancing CTL activity, HSP70 fragment including stimulatory domain P407-426 was amplified by PCR from mycobacterial HSP70 gene and cloned into pcDNA3.1(+). Then Multi-WT1 was fused to the N-terminal of pcDNA3.1-mHSP70(407-426) to make the multi-epitope fusion gene vaccine pcDNA3.1-WT1-mHSP70(407-426). HEK-293T cells were transfected with this vaccine and the expressed product was identified by RT-PCR. Enzyme-linked immunospot assay (ELISPOT) was used to evaluate the immunological responses elicited by vaccine. The results showed that the most of WT1 epitopes could be correctly cleaved which was confirmed by software Net Chop 3.1 and PAPROCIanalysis. RT-PCR showed correct expression of target gene in HEK293T cells and ELISPOT showed specific T-cell responses. It is concluded that the eukaryotic expression vector PcDNA3.1-WT1-mHSP70(407-426) fusion gene has been successfully constructed and the immunity response is also elicited, which is a good candidate for further research of DNA vaccine.

  5. Renal Cell Carcinoma Associated with Xp11.2 Translocation/TFE3 Gene Fusion: A Rare Case Report with Review of the Literature

    Directory of Open Access Journals (Sweden)

    Puneet Ahluwalia


    Full Text Available Introduction. The recently recognized renal cell carcinomas associated with Xp11.2 translocations are rare tumors predominantly reported in children. Chromosome Xp11.2 translocation results in gene fusion related to transcription factor E3 (TFE3 that plays an important role in proliferation and survival. Case Report. Herein, we present two cases of a TFE3 translocation-associated RCC in young female adults, one detected incidentally and the other one presenting with gross hematuria. Tumor is characterized by immunohistochemistry and a literature review with optimal treatment regimen is presented. Discussion. Xp11.2 translocation RCCs in adult patients are associated with advanced stages, large tumors, and extracapsular disease and usually have an aggressive clinical course. Conclusion. In TFE3 RCC, the genetic background may not only contribute to tumorigenesis, but also determine the response to chemotherapy and targeted therapy. Therefore it is necessary to diagnose this tumor entity accurately. Because of the small number of TFE3 gene fusion-related renal tumors described in the literature, the exact biologic behavior and impact of current treatment modalities remain to be uncertain.

  6. Improved immunogenicity of Newcastle disease virus inactivated vaccine following DNA vaccination using Newcastle disease virus hemagglutinin-neuraminidase and fusion protein genes. (United States)

    Firouzamandi, Masoumeh; Moeini, Hassan; Hosseini, Davood; Bejo, Mohd Hair; Omar, Abdul Rahman; Mehrbod, Parvaneh; Ideris, Aini


    The present study describes the development of DNA vaccines using the hemagglutinin-neuraminidase (HN) and fusion (F) genes from AF2240 Newcastle disease virus strain, namely pIRES/HN, pIRES/F and pIRES-F/HN. Transient expression analysis of the constructs in Vero cells revealed the successful expression of gene inserts in vitro. Moreover, in vivo experiments showed that single vaccination with the constructed plasmid DNA (pDNA) followed by a boost with inactivated vaccine induced a significant difference in enzyme-linked immunosorbent assay antibody levels (p < 0.05) elicited by either pIRES/F, pIRES/F+ pIRES/HN or pIRES-F/HN at one week after the booster in specific pathogen free chickens when compared with the inactivated vaccine alone. Taken together, these results indicated that recombinant pDNA could be used to increase the efficacy of the inactivated vaccine immunization procedure.

  7. Enhancement of DNA vaccine potency by vaccination with the CTLA-4-based fusion gene%CTLA-4融合基因免疫策略增强DNA疫苗的免疫应答效应

    Institute of Scientific and Technical Information of China (English)

    周承; 陈智


    DNA疫苗具有较大的临床应用前景,但其免疫效力还不够强大,尤其在大动物和人类中.CTLA-4融合基因免疫能同时增强机体的特异性体液和细胞免疫应答,在抗感染、抗肿瘤中显示巨大的应用前景,并且其免疫增强效应已在部分大动物体内得到证实.%DNA vaccine has shown peat prospects in clinical application, but its immune potency is limited, especially in large animals and mankind. Vaccination with the fusion gene encoding the extracellular domain of CTLA-4 fused to the specific antigen is able to enhance both humoral and cellular immune reponses in several kinds of large animals, which represent a promising approach in the treatment of infectious diseases and cancers.

  8. Retrovirus-mediated transfer of the fusion gene encoding EGFP-BMP_2 in mesenchymal stem cells

    Institute of Scientific and Technical Information of China (English)


    Bone marrow mesenchymal stemcells(MSCs)are pluripotential stemcells that have the capacitytodifferentiate into chondrocytes and osteoblasts[1].Ithas been well documented that bone morphogeneticproteins(BMPs),a group of proteins belonging tothe TGF-βsuperfamily,can induce bone for mationbothin vivoandin vitroas well as promote osteo-blastic differentiation of MSC[2].HeterologousBMP2is successfully transferred to MSCs and genetherapy is employed based on repairing bony andcartilage defects,spinal fusion[3-5]....

  9. Genetic variability available through cell fusion

    Energy Technology Data Exchange (ETDEWEB)

    Smith, H.H.; Mastrangelo-Hough, I.A.


    Results are reported for the following studies: plant hybridization through protoplast fusion using species of Nicotiana and Petunia; chromosome instability studies on culture-induced chromosome changes and chromosome elimination; chloroplast distribution in parasexual hybrids; chromosomal introgression following fusion; plant-animal fusion; and microcell-mediated chromosome transfer and chromosome-mediated gene transfer. (HLW)

  10. Involvement of human endogenous retroviral syncytin-1 in human osteoclast fusion

    DEFF Research Database (Denmark)

    Søe, Kent; Andersen, Thomas Lykke; Hobolt-Pedersen, Anne-Sofie


    fusion of the lipid bilayers of their cell membranes are still unknown. Syncytin-1 is a protein encoded by a human endogenous retroviral gene which was stably integrated into the human ancestor genome more than 24 million years ago. Upon activation, syncytin-1 is able to destabilize the lipid bilayer......Generation of osteoclasts through fusion of mono-nucleated precursors is a key event of bone physiology and bone resorption is inefficient without osteoclast fusion. Several factors playing a critical role in the fusion process have already been recognized, but the factors involved in the actual....... This was documented through Q-PCR, Western blot and immunofluorescence analyses. These in vitro findings were confirmed by immunohistochemical stainings in human iliac crest biopsies. A syncytin-1 inhibitory peptide reduced the number of nuclei per osteoclast by 30%, as well as TRACP activity. From a mechanistic...

  11. Delivery of human NKG2D-IL-15 fusion gene by chitosan nanoparticles to enhance antitumor immunity

    Energy Technology Data Exchange (ETDEWEB)

    Yan, Chen; Jie, Leng; Yongqi, Wang [Department of Immunology, School of Medicine, Yangzhou University, Yangzhou, 225009 (China); Weiming, Xiao [Department of Gastroenterology, The Second Clinical Medical College, Yangzhou University, Yangzhou, 225009 (China); Juqun, Xi [Jiangsu Key Laboratory of Integrated Traditional Chinese and Western Medicine for Prevention and Treatment of Senile Diseases, Yangzhou, 225009 (China); Yanbing, Ding [Department of Gastroenterology, The Second Clinical Medical College, Yangzhou University, Yangzhou, 225009 (China); Li, Qian [Department of Immunology, School of Medicine, Yangzhou University, Yangzhou, 225009 (China); Xingyuan, Pan [Jiangsu Key Laboratory of Zoonosis, Yangzhou University, Yangzhou, 225009 (China); Mingchun, Ji [Department of Immunology, School of Medicine, Yangzhou University, Yangzhou, 225009 (China); Weijuan, Gong, E-mail: [Department of Immunology, School of Medicine, Yangzhou University, Yangzhou, 225009 (China); Department of Gastroenterology, The Second Clinical Medical College, Yangzhou University, Yangzhou, 225009 (China); Jiangsu Key Laboratory of Integrated Traditional Chinese and Western Medicine for Prevention and Treatment of Senile Diseases, Yangzhou, 225009 (China); Jiangsu Key Laboratory of Zoonosis, Yangzhou University, Yangzhou, 225009 (China); Jiangsu Co-Innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Yangzhou, 225009 (China)


    Nanoparticles are becoming promising carriers for gene delivery because of their high capacity in gene loading and low cell cytotoxicity. In this study, a chitosan-based nanoparticle encapsulated within a recombinant pcDNA3.1-dsNKG2D-IL-15 plasmid was generated. The fused dsNKG2D-IL-15 gene fragment consisted of double extracellular domains of NKG2D with IL-15 gene at downstream. The average diameter of the gene nanoparticles ranged from 200 nm to 400 nm, with mean zeta potential value of 53.8 ± 6.56 mV. The nanoparticles which were loaded with the dsNKG2D-IL-15 gene were uptaken by tumor cells with low cytotoxicity. Tumor cells pre-transfected by gene nanopartilces stimulated NK and T cells in vitro. Intramuscular injection of gene nanoparticles suppressed tumor growth and prolonged survival of tumor-bearing mice through activation of NK and CD8{sup +} T cells. Thus, chitosan-based nanoparticle delivery of dsNKG2D-IL-15 gene vaccine can be potentially used for tumor therapy. - Highlights: • Generation of a nanoparticle for delivery of dsNKG2D-IL-15 gene. • Characterization of the gene nanoparticle. • Antitumor activity mediated by the gene nanoparticle.

  12. Mechanisms of influenza viral membrane fusion. (United States)

    Blijleven, Jelle S; Boonstra, Sander; Onck, Patrick R; van der Giessen, Erik; van Oijen, Antoine M


    Influenza viral particles are enveloped by a lipid bilayer. A major step in infection is fusion of the viral and host cellular membranes, a process with large kinetic barriers. Influenza membrane fusion is catalyzed by hemagglutinin (HA), a class I viral fusion protein activated by low pH. The exact nature of the HA conformational changes that deliver the energy required for fusion remains poorly understood. This review summarizes our current knowledge of HA structure and dynamics, describes recent single-particle experiments and modeling studies, and discusses their role in understanding how multiple HAs mediate fusion. These approaches provide a mechanistic picture in which HAs independently and stochastically insert into the target membrane, forming a cluster of HAs that is collectively able to overcome the barrier to membrane fusion. The new experimental and modeling approaches described in this review hold promise for a more complete understanding of other viral fusion systems and the protein systems responsible for cellular fusion.

  13. Induction of autophagy by Imatinib sequesters Bcr-Abl in autophagosomes and down-regulates Bcr-Abl protein.

    LENUS (Irish Health Repository)

    Elzinga, Baukje M


    Chronic Myeloid Leukemia (CML) is a disease of hematopoietic stem cells which harbor the chimeric gene Bcr-Abl. Expression levels of this constitutively active tyrosine kinase are critical for response to tyrosine kinase inhibitor treatment and also disease progression, yet the regulation of protein stability is poorly understood. We have previously demonstrated that imatinib can induce autophagy in Bcr-Abl expressing cells. Autophagy has been associated with the clearance of large macromolecular signaling complexes and abnormal proteins, however, the contribution of autophagy to the turnover of Bcr-Abl protein in imatinib treated cells is unknown. In this study, we show that following imatinib treatment, Bcr-Abl is sequestered into vesicular structures that co-localize with the autophagy marker LC3 or GABARAP. This association is inhibited by siRNA mediated knockdown of autophagy regulators (Beclin 1\\/ATG7). Pharmacological inhibition of autophagy also reduced Bcr-Abl\\/LC3 co-localization in both K562 and CML patient cells. Bcr-Abl protein expression was reduced with imatinib treatment. Inhibition of both autophagy and proteasome activity in imatinib treated cells was required to restore Bcr-Abl protein levels to those of untreated cells. This ability to down-regulate Bcr-Abl protein levels through the induction of autophagy may be an additional and important feature of the activity of imatinib.

  14. High-level SUMO-mediated fusion expression of ABP-dHC-cecropin A from multiple joined genes in Escherichia coli. (United States)

    Zhang, Jiaxin; Movahedi, Ali; Wei, Zhiheng; Sang, Ming; Wu, Xiaolong; Wang, Mengyang; Wei, Hui; Pan, Huixin; Yin, Tongming; Zhuge, Qiang


    The antimicrobial peptide ABP-dHC-cecropin A is a small cationic peptide with potent activity against a wide range of bacterial species. Evidence of antifungal activity has also been suggested; however, evaluation of this peptide has been limited due to the low expression of cecropin proteins in Escherichia coli. To improve the expression level of ABP-dHC-cecropin A in E. coli, tandem repeats of the ABP-dHC-cecropin A gene were constructed and expressed as fusion proteins (SUMO-nABP-dHC-cecropin, n = 1, 2, 3, 4) via pSUMO-nABP-dHC-cecropin A vectors (n = 1, 2, 3, 4). Comparison of the expression levels of soluble SUMO-nABP-dHC-cecropin A fusion proteins (n = 1, 2, 3, 4) suggested that BL21 (DE3)/pSUMO-3ABP-dHC-cecropin A is an ideal recombinant strain for ABP-dHC-cecropin A production. Under the selected conditions of cultivation and isopropylthiogalactoside (IPTG) induction, the expression level of ABP-dHC-cecropin A was as high as 65 mg/L, with ∼21.3% of the fusion protein in soluble form. By large-scale fermentation, protein production reached nearly 300 mg/L, which is the highest yield of ABP-dHC-cecropin A reported to date. In antibacterial experiments, the efficacy was approximately the same as that of synthetic ABP-dHC-cecropin A. This method provides a novel and effective means of producing large amounts of ABP-dHC-cecropin A.

  15. 实时定量荧光反转录-聚合酶链反应检测急性白血病AML1/ETO融合基因%Real-time RT-PCR for detection and quantification of AML1/ETO leukemia fusion gene

    Institute of Scientific and Technical Information of China (English)

    李斌; 喻镁佳; 梁洋; 李晓进; 胡杰; 陈琪; 李惠民


    Objective To set up real-time quantitative RT-PCR technique and measure leukemia fusion gene transcripts in patients with AML of FAB-M_2 subtype,and also to investigate the positive rate in patients and the relationship between the AMLI/ETO mRNA levels and the response rate after chemical therapy.Methods The plasmid containing the AMLI/ETO fusion gene sequences were constructed from myeloid cell lines Kasumi-1 (expressing AML1/ETO)to establish the standard curves,A TaqMan based realtime quantitative RT-PCR was performed to measure aberrant fusion gene transcripts in 45 samples of peripheral blood(PB) or bone marrow (BM) from 25 newly diagnosed patients with AML-M_2.All these 25 patients were diagnosed by the FCM(flow cytometry)and bone marrow molecular cytogenetics,and received the induce remission therapy with MA (Mitoxantrone+Ara-C).Results As a result,the AML1/ETO fusion gene transcripts were detected in 7(28%)out of 25 AML-M_2 patients (the ratios of AML1/ETO/ABL vary from 0.01 to 19.2),in which 5 patients were found t(8;21)(q22;q22).The transcript level of AML1/ETO fusion gene varied from the clinical situation of patients.These 7 patients with AML1/ETO fusion gene got complete remission(CR) after the first MA therapy,and the fusion gene reduced by 3 log in AML1/ETO/ABL.Only 11 patients got CR in 18 patients without AML1/ETO fusion gene.By following up these 7 patients with AML1/ ETO fusion gene kept persistent CR for 6 months.Conclusion It was concluded that real-time quantitative PCR is a reliable,innovative and promising technology with high sensitivity and speciality.It has potential clinical value for diagnosis,tumor typing,treatment selection,measuring the tumor load,monitoring fusion gene expression level and evaluating therapeutic strategy.It is worthy to apply in the clinical practice.%目的 建立实时定量荧光反转录-聚合酶链反应(RQ RT-PCR)的方法并用来检测急性髓系白血病(AML)-M_2:患者中 AML1/ETO融合基因的拷贝

  16. Outcomes of Children With BCR-ABL1–Like Acute Lymphoblastic Leukemia Treated With Risk-Directed Therapy Based on the Levels of Minimal Residual Disease (United States)

    Roberts, Kathryn G.; Pei, Deqing; Campana, Dario; Payne-Turner, Debbie; Li, Yongjin; Cheng, Cheng; Sandlund, John T.; Jeha, Sima; Easton, John; Becksfort, Jared; Zhang, Jinghui; Coustan-Smith, Elaine; Raimondi, Susana C.; Leung, Wing H.; Relling, Mary V.; Evans, William E.; Downing, James R.; Mullighan, Charles G.; Pui, Ching-Hon


    Purpose BCR-ABL1–like acute lymphoblastic leukemia (ALL) is a recently identified B-cell ALL (B-ALL) subtype with poor outcome that exhibits a gene expression profile similar to BCR-ABL1-positive ALL but lacks the BCR-ABL1 fusion protein. We examined the outcome of children with BCR-ABL1–like ALL treated with risk-directed therapy based on minimal residual disease (MRD) levels during remission induction. Patients and Methods Among 422 patients with B-ALL enrolled onto the Total Therapy XV study between 2000 and 2007, 344 had adequate samples for gene expression profiling. Next-generation sequencing and/or analysis of genes known to be altered in B-ALL were performed in patients with BCR-ABL1–like ALL who had available material. Outcome was compared between patients with and those without BCR-ABL1–like ALL. Results Forty (11.6%) of the 344 patients had BCR-ABL1–like ALL. They were significantly more likely to be male, have Down syndrome, and have higher MRD levels on day 19 and at the end of induction than did other patients with B-ALL. Among 25 patients comprehensively studied for genetic abnormalities, 11 harbored a genomic rearrangement of CRLF2, six had fusion transcripts responsive to ABL tyrosine kinase inhibitors or JAK inhibitors, and seven had mutations involving the Ras signaling pathway. There were no significant differences in event-free survival (90.0% ± 4.7% [SE] v 88.4% ± 1.9% at 5 years; P = .41) or in overall survival (92.5% ± 4.2% v 95.1% ± 1.3% at 5 years; P = .41) between patients with and without BCR-ABL1–like ALL. Conclusion Patients who have BCR-ABL1–like ALL with poor initial treatment response can be salvaged with MRD-based risk-directed therapy and may benefit from identification of kinase-activating lesions for targeted therapies. PMID:25049327

  17. Expression of γ-aminobutyric acid ρ1 and ρ1Δ450 as gene fusions with the green fluorescent protein (United States)

    Martínez-Torres, Ataúlfo; Miledi, Ricardo


    The functional characteristics and cellular localization of the γaminobutyric acid (GABA) ρ1 receptor and its nonfunctional isoform ρ1Δ450 were investigated by expressing them as gene fusions with the enhanced version of the green fluorescent protein (GFP). Oocytes injected with ρ1-GFP had receptors that gated chloride channels when activated by GABA. The functional characteristics of these receptors were the same as for those of wild-type ρ1 receptors. Fluorescence, because of the chimeric receptors expressed, was over the whole oocyte but was more intense near the cell surface and more abundant in the animal hemisphere. Similar to the wild type, ρ1Δ450-GFP did not lead to the expression of functional GABA receptors, and injected oocytes failed to generate currents even after exposure to high concentrations of GABA. Nonetheless, the fluorescence displayed by oocytes expressing ρ1Δ450-GFP was distributed similarly to that of ρ1-GFP. Mammalian cells transfected with the ρ1-GFP or ρ1Δ450-GFP constructs showed mostly intracellularly distributed fluorescence in confocal microscope images. A sparse localization of fluorescence was observed in the plasma membrane regardless of the cell line used. We conclude that ρ1Δ450 is expressed and transported close to, and perhaps incorporated into, the plasma membrane. Thus, ρ1- and ρ1Δ450-GFP fusions provide a powerful tool to visualize the traffic of GABA type C receptors. PMID:11172056

  18. Role of Abl kinase and the Wave2 signaling complex in HIV-1 entry at a post-hemifusion step.

    Directory of Open Access Journals (Sweden)

    Brooke Harmon

    Full Text Available Entry of human immunodeficiency virus type 1 (HIV-1 commences with binding of the envelope glycoprotein (Env to the receptor CD4, and one of two coreceptors, CXCR4 or CCR5. Env-mediated signaling through coreceptor results in Galphaq-mediated Rac activation and actin cytoskeleton rearrangements necessary for fusion. Guanine nucleotide exchange factors (GEFs activate Rac and regulate its downstream protein effectors. In this study we show that Env-induced Rac activation is mediated by the Rac GEF Tiam-1, which associates with the adaptor protein IRSp53 to link Rac to the Wave2 complex. Rac and the tyrosine kinase Abl then activate the Wave2 complex and promote Arp2/3-dependent actin polymerization. Env-mediated cell-cell fusion, virus-cell fusion and HIV-1 infection are dependent on Tiam-1, Abl, IRSp53, Wave2, and Arp3 as shown by attenuation of fusion and infection in cells expressing siRNA targeted to these signaling components. HIV-1 Env-dependent cell-cell fusion, virus-cell fusion and infection were also inhibited by Abl kinase inhibitors, imatinib, nilotinib, and dasatinib. Treatment of cells with Abl kinase inhibitors did not affect cell viability or surface expression of CD4 and CCR5. Similar results with inhibitors and siRNAs were obtained when Env-dependent cell-cell fusion, virus-cell fusion or infection was measured, and when cell lines or primary cells were the target. Using membrane curving agents and fluorescence microscopy, we showed that inhibition of Abl kinase activity arrests fusion at the hemifusion (lipid mixing step, suggesting a role for Abl-mediated actin remodeling in pore formation and expansion. These results suggest a potential utility of Abl kinase inhibitors to treat HIV-1 infected patients.

  19. Delivery of human NKG2D-IL-15 fusion gene by chitosan nanoparticles to enhance antitumor immunity. (United States)

    Yan, Chen; Jie, Leng; Yongqi, Wang; Weiming, Xiao; Juqun, Xi; Yanbing, Ding; Li, Qian; Xingyuan, Pan; Mingchun, Ji; Weijuan, Gong


    Nanoparticles are becoming promising carriers for gene delivery because of their high capacity in gene loading and low cell cytotoxicity. In this study, a chitosan-based nanoparticle encapsulated within a recombinant pcDNA3.1-dsNKG2D-IL-15 plasmid was generated. The fused dsNKG2D-IL-15 gene fragment consisted of double extracellular domains of NKG2D with IL-15 gene at downstream. The average diameter of the gene nanoparticles ranged from 200 nm to 400 nm, with mean zeta potential value of 53.8 ± 6.56 mV. The nanoparticles which were loaded with the dsNKG2D-IL-15 gene were uptaken by tumor cells with low cytotoxicity. Tumor cells pre-transfected by gene nanopartilces stimulated NK and T cells in vitro. Intramuscular injection of gene nanoparticles suppressed tumor growth and prolonged survival of tumor-bearing mice through activation of NK and CD8(+) T cells. Thus, chitosan-based nanoparticle delivery of dsNKG2D-IL-15 gene vaccine can be potentially used for tumor therapy.

  20. Spatial localization of genes determined by intranuclear DNA fragmentation with the fusion proteins lamin KRED and histone KRED und visible light. (United States)

    Waldeck, Waldemar; Mueller, Gabriele; Glatting, Karl-Heinz; Hotz-Wagenblatt, Agnes; Diessl, Nicolle; Chotewutmonti, Sasithorn; Langowski, Jörg; Semmler, Wolfhard; Wiessler, Manfred; Braun, Klaus


    The highly organized DNA architecture inside of the nuclei of cells is accepted in the scientific world. In the human genome about 3 billion nucleotides are organized as chromatin in the cell nucleus. In general, they are involved in gene regulation and transcription by histone modification. Small chromosomes are localized in a central nuclear position whereas the large chromosomes are peripherally positioned. In our experiments we inserted fusion proteins consisting of a component of the nuclear lamina (lamin B1) and also histone H2A, both combined with the light inducible fluorescence protein KillerRed (KRED). After activation, KRED generates reactive oxygen species (ROS) producing toxic effects and may cause cell death. We analyzed the spatial damage distribution in the chromatin after illumination of the cells with visible light. The extent of DNA damage was strongly dependent on its localization inside of nuclei. The ROS activity allowed to gain information about the location of genes and their functions via sequencing and data base analysis of the double strand breaks of the isolated DNA. A connection between the damaged gene sequences and some diseases was found.

  1. Phylogenetic characterization of the fusion genes of the Newcastle disease viruses isolated in Fars province poultry farms during 2009-2011. (United States)

    Mehrabanpour, Mohammad Javad; Khoobyar, Setareh; Rahimian, Abdollah; Nazari, Mohammad Bagher; Keshtkar, Mohammad Reza


    Despite routine vaccination programs against Newcastle disease (ND), sporadic cases have occasionally occurred that remain a constant threat to commercial poultry. Ten isolates of Newcastle disease viruses (NDV) from infected broiler chicken cases were obtained from various locations in Fars province during 2009-2011 and genetically analyzed using reverse transcription polymerase chain reaction (RT- PCR) with primers specific to the viral fusion (F) protein- gene. The viruses were confirmed as NDV by hemagglutination inhibition assay and RT- PCR. The isolates based on the sequence and phylogenetic analyses of partial F gene were genotypically analyzed by RT PCR. In the present investigation, the pathogenicity of NDV strains was determined by internationally recognized test mean death time (MDT). Analysis based on F gene showed that characterized isolates possess three different types of protease cleavage site motifs and appear to show maximum identities with isolates in the region. The subsequent phylogenetic analysis was implemented using MEGA and the phylogenetic tree. The results of RT-PCR and MDT showed that 10 isolates were positive for NDV, (60% velogenic, 30% mesogenic and 10% lentogenic). The results of the phylogenetic analysis showed that 10 NDV isolates from Iran belong to the class II, genotype III viruses. This information is fundamental to improve the efficacy of controlling strategies and vaccine development for NDV.

  2. DNA immunization with fusion genes encoding different regions of hepatitis C virus E2 fused to the gene for hepatitis B surface antigen elicits immune responses to both HCV and HBV

    Institute of Scientific and Technical Information of China (English)

    Jing Jin; Jian-Ying Yang; Jing Liu; Yu-Ying Kong; Yuan Wang; Guang-Di Li


    AIM: Both Hepatitis B virus (HBV) and Hepatitis C virus(HCV) are major causative agents of transfusion-associatedand community-acquired hepatitis worldwide. Developmentof a HCV vaccine as well as more effective HBV vaccines isan urgent task. DNA immunization provides a promisingapproach to elicit protective humoral and cellular immuneresponses against viral infection. The aim of this study is toachieve immune responses against both HCV and HBV by DNAimmunization with fusion constructs comprising various HCVE2 gene fragments fused to HBsAg gane of HBV.METHODS: C57BL/6 mice were immunized with plasmid DNAexpressing five fragments of HCV E2 fused to the gene forHBsAg respectively. After one primary and one boostingimmunizations, antibodies against HCV E2 and HBsAg weretested and subtyped in ELISA. Splenic cytokine expressionof IFN-γ and IL-10 was analyzed using an RT-PCR assay.Post-immune mouse antisera also were tested for theirability to capture HCV viruses in the serum of a hepatitis Cpatient in vitro.RESUTLTS: After immunization, antibodies against bothHBsAg and HCV E2 were detected in mouse sera, withIgG2a being the dominant immunoglobulin sub-class. High-level expression of INF-γ was deuetected in cultured splenic cells.Mouse antisera against three of the five fusion constructs wereable to capture HCV viruses in an in vitro assay.CONCLUSION: The results indicate that these fusionconstructs could efficiently elicit humoral and Th1 dominantcellular immune responses against both HBV S and HCV E2antigens in DNA-immunized mice. They thus could serve ascandidates for a bivalent vaccine against HBV and HCVinfection. In addition, the capacity of mouse antisera againstthree of the five fusion constnucts to capture HCV virusses invitro suggested that neutralizing epitopes may be present inother regions of E2 besides the hypervariable region 1.

  3. An MSC2 Promoter-lacZ Fusion Gene Reveals Zinc-Responsive Changes in Sites of Transcription Initiation That Occur across the Yeast Genome (United States)

    Wu, Yi-Hsuan; Taggart, Janet; Song, Pamela Xiyao; MacDiarmid, Colin; Eide, David J.


    The Msc2 and Zrg17 proteins of Saccharomyces cerevisiae form a complex to transport zinc into the endoplasmic reticulum. ZRG17 is transcriptionally induced in zinc-limited cells by the Zap1 transcription factor. In this report, we show that MSC2 mRNA also increases (~1.5 fold) in zinc-limited cells. The MSC2 gene has two in-frame ATG codons at its 5’ end, ATG1 and ATG2; ATG2 is the predicted initiation codon. When the MSC2 promoter was fused at ATG2 to the lacZ gene, we found that unlike the chromosomal gene this reporter showed a 4-fold decrease in lacZ mRNA in zinc-limited cells. Surprisingly, β-galactosidase activity generated by this fusion gene increased ~7 fold during zinc deficiency suggesting the influence of post-transcriptional factors. Transcription of MSC2ATG2-lacZ was found to start upstream of ATG1 in zinc-replete cells. In zinc-limited cells, transcription initiation shifted to sites just upstream of ATG2. From the results of mutational and polysome profile analyses, we propose the following explanation for these effects. In zinc-replete cells, MSC2ATG2-lacZ mRNA with long 5’ UTRs fold into secondary structures that inhibit translation. In zinc-limited cells, transcripts with shorter unstructured 5’ UTRs are generated that are more efficiently translated. Surprisingly, chromosomal MSC2 did not show start site shifts in response to zinc status and only shorter 5’ UTRs were observed. However, the shifts that occur in the MSC2ATG2-lacZ construct led us to identify significant transcription start site changes affecting the expression of ~3% of all genes. Therefore, zinc status can profoundly alter transcription initiation across the yeast genome. PMID:27657924

  4. Cold fusion

    Energy Technology Data Exchange (ETDEWEB)

    Suh, Suk Yong; Sung, Ki Woong; Kang, Joo Sang; Lee, Jong Jik [Korea Atomic Energy Research Institute, Taejon (Korea, Republic of)


    So called `cold fusion phenomena` are not confirmed yet. Excess heat generation is very delicate one. Neutron generation is most reliable results, however, the records are erratic and the same results could not be repeated. So there is no reason to exclude the malfunction of testing instruments. The same arguments arise in recording {sup 4}He, {sup 3}He, {sup 3}H, which are not rich in quantity basically. An experiment where plenty of {sup 4}He were recorded is attached in appendix. The problem is that we are trying to search cold fusion which is permitted by nature or not. The famous tunneling effect in quantum mechanics will answer it, however, the most fusion rate is known to be negligible. The focus of this project is on the theme that how to increase that negligible fusion rate. 6 figs, 4 tabs, 1512 refs. (Author).

  5. Spinal Fusion (United States)

    ... results in predictable healing. Autograft is currently the “gold standard” source of bone for a fusion. The ... pump. With this technique, the patient presses a button that delivers a predetermined amount of narcotic pain ...

  6. Fusion of the Tumor-Suppressor Gene CHEK2 and the Gene for the Regulatory Subunit B of Protein Phosphatase 2 PPP2R2A in Childhood Teratoma

    Directory of Open Access Journals (Sweden)

    Yuesheng Jin


    Full Text Available We characterized the molecular genetic consequences of a balanced chromosome translocation t(8;22(p21; q12, which occurred as the sole cytogenetic aberration in short-term cultured cells from an intrathoracic mature teratoma in a 15-year-old girl. Fluorescence in situ hybridization and reverse transcription- polymerase chain reaction disclosed that t(8;22 resulted in the fusion of the genes PPP2R2A and CHEK2, with an inserted fragment belonging to class I endogenous retrovirus-related sequences at the junction. Sequencing of the two genes did not reveal any additional mutation. None of the three detected PPP2R2A/CHEK2 fusion transcripts resulted in an in-frame PPP2R2A/CHEK2 chimerical open reading frame; however, in all of them, the known open reading frame of CHEK2 was preserved. Thus, promoter swapping leading to deregulated CHEK2 expression would be the most likely oncogenic mechanism. Whereas inactivating mutations of CHEK2 previously have been described in a variety of sporadic tumors and in inherited cancer-predisposing syndromes, PPP2R2A, encoding a regulatory subunit of the multimeric enzyme phosphatase 2, has not been directly implicated in tumorigenesis. Our findings suggest that deregulation of CHEK2 and/or PPP2R2A is of pathogenetic importance in at least a subset of germ cell tumors.

  7. Myoblast fusion in Drosophila

    Energy Technology Data Exchange (ETDEWEB)

    Haralalka, Shruti [Stowers Institute for Medical Research, Kansas City, MO 64110 (United States); Abmayr, Susan M., E-mail: [Stowers Institute for Medical Research, Kansas City, MO 64110 (United States); Department of Anatomy and Cell Biology, University of Kansas Medical Center, Kansas City, MO 66160 (United States)


    The body wall musculature of a Drosophila larva is composed of an intricate pattern of 30 segmentally repeated muscle fibers in each abdominal hemisegment. Each muscle fiber has unique spatial and behavioral characteristics that include its location, orientation, epidermal attachment, size and pattern of innervation. Many, if not all, of these properties are dictated by founder cells, which determine the muscle pattern and seed the fusion process. Myofibers are then derived from fusion between a specific founder cell and several fusion competent myoblasts (FCMs) fusing with as few as 3-5 FCMs in the small muscles on the most ventral side of the embryo and as many as 30 FCMs in the larger muscles on the dorsal side of the embryo. The focus of the present review is the formation of the larval muscles in the developing embryo, summarizing the major issues and players in this process. We have attempted to emphasize experimentally-validated details of the mechanism of myoblast fusion and distinguish these from the theoretically possible details that have not yet been confirmed experimentally. We also direct the interested reader to other recent reviews that discuss myoblast fusion in Drosophila, each with their own perspective on the process . With apologies, we use gene nomenclature as specified by Flybase ( but provide Table 1 with alternative names and references.

  8. Switch-like reprogramming of gene expression after fusion of multinucleate plasmodial cells of two Physarum polycephalum sporulation mutants

    Energy Technology Data Exchange (ETDEWEB)

    Walter, Pauline; Hoffmann, Xenia-Katharina; Ebeling, Britta; Haas, Markus; Marwan, Wolfgang, E-mail:


    Highlights: •We investigate reprogramming of gene expression in multinucleate single cells. •Cells of two differentiation control mutants are fused. •Fused cells proceed to alternative gene expression patterns. •The population of nuclei damps stochastic fluctuations in gene expression. •Dynamic processes of cellular reprogramming can be observed by repeated sampling of a cell. -- Abstract: Nonlinear dynamic processes involving the differential regulation of transcription factors are considered to impact the reprogramming of stem cells, germ cells, and somatic cells. Here, we fused two multinucleate plasmodial cells of Physarum polycephalum mutants defective in different sporulation control genes while being in different physiological states. The resulting heterokaryons established one of two significantly different expression patterns of marker genes while the plasmodial halves that were fused to each other synchronized spontaneously. Spontaneous synchronization suggests that switch-like control mechanisms spread over and finally control the entire plasmodium as a result of cytoplasmic mixing. Regulatory molecules due to the large volume of the vigorously streaming cytoplasm will define concentrations in acting on the population of nuclei and in the global setting of switches. Mixing of a large cytoplasmic volume is expected to damp stochasticity when individual nuclei deliver certain RNAs at low copy number into the cytoplasm. We conclude that spontaneous synchronization, the damping of molecular noise in gene expression by the large cytoplasmic volume, and the option to take multiple macroscopic samples from the same plasmodium provide unique options for studying the dynamics of cellular reprogramming at the single cell level.

  9. Structural and functional studies of FKHR-PAX3, a reciprocal fusion gene of the t(2;13 chromosomal translocation in alveolar rhabdomyosarcoma.

    Directory of Open Access Journals (Sweden)

    Qiande Hu

    Full Text Available Alveolar rhabdomyosarcoma (ARMS is an aggressive pediatric cancer of skeletal muscle. More than 70% of ARMS tumors carry balanced t(2;13 chromosomal translocation that leads to the production of two novel fusion genes, PAX3-FKHR and FKHR-PAX3. While the PAX3-FKHR gene has been intensely studied, the reciprocal FKHR-PAX3 gene has rarely been described. We report here the cloning and functional characterization of the FKHR-PAX3 gene as the first step towards a better understanding of its potential impact on ARMS biology. From RH30 ARMS cells, we detected and isolated three versions of FKHR-PAX3 cDNAs whose C-terminal sequences corresponded to PAX3c, PAX3d, and PAX3e isoforms. Unlike the nuclear-specific localization of PAX3-FKHR, the reciprocal FKHR-PAX3 proteins stayed predominantly in the cytoplasm. FKHR-PAX3 potently inhibited myogenesis in both non-transformed myoblast cells and ARMS cells. We showed that FKHR-PAX3 was not a classic oncogene but could act as a facilitator in oncogenic pathways by stabilizing PAX3-FKHR expression, enhancing cell proliferation, clonogenicity, anchorage-independent growth, and matrix adhesion in vitro, and accelerating the onset of tumor formation in xenograft mouse model in vivo. In addition to these pro-oncogenic behaviors, FKHR-PAX3 also negatively affected cell migration and invasion in vitro and lung metastasis in vivo. Taken together, these functional characteristics suggested that FKHR-PAX3 might have a critical role in the early stage of ARMS development.

  10. Contruction of the Genetic Engineering Strain Expressed Nontoxic ST1-LTB Fusion Protein Against Enterotoxigenic Eschenichia coli

    Institute of Scientific and Technical Information of China (English)

    BAI Jia-ning; SUN Yi-min; BIAN Yan-qing; ZHAO Bao-hua


    Thermostable enterotoxinⅠ(ST1)mutant genes and thermolabile enterotoxin B subunit(LTB)genes were amplified by PCR from plasmids of Eschenichia coli C83902.The recombinant expression plasmid pZST3LTB containing ST1-LTB fusion gene was constructed by recombinant DNA technique and then transformed into Escherichia coli BL21(DE3).The ST1-LTB fusion protein was highly expressed in recombinant strain BL21(DE3)(pZST3LTB)and the fusion protein was about 38.53% of total cellular protein by SDS-PAGE and thin-layer gel scanning analysis.More important,mice immunized with crude preparation containing the fusion protein inclusion bodies or inactivated recombinant strain produced antibodies that were able to recognize ST1 in vitro.These sera antibodies were able to neutralize the biological activity of native ST1 in the suckling mouse assay.Hence the ST1-LTB fusion protein was nontoxic and immunogenic,the constructed recombinant strain BL21(DE3)(pZST3LTB)could be used as a candidate of vaccine strain.

  11. An ancient history of gene duplications, fusions and losses in the evolution of APOBEC3 mutators in mammals

    Directory of Open Access Journals (Sweden)

    Münk Carsten


    Full Text Available Abstract Background The APOBEC3 (A3 genes play a key role in innate antiviral defense in mammals by introducing directed mutations in the DNA. The human genome encodes for seven A3 genes, with multiple splice alternatives. Different A3 proteins display different substrate specificity, but the very basic question on how discerning self from non-self still remains unresolved. Further, the expression of A3 activity/ies shapes the way both viral and host genomes evolve. Results We present here a detailed temporal analysis of the origin and expansion of the A3 repertoire in mammals. Our data support an evolutionary scenario where the genome of the mammalian ancestor encoded for at least one ancestral A3 gene, and where the genome of the ancestor of placental mammals (and possibly of the ancestor of all mammals already encoded for an A3Z1-A3Z2-A3Z3 arrangement. Duplication events of the A3 genes have occurred independently in different lineages: humans, cats and horses. In all of them, gene duplication has resulted in changes in enzyme activity and/or substrate specificity, in a paradigmatic example of convergent adaptive evolution at the genomic level. Finally, our results show that evolutionary rates for the three A3Z1, A3Z2 and A3Z3 motifs have significantly decreased in the last 100 Mya. The analysis constitutes a textbook example of the evolution of a gene locus by duplication and sub/neofunctionalization in the context of virus-host arms race. Conclusions Our results provide a time framework for identifying ancestral and derived genomic arrangements in the APOBEC loci, and to date the expansion of this gene family for different lineages through time, as a response to changes in viral/retroviral/retrotransposon pressure.

  12. Proteome-wide identification of novel binding partners to the oncogenic fusion gene protein, NPM-ALK, using tandem affinity purification and mass spectrometry. (United States)

    Wu, Fang; Wang, Peng; Young, Leah C; Lai, Raymond; Li, Liang


    Nucleophosmin-anaplastic lymphoma kinase (NPM-ALK), an oncogenic fusion gene protein that is characteristically found in a subset of anaplastic large cell lymphomas, promotes tumorigenesis through its functional and physical interactions with various biologically important proteins. The identification of these interacting proteins has proven to be useful to further our understanding of NPM-ALK-mediated tumorigenesis. For the first time, we performed a proteome-wide identification of NPM-ALK-binding proteins using tandem affinity purification and a highly sensitive mass spectrometric technique. Tandem affinity purification is a recently developed method that carries a lower background and higher sensitivity compared with the conventional immunoprecipitation-based protein purification protocols. The NPM-ALK gene was cloned into an HB-tagged vector and expressed in GP293 cells. Three independent experiments were performed and the reproducibility of the data was 68%. The vast majority of the previously reported NPM-ALK-binding proteins were detected. We also identified proteins that are involved in various cellular processes that were not previously described in association with NPM-ALK, such as MCM6 and MSH2 (DNA repair), Nup98 and importin 8 (subcellular protein transport), Stim1 (calcium signaling), 82Fip (RNA regulation), and BAG2 (proteosome degradation). We believe that these data highlight the functional diversity of NPM-ALK and provide new research directions for the study of the biology of this oncoprotein.

  13. A new system for regulated functional gene expression for gene therapy applications: nuclear delivery of a p16INK4A-estrogen receptor carboxy terminal fusion protein only in the presence of estrogen. (United States)

    Tamura, Tomohiro; Kanuma, Tatsuya; Nakazato, Tomoko; Faried, Leri S; Aoki, Hiroshi; Minegishi, Takashi


    The clinical use of gene therapy requires tight regulation of the gene of interest and functional expression only when it is needed. Thus, it is necessary to develop ways of regulating functional gene expression with exogenous stimuli. Many regulatable systems are currently under development. For example, the tetracycline-dependent transcriptional switch has been successfully employed for in vivo preclinical applications. However, there are no examples of regulatable systems that have been employed in human clinical trials. In the present study, we established an adenovirus-delivered functional gene expression system that is regulated by estrogen. This system uses p16INK4A fused at its C-terminus to the ligand-binding domain of the estrogen receptor (DeltaERalpha). We were able to establish cell lines expressing this gene wherein the functional expression of p16INK4A is estrogen-dependent and causes the arrest of several ovarian cancer cell lines. This inducible and adenovirus-mediated gene transfer system may allow gene therapy using nuclear functioning genes in postmenopausal or ovariectomized women.

  14. Trophoblast fusion. (United States)

    Huppertz, Berthold; Gauster, Martin


    The villous trophoblast of the human placenta is the epithelial cover of the fetal chorionic villi floating in maternal blood. This epithelial cover is organized in two distinct layers, the multinucleated syncytiotrophoblast directly facing maternal blood and a second layer of mononucleated cytotrophoblasts. During pregnancy single cytotrophoblasts continuously fuse with the overlying syncytiotrophoblast to preserve this end-differentiated layer until delivery. Syncytial fusion continuously supplies the syncytiotrophoblast with compounds of fusing cytotrophoblasts such as proteins, nucleic acids and lipids as well as organelles. At the same time the input of cytotrophoblastic components is counterbalanced by a continuous release of apoptotic material from the syncytiotrophoblast into maternal blood. Fusion is an essential step in maintaining the syncytiotrophoblast. Trophoblast fusion was shown to be dependant on and regulated by multiple factors such as fusion proteins, proteases and cytoskeletal proteins as well as cytokines, hormones and transcription factors. In this chapter we focus on factors that may be involved in the fusion process of trophoblast directly or that may prepare the cytotrophoblast to fuse.

  15. On not being able to dream. (United States)

    Ogden, Thomas H


    In this paper, the author explores the phenomenon of not being able to dream (as opposed to not being able to remember one's dreams) from three different vantage points. First, from the point of view of psychoanalytic theory, he discusses Bion's idea that the work of dreaming creates the conscious and unconscious mind (and not the other way around). A person who cannot dream is unable to generate differentiable conscious and unconscious experience and, consequently, lives in a psychic state in which he is unable to differentiate waking from sleeping, dreaming from perceiving. The author then approaches the problem of the inability to dream from the perspective achieved by a literary work. He discusses a Borges fiction that creates, in a singularly artful way, the experience of not being able to dream. Finally, the author utilises the vantage point of a detailed account of a clinical experience to explore what it means not to be able to dream. He describes an initial state characterised by the patient's proliferation of unutilisable 'psychic noise' which, over a period of years, led to the analyst's experiencing 'reverie-deprivation' and brief periods of countertransference psychosis. Two analytic sessions are presented and discussed in which psychological work was done that contributed to an enhanced capacity on the part of both patient and analyst for genuine dreaming - both in sleep and in analytic reverie states.

  16. A new transcriptional variant and small azurophilic granules in an acute promyelocytic leukemia case with NPM1/RARA fusion gene. (United States)

    Kikuma, Tomoe; Nakamachi, Yuji; Noguchi, Yoriko; Okazaki, Yoko; Shimomura, Daisuke; Yakushijin, Kimikazu; Yamamoto, Katsuya; Matsuoka, Hiroshi; Minami, Hironobu; Itoh, Tomoo; Kawano, Seiji


    We report here the first case of NPM1/RARA-positive acute promyelocytic leukemia (APL) preceded by myeloid sarcoma (MS) in the vertebra. A 52-year-old man was diagnosed with MS, as the tumor cells were positive for myeloperoxidase and CD68 but negative for CD163. After treatment with steroids and radiation, the size of the tumor was markedly reduced and peripheral blood count was normal. Bone marrow examination showed 89.2% consisted of unclassified promyelocytes characterized by round nuclei and abundant small azurophilic granules but no Auer rods. The results of chromosome analysis showed 46,XY,t(5;17)(q35;q12). Reverse-transcription polymerase chain reaction amplified the NPM1/RARA fusion transcripts derived from a combination of NPM1 exon 4 and RARA exon 5, or of NPM1 exon 1 and RARA exon 5; the latter of these has not been reported previously. Electron microscopic examination of the promyelocyte nuclei showed they were oval with mild nuclear chromatin condensation and small- to medium-sized nucleoli. Hematological and molecular complete remission was attained after induction therapy including all-trans retinoic acid. As MS was also diagnosed in two of the seven other reported cases of APL with NPM1/RARA, MS may occur more frequently in APL with NPM1/RARA than APL with PML/RARA.

  17. Crizotinib Treatment in a Lung Adenocarcinoma Harboring ALK Fusion Gene with Bone Marrow Metastasis: Case Report and Literature Review

    Directory of Open Access Journals (Sweden)

    Xiaoyan LI


    Full Text Available Background and objective Distant metastasis was common in lung cancer, especially patients with bone marrow metastasis had poor prognosis and there were few effective methods. Crizotinib had been confirmed to be used in anaplastic lymphoma kinase (ALK positive lung adenocarcinoma, but the efficacy in lung cancer with bone marrow metastasis was unknown. In the present study, we reported one case of ALK-positive lung adenocarcinoma with bone marrow matastasis given crizotinib treatment, the safety and efficacy was summarized. Methods ALK fusion was tested by fluorescence in situ hybridization (FISH. Crizotinib was given with dose of 250 mg, bid. Objective response was evaluated by Response Evaluation Criteriation in Solid Tumours (RECIST v1.1 and bone marrow response was evaluated by bone marrow puncture and biopsy. Adeverse events were evaluated according to Common Terminology Criteria for Adverse Events (CTC AE v4.0. Results The patient achieved partial response (PR after 6 weeks of crizotinib, especially the objective response of bone marrow metastasis was complete response (CR. The patient stopped crizotinib because of pneumonia. The progression free survival (PFS and overall survival (OS was 20 weeks and 22 weeks respectively. Conclusion Crizotinib could be an effective method for ALK-positive lung cancer with bone marrow metastasis and showed good tolerance.

  18. Membrane fusion during poxvirus entry. (United States)

    Moss, Bernard


    Poxviruses comprise a large family of enveloped DNA viruses that infect vertebrates and invertebrates. Poxviruses, unlike most DNA viruses, replicate in the cytoplasm and encode enzymes and other proteins that enable entry, gene expression, genome replication, virion assembly and resistance to host defenses. Entry of vaccinia virus, the prototype member of the family, can occur at the plasma membrane or following endocytosis. Whereas many viruses encode one or two proteins for attachment and membrane fusion, vaccinia virus encodes four proteins for attachment and eleven more for membrane fusion and core entry. The entry-fusion proteins are conserved in all poxviruses and form a complex, known as the Entry Fusion Complex (EFC), which is embedded in the membrane of the mature virion. An additional membrane that encloses the mature virion and is discarded prior to entry is present on an extracellular form of the virus. The EFC is held together by multiple interactions that depend on nine of the eleven proteins. The entry process can be divided into attachment, hemifusion and core entry. All eleven EFC proteins are required for core entry and at least eight for hemifusion. To mediate fusion the virus particle is activated by low pH, which removes one or more fusion repressors that interact with EFC components. Additional EFC-interacting fusion repressors insert into cell membranes and prevent secondary infection. The absence of detailed structural information, except for two attachment proteins and one EFC protein, is delaying efforts to determine the fusion mechanism.

  19. Heterogeneous breakpoints on the immunoglobulin genes are involved in fusion with the 5' region of BCL2 in B-cell tumors. (United States)

    Yonetani, N; Ueda, C; Akasaka, T; Nishikori, M; Uchiyama, T; Ohno, H


    The 5' flanking region of the BCL2 gene (5'-BCL2) is a breakpoint cluster of rearrangements with immunoglobulin genes (IGs). In contrast to t(14;18)(q32;q21) affecting the 3' region of BCL2, 5'-BCL2 can fuse to not only the heavy chain gene (IGH), but also two light chain gene (IGL) loci. We report here cloning and sequencing of a total of eleven 5'-BCL2 / IGs junctional areas of B-cell tumors, which were amplified by long-distance polymerase chain reaction-based assays. The breakpoints on 5'-BCL2 were distributed from 378 to 2312 bp upstream of the translational initiation site and, reflecting the alteration of regulatory sequences of BCL2, 5'-BCL2 / IGs-positive cells showed markedly higher levels of BCL2 expression than those of t(14;18)-positive cells. In contrast, the breakpoints on the IGs were variable. Two 5'-BCL2 / IGH and two 5'-BCL2 / IGLkappa junctions occurred 5' of the joining (J) segments, suggesting operation of an erroneous variable (V) / diversity (D) / J and V / J rearrangement mechanism. However, two other 5'-BCL2 / IGH junctions affected switch regions, and the kappa-deleting element, which is located 24 kb downstream of the constant region of IGLkappa, followed the 5'-BCL2 in another case. One 5'-BCL2 / IGLkappa and two 5'-BCL2 / IGLlambda junctions involved intronic regions where the normal recombination process does not occur. In the remaining one case, the 5'-BCL2 fused 3' of a Vlambda gene that was upstream of another Vlambda / Jlambda complex carrying a non-producing configuration, indicating that the receptor editing mechanism was likely involved in this rearrangement. Our study revealed heterogeneous anatomy of the 5'-BCL2 / IGs fusion gene leading to transcriptional activation of BCL2, and suggested that the mechanisms underlying the formation of this particular oncogene / IGs recombination are not identical to those of t(14;18).

  20. Fusion Machinery

    DEFF Research Database (Denmark)

    Sørensen, Jakob Balslev; Milosevic, Ira


    the vesicular SNARE VAMP2/synaptobrevin-2 and the target (plasma membrane) SNAREs SNAP25 and syntaxin-1 results in fusion and release of neurotransmitter, synchronized to the electrical activity of the cell by calcium influx and binding to synaptotagmin. Formation of the SNARE complex is tightly regulated...... and appears to start with syntaxin-1 bound to an SM (Sec1/Munc18-like) protein. Proteins of the Munc13-family are responsible for opening up syntaxin and allowing sequential binding of SNAP-25 and VAMP2/synaptobrevin-2. N- to C-terminal “zippering” of the SNARE domains leads to membrane fusion...

  1. Repair of Staphylococcus aureus-infected wound with gene-modified C3H10T1/2 cells expressing BPI-BD3 fusion antibiotic peptide

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    Xin-ran ZHANG


    Full Text Available Objective To study the antibacterial and tissue reparative effect of BPI-BD3 gene-modified mesenchymal stem cells in a mouse model of wound infection. Methods C3H10T1/2 cells were transfected with recombinant adenovirus vector pAdxsi-BPI-BD3, the expression of BPI-BD3 fusion protein was verified by RT-PCR and Western blotting. Excision wound with a diameter of 1cm was inoculated with Staphylococcus aureuswas made on the back of 30 mice. The mice were randomly divided into 3 groups (10 each. Mice in group T were injected with BPI-BD3 gene-modified C3H10T1/2 cells through caudal vein, those in group C were injected with unmodified C3H10T1/2 cells, and in group N were injected with PBS as control. The wound repair result was evaluated by estimation of the percentage of remaining wound area and the amount of wound bacteria under the scar, followed by observation of pathological changes. Inflammatory reactions of the wounds were assessed accordingly. Results The amount of bacteria under the scar was less in group T than in the other two groups (P<0.05. It was also found that the wound healing process was faster in group T than in group C and group N. Pathological observation showed that the inflammatory reaction in group T was also significantly milder than in the other two groups. Conclusion BPI-BD3 gene-modified mesenchymal stem cells may enhance wound repair by controlling infection and promoting tissue regeneration, thus it may be promising in clinical application. DOI: 10.11855/j.issn.0577-7402.2015.09.07

  2. BCR-ABL1 kinase inhibits uracil DNA glycosylase UNG2 to enhance oxidative DNA damage and stimulate genomic instability. (United States)

    Slupianek, A; Falinski, R; Znojek, P; Stoklosa, T; Flis, S; Doneddu, V; Pytel, D; Synowiec, E; Blasiak, J; Bellacosa, A; Skorski, T


    Tyrosine kinase inhibitors (TKIs) revolutionized the treatment of chronic myeloid leukemia in chronic phase (CML-CP). Unfortunately, 25% of TKI-naive patients and 50-90% of patients developing TKI-resistance carry CML clones expressing TKI-resistant BCR-ABL1 kinase mutants. We reported that CML-CP leukemia stem and progenitor cell populations accumulate high amounts of reactive oxygen species, which may result in accumulation of uracil derivatives in genomic DNA. Unfaithful and/or inefficient repair of these lesions generates TKI-resistant point mutations in BCR-ABL1 kinase. Using an array of specific substrates and inhibitors/blocking antibodies we found that uracil DNA glycosylase UNG2 were inhibited in BCR-ABL1-transformed cell lines and CD34(+) CML cells. The inhibitory effect was not accompanied by downregulation of nuclear expression and/or chromatin association of UNG2. The effect was BCR-ABL1 kinase-specific because several other fusion tyrosine kinases did not reduce UNG2 activity. Using UNG2-specific inhibitor UGI, we found that reduction of UNG2 activity increased the number of uracil derivatives in genomic DNA detected by modified comet assay and facilitated accumulation of ouabain-resistant point mutations in reporter gene Na(+)/K(+)ATPase. In conclusion, we postulate that BCR-ABL1 kinase-mediated inhibition of UNG2 contributes to accumulation of point mutations responsible for TKI resistance causing the disease relapse, and perhaps also other point mutations facilitating malignant progression of CML.

  3. Development of novel prime-boost strategies based on a tri-gene fusion recombinant L. tarentolae vaccine against experimental murine visceral leishmaniasis.

    Directory of Open Access Journals (Sweden)

    Noushin Saljoughian

    Full Text Available Visceral leishmaniasis (VL is a vector-borne disease affecting humans and domestic animals that constitutes a serious public health problem in many countries. Although many antigens have been examined so far as protein- or DNA-based vaccines, none of them conferred complete long-term protection. The use of the lizard non-pathogenic to humans Leishmania (L. tarentolae species as a live vaccine vector to deliver specific Leishmania antigens is a recent approach that needs to be explored further. In this study, we evaluated the effectiveness of live vaccination in protecting BALB/c mice against L. infantum infection using prime-boost regimens, namely Live/Live and DNA/Live. As a live vaccine, we used recombinant L. tarentolae expressing the L. donovani A2 antigen along with cysteine proteinases (CPA and CPB without its unusual C-terminal extension (CPB(-CTE as a tri-fusion gene. For DNA priming, the tri-fusion gene was encoded in pcDNA formulated with cationic solid lipid nanoparticles (cSLN acting as an adjuvant. At different time points post-challenge, parasite burden and histopathological changes as well as humoral and cellular immune responses were assessed. Our results showed that immunization with both prime-boost A2-CPA-CPB(-CTE-recombinant L. tarentolae protects BALB/c mice against L. infantum challenge. This protective immunity is associated with a Th1-type immune response due to high levels of IFN-γ production prior and after challenge and with lower levels of IL-10 production after challenge, leading to a significantly higher IFN-γ/IL-10 ratio compared to the control groups. Moreover, this immunization elicited high IgG1 and IgG2a humoral immune responses. Protection in mice was also correlated with a high nitric oxide production and low parasite burden. Altogether, these results indicate the promise of the A2-CPA-CPB(-CTE-recombinant L. tarentolae as a safe live vaccine candidate against VL.

  4. Switch-like reprogramming of gene expression after fusion of multinucleate plasmodial cells of two Physarum polycephalum sporulation mutants. (United States)

    Walter, Pauline; Hoffmann, Xenia-Katharina; Ebeling, Britta; Haas, Markus; Marwan, Wolfgang


    Nonlinear dynamic processes involving the differential regulation of transcription factors are considered to impact the reprogramming of stem cells, germ cells, and somatic cells. Here, we fused two multinucleate plasmodial cells of Physarum polycephalum mutants defective in different sporulation control genes while being in different physiological states. The resulting heterokaryons established one of two significantly different expression patterns of marker genes while the plasmodial halves that were fused to each other synchronized spontaneously. Spontaneous synchronization suggests that switch-like control mechanisms spread over and finally control the entire plasmodium as a result of cytoplasmic mixing. Regulatory molecules due to the large volume of the vigorously streaming cytoplasm will define concentrations in acting on the population of nuclei and in the global setting of switches. Mixing of a large cytoplasmic volume is expected to damp stochasticity when individual nuclei deliver certain RNAs at low copy number into the cytoplasm. We conclude that spontaneous synchronization, the damping of molecular noise in gene expression by the large cytoplasmic volume, and the option to take multiple macroscopic samples from the same plasmodium provide unique options for studying the dynamics of cellular reprogramming at the single cell level.

  5. [Is fetus able to feel pain?]. (United States)

    Kosińska-Kaczyńska, Katarzyna; Wielgoś, Mirosław


    On the basis of fetal hormonal and hemodynamic responses to pain related stimuli, neuroanatomy and observations of preterm babies, it was concluded that human fetus is able to feel pain after 24 weeks gestation. However it is possible that the fetus may feel pain even before that time. With the development of intrauterine diagnostic and therapeutic procedures, it is crucial to provide fetuses undergoing painful procedures not only with anesthesia but also analgesia. The article presents fetal pain research history and its implications for medicine.

  6. Protocol standardization for detection of TMPRSS2 fusions: ERG and gene expression of EZH2, SPINK-1 and NKX3.1 in prostate cancer (PCa

    Directory of Open Access Journals (Sweden)

    Yenifer Yamile Segura Moreno


    Full Text Available At present doesn't exist tool to differentiate patients with prostate cancer (PCa of poor prognosis of those with indolent disease that only require a controlled monitoring of the disease. Because of the coexistence of different premalignant and malignant foci in CaP, the understanding of the carcinogenesis process requires a better understanding. Currently, the morphological heterogeneity in PCa is evaluated with Gleason score, which is closely related to the prognosis of the disease, but this is insufficient so it is currently to work on identifying molecular alterations to identify subtypes that can establish more precisely the patient's prognosis. This preliminary study aimed to standardization of the method of quantification in prostatic samples of FFPE of expression of transcripts of possible biomarkers, such as the oncogenes, SPINK-1 y EZH2, the tumour suppressor, NKX3.1, together with the determination of the presence/absence of gene fusion, TMPRSS2:ERG, being that these transcripts are involved in apparent exclusive events of the natural evolution of PCa, that support the possibility of a molecular classification for this disease.

  7. Increased production of wax esters in transgenic tobacco plants by expression of a fatty acid reductase:wax synthase gene fusion. (United States)

    Aslan, Selcuk; Hofvander, Per; Dutta, Paresh; Sun, Chuanxin; Sitbon, Folke


    Wax esters are hydrophobic lipids consisting of a fatty acid moiety linked to a fatty alcohol with an ester bond. Plant-derived wax esters are today of particular concern for their potential as cost-effective and sustainable sources of lubricants. However, this aspect is hampered by the fact that the level of wax esters in plants generally is too low to allow commercial exploitation. To investigate whether wax ester biosynthesis can be increased in plants using transgenic approaches, we have here exploited a fusion between two bacterial genes together encoding a single wax ester-forming enzyme, and targeted the resulting protein to chloroplasts in stably transformed tobacco (Nicotiana benthamiana) plants. Compared to wild-type controls, transgenic plants showed both in leaves and stems a significant increase in the total level of wax esters, being eight-fold at the whole plant level. The profiles of fatty acid methyl ester and fatty alcohol in wax esters were related, and C16 and C18 molecules constituted predominant forms. Strong transformants displayed certain developmental aberrations, such as stunted growth and chlorotic leaves and stems. These negative effects were associated with an accumulation of fatty alcohols, suggesting that an adequate balance between formation and esterification of fatty alcohols is crucial for a high wax ester production. The results show that wax ester engineering in transgenic plants is feasible, and suggest that higher yields may become achieved in the near future.

  8. Genetic diversity, seasonality and transmission network of human metapneumovirus: identification of a unique sub-lineage of the fusion and attachment genes (United States)

    Chow, Wei Zhen; Chan, Yoke Fun; Oong, Xiang Yong; Ng, Liang Jie; Nor’E, Siti Sarah; Ng, Kim Tien; Chan, Kok Gan; Hanafi, Nik Sherina; Pang, Yong Kek; Kamarulzaman, Adeeba; Tee, Kok Keng


    Human metapneumovirus (HMPV) is an important viral respiratory pathogen worldwide. Current knowledge regarding the genetic diversity, seasonality and transmission dynamics of HMPV among adults and children living in tropical climate remains limited. HMPV prevailed at 2.2% (n = 86/3,935) among individuals presented with acute respiratory tract infections in Kuala Lumpur, Malaysia between 2012 and 2014. Seasonal peaks were observed during the northeast monsoon season (November–April) and correlated with higher relative humidity and number of rainy days (P < 0.05). Phylogenetic analysis of the fusion and attachment genes identified the co-circulation of three known HMPV sub-lineages, A2b and B1 (30.2% each, 26/86) and B2 (20.9%, 18/86), with genotype shift from sub-lineage B1 to A2b observed in 2013. Interestingly, a previously unrecognized sub-lineage of A2 was identified in 18.6% (16/86) of the population. Using a custom script for network construction based on the TN93 pairwise genetic distance, we identified up to nine HMPV transmission clusters circulating as multiple sub-epidemics. Although no apparent major outbreak was observed, the increased frequency of transmission clusters (dyads) during seasonal peaks suggests the potential roles of transmission clusters in driving the spread of HMPV. Our findings provide essential information for therapeutic research, prevention strategies, and disease outbreak monitoring of HMPV. PMID:27279080

  9. Optimizing expression and purification of an ATP-binding gene gsiA from Escherichia coli k-12 by using GFP fusion

    Directory of Open Access Journals (Sweden)

    Zhongshan Wang


    Full Text Available The cloning, expression and purification of the glutathione (sulfur import system ATP-binding protein (gsiA was carried out. The coding sequence of Escherichia coli gsiA, which encodes the ATP-binding protein of a glutathione importer, was amplified by PCR, and then inserted into a prokaryotic expression vector pWaldo-GFPe harboring green fluorescent protein (GFP reporter gene. The resulting recombinant plasmid pWaldo-GFP-GsiA was transformed into various E. coli strains, and expression conditions were optimized. The effect of five E. coli expression strains on the production of the recombinant gsiA protein was evaluated. E. coli BL21 (DE3 was found to be the most productive strain for GsiA-GFP fusion-protein expression, most of which was insoluble fraction. However, results from in-gel and Western blot analysis suggested that expression of recombinant GsiA in Rosetta (DE3 provides an efficient source in soluble form. By using GFP as reporter, the most suitable host strain was conveniently obtained, whereby optimizing conditions for overexpression and purification of the proteins for further functional and structural studies, became, not only less laborious, but also time-saving.

  10. Magnetic fusion; La fusion magnetique

    Energy Technology Data Exchange (ETDEWEB)



    This document is a detailed lecture on thermonuclear fusion. The basic physics principles are recalled and the technological choices that have led to tokamaks or stellarators are exposed. Different aspects concerning thermonuclear reactors such as safety, economy and feasibility are discussed. Tore-supra is described in details as well as the ITER project.

  11. Mitochondrial fusion is increased by the nuclear coactivator PGC-1beta.

    Directory of Open Access Journals (Sweden)

    Marc Liesa

    Full Text Available BACKGROUND: There is no evidence to date on whether transcriptional regulators are able to shift the balance between mitochondrial fusion and fission events through selective control of gene expression. METHODOLOGY/PRINCIPAL FINDINGS: Here, we demonstrate that reduced mitochondrial size observed in knock-out mice for the transcriptional regulator PGC-1beta is associated with a selective reduction in Mitofusin 2 (Mfn2 expression, a mitochondrial fusion protein. This decrease in Mfn2 is specific since expression of the remaining components of mitochondrial fusion and fission machinery were not affected. Furthermore, PGC-1beta increases mitochondrial fusion and elongates mitochondrial tubules. This PGC-1beta-induced elongation specifically requires Mfn2 as this process is absent in Mfn2-ablated cells. Finally, we show that PGC-1beta increases Mfn2 promoter activity and transcription by coactivating the nuclear receptor Estrogen Related Receptor alpha (ERRalpha. CONCLUSIONS/SIGNIFICANCE: Taken together, our data reveal a novel mechanism by which mammalian cells control mitochondrial fusion. In addition, we describe a novel role of PGC-1beta in mitochondrial physiology, namely the control of mitochondrial fusion mainly through Mfn2.

  12. The significance of TMPRSS2-ETS fusion gene in urine in diagnosis of prostate cancer%尿沉淀细胞中TMPRSS2-ETS融合基因对诊断前列腺癌的意义

    Institute of Scientific and Technical Information of China (English)

    丁奕星; 齐隽; 马妍慧; 沈立松


    目的:检测尿沉淀细胞中TMPRSS2-ETS融合基因对诊断前列腺癌的意义.方法:将2010~2011年我院收治的经病理检查证实为前列腺癌者归为前列腺癌组,经病理检查证实为BPH者归为BPH组.收集两组患者前列腺按摩后首次尿液标本,用FISH检测其TMPRSS2-ERG、TMPRSS2-ETV1和TMPRSS2-ETV4融合基因的表达.结果:纳入本次研究的前列腺癌组和BPH组分别为51例和20例,TMPRSS2-ERG(+)的病例分别为26例(50.98%)和4例(20%),差异有显著统计学意义(P<0.05).其敏感度为50.98%,特异度为80%,假阳性率为20%,假阴性率为49.02%,阳性似然比为2.549,阴性似然比为0.613,Youden指数为0.3098.两组中均未发现有TMPRSS2-ETV1或TMPRSS2-ETV4融合基因阳性患者.结论:用FISH方法检测尿沉淀细胞TMPRSS2 ERG融合基因有助于区分前列腺癌与BPH.TMPRSS2-ETV1和TMPRSS2-ETV4融合基因在前列腺癌中出现率较低,故不推荐用于前列腺癌的诊断检测.%Objective:To investigate the significance of detecting TMRPSS2-ETS fusion gene in urine in diagnosis of prostate cancer. Methods: The patients who was treated in the department of urology in our hospital from 2010-2011 were divided into 2 groups. The prostate cancer group including the patients who was diagnosed as prostate cancer by pathology while the benign prostate hyperplasia group including the patients who was diagnosed as benign prostate hyperplasia by pathology. The first voiding urine after prostate massage of the patients in 2 groups was collected and was detected the expression of TMPRSS2-ERG, TMPRSS2-ETV1 and TMPRSS2-ETV4 fusion gene by FISH. Results: There are 51 cases in the prostate cancer group while TMPRSSW-ERG fusion gene in 26 cases(50. 98%)was positive and 20 cases in the benign prostate hyperplasia group while the fusion gene in 4 cases(20%)was positive, which has showing significant difference statistically(P<0. 05). The sensitivity of detection of TMPRSS2-ERG fusion

  13. Silencing of the PiAvr3a effector-encoding gene from Phytophthora infestans by transcriptional fusion to a short interspersed element. (United States)

    Vetukuri, Ramesh R; Tian, Zhendong; Avrova, Anna O; Savenkov, Eugene I; Dixelius, Christina; Whisson, Stephen C


    Phytophthora infestans is the notorious oomycete causing late blight of potato and tomato. A large proportion of the P. infestans genome is composed of transposable elements, the activity of which may be controlled by RNA silencing. Accumulation of small RNAs is one of the hallmarks of RNA silencing. Here we demonstrate the presence of small RNAs corresponding to the sequence of a short interspersed retrotransposable element (SINE) suggesting that small RNAs might be involved in silencing of SINEs in P. infestans. This notion was exploited to develop novel tools for gene silencing in P. infestans by engineering transcriptional fusions of the PiAvr3a gene, encoding an RXLR avirulence effector, to the infSINEm retroelement. Transgenic P. infestans lines expressing either 5'-infSINEm::PiAvr3a-3' or 5'-PiAvr3a::SINEm-3' chimeric transcripts initially exhibited partial silencing of PiAvr3a. Over time, PiAvr3a either recovered wild type transcript levels in some lines, or became fully silenced in others. Introduction of an inverted repeat construct was also successful in yielding P. infestans transgenic lines silenced for PiAvr3a. In contrast, constructs expressing antisense or aberrant RNA transcripts failed to initiate silencing of PiAvr3a. Lines exhibiting the most effective silencing of PiAvr3a were either weakly or non-pathogenic on susceptible potato cv. Bintje. This study expands the repertoire of reverse genetics tools available for P. infestans research, and provides insights into a possible mode of variation in effector expression through spread of silencing from adjacent retroelements.

  14. Unintended Changes in Genetically Modified Rice Expressing the Lysine-Rich Fusion Protein Gene Revealed by a Proteomics Approach

    Institute of Scientific and Technical Information of China (English)

    ZHAO Xiang-xiang; TANG Tang; LIU Fu-xia; LU Chang-li; HU Xiao-lan; JI Li-lian; LIU Qiao-quan


    Development of new technologies for evaluating genetically modiifed (GM) crops has revealed that there are unintended insertions and expression changes in GM crops. Proifling techniques are non-targeted approaches and are capable of detecting more unintended changes in GM crops. Here, we report the application of a comparative proteomic approach to investigate the protein proifle differences between a GM rice line, which has a lysine-rich protein gene, and its non-transgenic parental line. Proteome analysis by two-dimensional gel electrophoresis (2-DE) and mass spectrum analysis of the seeds identiifed 22 differentially expressed protein spots. Apart from a number of glutelins that were detected as targeted proteins in the GM line, the majority of the other changed proteins were involved in carbohydrate metabolism, protein synthesis and stress responses. These results indicated that the altered proteins were not associated with plant allergens or toxicity.

  15. Response of the Gypsy Moth, Lymantria dispar to Transgenic Poplar, Populus simonii x P. nigra, Expressing Fusion Protein Gene of the Spider Insecticidal Peptide and Bt-toxin C-peptide


    Cao, Chuan-Wang; Liu, Gui-Feng; Wang, Zhi-Ying; Yan, Shan-Chun; Ma, Ling; Yang, Chuan-Ping


    The response of the Asian gypsy moth Lymantria dispar (L.) (Lepidoptera: Lymantriidae) to a fusion gene consisting of the spider, Atrax robustus Simon (Araneae: Hexanthelidae) ω?-ACTX-Ar1 sequence coding for an ω?-atracotoxin and a sequence coding for the Bt-toxin C-peptide, expressed in transgenic poplar Populus simonii x P. nigra L. (Malphigiales: Salicaceae) was investigated. Individual performance, feeding selection, midgut proteinase activity and nutrition utilization were monitored. The...

  16. Monitoring of bcr/abl Fusion Gene by Interphase-Dual-Color and Dual-Fusion Fluorescence in situ Hybridization in CML after Allo-HSCT%间期双色双融合荧光原位杂交监测慢性髓系白血病异基因造血干细胞移植后bcr/abl融合基因

    Institute of Scientific and Technical Information of China (English)

    钱思轩; 李建勇; 张闰; 洪鸣; 仇海荣; 李丽; 徐卫; 盛瑞兰; 吴汉新



  17. Tame Fusion

    Institute of Scientific and Technical Information of China (English)

    S.D. Scott


    The first section of this paper covers preliminaries. Essentially, the next four cover units. It is shown that a compatible nearring with DCCR is Nnilpotent if and only if every maximal right N-subgroup is a right ideal. The last five sections relate to fusion (I.e., N-groups minimal for being generated by Nsubgroups, where each is N-isomorphic to a given N-group). Right N-subgroups of a tame nearring N with DCCR, minimal for not annihilating a minimal ideal from the left, are self monogenic and N-isomorphic. That this holds for any collection of minimal ideals is significant. Here, the right N-subgroup involved is a 'fusion product' of the 'components'.

  18. Carpal Fusion



    Carpal fusion may be seen in hereditary and nonhereditary conditions such as acrocallosal syndrome,acromegaly, Apert syndrome, arthrogryposis, Carpenter syndrome, chromosomal abnormalities, ectrodactyly-ectodermal dysplasia-cleft (EEC) syndrome, the F form of acropectorovertebral dysgenesis or the F syndrome, fetal alcohol syndrome, Holt-Oram syndrome, Leopard syndrome, multiple synostosis syndrome, oligosyndactyly syndrome, Pfeiffer-like syndrome, scleroderma, split hand and foot malformatio...

  19. Fusion rules of equivariantizations of fusion categories



    We determine the fusion rules of the equivariantization of a fusion category $\\mathcal{C}$ under the action of a finite group $G$ in terms of the fusion rules of $\\mathcal{C}$ and group-theoretical data associated to the group action. As an application we obtain a formula for the fusion rules in an equivariantization of a pointed fusion category in terms of group-theoretical data. This entails a description of the fusion rules in any braided group-theoretical fusion category.

  20. Quantitative PCR detection of NPM/ALK fusion gene and CD30 gene expression in patients with anaplastic large cell lymphoma--residual disease monitoring and a correlation with the disease status. (United States)

    Kalinova, Marketa; Krskova, Lenka; Brizova, Helena; Kabickova, Edita; Kepak, Tomas; Kodet, Roman


    Anaplastic large cell lymphoma (ALCL) represents a heterogeneous group of malignant lymphoproliferative diseases with a consistent expression of the cytokine receptor CD30. ALCL is frequently associated with a NPM/ALK fusion gene which is found in up to 75% of pediatric ALCLs. Real-time quantitative RT-PCR (RQ-RT-PCR) of NPM/ALK and CD30 gene expression was employed to analyze minimal residual disease (MRD) in 10 patients with NPM/ALK positive ALCL in 79 follow-up bone marrow (BM) and/or peripheral blood (PB) samples. In all BM samples from relapses and/or closely before a relapse, BM samples revealed NPM/ALK and CD30 positivity in at least one of the iliac BM trephines. Five out of nine relapses were preceded or were accompanied by minimally half log increased NPM/ALK levels in the BM. We found that RQ-RT-PCR of the CD30 expression is not suitable for MRD detection--only two relapses were accompanied by an increase of the CD30 level above a level which was detected in BM/PB samples from healthy individuals. RQ-RT-PCR of NPM/ALK expression is a promising and rapid approach for monitoring MRD.

  1. The Ewing's sarcoma EWS/FLI-1 fusion gene encodes a more potent transcriptional activator and is a more powerful transforming gene than FLI-1.


    May, W A; Lessnick, S L; Braun, B S; Klemsz, M; Lewis, B. C.; Lunsford, L B; Hromas, R; Denny, C T


    EWS/FLI-1 is a chimeric protein formed by a tumor-specific 11;22 translocation found in both Ewing's sarcoma and primitive neuroectodermal tumor of childhood. EWS/FLI-1 has been shown to be a potent transforming gene, suggesting that it plays an important role in the genesis of these human tumors. We now demonstrate that EWS/FLI-1 has the characteristics of an aberrant transcription factor. Subcellular fractionation experiments localized the EWS/FLI-1 protein to the nucleus of primitive neuro...

  2. Understanding of decreased sialylation of Fc-fusion protein in hyperosmotic recombinant Chinese hamster ovary cell culture: N-glycosylation gene expression and N-linked glycan antennary profile. (United States)

    Lee, Jong Hyun; Jeong, Yeong Ran; Kim, Yeon-Gu; Lee, Gyun Min


    To understand the effects of hyperosmolality on protein glycosylation, recombinant Chinese hamster ovary (rCHO) cells producing the Fc-fusion protein were cultivated in hyperosmolar medium resulting from adding NaCl (415 mOsm/kg). The hyperosmotic culture showed increased specific Fc-fusion protein productivity (qFc ) but a decreased proportion of acidic isoforms and sialic acid content of the Fc-fusion protein. The intracellular and extracellular sialidase activities in the hyperosmotic cultures were similar to those in the control culture (314 mOsm/kg), indicating that reduced sialylation of Fc-fusion protein at hyperosmolality was not due to elevated sialidase activity. Expression of 52 N-glycosylation-related genes was assessed by the NanoString nCounter system, which provides a direct digital readout using custom-designed color-coded probes. After three days of hyperosmotic culture, nine genes (ugp, slc35a3, slc35d2, gcs1, manea, mgat2, mgat5b, b4galt3, and b4galt4) were differentially expressed over 1.5-fold of the control, and all these genes were down-regulated. N-linked glycan analysis by anion exchange and hydrophilic interaction HPLC showed that the proportion of highly sialylated (di-, tri-, tetra-) and tetra-antennary N-linked glycans was significantly decreased upon hyperosmotic culture. Addition of betaine, an osmoprotectant, to the hyperosmotic culture significantly increased the proportion of highly sialylated and tetra-antennary N-linked glycans (P ≤ 0.05), while it increased the expression of the N-glycan branching/antennary genes (mgat2 and mgat4b). Thus, decreased expression of the genes with roles in the N-glycan biosynthesis pathway correlated with reduced sialic acid content of Fc-fusion protein caused by hyperosmolar conditions. Taken together, the results obtained in this study provide a better understanding of the detrimental effects of hyperosmolality on N-glycosylation, especially sialylation, in rCHO cells. This article is protected

  3. LDL receptor-GFP fusion proteins: new tools for the characterization of disease-causing mutations in the LDL receptor gene

    DEFF Research Database (Denmark)

    Holst, Henrik Uffe; Dagnæs-Hansen, Frederik; Corydon, Thomas Juhl;


    The function of a series of LDL receptor GFP fusion proteins with different, flexible, unstructured spacer regions was analysed. An optimised version of the fusion protein was used to analyse the effect of a LDL receptor mutation (W556S) found in FH patients and characterized as transport defective....... In cultured liver cells this mutation was found to inhibit the transport of LDL receptor GFP fusion protein to the cell surface, thus leading to impaired internalisation of fluorescent labelled LDL. Co-locallisation studies confirmed the retention of the mutant protein in the endoplasmic reticulum....


    Institute of Scientific and Technical Information of China (English)

    Caroline; 黄颖(翻译)


    Fusion World”科技展示体验中心是英国设计公司MET Studio为新加坡科技研究局(A*Star)的科学工程委员会(SERC)所设计的,位于启汇城的办公地点,用于展示该委员会的精选技术作品,以吸引潜在的客户和启汇城内的学生购买群体。

  5. Novel Hydrophobin Fusion Tags for Plant-Produced Fusion Proteins (United States)

    Ritala, Anneli; Linder, Markus; Joensuu, Jussi


    Hydrophobin fusion technology has been applied in the expression of several recombinant proteins in plants. Until now, the technology has relied exclusively on the Trichoderma reesei hydrophobin HFBI. We screened eight novel hydrophobin tags, T. reesei HFBII, HFBIII, HFBIV, HFBV, HFBVI and Fusarium verticillioides derived HYD3, HYD4 and HYD5, for production of fusion proteins in plants and purification by two-phase separation. To study the properties of the hydrophobins, we used N-terminal and C-terminal GFP as a fusion partner. Transient expression of the hydrophobin fusions in Nicotiana benthamiana revealed large variability in accumulation levels, which was also reflected in formation of protein bodies. In two-phase separations, only HFBII and HFBIV were able to concentrate GFP into the surfactant phase from a plant extract. The separation efficiency of both tags was comparable to HFBI. When the accumulation was tested side by side, HFBII-GFP gave a better yield than HFBI-GFP, while the yield of HFBIV-GFP remained lower. Thus we present here two alternatives for HFBI as functional fusion tags for plant-based protein production and first step purification. PMID:27706254

  6. Carpal Fusion

    Directory of Open Access Journals (Sweden)

    Jalal Jalalshokouhi*


    Full Text Available Carpal fusion may be seen in hereditary and nonhereditary conditions such as acrocallosal syndrome,acromegaly, Apert syndrome, arthrogryposis, Carpenter syndrome, chromosomal abnormalities, ectrodactyly-ectodermal dysplasia-cleft (EEC syndrome, the F form of acropectorovertebral dysgenesis or the F syndrome, fetal alcohol syndrome, Holt-Oram syndrome, Leopard syndrome, multiple synostosis syndrome, oligosyndactyly syndrome, Pfeiffer-like syndrome, scleroderma, split hand and foot malformation, Stickler syndrome, thalidomide embryopathy, Turner syndrome and many other conditions as mentioned in Rubinstein-Taybi's book. Sometimes there is no known causative disease.Diagnosis is usually made by plain X-ray during studying a syndrome or congenital disease or could be an incidental finding like our patients. Hand bone anomalies are more common in syndromes or other congenital or non-hereditary conditions, but polydactyly, syndactyly or oligodactyly and carpal fusions are interesting. X-ray is the modality of choice, but MRI and X-ray CT with multiplanar reconstructions may be used for diagnosis.

  7. BCR/ABL stimulates WRN to promote survival and genomic instability (United States)

    Slupianek, Artur; Poplawski, Tomasz; Jozwiakowski, Stanislaw K.; Cramer, Kimberly; Pytel, Dariusz; Stoczynska, Ewelina; Nowicki, Michal O.; Blasiak, Janusz; Skorski, Tomasz


    BCR/ABL-transformed chronic myeloid leukemia (CML) cells accumulate numerous DNA double-strand breaks (DSBs) induced by reactive oxygen species (ROS) and genotoxic agents. To repair these lesions BCR/ABL stimulate unfaithful DSB repair pathways, homologous recombination repair (HRR), non-homologous end-joining (NHEJ) and single-strand annealing (SSA). Here we show that BCR/ABL enhances the expression and increase nuclear localization of WRN (mutated in Werner syndrome), which is required for processing DSB ends during the repair. Other fusion tyrosine kinases (FTKs) such as TEL/ABL, TEL/JAK2, TEL/PDGFβR, and NPM/ALK also elevate WRN. BCR/ABL induces WRN mRNA and protein expression in part by c-MYC -mediated activation of transcription and Bcl-xL –dependent inhibition of caspase-dependent cleavage, respectively. WRN is in complex with BCR/ABL resulting in WRN tyrosine phosphorylation and stimulation of its helicase and exonuclease activities. Activated WRN protects BCR/ABL-positive cells from the lethal effect of oxidative and genotoxic stresses, which causes DSBs. In addition, WRN promotes unfaithful recombination-dependent repair mechanisms HRR and SSA, and enhances the loss of DNA bases during NHEJ in leukemia cells. In summary, we postulate that BCR/ABL-mediated stimulation of WRN modulates the efficiency and fidelity of major DSB repair mechanisms to protect leukemia cells from apoptosis and to facilitate genomic instability. PMID:21123451

  8. BCR/ABL stimulates WRN to promote survival and genomic instability. (United States)

    Slupianek, Artur; Poplawski, Tomasz; Jozwiakowski, Stanislaw K; Cramer, Kimberly; Pytel, Dariusz; Stoczynska, Ewelina; Nowicki, Michal O; Blasiak, Janusz; Skorski, Tomasz


    BCR/ABL-transformed chronic myeloid leukemia (CML) cells accumulate numerous DNA double-strand breaks (DSB) induced by reactive oxygen species (ROS) and genotoxic agents. To repair these lesions BCR/ABL stimulate unfaithful DSB repair pathways, homologous recombination repair (HRR), nonhomologous end-joining (NHEJ), and single-strand annealing (SSA). Here, we show that BCR/ABL enhances the expression and increase nuclear localization of WRN (mutated in Werner syndrome), which is required for processing DSB ends during the repair. Other fusion tyrosine kinases (FTK), such as TEL/ABL, TEL/JAK2, TEL/PDGFβR, and NPM/ALK also elevate WRN. BCR/ABL induces WRN mRNA and protein expression in part by c-MYC-mediated activation of transcription and Bcl-xL-dependent inhibition of caspase-dependent cleavage, respectively. WRN is in complex with BCR/ABL resulting in WRN tyrosine phosphorylation and stimulation of its helicase and exonuclease activities. Activated WRN protects BCR/ABL-positive cells from the lethal effect of oxidative and genotoxic stresses, which causes DSBs. In addition, WRN promotes unfaithful recombination-dependent repair mechanisms HRR and SSA, and enhances the loss of DNA bases during NHEJ in leukemia cells. In summary, we postulate that BCR/ABL-mediated stimulation of WRN modulates the efficiency and fidelity of major DSB repair mechanisms to protect leukemia cells from apoptosis and to facilitate genomic instability.

  9. Transgenic Carrot Expressing Fusion Protein Comprising M. tuberculosis Antigens Induces Immune Response in Mice

    Directory of Open Access Journals (Sweden)

    Natalia V. Permyakova


    Full Text Available Tuberculosis remains one of the major infectious diseases, which continues to pose a major global health problem. Transgenic plants may serve as bioreactors to produce heterologous proteins including antibodies, antigens, and hormones. In the present study, a genetic construct has been designed that comprises the Mycobacterium tuberculosis genes cfp10, esat6 and dIFN gene, which encode deltaferon, a recombinant analog of the human γ-interferon designed for expression in plant tissues. This construct was transferred to the carrot (Daucus carota L. genome by Agrobacterium-mediated transformation. This study demonstrates that the fusion protein CFP10-ESAT6-dIFN is synthesized in the transgenic carrot storage roots. The protein is able to induce both humoral and cell-mediated immune responses in laboratory animals (mice when administered either orally or by injection. It should be emphasized that M. tuberculosis antigens contained in the fusion protein have no cytotoxic effect on peripheral blood mononuclear cells.

  10. Quantification of FML/RARα fusion gene transcripts in patients with acute promyelocytic leukemia by using realtime quantitative PCR%建立实时定量PCR检测急性早幼粒细胞白血病PML/RARa融合基因转录本

    Institute of Scientific and Technical Information of China (English)

    林江; 韩兰秀; 钱军; 王雅丽; 钱震; 杨小飞; 盛晓静


    目的 建立实时定量PCR(real-time quantitative PCR,RQ-PCR)法榆测急性早幼粒细胞白血病(acute promyelocytic leukemia,APL)PML/RARa融合基因转录本,评价其在APL患者诊断和微小残留病(minimal residual disease,MRD)监测中的意义.方法 设计TaqMan探针和引物,建立RQ-PCR法对长型(L-form)、短型(S-form)、变异型(V-form)不同类型PML/RARa阳性模板进行扩增,并检测6例APL患者PML/RARa融合基因转录本含量.结果 RQ-PCR法最低可检测到10个拷贝/μL的阳性模板,但重复性较差,而108~102拷贝/μL阳模的重复性良好,止常对照无扩增信号.6例初诊APL患者不同类型PML/RARa融合基因转录本绝对含量为4.27×103~3.36×105拷贝/50 ng(中位4.33×104拷贝/50 ng),ABL纠正后的相对含量为29.38%~600.53%(中位48.12%).1例患者诱导缓解后PML/RAHa融合基因转录本下降,复发时义明显升高.结论 RQ-PCR方法敏感、可靠,可用于APL患者的诊断和MRD监测.%Objective To establish and evaluate a real time quantitative PCR(RQ-PCR)method for detection and quantifcation the PML/RARa fusion gene transcripts in patients with acute promyelocytic leukemia(APL).Methods Three pairs ofprmers and TaqMan Probe were designed for detecting the most frequent PML/RAPa transcripts(Lfrom,S-from and V-form)and normal ABL was used as an internal control.A real time PCR condition Was established to detect PML/RARa and ABL positive templates with a series of dilutions.To evaluate this assay,bone HlalTOW samples from 6 API,patients were detected.Results In repeated tests,mammal sensitivities of 10 copies/μL were obtained,while reproducible maximal sensitivity achieved 100 copieμL.In 10 normal controls,no amplified fluorescent signdS Were dctectcd.The median absolute and normalized amount of PML/RARtx fusion gene transcripts were 4.27 X 103-3.36x 105 copies/50 ng(median 4.33 X 104 eopies/50ng)and 29.38%.600.53%(median 48.12%)respectively.Onecose showcxt significant

  11. Detection of EML4-ALK fusion gene in patient with lung adenocarcinoma%检测EML4-ALK融合基因对肺腺癌患者的临床意义

    Institute of Scientific and Technical Information of China (English)

    金涛; 朱理; 赵琼; 方维嘉; 曾蕾; 彭玲; 王一青


      目的探讨检测EML4-ALK融合基因对肺腺癌患者的临床意义。方法收集2012年7至12月手术或活检肺腺癌组织标本37例,所有标本均经病理证实为腺癌,用RT-PCR方法检测EML4-ALK融合基因的表达,设计9种探针检测EML4-ALK融合基因最常见的9种类型,并随访阳性患者的治疗结果。结果37例腺癌标本中,1例检出EML4-ALK融合基因表达阳性,并经测序法证实为1型变异,检出率2.70%。为女性患者,左上肺肿块,T1N0M0,行左肺上叶肺癌根治术,术后给予克唑替尼,随访6个月无复发。结论使用RT-PCR结合测序是一种快速、简便、容易推广的EML4-ALK融合基因检测技术,可以在肺腺癌患者检测中推广使用。%Objective To detect echinoderm microtubule-associated protein-like 4 (EML4)-anaplastic lymphoma ki-nase (ALK) fusion gene in patients with non-smal cel lung cancer (NSCLC). Methods Biopsy or surgical specimens were col-lected from 37 patients with pathological y confirmed adenocarcinoma from July 2012 to Dec 2012. The expression of ELM4-ALK fusion gene was detected by RT-PCR with primers designed for 9 subtypes of the fusion gene, and the detected gene was veri-fied by sequencing. Results The expression of ELM4-ALK fusion gene was detected and verified in the surgical specimen from a 36y female patient with T1N0M0 lung adenocarcinoma with a detection rate of 2.70%. The radical resection of upper lobe of the left lung was performed and XALKORI was given one month after the surgery. No metastasis was found after 6-months fol-low-up. Conclusion The detection rate for ELM4-ALK fusion gene is similar as reported in patients with lung adenocarcinoma, and the RT-PCR combined with sequencing is a sensitive and effective method for detection of EML4-ALK fusion gene in surgi-cal or biopsy specimens of patients with lung adenocarninoma.

  12. Catalysed fusion

    CERN Document Server

    Farley, Francis


    A sizzling romance and a romp with subatomic particles at CERN. Love, discovery and adventure in the city where nations meet and beams collide. Life in a large laboratory. As always, the challenges are the same. Who leads? Who follows? Who succeeds? Who gets the credit? Who gets the women or the men? Young Jeremy arrives in CERN and joins the quest for green energy. Coping with baffling jargon and manifold dangers, he is distracted by radioactive rats, lovely ladies and an unscrupulous rival. Full of doubts and hesitations, he falls for a dazzling Danish girl, who leads him astray. His brilliant idea leads to a discovery and a new route to cold fusion. But his personal life is scrambled. Does it bring fame or failure? Tragedy or triumph?

  13. Rapid Evolution to Blast Crisis Associated with a Q252H ABL1 Kinase Domain Mutation in e19a2 BCR-ABL1 Chronic Myeloid Leukaemia

    Directory of Open Access Journals (Sweden)

    Sarah L. McCarron


    Full Text Available A minority of chronic myeloid leukaemia (CML patients express variant transcripts of which the e19a2 BCR-ABL1 fusion is the most common. Instances of tyrosine kinase inhibitor (TKI resistance in e19a2 BCR-ABL1 CML patients have rarely been reported. A case of e19a2 BCR-ABL1 CML is described in whom imatinib resistance, associated with a Q252H ABL1 kinase domain mutation, became apparent soon after initiation of TKI therapy. The patient rapidly transformed to myeloid blast crisis (BC with considerable bone marrow fibrosis and no significant molecular response to a second generation TKI. The clinical course was complicated by comorbidities with the patient rapidly succumbing to advanced disease. This scenario of Q252H-associated TKI resistance with rapid BC transformation has not been previously documented in e19a2 BCR-ABL1 CML. This case highlights the considerable challenges remaining in the management of TKI-resistant BC CML, particularly in the elderly patient.

  14. Preparation, characterization, and in ovo vaccination of dextran-spermine nanoparticle DNA vaccine coexpressing the fusion and hemagglutinin genes against Newcastle disease. (United States)

    Firouzamandi, Masoumeh; Moeini, Hassan; Hosseini, Seyed Davood; Bejo, Mohd Hair; Omar, Abdul Rahman; Mehrbod, Parvaneh; El Zowalaty, Mohamed E; Webster, Thomas J; Ideris, Aini


    Plasmid DNA (pDNA)-based vaccines have emerged as effective subunit vaccines against viral and bacterial pathogens. In this study, a DNA vaccine, namely plasmid internal ribosome entry site-HN/F, was applied in ovo against Newcastle disease (ND). Vaccination was carried out using the DNA vaccine alone or as a mixture of the pDNA and dextran-spermine (D-SPM), a nanoparticle used for pDNA delivery. The results showed that in ovo vaccination with 40 μg pDNA/egg alone induced high levels of antibody titer (P0.05). Higher antibody titer was observed in the group immunized with 40 μg pDNA/egg at 4 weeks postvaccination. The findings also showed that vaccination with 40 μg pDNA/egg alone was able to confer protection against Newcastle disease virus strain NDIBS002 in two out of seven SPF chickens. Although the chickens produced antibody titers 3 weeks after in ovo vaccination, it was not sufficient to provide complete protection to the chickens from lethal viral challenge. In addition, vaccination with pDNA/D-SPM complex did not induce high antibody titer when compared with naked pDNA. Therefore, it was concluded that DNA vaccination with plasmid internal ribosome entry site-HN/F can be suitable for in ovo application against ND, whereas D-SPM is not recommended for in ovo gene delivery.

  15. Neuronal c-Abl activation leads to induction of cell cycle and interferon signaling pathways

    Directory of Open Access Journals (Sweden)

    Schlatterer Sarah D


    Full Text Available Abstract Background Expression of active c-Abl in adult mouse forebrain neurons in the AblPP/tTA mice resulted in severe neurodegeneration, particularly in the CA1 region of the hippocampus. Neuronal loss was preceded and accompanied by substantial microgliosis and astrocytosis. In contrast, expression of constitutively active Arg (Abl-related gene in mouse forebrain neurons (ArgPP/tTA mice caused no detectable neuronal loss or gliosis, although protein expression and kinase activity were at similar levels to those in the AblPP/tTA mice. Methods To begin to elucidate the mechanism of c-Abl-induced neuronal loss and gliosis, gene expression analysis of AblPP/tTA mouse forebrain prior to development of overt pathology was performed. Selected results from gene expression studies were validated with quantitative reverse transcription PCR , immunoblotting and bromodeoxyuridine (BrdU labeling, and by immunocytochemistry. Results Two of the top pathways upregulated in AblPP/tTA mice with c-Abl expression for 2 weeks were cell cycle and interferon signaling. However, only the expression of interferon signaling pathway genes remained elevated at 4 weeks of c-Abl induction. BrdU incorporation studies confirm that, while the cell cycle pathway is upregulated in AblPP/tTA mice at 2 weeks of c-Abl induction, the anatomical localization of the pathway is not consistent with previous pathology seen in the AblPP/tTA mice. Increased expression and activation of STAT1, a known component of interferon signaling and interferon-induced neuronal excitotoxicity, is an early consequence of c-Abl activation in AblPP/tTA mice and occurs in the CA1 region of the hippocampus, the same region that goes on to develop severe neurodegenerative pathology and neuroinflammation. Interestingly, no upregulation of gene expression of interferons themselves was detected. Conclusions Our data suggest that the interferon signaling pathway may play a role in the pathologic processes

  16. Endocrine Disrupters: the new players able to affect the epigenome.

    Directory of Open Access Journals (Sweden)

    Lavinia eCasati


    Full Text Available Epigenetics represents the way by which the environment is able to program the genome; there are three main levels of epigenetic control on genome: DNA methylation, post-translational histone modification and microRNA expression. The term Epigenetics has been widened by NIH to include both heritable changes in gene activity and expression but also stable, long-term alterations in the transcriptional potential of a cell that are not necessarily heritable. These changes might be produced mostly by the early life environment and might affect health influencing the susceptibility to develop diseases, from cancer to mental disorder, during the entire life span. The most studied environmental influences acting on epigenome are diet, infections, wasting, child care, smoking and environmental pollutants, in particular endocrine disrupters (EDs. These are environmental xenobiotics able to interfere with the normal development of the male and female reproductive systems of wildlife, of experimental animals and possibly of humans, disrupting the normal reproductive functions. Data from literature indicate that EDs can act at different levels of epigenetic control, in some cases transgenerationally, in particular when the exposure to these compounds occurs during the prenatal and earliest period of life. Some of the best characterized EDs will be considered in this review. Among the EDs, vinclozolin (VZ and methoxychlor (MXC promote epigenetic transgenerational effects. Polychlorinated biphenils (PCBs, the most widespread environmental EDs, affect histone post-translational modifications in a dimorphic way, possibly as the result of an alteration of gene expression of the enzymes involved in histone modification, as the demethylase Jarid1b, an enzyme also involved in regulating the interaction of androgens with their receptor.

  17. 急性早幼粒细胞白血病PML/RARα融合基因检测的临床意义%Clinical significance of detecting PML/RARα fusion gene in acute promyelocytic leukemia

    Institute of Scientific and Technical Information of China (English)

    刘小宁; 李元媛; 甘宜敏; 陈凤丽; 衡春; 于亮


    目的:探讨荧光原位杂交技术(FISH)检测PML/RARα融合基因在急性早幼粒细胞白血病(APL)诊断中的应用价值。方法应用常规细胞遗传学分析28例APL患者的染色体核型,同时用FISH法检测PML/RARα融合基因。结果28例初诊的APL患者254例常规核型分析检出t(15;17)(q22;q12);2例患者核型正常;另1例未检出t(15;17),核型分析结果为涉及15和17号染色体的复杂异常;FISH检测所有病例均存在PML/RARα融合基因。结论 FISH法行PML/RARα融合基因检测特异性和敏感性好,是诊断APL的可靠方法。%Objective To explore the application value of detecting PML/RARα fusion gene in the diagnosis of acute promye-locytic leukemia(APL) with fluorescence in situ hybridization(FISH). Methods The chromosome karyotype of 28 cases of APL pa-tients was analyzed with the conventional cytogenetic analysis method and the PML/RARα fusion gene was detected with FISH method. Results Among 28 first diagnosed APL patients,the t (15;17)(q22;q12) was detected in 24 25 patients with the conven-tional karyotype analysis;Two patients were detected to be normal karyotype;And the t (15;17) was not detected in one case. The karyotype analysis results showed the complex abnormalities involving in the chromosome 15 and 17;The PML/RARα fusion gene was detected to exist in all cases with FISH detection. Conclusion The PML/RARα fusion gene detection with FISH method is a reliable way for the diagnosis of APL with fine specificity and sensitivity.

  18. Correcting mitochondrial fusion by manipulating mitofusin conformations (United States)

    Franco, Antonietta; Kitsis, Richard N.; Fleischer, Julie A.; Gavathiotis, Evripidis; Kornfeld, Opher S.; Gong, Guohua; Biris, Nikolaos; Benz, Ann; Qvit, Nir; Donnelly, Sara K; Chen, Yun; Mennerick, Steven; Hodgson, Louis; Mochly-Rosen, Daria; Dorn, Gerald W


    Summary Mitochondria are dynamic organelles, remodeling and exchanging contents during cyclic fusion and fission. Genetic mutations of mitofusin (Mfn) 2 interrupt mitochondrial fusion and cause the untreatable neurodegenerative condition, Charcot Marie Tooth disease type 2A (CMT2A). It has not been possible to directly modulate mitochondrial fusion, in part because the structural basis of mitofusin function is incompletely understood. Here we show that mitofusins adopt either a fusion-constrained or fusion-permissive molecular conformation directed by specific intramolecular binding interactions, and demonstrate that mitofusin-dependent mitochondrial fusion can be regulated by targeting these conformational transitions. Based on this model we engineered a cell-permeant minipeptide to destabilize fusion-constrained mitofusin and promote the fusion-permissive conformation, reversing mitochondrial abnormalities in cultured fibroblasts and neurons harboring CMT2A gene defects. The relationship between mitofusin conformational plasticity and mitochondrial dynamism uncovers a central mechanism regulating mitochondrial fusion whose manipulation can correct mitochondrial pathology triggered by defective or imbalanced mitochondrial dynamics. PMID:27775718

  19. 原发性胃淋巴瘤凋亡抑制蛋白2-黏膜相关淋巴瘤转位基因1融合基因检测的意义%The clinical value of apoptosis protein 2-mucosa associated lymphoid tissue lymphoma translocation gene 1 fusion gene detection in tissue of primary gastric lymphoma

    Institute of Scientific and Technical Information of China (English)

    许国强; 陈妙辉; 陈洪潭; 单国栋; 胡凤玲; 杨铭


    Objective To explore the feasibility of inhibitor of apoptosis protein 2-mucosa associated lymphoid tissue lymphoma translocation gene 1 (API2-MALT1) fusion gene detection in endoscopic biopsy tissue of primary gastric lymphoma (PGL), and to study the expression of this fusion gene in PGL and its clinical values in diagnosis and treatment of PGL.Methods A total of 32 suspicious PGL patients underwent endoscopic ultrasonography (EUS) examination and mucosal biopsy.The biopsy specimens were conducted histopathology examination, immunohistochemistry detection and real time RT-PCR for API2-MALT1 fusion gene expression.The expression of API2MALT1 fusion gene in clearly diagnosed PGL and its relation with PGL diagnosis, classification and treatment were analyzed and summarized.Results A total of 14 cases of 32 suspicious PGL patients were diagnosed as PGL by histopathology and immunohistochemistry examination, which including 11 cases of gastric mucosa-associated lymphoid tissue (MALT) lymphoma and 3 cases of gastric diffuse large B-cell lymphoma (DLBCL).There were 5 API2-MALT1 fusion gene positive cases, which were all MALT lymphoma, about 5 out of total 11 MALT lymphoma patients.API2-MALT1 fusion gene expression was negative in all 3 gastric DLBCL cases.The depth of lesion invasion and lymph nodes metastasis were more severe in API2-MALT1 fusion gene negative group than those of positive group.There were 2 of 5 API2-MALT1 fusion gene positive cases were Hp positive, and 5 of 9 negative cases were Hp positive.The anti-Hp therapy in 5 API2-MALT1 fusion gene positive cases was ineffective,however, chemotherapy was effective.In negative group, 2 of 5 Hp positive cases was complete remission and 4 Hp negative cases were ineffective with anti-Hp therapy.Concltsions API2-MALT1 fusion gene is common genetic abnormality in gastric MALT lymphoma.The detection of this fusion gene expression in endoscopic biopsy specimen by real time RT-PCR is clinically practical, and the

  20. An improved vector system for constructing transcriptional lacZ fusions: analysis of regulation of the dnaA, dnaN, recF and gyrB genes of Escherichia coli. (United States)

    Macián, F; Pérez-Roger, I; Armengod, M E


    We describe a new vector system for the in vitro construction of transcriptional fusions to the lacZ gene, which is expressed from the translational start signals of galK. The galK ribosome-binding site (RBS) and its natural preceding region ensure a constant efficiency for lacZ translation and, thus, the beta-galactosidase (beta Gal) production of a given fusion is directly proportional to the in vivo transcriptional activity of the inserted DNA fragment. Single-copy lambda prophage versions of multicopy constructs can be made by in vivo recombination. We use this system to compare the transcriptional activities of the promoters present in the dnaA-dnaN-recF-gyrB cluster. The order of strength of these promoters is gyrB > dnaA > recF > dnaN. It is assumed that gyrB belongs to the dnaA-dnaN-recF operon, because the short recF-gyrB intercistronic region does not contain a terminator. By using this new vector system, we have detected strong termination signals within recF that are functional even when recF is translated at its normal rate. The low level of transcription coming to the end of recF, and the highest activity of the gyrB promoter, as well as results obtained with several gyrB::lacZ translational fusions, support the conclusion that gyrB is predominantly expressed from its own promoter under standard growth conditions. Finally, we have found that transcription from the dnaA promoters is constant at different growth rates. This supports the idea that autoregulation of the dnaA gene is responsible for the coupling of the DnaA protein synthesis to cell mass increase, and accumulation of DnaA protein governs the initiation of chromosome replication.

  1. Cold fusion research

    Energy Technology Data Exchange (ETDEWEB)



    I am pleased to forward to you the Final Report of the Cold Fusion Panel. This report reviews the current status of cold fusion and includes major chapters on Calorimetry and Excess Heat, Fusion Products and Materials Characterization. In addition, the report makes a number of conclusions and recommendations, as requested by the Secretary of Energy.

  2. Effects of Character Education on the Self-Esteem of Intellectually Able and Less Able Elementary Students in Kuwait (United States)

    Tannir, Abir; Al-Hroub, Anies


    This research study investigates effects of character education activities on the self-esteem of intellectually able and less able students in the lower elementary level in Kuwait. The participants were 39 students in grade three with an average age of eight years old. Students were first divided into two ability subgroups (intellectually able vs.…

  3. Viral membrane fusion

    Energy Technology Data Exchange (ETDEWEB)

    Harrison, Stephen C., E-mail:


    Membrane fusion is an essential step when enveloped viruses enter cells. Lipid bilayer fusion requires catalysis to overcome a high kinetic barrier; viral fusion proteins are the agents that fulfill this catalytic function. Despite a variety of molecular architectures, these proteins facilitate fusion by essentially the same generic mechanism. Stimulated by a signal associated with arrival at the cell to be infected (e.g., receptor or co-receptor binding, proton binding in an endosome), they undergo a series of conformational changes. A hydrophobic segment (a “fusion loop” or “fusion peptide”) engages the target-cell membrane and collapse of the bridging intermediate thus formed draws the two membranes (virus and cell) together. We know of three structural classes for viral fusion proteins. Structures for both pre- and postfusion conformations of illustrate the beginning and end points of a process that can be probed by single-virion measurements of fusion kinetics. - Highlights: • Viral fusion proteins overcome the high energy barrier to lipid bilayer merger. • Different molecular structures but the same catalytic mechanism. • Review describes properties of three known fusion-protein structural classes. • Single-virion fusion experiments elucidate mechanism.

  4. Increased retention of functional fusions to toxic genes in new two-hybrid libraries of the E. coli strain MG1655 and B. subtilis strain 168 genomes, prepared without passaging through E. coli

    Directory of Open Access Journals (Sweden)

    Projan Steve


    Full Text Available Abstract Background Cloning of genes in expression libraries, such as the yeast two-hybrid system (Y2H, is based on the assumption that the loss of target genes is minimal, or at worst, managable. However, the expression of genes or gene fragments that are capable of interacting with E. coli or yeast gene products in these systems has been shown to be growth inhibitory, and therefore these clones are underrepresented (or completely lost in the amplified library. Results Analysis of candidate genes as Y2H fusion constructs has shown that, while stable in E. coli and yeast for genetic studies, they are rapidly lost in growth conditions for genomic libraries. This includes the rapid loss of a fragment of the E. coli cell division gene ftsZ which encodes the binding site for ZipA and FtsA. Expression of this clone causes slower growth in E. coli. This clone is also rapidly lost in yeast, when expressed from a GAL1 promoter, relative to a vector control, but is stable when the promoter is repressed. We have demonstrated in this report that the construction of libraries for the E. coli and B. subtilis genomes without passaging through E. coli is practical, but the number of transformants is less than for libraries cloned using E. coli as a host. Analysis of several clones in the libraries that are strongly growth inhibitory in E. coli include genes for many essential cellular processes, such as transcription, translation, cell division, and transport. Conclusion Expression of Y2H clones capable of interacting with E. coli and yeast targets are rapidly lost, causing a loss of complexity. The strategy for preparing Y2H libraries described here allows the retention of genes that are toxic when inappropriately expressed in E. coli, or yeast, including many genes that represent potential antibacterial targets. While these methods are generally applicable to the generation of Y2H libraries from any source, including mammalian and plant genomes, the

  5. Fusion expression of mutated cecropin CMIV in E. coli

    Institute of Scientific and Technical Information of China (English)

    谢维; 邱奇峰; 董雪吟; 华子春; 徐贤秀


    A cDNA coding mutated cecropin CMIV from Bombyx mori was synthesized according to its amino acid sequense using E .coli biased codons .The gene was cloned into the fusion expression vector pEZZ318 and was expressed in E .coli HB101.The fusion protein produced was purified by affinity chromatography to yield 26 mg/L fusion product .The anti-bacterial activities of recombinant cecropin CMIV were recovered after cleavage by chemical method.

  6. Protein-protein fusion catalyzed by sortase A. (United States)

    Levary, David A; Parthasarathy, Ranganath; Boder, Eric T; Ackerman, Margaret E


    Chimeric proteins boast widespread use in areas ranging from cell biology to drug delivery. Post-translational protein fusion using the bacterial transpeptidase sortase A provides an attractive alternative when traditional gene fusion fails. We describe use of this enzyme for in vitro protein ligation and report the successful fusion of 10 pairs of protein domains with preserved functionality--demonstrating the robust and facile nature of this reaction.

  7. Protein-protein fusion catalyzed by sortase A.

    Directory of Open Access Journals (Sweden)

    David A Levary

    Full Text Available Chimeric proteins boast widespread use in areas ranging from cell biology to drug delivery. Post-translational protein fusion using the bacterial transpeptidase sortase A provides an attractive alternative when traditional gene fusion fails. We describe use of this enzyme for in vitro protein ligation and report the successful fusion of 10 pairs of protein domains with preserved functionality--demonstrating the robust and facile nature of this reaction.

  8. Abl suppresses cell extrusion and intercalation during epithelium folding. (United States)

    Jodoin, Jeanne N; Martin, Adam C


    Tissue morphogenesis requires control over cell shape changes and rearrangements. In the Drosophila mesoderm, linked epithelial cells apically constrict, without cell extrusion or intercalation, to fold the epithelium into a tube that will then undergo epithelial-to-mesenchymal transition (EMT). Apical constriction drives tissue folding or cell extrusion in different contexts, but the mechanisms that dictate the specific outcomes are poorly understood. Using live imaging, we found that Abelson (Abl) tyrosine kinase depletion causes apically constricting cells to undergo aberrant basal cell extrusion and cell intercalation. abl depletion disrupted apical-basal polarity and adherens junction organization in mesoderm cells, suggesting that extruding cells undergo premature EMT. The polarity loss was associated with abnormal basolateral contractile actomyosin and Enabled (Ena) accumulation. Depletion of the Abl effector Enabled (Ena) in abl-depleted embryos suppressed the abl phenotype, consistent with cell extrusion resulting from misregulated ena Our work provides new insight into how Abl loss and Ena misregulation promote cell extrusion and EMT.

  9. Response of the gypsy moth, Lymantria dispar to transgenic poplar, Populus simonii x P. nigra, expressing fusion protein gene of the spider insecticidal peptide and Bt-toxin C-peptide. (United States)

    Cao, Chuan-Wang; Liu, Gui-Feng; Wang, Zhi-Ying; Yan, Shan-Chun; Ma, Ling; Yang, Chuan-Ping


    The response of the Asian gypsy moth Lymantria dispar (L.) (Lepidoptera: Lymantriidae) to a fusion gene consisting of the spider, Atrax robustus Simon (Araneae: Hexanthelidae) ω-ACTX-Ar1 sequence coding for an ω-atracotoxin and a sequence coding for the Bt-toxin C-peptide, expressed in transgenic poplar Populus simonii x P. nigra L. (Malphigiales: Salicaceae) was investigated. Individual performance, feeding selection, midgut proteinase activity and nutrition utilization were monitored. The growth and development of L. dispar were significantly affected by continually feeding on the transgenic poplar, with the larval instars displaying significantly shorter developmental times than those fed on nontransgenic poplar, but pupation was delayed. Mortality was higher in populations fed transgenic poplar leaves, than for larvae fed nontransgenic poplar leaves. The cumulative mortality during all stages of larvae fed transgenic leaves was 92% compared to 16.7% of larvae on nontransgenic leaves. The highest mortality observed was 71.7% in the last larval instar stage. A two-choice test showed that fifth-instar larvae preferred to feed on nontransgenic leaves at a ratio of 1:1.4. Feeding on transgenic leaves had highly significant negative effects on relative growth of larvae, and the efficiency of conversion of ingested and digested food. Activity of major midgut proteinases was measured using substrates TAME and BTEE showed significant increases in tryptase and chymotrypsinlike activity (9.2- and 9.0-fold, respectively) in fifth-instar larvae fed on transgenic leaves over control. These results suggest transgenic poplar is resistant to L. dispar, and the mature L. dispar may be weakened by the transgenic plants due to Bt protoxins activated by elevated major midgut proteinase activity. The new transgenic poplar expressing fusion protein genes of Bt and a new spider insecticidal peptide are good candidates for managing gypsy moth.

  10. A New Target in Non-small Cell Lung Cancer: EML4-ALK Fusion Gene%非小细胞肺癌治疗的新靶点:EML4-ALK融合基因

    Institute of Scientific and Technical Information of China (English)

    王慧娟; 袁静


    It was only 3 years ago that the fusion gene between echinoderm microtubule-associated protein-like4 (EML4) and anaplastic lymphoma kinase (ALK) has been identified in a subset of non-small cell lung cancer (NSCLC).EML4-ALK is most often detected in never smokers with lung adenocarcinoma and has unique pathologic features.EML4ALK fusion gene is oncogenic, which could be suppressed by ALK-inhibitor through blocking the downstream signaling passway of EML4-ALK.This review will focus on the molecular structure, function, biology, detection method and the diagnostic and therapeutic meaning of EML4-ALK of lung cancer.%3年前棘皮动物微管相关类蛋白4 (echinoderm microtubule-associated protein-like4,EML4)与间变性淋巴瘤激酶 (anaplastic lymphoma kinase,ALK)融合基因被发现存在于部分非小细胞肺癌(non-small cell lung cancer,NSCLC)中.该融合基因常见于不吸烟的肺腺癌患者,有其独特的病理学特征,可以诱导肿瘤生成.ALK抑制剂能够作用于该基因的下游信号传导通路并拮抗其促肿瘤生成活性.本文旨在介绍EML4-ALK基因的结构、功能、生物学特征、检测方法 及其在肺癌诊断治疗中的意义.

  11. Promoter hypermethylation of the retinoic acid receptor beta2 gene is frequent in acute myeloid leukaemia and associated with the presence of CBFβ-MYH11 fusion transcripts

    DEFF Research Database (Denmark)

    Rethmeier, Anita; Aggerholm, Anni; Olesen, Lene Hyldahl;


    was unmethylated in 10/10 bone marrow and 7/7 blood samples from healthy individuals, the gene was hypermethylated in 43% of the AML patients. The RARbeta2 degree of promoter methylation differed between and within individuals, and the mRNA transcription levels of the gene varied inter-individually by a factor...

  12. Economics of fusion research

    Energy Technology Data Exchange (ETDEWEB)

    None, None


    This report provides the results of a study of methods of economic analysis applied to the evaluation of fusion research. The study recognizes that a hierarchy of economic analyses of research programs exists: standard benefit-cost analysis, expected value of R and D information, and expected utility analysis. It is shown that standard benefit-cost analysis, as commonly applied to research programs, is inadequate for the evaluation of a high technology research effort such as fusion research. A methodology for performing an expected value analysis is developed and demonstrated and an overview of an approach to perform an expected utility analysis of fusion research is presented. In addition, a potential benefit of fusion research, not previously identified, is discussed and rough estimates of its magnitude are presented. This benefit deals with the effect of a fusion research program on optimal fossil fuel consumption patterns. The results of this study indicate that it is both appropriate and possible to perform an expected value analysis of fusion research in order to assess the economics of a fusion research program. The results indicate further that the major area of benefits of fusion research is likely due to the impact of a fusion research program on optimal fossil fuel consumption patterns and it is recommended that this benefit be included in future assessments of fusion research economics.

  13. Materials research for fusion (United States)

    Knaster, J.; Moeslang, A.; Muroga, T.


    Fusion materials research started in the early 1970s following the observation of the degradation of irradiated materials used in the first commercial fission reactors. The technological challenges of fusion energy are intimately linked with the availability of suitable materials capable of reliably withstanding the extremely severe operational conditions of fusion reactors. Although fission and fusion materials exhibit common features, fusion materials research is broader. The harder mono-energetic spectrum associated with the deuterium-tritium fusion neutrons (14.1 MeV compared to hydrogen and helium as transmutation products that might lead to a (at present undetermined) degradation of structural materials after a few years of operation. Overcoming the historical lack of a fusion-relevant neutron source for materials testing is an essential pending step in fusion roadmaps. Structural materials development, together with research on functional materials capable of sustaining unprecedented power densities during plasma operation in a fusion reactor, have been the subject of decades of worldwide research efforts underpinning the present maturity of the fusion materials research programme.

  14. Cell fusion in tumor progression: the isolation of cell fusion products by physical methods

    Directory of Open Access Journals (Sweden)

    Vincitorio Massimo


    Full Text Available Abstract Background Cell fusion induced by polyethylene glycol (PEG is an efficient but poorly controlled procedure for obtaining somatic cell hybrids used in gene mapping, monoclonal antibody production, and tumour immunotherapy. Genetic selection techniques and fluorescent cell sorting are usually employed to isolate cell fusion products, but both procedures have several drawbacks. Results Here we describe a simple improvement in PEG-mediated cell fusion that was obtained by modifying the standard single-step procedure. We found that the use of two PEG undertreatments obtains a better yield of cell fusion products than the standard method, and most of these products are bi- or trinucleated polykaryocytes. Fusion rate was quantified using fluorescent cell staining microscopy. We used this improved cell fusion and cell isolation method to compare giant cells obtained in vitro and giant cells obtained in vivo from patients with Hodgkin's disease and erythroleukemia. Conclusions In the present study we show how to improve PEG-mediated cell fusion and that cell separation by velocity sedimentation offers a simple alternative for the efficient purification of cell fusion products and to investigate giant cell formation in tumor development.

  15. 检测循环前列腺癌细胞中TMPRSS2:ERG融合基因%Detection of TMPRSS2:ERG fusion gene in circulating prostate cancer cells

    Institute of Scientific and Technical Information of China (English)

    Xueying Mao; Dan Berney; David M. Prowse; Yong-Jie Lu; Greg Shaw; Sharon Y. James; Patraicia Purkis; Sakunthala C. Kudahetti; Theodora Tsigani; Saname Kia; Bryan D. Young; R. Tim D. Oliver


    目的:研究前列腺癌病人的循环癌细胞(CTC)中TMPRSS2:ERG融合基因的存在及其与肿瘤转移之间的潜在关系.方法:利用RT-PCR,在27例前列腺切除术中得到的前列腺癌活检标本中检测TMPRSS2:ERG和TMPRSS2:ETV1转录子出现的频率,在15名晚期雄激素非依赖病人的循环癌细胞中检测TMPRSS2:ERGG转录子出现的频率.利用荧光原位杂交技术(FISH)分析10个CTC样本(取自15个CTC样本)中导致TMPRSS2:ERG融合的ERG基因组截短情况.结果:在44%的样本中发现了TMPRSS2:ERGG转录子,但是没有检测到TMPRSS2:ETV1转录子的表达.FISH分析结果显示在10例CTC样本的6例中发现染色体重组影响了ERG基因,包括一例在原癌位置上发生了TMPRSS2:ERG融合.可是,在15例CTC样本中没有检测到TMPRSS2:ERG转录子,包括用FISH检测的10例.结论:虽然需要进一步研究来确认TMPRSS2:ERG融合与前列腺癌转移之间的关系,但是通过FISH分析ERG基因基因组截短是一种有效的监测CTC的出现和前列腺癌潜在转移的方法.%Aim: To investigate the existence of TMPRSS2:ERG fusion gene in circulating tumor cells (CTC) from prostate cancer patients and its potential in monitoring tumor metastasis. Methods: We analyzed the frequency of TMPRSS2:ERG and TMPRSS2:ETV1 transcripts in 27 prostate cancer biopsies from prostatectomies, and TMPRSS2:ERG transcripts in CTC isolated from 15 patients with advanced androgen independent disease using reverse transcription polymerase chain reaction (RT-PCR). Fluorescence in situ hybridization (FISH) was applied to analyze the genomic truncation of ERG, which is the result of TMPRSS2:ERG fusion in 10 of the 15 CTC samples. Results: TMPRSS2:ERG transcripts were found in 44% of our samples, but we did not detect expression of TMPRSS2:ETV1. Using FISH analysis we detected chromosomal rearrangements affecting the ERG gene in 6 of 10 CTC samples, including 1 case with associated TMPRSS2:ERG fusion at the

  16. [Sequence analysis of the matrix protein and fusion protein genes of a field peste des petits ruminants virus strain from Tibet, China]. (United States)

    Bao, Jing-Yue; Zhao, Wen-Ji; Wang, Zhi-Liang; Li, Lin; Wu, Guo-Zhen; Wu, Xiao-Dong; Liu, Chun-Ju; Wang, Jun-Wei; Liu, Yu-Tian; Li, Jin-Ming; Wang, Ying-Li


    The nucleotide sequences of M and F genes from a field strain of peste des petits ruminants virus (PPRV) ("China/Tib/Gej/07-30") was firstly determined. The M gene was 1 483 nucleotides in length with a single open reading frame (ORF), encoding a protein of 335 amino acids. The F gene was 2411 nucleotides in length, encoding a protein of 546 amino acids. The resulting nucleotide sequence and the deduced amino acid sequences were compared with the homologous regions of other PPRV isolates. The nucleotide sequences of M and F genes of the "China/Tib/Gej/07-30" was 92.4%-97.7% and 85.5%-96.1% identical to other PPRV isolates, respectively, while a homology of 97.0%-98.2% and 94.3%-98.2% could be observed at the amino acids level respectively. Several sequence motifs in the M and F genes had been identified on the basis of conservation in the PPRVs and the morbilliviruses. The 3' untranslated region of M gene was 443 nucleotides in length with 82.4%-93.5% identical to other PPRV isolates. The 5' untranslated region of F gene was 634 nucleotides in length with 76.2%-91.7% identical to other PPRV isolates.

  17. Muon Catalyzed Fusion (United States)

    Armour, Edward A.G.


    Muon catalyzed fusion is a process in which a negatively charged muon combines with two nuclei of isotopes of hydrogen, e.g, a proton and a deuteron or a deuteron and a triton, to form a muonic molecular ion in which the binding is so tight that nuclear fusion occurs. The muon is normally released after fusion has taken place and so can catalyze further fusions. As the muon has a mean lifetime of 2.2 microseconds, this is the maximum period over which a muon can participate in this process. This article gives an outline of the history of muon catalyzed fusion from 1947, when it was first realised that such a process might occur, to the present day. It includes a description of the contribution that Drachrnan has made to the theory of muon catalyzed fusion and the influence this has had on the author's research.

  18. A sensitive HIV-1 envelope induced fusion assay identifies fusion enhancement of thrombin

    Energy Technology Data Exchange (ETDEWEB)

    Cheng, De-Chun; Zhong, Guo-Cai; Su, Ju-Xiang [Department of Microbiology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Liu, Yan-Hong [Second Affiliated Hospital of Harbin Medical University, 246 Xuefu Road, Harbin, Heilongjiang 150081 (China); Li, Yan; Wang, Jia-Ye [Department of Microbiology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Hattori, Toshio [Department of Emerging Infectious Diseases, Division of Internal Medicine, Graduate School of Medicine, Tohoku University, Sendai 9808574 (Japan); Ling, Hong, E-mail: [Department of Microbiology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Department of Parasitology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Key Lab of Heilongjiang Province for Infection and Immunity, Key Lab of Heilongjiang Province Education Bureau for Etiology, Harbin, Heilongjiang 150081 (China); Zhang, Feng-Min, E-mail: [Department of Microbiology, Harbin Medical University, 194 Xuefu Road, Harbin, Heilongjiang 150081 (China); Key Lab of Heilongjiang Province for Infection and Immunity, Key Lab of Heilongjiang Province Education Bureau for Etiology, Harbin, Heilongjiang 150081 (China)


    To evaluate the interaction between HIV-1 envelope glycoprotein (Env) and target cell receptors, various cell-cell-fusion assays have been developed. In the present study, we established a novel fusion system. In this system, the expression of the sensitive reporter gene, firefly luciferase (FL) gene, in the target cells was used to evaluate cell fusion event. Simultaneously, constitutively expressed Renilla luciferase (RL) gene was used to monitor effector cell number and viability. FL gave a wider dynamic range than other known reporters and the introduction of RL made the assay accurate and reproducible. This system is especially beneficial for investigation of potential entry-influencing agents, for its power of ruling out the false inhibition or enhancement caused by the artificial cell-number variation. As a case study, we applied this fusion system to observe the effect of a serine protease, thrombin, on HIV Env-mediated cell-cell fusion and have found the fusion enhancement activity of thrombin over two R5-tropic HIV strains.

  19. Magnetic fusion technology

    CERN Document Server

    Dolan, Thomas J


    Magnetic Fusion Technology describes the technologies that are required for successful development of nuclear fusion power plants using strong magnetic fields. These technologies include: ? magnet systems, ? plasma heating systems, ? control systems, ? energy conversion systems, ? advanced materials development, ? vacuum systems, ? cryogenic systems, ? plasma diagnostics, ? safety systems, and ? power plant design studies. Magnetic Fusion Technology will be useful to students and to specialists working in energy research.

  20. Fusion research principles

    CERN Document Server

    Dolan, Thomas James


    Fusion Research, Volume I: Principles provides a general description of the methods and problems of fusion research. The book contains three main parts: Principles, Experiments, and Technology. The Principles part describes the conditions necessary for a fusion reaction, as well as the fundamentals of plasma confinement, heating, and diagnostics. The Experiments part details about forty plasma confinement schemes and experiments. The last part explores various engineering problems associated with reactor design, vacuum and magnet systems, materials, plasma purity, fueling, blankets, neutronics

  1. Activation of tyrosine kinase c-Abl contributes to α-synuclein–induced neurodegeneration (United States)

    Lee, Su Hyun; Kim, Donghoon; Karuppagounder, Senthilkumar S.; Kumar, Manoj; Mao, Xiaobo; Shin, Joo Ho; Lee, Yunjong; Pletnikova, Olga; Troncoso, Juan C.; Dawson, Valina L.; Dawson, Ted M.; Ko, Han Seok


    Aggregation of α-synuclein contributes to the formation of Lewy bodies and neurites, the pathologic hallmarks of Parkinson disease (PD) and α-synucleinopathies. Although a number of human mutations have been identified in familial PD, the mechanisms that promote α-synuclein accumulation and toxicity are poorly understood. Here, we report that hyperactivity of the nonreceptor tyrosine kinase c-Abl critically regulates α-synuclein–induced neuropathology. In mice expressing a human α-synucleinopathy–associated mutation (hA53Tα-syn mice), deletion of the gene encoding c-Abl reduced α-synuclein aggregation, neuropathology, and neurobehavioral deficits. Conversely, overexpression of constitutively active c-Abl in hA53Tα-syn mice accelerated α-synuclein aggregation, neuropathology, and neurobehavioral deficits. Moreover, c-Abl activation led to an age-dependent increase in phosphotyrosine 39 α-synuclein. In human postmortem samples, there was an accumulation of phosphotyrosine 39 α-synuclein in brain tissues and Lewy bodies of PD patients compared with age-matched controls. Furthermore, in vitro studies show that c-Abl phosphorylation of α-synuclein at tyrosine 39 enhances α-synuclein aggregation. Taken together, this work establishes a critical role for c-Abl in α-synuclein–induced neurodegeneration and demonstrates that selective inhibition of c-Abl may be neuroprotective. This study further indicates that phosphotyrosine 39 α-synuclein is a potential disease indicator for PD and related α-synucleinopathies. PMID:27348587

  2. Scaphoid excision with four-corner fusion. (United States)

    Enna, Matthew; Hoepfner, Peter; Weiss, Arnold-Peter C


    The scaphoid plays a critical role in maintain-ing normal carpal kinematics. SLAC and SNAC wrist arthritis demonstrate the ramifications ofscaphoid pathology on wrist biomechanics. In the past, symptomatic SLAC or SNAC pathology spelled total wrist arthrodesis. Over the past 20 years there has been a movement toward limited wrist arthrodesis in the treatment of SLAC/SNAC wrists. In the long-term follow-up of four-corner fusions, patient satisfaction is high, patients are able to return to their previous vocation, and wrist function averages 60%-70% of the contralateral wrist. The Spider plate is a recent advancement in the four-corner fusion armamentarium that has thus far shown great promise in respect to fusion rates (100% in the first documented series [36]),functional range of motion, intercarpal stability[37], and patient satisfaction.

  3. Magnetic fusion reactor economics

    Energy Technology Data Exchange (ETDEWEB)

    Krakowski, R.A.


    An almost primordial trend in the conversion and use of energy is an increased complexity and cost of conversion systems designed to utilize cheaper and more-abundant fuels; this trend is exemplified by the progression fossil fission {yields} fusion. The present projections of the latter indicate that capital costs of the fusion ``burner`` far exceed any commensurate savings associated with the cheapest and most-abundant of fuels. These projections suggest competitive fusion power only if internal costs associate with the use of fossil or fission fuels emerge to make them either uneconomic, unacceptable, or both with respect to expensive fusion systems. This ``implementation-by-default`` plan for fusion is re-examined by identifying in general terms fusion power-plant embodiments that might compete favorably under conditions where internal costs (both economic and environmental) of fossil and/or fission are not as great as is needed to justify the contemporary vision for fusion power. Competitive fusion power in this context will require a significant broadening of an overly focused program to explore the physics and simbiotic technologies leading to more compact, simplified, and efficient plasma-confinement configurations that reside at the heart of an attractive fusion power plant.

  4. Magnetic-confinement fusion (United States)

    Ongena, J.; Koch, R.; Wolf, R.; Zohm, H.


    Our modern society requires environmentally friendly solutions for energy production. Energy can be released not only from the fission of heavy nuclei but also from the fusion of light nuclei. Nuclear fusion is an important option for a clean and safe solution for our long-term energy needs. The extremely high temperatures required for the fusion reaction are routinely realized in several magnetic-fusion machines. Since the early 1990s, up to 16 MW of fusion power has been released in pulses of a few seconds, corresponding to a power multiplication close to break-even. Our understanding of the very complex behaviour of a magnetized plasma at temperatures between 150 and 200 million °C surrounded by cold walls has also advanced substantially. This steady progress has resulted in the construction of ITER, a fusion device with a planned fusion power output of 500 MW in pulses of 400 s. ITER should provide answers to remaining important questions on the integration of physics and technology, through a full-size demonstration of a tenfold power multiplication, and on nuclear safety aspects. Here we review the basic physics underlying magnetic fusion: past achievements, present efforts and the prospects for future production of electrical energy. We also discuss questions related to the safety, waste management and decommissioning of a future fusion power plant.

  5. Frontiers in fusion research

    CERN Document Server

    Kikuchi, Mitsuru


    Frontiers in Fusion Research provides a systematic overview of the latest physical principles of fusion and plasma confinement. It is primarily devoted to the principle of magnetic plasma confinement, that has been systematized through 50 years of fusion research. Frontiers in Fusion Research begins with an introduction to the study of plasma, discussing the astronomical birth of hydrogen energy and the beginnings of human attempts to harness the Sun's energy for use on Earth. It moves on to chapters that cover a variety of topics such as: * charged particle motion, * plasma kinetic theory, *

  6. Gene ranking and signaling pathway of schizophrenia susceptible genes based on data fusion arithmetic%基于数据融合算法的精神分裂症易感基因排序及其信号通路研究

    Institute of Scientific and Technical Information of China (English)

    杨铠冰; 冀燃; 王美琴; 张敏; 张大保


    目的:利用已有的研究结果,采用数据融合分析方法,对与已知的精神分裂症易感基因有密切关系的候选基因进行统计分析,得出基因排序及其信号通路的结果,以此证明数据融合算法能够有效筛选精神分裂症易感基因。方法根据已知的与精神分裂症相关的基因,利用 Endeavour工具,通过数据融合算法将候选基因排序,并用 DAVID 数据库对基因排序结果进行功能注释和富集分析。结果利用数据融合算法获得的排序较高的基因与已有研究进行比较,证实排名第二的 NRG3和排名第三的AKT1与精神分裂症有密切联系。其他排名较高的基因可以作为潜在的疾病易感基因,为精神分裂症的研究提供参考。结论数据融合算法能够准确评价候选基因与精神分裂症的联系,使用该方法能有效地对精神分裂症易感基因进行筛选和判定。%Objective Based on previous studies,this paper applies data fusion arithmetic to extract the potential susceptible genes and signaling pathway related to schizophrenia, which proves that data fusion algorithm is an effective method to filter schizophrenia susceptible genes. Methods By making use of Endeavour tool which is based on data fusion method,the candidate genes were ranked according to their similarity with the tralning genes. Then DAVID database was used to implement the functional annotation and enrichment analysis for the extracted genes. Results Compared with previous studies,the second NRG3 and third AKT1 which are obtalned from higher ranking candidate genes,are closely related to schizophrenia. Other higher ranking genes could be used as potential disease-susceptibility genes to provide reference for the research of schizophrenia. Conclusions Data fusion arithmetic can help to detect the potential schizophrenia candidate genes objectively and correctly,and this method is a powerful tool for identifying schizophrenia

  7. Incidence and clinical relevance of TEL/AML1 fusion genes in children with acute lymphoblastic leukemia enrolled in the German and Italian multicenter therapy trials. Associazione Italiana Ematologia Oncologia Pediatrica and the Berlin-Frankfurt-Münster Study Group. (United States)

    Borkhardt, A; Cazzaniga, G; Viehmann, S; Valsecchi, M G; Ludwig, W D; Burci, L; Mangioni, S; Schrappe, M; Riehm, H; Lampert, F; Basso, G; Masera, G; Harbott, J; Biondi, A


    The molecular approach for the analysis of leukemia associated chromosomal translocations has led to the identification of prognostic relevant subgroups. In pediatric acute lymphoblastic leukemia (ALL), the most common translocations, t(9;22) and t(4;11), have been associated with a poorer clinical outcome. Recently the TEL gene at chromosome 12p13 and the AML1 gene at chromosome 21q22 were found to be involved in the translocation t(12;21)(p13;q22). By conventional cytogenetics, however, this chromosomal abnormality is barely detectable and occurs in less than 0.05% of childhood ALL. To investigate the frequency of the molecular equivalent of the t(12;21), the TEL/AML1 gene fusion, we have undertaken a prospective screening in the running German Berlin-Frankfurt-Münster (BFM) and Italian Associazione Italiana Ematologia Oncologia Pediatrica (AIEOP) multicenter ALL therapy trials. We have analyzed 334 unselected cases of pediatric ALL patients consecutively referred over a period of 5 and 9 months, respectively. The overall incidence of the t(12;21) in pediatric ALL is 18.9%. The 63 cases positive for the TEL/AML1 chimeric products ranged in age between 1 and 12 years, and all but one showed CD10 and pre-B immunophenotype. Interestingly, one case displayed a pre-pre-B immunophenotype. Among the B-lineage subgroup, the t(12;21) occurs in 22.0% of the cases. Fifteen of 61 (24.6%) cases coexpressed at least two myeloid antigens (CD13, CD33, or CDw65) in more than 20% of the gated blast cells. DNA index was available for 59 of the 63 TEL/AML1 positive cases; a hyperdiploid DNA content (> or = 1.16) was detected in only four patients, being nonhyperdiploid in the remaining 55. Based on this prospective analysis, we retrospectively evaluated the impact of TEL/AML1 in prognosis by identifying the subset of B-lineage ALL children enrolled in the closed German ALL-BFM-90 and Italian ALL-AIEOP-91 protocols who had sufficient material for analysis. A total of 342 children

  8. 融合基因VH-mms13的构建及其蛋白表达鉴定%Construction of fusion gene VH-mms13 and identification of its protein

    Institute of Scientific and Technical Information of China (English)

    杨柳青; 孔登; 王雪耘; 王晓红; 孟丽; 王小柯


    Objective To construct the prokaryotic expression vector pET30a( +)-VH-mms13 and identification of its protein after induced with IPTG.Method Heavy chain variable region VH gene of typeⅣcollagenase monoclonal antibody and magnetosome membrane protein gene mms13 were amplified separately,the fusion gene VH-linker-mms13 were synthesized by SOE-PCR technique and inserted into pET30a ( +) plasmid, which was confirmed by restriction enzyme digest and sequencing.Then the recombinant plasmid pET30a ( +)-VH-mms13 was transform into E.coli DE3 and induced with 0.4 mmol/L IPTG.The fused protein was identified by SDS-PAGE and Western blot.Results The length of fusion gene VH-mms13 was 738 bp,and the sequence was correct.After induced with IPTG,the fused protein was found in the inclusion body and Western blot results suggested that the fused protein can bind with His-tag antibody specifically.Conclusion Expression vector pET30a ( +)-VH-mms13 is successfully constructed and the fusion protein has good immunogenicity,which lay the foundation for the development of biomagnetism-targeted drug.%目的:构建原核表达载体pET30a(+)-VH-mms13,诱导表达后鉴定融合蛋白表达。方法分别扩增单克隆抗体基因的重链可变区VH基因和细菌磁小体膜蛋白基因mms13基因,采用重叠延伸PCR技术(splicing by overlap extension ,SOE-PCR)构建融合基因VH-linker-mms13,并将融合基因插入pET30a(+)载体,酶切、测序验证;将重组质粒导入大肠杆菌DE3中,0.4 mmol/L异丙基疏代半乳糖苷( Isopropyl β-D-thiogalactoside ,IPTG)诱导表达,产物经SDS-PAGE电泳和Western blot 双重鉴定。结果 PCR鉴定构建的融合基因VH-mms13大小为738 bp,与理论值相符,测序结果表明序列无误;转入DE3经IPTG诱导,在包含体中检测到融合蛋白表达;Western blot结果显示该表达蛋白可与His-tag抗体特异性结合,蛋白大小符合融合

  9. Cell fusion and nuclear fusion in plants. (United States)

    Maruyama, Daisuke; Ohtsu, Mina; Higashiyama, Tetsuya


    Eukaryotic cells are surrounded by a plasma membrane and have a large nucleus containing the genomic DNA, which is enclosed by a nuclear envelope consisting of the outer and inner nuclear membranes. Although these membranes maintain the identity of cells, they sometimes fuse to each other, such as to produce a zygote during sexual reproduction or to give rise to other characteristically polyploid tissues. Recent studies have demonstrated that the mechanisms of plasma membrane or nuclear membrane fusion in plants are shared to some extent with those of yeasts and animals, despite the unique features of plant cells including thick cell walls and intercellular connections. Here, we summarize the key factors in the fusion of these membranes during plant reproduction, and also focus on "non-gametic cell fusion," which was thought to be rare in plant tissue, in which each cell is separated by a cell wall.

  10. Study on Agrobacterium tumefaciens-mediated Transformation of Brassica campestris L. with Fusion Gene Ycoil-bFGF%农杆菌介导油菜油体基因与碱性成纤维细胞生长因子融合基因转化油菜研究

    Institute of Scientific and Technical Information of China (English)

    徐岩; 肖艳双; 杜金霞; 汪洪; 郑伟; 李营; 庞实锋


    [Objective] The study is to generate pharmaceutical protein via plant transgenic technique. [Method] Using the cotyledons with petiole as transformation receptor, the fusion gene of rapeseed oil-body gene and bFGF was introduced into the rapeseed (Brassica campestris L.) by Agrobacterium tumefaciens-mediated transformation; meanwhile regeneration conditions of rapeseed were also optimized, and the regenerated resistant plantlets were detected by PCR and Southern blot. [Result] This fusion gene had been integrated into rapeseed genome successfully, and the optimized conditions of transformation and regeneration were as follows: explants pre-culture for 2 d, co-culture for 3 d, bacteria solution OD600 for 0.3 and infection time for 5 min. [Conclusion] The results laid a solid foundation for extraction, isolation and purification of protein in transgenic plant seeds.

  11. Nuclear fusion inside condense matters

    Institute of Scientific and Technical Information of China (English)

    HE Jing-tang


    This article describes in detail the nuclear fusion inside condense matters--the Fleischmann-Pons effect, the reproducibility of cold fusions, self-consistentcy of cold fusions and the possible applications.

  12. The CRISPR/Cas9 system efficiently reverts the tumorigenic ability of BCR/ABL in vitro and in a xenograft model of chronic myeloid leukemia. (United States)

    García-Tuñón, Ignacio; Hernández-Sánchez, María; Ordoñez, José Luis; Alonso-Pérez, Veronica; Álamo-Quijada, Miguel; Benito, Rocio; Guerrero, Carmen; Hernández-Rivas, Jesús María; Sánchez-Martín, Manuel


    CRISPR/Cas9 technology was used to abrogate p210 oncoprotein expression in the Boff-p210 cell line, a pro-B line derived from interlukin-3-dependent Baf/3, that shows IL-3-independence arising from the constitutive expression of BCR-ABL p210. Using this approach, pools of Boff-p210-edited cells and single edited cell-derived clones were obtained and functionally studied in vitro. The loss of p210 expression in Boff-p210 cells resulted in the loss of ability to grow in the absence of IL-3, as the Baf/3 parental line, showing significantly increased apoptosis levels. Notably, in a single edited cell-derived clone carrying a frame-shift mutation that prevents p210 oncoprotein expression, the effects were even more drastic, resulting in cell death. These edited cells were injected subcutaneously in immunosuppressed mice and tumor growth was followed for three weeks. BCR/ABL-edited cells developed smaller tumors than those originating from unedited Boff-p210 parental cells. Interestingly, the single edited cell-derived clone was unable to develop tumors, similar to what is observed with the parental Baf/3 cell line.CRISPR/Cas9 genomic editing technology allows the ablation of the BCR/ABL fusion gene, causing an absence of oncoprotein expression, and blocking its tumorigenic effects in vitro and in the in vivo xenograft model of CML. The future application of this approach in in vivo models of CML will allow us to more accurately assess the value of CRISPR/Cas9 technology as a new therapeutic tool that overcomes resistance to the usual treatments for CML patients.

  13. Complementary Advanced Fusion Exploration (United States)


    homographic computer vision image fusion, out-of-sequence measurement and track data handling, Nash bargaining approaches to sensor management... homographic fusion notions are identified together with the Nash approach, the pursuit-evasion approach to threat situation outcome determination, and the

  14. Controlled Nuclear Fusion. (United States)

    Glasstone, Samuel

    This publication is one of a series of information booklets for the general public published by The United States Atomic Energy Commission. Among the topics discussed are: Importance of Fusion Energy; Conditions for Nuclear Fusion; Thermonuclear Reactions in Plasmas; Plasma Confinement by Magnetic Fields; Experiments With Plasmas; High-Temperature…

  15. Fusion helps diversification

    NARCIS (Netherlands)

    S. Liang; Z. Ren; M. de Rijke


    A popular strategy for search result diversification is to first retrieve a set of documents utilizing a standard retrieval method and then rerank the results. We adopt a different perspective on the problem, based on data fusion. Starting from the hypothesis that data fusion can improve performance

  16. Fusion of biological membranes

    Indian Academy of Sciences (India)

    K Katsov; M Müller; M Schick


    The process of membrane fusion has been examined by Monte Carlo simulation, and is found to be very different than the conventional picture. The differences in mechanism lead to several predictions, in particular that fusion is accompanied by transient leakage. This prediction has recently been verified. Self-consistent field theory is applied to examine the free energy barriers in the different scenarios.

  17. Controlled thermonuclear fusion

    CERN Document Server

    Bobin, Jean Louis


    The book is a presentation of the basic principles and main achievements in the field of nuclear fusion. It encompasses both magnetic and inertial confinements plus a few exotic mechanisms for nuclear fusion. The state-of-the-art regarding thermonuclear reactions, hot plasmas, tokamaks, laser-driven compression and future reactors is given.

  18. Changes in Parthenogenetic Imprinting Patterns during Reprogramming by Cell Fusion.

    Directory of Open Access Journals (Sweden)

    Hyun Sik Jang

    Full Text Available Differentiated somatic cells can be reprogrammed into the pluripotent state by cell-cell fusion. In the pluripotent state, reprogrammed cells may then self-renew and differentiate into all three germ layers. Fusion-induced reprogramming also epigenetically modifies the somatic cell genome through DNA demethylation, X chromosome reactivation, and histone modification. In this study, we investigated whether fusion with embryonic stem cells (ESCs also reprograms genomic imprinting patterns in somatic cells. In particular, we examined imprinting changes in parthenogenetic neural stem cells fused with biparental ESCs, as well as in biparental neural stem cells fused with parthenogenetic ESCs. The resulting hybrid cells expressed the pluripotency markers Oct4 and Nanog. In addition, methylation of several imprinted genes except Peg3 was comparable between hybrid cells and ESCs. This finding indicates that reprogramming by cell fusion does not necessarily reverse the status of all imprinted genes to the state of pluripotent fusion partner.

  19. Cloning and fusion protein expression of the S100A4 gene in sika deer%梅花鹿S100A4基因的克隆及融合蛋白的表达

    Institute of Scientific and Technical Information of China (English)

    王大涛; 赵海平; 褚文辉; 杨福合; 邢秀梅; 王桂武; 李春义


    为了研究梅花鹿S100A4(S100 calcium binding protein A4)基因在鹿茸生长过程中的作用.用RT-PCR法从生茸骨膜细胞总RNA中克隆了梅花鹿S100A4基因,在NCBI中对基因序列进行比对;将完整的基因序列与逆转录病毒表达载体pLEGFP-Ci重组,获得了重组质粒pLEGFP-S100;用脂质体法将pLEGFP-S100与pVSV-G(被膜载体)共转染包装细胞GP2-293,获得重组病毒上清液,感染角柄骨膜细胞后逆转录病毒携带的基因进入宿主细胞.结果显示:S100A4基因是一个相对保守的基因,与多个物种的匹配度达到90%;重组逆转录病毒载体pLEGFP-S100可以形成重组逆转录病毒粒子,将S100A4基因导入靶细胞,并表达S100A4与GFP(Green fluorescent protein)的融合蛋白.%In order to better understand the function of S100 calcium binding protein A4 in antler development of sika deer ( Cervus nippon), we cloned S100A4 genes from total RNA of cultured antlerogenic periosteum cells using reverse transcription polymerase chain reaction ( RT-PCR), S100A4 gene sequences were compared with closely related animal species in NCBI. Full lengths of S100A4 genes were inserted into a vector plasmid pLEGFP-C1 ( retroviral express vector). The recombinant plasmid pLEGFP-S100 and pVSV-G (envelope vector) were co-transfected into GP2 -293 cells (packaging cell line ) using lipofectimin 2000, and the resultant viral supernatants were collected. The cultured pedicle periosteum cells were then infected with virus in the supernatants. Results showed that S100A4 gene was a relatively conserved gene, and had about 90% homology with several species. Recombinant retroviral vector pLEGFP-S100 could effectively deliver a gene into target cell line, and express a fusion GFP ( green fluorescent protein ) protein.

  20. Fusion of a viral antigen to invariant chain leads to augmented T-cell immunity and improved protection in gene-gun DNA-vaccinated mice

    DEFF Research Database (Denmark)

    Grujic, Mirjana; Holst, Peter J; Christensen, Jan P


    against lethal peripheral challenge. The current study questioned whether the same strategy, i.e. linkage of GP to an Ii chain, could be applied to a naked DNA vaccine. Following gene-gun immunization with the linked construct (DNA-IiGP), GP-specific CD4(+) T cells could not be detected by flow cytometry...

  1. Compact fusion reactors

    CERN Document Server

    CERN. Geneva


    Fusion research is currently to a large extent focused on tokamak (ITER) and inertial confinement (NIF) research. In addition to these large international or national efforts there are private companies performing fusion research using much smaller devices than ITER or NIF. The attempt to achieve fusion energy production through relatively small and compact devices compared to tokamaks decreases the costs and building time of the reactors and this has allowed some private companies to enter the field, like EMC2, General Fusion, Helion Energy, Lawrenceville Plasma Physics and Lockheed Martin. Some of these companies are trying to demonstrate net energy production within the next few years. If they are successful their next step is to attempt to commercialize their technology. In this presentation an overview of compact fusion reactor concepts is given.

  2. Fusion transcriptome profiling provides insights into alveolar rhabdomyosarcoma. (United States)

    Xie, Zhongqiu; Babiceanu, Mihaela; Kumar, Shailesh; Jia, Yuemeng; Qin, Fujun; Barr, Frederic G; Li, Hui


    Gene fusions and fusion products were thought to be unique features of neoplasia. However, more and more studies have identified fusion RNAs in normal physiology. Through RNA sequencing of 27 human noncancer tissues, a large number of fusion RNAs were found. By analyzing fusion transcriptome, we observed close clusterings between samples of same or similar tissues, supporting the feasibility of using fusion RNA profiling to reveal connections between biological samples. To put the concept into use, we selected alveolar rhabdomyosarcoma (ARMS), a myogenic pediatric cancer whose exact cell of origin is not clear. PAX3-FOXO1 (paired box gene 3 fused with forkhead box O1) fusion RNA, which is considered a hallmark of ARMS, was recently found during normal muscle cell differentiation. We performed and analyzed RNA sequencing from various time points during myogenesis and uncovered many chimeric fusion RNAs. Interestingly, we found that the fusion RNA profile of RH30, an ARMS cell line, is most similar to the myogenesis time point when PAX3-FOXO1 is expressed. In contrast, full transcriptome clustering analysis failed to uncover this connection. Strikingly, all of the 18 chimeric RNAs in RH30 cells could be detected at the same myogenic time point(s). In addition, the seven chimeric RNAs that follow the exact transient expression pattern as PAX3-FOXO1 are specific to rhabdomyosarcoma cells. Further testing with clinical samples also confirmed their specificity to rhabdomyosarcoma. These results provide further support for the link between at least some ARMSs and the PAX3-FOXO1-expressing myogenic cells and demonstrate that fusion RNA profiling can be used to investigate the etiology of fusion-gene-associated cancers.

  3. ALK protein expression and gene fusion in bronchoscopic specimens of lung adenocarcinoma%检测肺腺癌活检标本间变性淋巴瘤激酶蛋白表达和基因融合

    Institute of Scientific and Technical Information of China (English)

    梁小龙; 王孟昭; 张静; 罗玉凤; 张淑英; 武莎斐; 刘媛媛; 曾瑄


    Objective To explore ALK protein expression and gene fusion in formalin-fixed and paraffin-embedded (FFPE) specimens obtained from lung cancer by bronchoscopy,and to investigate the relationship between ALK status and clinicopathological characteristics of the patients.Methods Seventyfour FFPE samples obtained from lung adenocarcinoma by bronchoscopy were tested for ALK protein expression and gene fusion respectively by immunohistochemistry (IHC) using Ventana D5F3 antibody and fluorescence in situ hybridization (FISH) using ALK break apart probe.Results sixty-five of the 74 samples were successfully tested by FISH (87.8%,65/74).There were 5 FISH-positive cases (7.7%,5/65),all with advanced stage carcinoma.Among these five FISH-positive cases,3 were IHC-positive (4.1%,3/74) and 2 IHC-negative cases.All the other 69 samples were IHC-negative,including nine FISH-uninformative samples (7 samples were less than 50 tumor cells and 2 samples with weak FISH signal).Both ALK IHC and FISH results were not correlated with age,sex,history of smoking,histological classification,differentiation and lymph node metastasis.Conclusions Bronchoscopic specimens of lung cancer can be used to detect ALK expression and gene fusion.Inmunohistochemistry in combination with FISH test may be more favorable for ALK test.%目的 探讨采用肺腺癌支气管镜活检标本检测间变性淋巴瘤激酶(ALK)蛋白表达和基因融合的可行性,及其与临床病理特征的关系.方法 74例福尔马林固定石蜡包埋的肺腺癌支气管镜活检标本,采用Bench Mark全自动免疫组化染色机和D5F3抗体试剂盒,以免疫组化法检测ALK蛋白的表达,采用Abbott ALK分离探针,以荧光原位杂交(FISH)法检测ALK基因融合.结果 74例标本中,65例成功地进行了FISH检测,成功检测率为87.8% (65/74);FISH阳性5例,阳性率为7.7%(5/65),均为中晚期低分化腺癌;其中免疫组化阳性3例,阳性率为4.1% (3/74);另2

  4. Transformation of an Unclassified Myeloproliferative Neoplasm with a Rare BCR-JAK2 Fusion Transcript Resulting from the Translocation (9;22)(p24;q11) (United States)

    Chamseddine, A. N.; Etancelin, P.; Penther, D.; Parmentier, F.; Kuadjovi, C.; Camus, V.; Contentin, N.; Lenain, P.; Bastard, C.; Tilly, H.; Jardin, F.


    BCR-ABL1 negative myeloproliferative neoplasms (MPNs) are known to contain alterations of the tyrosine kinase JAK2 (located on 9p24) that result in constitutive activation of the encoded protein. JAK2 fusions are reported in acute and chronic leukemias of myeloid and lymphoid phenotypes. Here, we report an unclassified case of MPN (MPN-U) showing a t(9;22)(p24;q11), which generates a BCR-JAK2 fusion gene by fusing the BCR at intron 13 to JAK2 at intron 17 on the derivative chromosome 22. Most reported JAK2 fusions cases reveal an aggressive clinical course and long-term remissions have only been achieved after allogeneic stem cell transplantation (ASCT). To the best of our knowledge, this is the thirteenth case reported worldwide to describe a BCR-JAK2 fusion transcript in MPN-U. The present report revealed a sustained complete clinical, hematologic, and cytogenetic remission 35 months after diagnosis and ~24 months after ASCT. Regarding BCR-ABL1  negative MPN patients this case report provides strong support for a role of JAK2 activation in the oncogenesis and suggests a possible diagnostic and therapeutic target that should be investigated. PMID:25789185

  5. Transformation of an Unclassified Myeloproliferative Neoplasm with a Rare BCR-JAK2 Fusion Transcript Resulting from the Translocation (9;22(p24;q11

    Directory of Open Access Journals (Sweden)

    A. N. Chamseddine


    Full Text Available BCR-ABL1 negative myeloproliferative neoplasms (MPNs are known to contain alterations of the tyrosine kinase JAK2 (located on 9p24 that result in constitutive activation of the encoded protein. JAK2 fusions are reported in acute and chronic leukemias of myeloid and lymphoid phenotypes. Here, we report an unclassified case of MPN (MPN-U showing a t(9;22(p24;q11, which generates a BCR-JAK2 fusion gene by fusing the BCR at intron 13 to JAK2 at intron 17 on the derivative chromosome 22. Most reported JAK2 fusions cases reveal an aggressive clinical course and long-term remissions have only been achieved after allogeneic stem cell transplantation (ASCT. To the best of our knowledge, this is the thirteenth case reported worldwide to describe a BCR-JAK2 fusion transcript in MPN-U. The present report revealed a sustained complete clinical, hematologic, and cytogenetic remission 35 months after diagnosis and ~24 months after ASCT. Regarding BCR-ABL1  negative MPN patients this case report provides strong support for a role of JAK2 activation in the oncogenesis and suggests a possible diagnostic and therapeutic target that should be investigated.

  6. DNA fusion-gene vaccination in patients with prostate cancer induces high-frequency CD8(+) T-cell responses and increases PSA doubling time. (United States)

    Chudley, Lindsey; McCann, Katy; Mander, Ann; Tjelle, Torunn; Campos-Perez, Juan; Godeseth, Rosemary; Creak, Antonia; Dobbyn, James; Johnson, Bernadette; Bass, Paul; Heath, Catherine; Kerr, Paul; Mathiesen, Iacob; Dearnaley, David; Stevenson, Freda; Ottensmeier, Christian


    We report on the immunogenicity and clinical effects in a phase I/II dose escalation trial of a DNA fusion vaccine in patients with prostate cancer. The vaccine encodes a domain (DOM) from fragment C of tetanus toxin linked to an HLA-A2-binding epitope from prostate-specific membrane antigen (PSMA), PSMA(27-35). We evaluated the effect of intramuscular vaccination without or with electroporation (EP) on vaccine potency. Thirty-two HLA-A2(+) patients were vaccinated and monitored for immune and clinical responses for a follow-up period of 72 weeks. At week 24, cross-over to the immunologically more effective delivery modality was permitted; this was shown to be with EP based on early antibody data, and subsequently, 13/15 patients crossed to the +EP arm. Thirty-two HLA-A2(-) control patients were assessed for time to next treatment and overall survival. Vaccination was safe and well tolerated. The vaccine induced DOM-specific CD4(+) and PSMA(27)-specific CD8(+) T cells, which were detectable at significant levels above baseline at the end of the study (p = 0.0223 and p = 0.00248, respectively). Of 30 patients, 29 had a measurable CD4(+) T-cell response and PSMA(27)-specific CD8(+) T cells were detected in 16/30 patients, with or without EP. At week 24, before cross-over, both delivery methods led to increased CD4(+) and CD8(+) vaccine-specific T cells with a trend to a greater effect with EP. PSA doubling time increased significantly from 11.97 months pre-treatment to 16.82 months over the 72-week follow-up (p = 0.0417), with no clear differential effect of EP. The high frequency of immunological responses to DOM-PSMA(27) vaccination and the clinical effects are sufficiently promising to warrant further, randomized testing.

  7. Colorimetric assessment of BCR-ABL1 transcripts in clinical samples via gold nanoprobes. (United States)

    Vinhas, Raquel; Correia, Cláudia; Ribeiro, Patricia; Lourenço, Alexandra; Botelho de Sousa, Aida; Fernandes, Alexandra R; Baptista, Pedro V


    Gold nanoparticles functionalized with thiolated oligonucleotides (Au-nanoprobes) have been used in a range of applications for the detection of bioanalytes of interest, from ions to proteins and DNA targets. These detection strategies are based on the unique optical properties of gold nanoparticles, in particular, the intense color that is subject to modulation by modification of the medium dieletric. Au-nanoprobes have been applied for the detection and characterization of specific DNA sequences of interest, namely pathogens and disease biomarkers. Nevertheless, despite its relevance, only a few reports exist on the detection of RNA targets. Among these strategies, the colorimetric detection of DNA has been proven to work for several different targets in controlled samples but demonstration in real clinical bioanalysis has been elusive. Here, we used a colorimetric method based on Au-nanoprobes for the direct detection of the e14a2 BCR-ABL fusion transcript in myeloid leukemia patient samples without the need for retro-transcription. Au-nanoprobes directly assessed total RNA from 38 clinical samples, and results were validated against reverse transcription-nested polymerase chain reaction (RT-nested PCR) and reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The colorimetric Au-nanoprobe assay is a simple yet reliable strategy to scrutinize myeloid leukemia patients at diagnosis and evaluate progression, with obvious advantages in terms of time and cost, particularly in low- to medium-income countries where molecular screening is not routinely feasible. Graphical abstract Gold nanoprobe for colorimetric detection of BCR-ABL1 fusion transcripts originating from the Philadelphia chromosome.

  8. Introducing the advanced burn life support (ABLS) course in Italy. (United States)

    D'Asta, F; Homsi, J; Clark, P; Buffalo, M C; Melandri, D; Carboni, A; Pinzauti, E; Graziano, A; Masellis, A; Bussolin, L; Messineo, A


    Systematic education based on internationally standardized programs is a well-established practice in Italy, especially in the emergency health care system. However, until recently, a specific program to treat burns was not available to guide emergency physicians, nurses, or volunteers acting as first responders. In 2010, two national faculty members, acting as ABA observers, and one Italian course coordinator, trained and certified in the United States, conducted a week-long training program which fully certified 10 Italian instructors. Authorized ABLS provider courses were conducted in Italy between 2010 and 2012, including one organized prior to the 20th annual meeting of the Italian Society of Burns (SIUst). In order to increase the effectiveness and diffusion of the course in Italy, changes were approved by the ABA to accommodate societal differences, including the translation of the manual into Italian. The ABA has also approved the creation and publication of a bilingual ABLS Italian website for the purpose of promoting the ABLS course in Italy. In response to high demand, a second ABLS Instructor course was organized in 2012 and has been attended by physicians and nurses from several Italian burn centers. In the following discourse the experiences of the first 15 Italian ABLS courses will be discussed.

  9. Globular adiponectin induces differentiation and fusion of skeletal muscle cells

    Institute of Scientific and Technical Information of China (English)

    Tania Fiaschi; Domenico Cirelli; Giuseppina Comito; Stefania Gelmini; Giampietro Ramponi; Maria Serio; Paola Chiarugi


    The growing interest in skeletal muscle regeneration is associated with the opening of new therapeutic strategies for muscle injury after trauma, as well as several muscular degenerative pathologies, including dystrophies, muscu-lar atrophy, and cachexia. Studies focused on the ability of extracellular factors to promote myogenesis are therefore highly promising. We now report that an adipocyte-derived factor, globular adiponectin (gAd), is able to induce mus-cle gene expression and cell differentiation, gAd, besides its well-known ability to regulate several metabolic func-tions in muscle, including glucose uptake and consumption and fatty acid catabolism, is able to block cell cycle entry of myoblasts, to induce the expression of specific skeletal muscle markers such as myosin heavy chain or eaveolin-3, as well as to provoke cell fusion into multinucleated syneytia and, finally, muscle fibre formation, gAd exerts its pro-differentiative activity through redox-dependent activation of p38, Akt and 5'-AMP-activated protein kinase path-ways. Interestingly, differentiating myoblasts are autocrine for adiponectiu, and the mimicking of pro-inflammatory settings or exposure to oxidative stress strongly increases the production of the hormone from differentiating cells. These data suggest a novel function of adiponectin, directly coordinating the myogenic differentiation program and serving an autocrine function during skeletal myogenesis.

  10. Prokaryotic Expression of the p23-IL-18 Fusion Gene of Theileria sergenti%牛瑟氏泰勒虫p23-IL-18融合基因的原核表达

    Institute of Scientific and Technical Information of China (English)

    白杨; 许应天; 吴艳丽; 于志云


    In order to prepare the genetically engineering subunit vaccine of brovine Theileria sergenti, the p23-IL-18 fusion gene was cloned into prokaryotic pET-28a expression vector in this experiment. It can construct recombinant prokaryotic expression plasmid pET-28a-p23-IL-18 which induced expression by IPTG. The expression products were examined by SDS-PAGE and Western-blotting. The results showed that the prokaryotic expression plasmid pET-28a-p23-IL-18 was built successfully and the objective gene was successfully expressed in Escherichia coli. The molecular weight of fusion protein was 48 kDa and could react with Theileria sergenti positive serum with good antigencity. The results of this research laid the foundations for genetically engineering subunit vaccine of brovine Theileria sergenti.%为探索牛瑟氏泰勒虫病基因工程亚单位疫苗的可行性,以p23-IL-18融合基因克隆到原核表达载体pET-28a,构建原核重组质粒pET-28a-p23-IL-18,用IPTG诱导表达,对表达产物进行SDS-PAGE和Western-blotting分析。结果表明,构建牛瑟氏泰勒虫pET-28a-p23-IL-18原核表达质粒,目的基因在大肠杆菌中获得表达,融合蛋白的分子量约为48 kDa,并被牛瑟氏泰勒虫阳性血清所识别,具有良好的反应原性。此结果为牛瑟氏泰勒虫病基因工程亚单位疫苗的制备提供了理论依据。

  11. Genetic variability of attachment (G and Fusion (F protein genes of human metapneumovirus strains circulating during 2006-2009 in Kolkata, Eastern India

    Directory of Open Access Journals (Sweden)

    Chawla-Sarkar Mamta


    Full Text Available Abstract Background Human metapneumovirus (hMPV is associated with the acute respiratory tract infection (ARTI in all the age groups. However, there is limited information on prevalence and genetic diversity of human metapneumovirus (hMPV strains circulating in India. Objective To study prevalence and genomic diversity of hMPV strains among ARTI patients reporting in outpatient departments of hospitals in Kolkata, Eastern India. Methods Nasal and/or throat swabs from 2309 patients during January 2006 to December 2009, were screened for the presence of hMPV by RT-PCR of nucleocapsid (N gene. The G and F genes of representative hMPV positive samples were sequenced. Results 118 of 2309 (5.11% clinical samples were positive for hMPV. The majority (≈80% of the positive cases were detected during July−November all through the study period. Genetic analysis revealed that 77% strains belong to A2 subgroup whereas rest clustered in B1 subgroup. G sequences showed higher diversity at the nucleotide and amino acid level. In contrast, less than 10% variation was observed in F gene of representative strains of all four years. Sequence analysis also revealed changes in the position of stop codon in G protein, which resulted in variable length (217-231 aa polypeptides. Conclusion The study suggests that approximately 5% of ARTI in the region were caused by hMPV. This is the first report on the genetic variability of G and F gene of hMPV strains from India which clearly shows that the G protein of hMPV is continuously evolving. Though the study partially fulfills lacunae of information, further studies from other regions are necessary for better understanding of prevalence, epidemiology and virus evolution in Indian subcontinent.

  12. USA sender Able og Baker en tur i rummet

    DEFF Research Database (Denmark)

    Alstrup, Aage Kristian Olsen



  13. Fusion Studies in Japan (United States)

    Ogawa, Yuichi


    A new strategic energy plan decided by the Japanese Cabinet in 2014 strongly supports the steady promotion of nuclear fusion development activities, including the ITER project and the Broader Approach activities from the long-term viewpoint. Atomic Energy Commission (AEC) in Japan formulated the Third Phase Basic Program so as to promote an experimental fusion reactor project. In 2005 AEC has reviewed this Program, and discussed on selection and concentration among many projects of fusion reactor development. In addition to the promotion of ITER project, advanced tokamak research by JT-60SA, helical plasma experiment by LHD, FIREX project in laser fusion research and fusion engineering by IFMIF were highly prioritized. Although the basic concept is quite different between tokamak, helical and laser fusion researches, there exist a lot of common features such as plasma physics on 3-D magnetic geometry, high power heat load on plasma facing component and so on. Therefore, a synergetic scenario on fusion reactor development among various plasma confinement concepts would be important.

  14. Non-Fusion and Fusion Expression of β-Galactosidase from Lactobacillus bulgaricus in Lactococcus lactis

    Institute of Scientific and Technical Information of China (English)



    Objective To construct four recombinant Lactococcus lactis strains exhibiting high β-galactosidase activity in fusion or non-fusion ways, and to study the influence factors for their protein expression and secretion. Methods The gene fragments encoding β-galactosidase from two strains of Lactobacillus bulgaricus, wch9901 isolated from yogurt and 1.1480 purchased from the Chinese Academy of Sciences, were amplified and inserted into lactococcal expression vector pMG36e. For fusion expression, the open reading frame of the β-galactosidase gene was amplified, while for non-fusion expression, the open reading frame of the β-galactosidase gene was amplified with its native Shine-Dalgarno sequence upstream. The start codon of the β-galactosidase gene partially overlapped with the stop codon of vector origin open reading frame. Then, the recombinant plasmids were transformed into Escherichia coli DH5α and Lactococcus lactis subsp, lactis MG1363 and confirmed by determining β-galactosidase activities. Results The non-fusion expression plasmids showed a significantly higher β-galactosidase activity in transformed strains than the fusion expression plasmids. The highest enzyme activity was observed in Lactococcus lactis transformed with the non-fusion expression plasmids which were inserted into the β-galactosidase gene from Lactobacillus bulgaricus wch9901. The β-galactosidase activity was 2.75 times as high as that of the native counterpart. In addition, β-galactosidase expressed by recombinant plasmids in Lactococcus lactis could be secreted into the culture medium. The highest secretion rate (27.1%) was observed when the culture medium contained 20 g/L of lactose. Conclusion Different properties of the native bacteria may have some effects on the protein expression of recombinant plasmids. Non-fusion expression shows a higher enzyme activity in host bacteria. There may be a ost-related weak secretion signal peptide gene within the structure gene of Lb

  15. Construction of Eukaryotic Expression Vector with mBD1-mBD3 Fusion Genes and Exploring Its Activity against Influenza A Virus

    Directory of Open Access Journals (Sweden)

    Wanyi Li


    Full Text Available Influenza (flu pandemics have exhibited a great threat to human health throughout history. With the emergence of drug-resistant strains of influenza A virus (IAV, it is necessary to look for new agents for treatment and transmission prevention of the flu. Defensins are small (2–6 kDa cationic peptides known for their broad-spectrum antimicrobial activity. Beta-defensins (β-defensins are mainly produced by barrier epithelial cells and play an important role in attacking microbe invasion by epithelium. In this study, we focused on the anti-influenza A virus activity of mouse β-defensin 1 (mBD1 and β defensin-3 (mBD3 by synthesizing their fusion peptide with standard recombinant methods. The eukaryotic expression vectors pcDNA3.1(+/mBD1-mBD3 were constructed successfully by overlap-PCR and transfected into Madin-Darby canine kidney (MDCK cells. The MDCK cells transfected by pcDNA3.1(+/mBD1-mBD3 were obtained by G418 screening, and the mBD1-mBD3 stable expression pattern was confirmed in MDCK cells by RT-PCR and immunofluorescence assay. The acquired stable transfected MDCK cells were infected with IAV (A/PR/8/34, H1N1, 0.1 MOI subsequently and the virus titers in cell culture supernatants were analyzed by TCID50 72 h later. The TCID50 titer of the experimental group was clearly lower than that of the control group (p < 0.001. Furthermore, BALB/C mice were injected with liposome-encapsulated pcDNA3.1(+/mBD1-mBD3 through muscle and then challenged with the A/PR/8/34 virus. Results showed the survival rate of 100% and lung index inhibitory rate of 32.6% in pcDNA3.1(+/mBD1-mBD3group; the TCID50 titer of lung homogenates was clearly lower than that of the control group (p < 0.001. This study demonstrates that mBD1-mBD3 expressed by the recombinant plasmid pcDNA3.1(+/mBD1-mBD3 could inhibit influenza A virus replication both in vitro and in vivo. These observations suggested that the recombinant mBD1-mBD3 might be developed into an agent for

  16. Construction of a fusion gene encoding h1a-spaO of Salmonella paratyphi A and analysis of immuno-protective effects of the recombinant protein%甲型副伤寒沙门菌 h1 a-spaO 融合基因构建及其表达产物免疫保护作用

    Institute of Scientific and Technical Information of China (English)

    金蕾; 蒋锦琴; 方佳琪; 林旭瑷; 严杰; 孙爱华


    -SpaO was examined by SDS-PAGE.The antigenicity and immunoreactivity of rH1a-SpaO protein were deter-mined by Western blot assay .The ability of rH1a-SpaO antiserum agglutinating S.paratyphi A strains was detected by micro-Widal′s test.The immunoprotective effects of rH 1a-SpaO against the lethal dose challenge of S.paratyphi A strains were analyzed in a mouse model and that were compared with those by using equal dose of individual recombinant protein H1a and SpaO (rH1a and rSpaO) as the immunogens, respectively. Results The h1a-spaO fusion gene was 100%identical with the individual h1a or spaO gene in nucleotide and amino acid sequences .The constructed prokaryotic expression system could express the recombinant pro-tein rH1a-SpaO with an advantage of high efficiency .rH1a-SpaO protein was able to react with rH 1a or rSpaO antiserum.Moreover, rH1a-SpaO antiserum also could efficiently recognize rH 1a and rSpaO as well as agglutinate Salmonella paratyphi A strains by binding with H-antigen.The immunoprotective rate (93.3%) in mice pre-immunized with 100 μg of rH1a-SpaO protein was significantly higher than that in those pre-immunized with equal dose of rH1a (60.0%) protein or rSpaO protein(53.3%) (P<0.05).Conclusion The recombinant fusion protein rH 1a-SpaO showed more stronger immunoprotective function than the individ-ual rH1a or rSpaO protein , which could be used as an effective antigen for the development of bi -valent para-typhoid A vaccine or typhoid/paratyphoid capsular polysaccharide-protein combined vaccine .

  17. Control of Fusion and Solubility in Fusion Systems

    CERN Document Server

    Craven, David A


    In this article, we consider the control of fusion in fusion systems, proving three previously known, non-trivial results in a new, largely elementary way. We then reprove a result of Aschbacher, that the product of two strongly closed subgroups is strongly closed; to do this, we consolidate the theory of quotients of fusion systems into a consistent theory. We move on considering p-soluble fusion systems, and prove that they are constrained, allowing us to effectively characterize fusion systems of p-soluble groups. This leads us to recast Thompson Factorization for Qd(p)-free fusion systems, and consider Thompson Factorization for more general fusion systems.

  18. Remote sensing image fusion

    CERN Document Server

    Alparone, Luciano; Baronti, Stefano; Garzelli, Andrea


    A synthesis of more than ten years of experience, Remote Sensing Image Fusion covers methods specifically designed for remote sensing imagery. The authors supply a comprehensive classification system and rigorous mathematical description of advanced and state-of-the-art methods for pansharpening of multispectral images, fusion of hyperspectral and panchromatic images, and fusion of data from heterogeneous sensors such as optical and synthetic aperture radar (SAR) images and integration of thermal and visible/near-infrared images. They also explore new trends of signal/image processing, such as

  19. Molecular measurement of BCR-ABL transcript variations in chronic myeloid leukemia patients in cytogenetic remission

    Directory of Open Access Journals (Sweden)

    Costa Juliana


    Full Text Available Abstract Background The monitoring of BCR-ABL transcript levels by real-time quantitative polymerase chain reaction (RT-qPCR has become important to assess minimal residual disease (MRD and standard of care in the treatment of chronic myeloid leukemia (CML. In this study, we performed a prospective, sequential analysis using RT-qPCR monitoring of BCR-ABL gene rearrangements in blood samples from 91 CML patients in chronic phase (CP who achieved complete cytogenetic remission (CCyR and major molecular remission (MMR throughout imatinib treatment. Methods The absolute level of BCR-ABL transcript from peripheral blood was serially measured every 4 to 12 weeks by RT-qPCR. Only level variations > 0.5%, according to the international scale, was considered positive. Sequential cytogenetic analysis was also performed in bone marrow samples from all patients using standard protocols. Results Based on sequential analysis of BCR-ABL transcripts, the 91 patients were divided into three categories: (A 57 (62.6% had no variation on sequential analysis; (B 30 (32.9% had a single positive variation result obtained in a single sample; and (C 4 (4.39% had variations of BCR-ABL transcripts in at least two consecutive samples. Of the 34 patients who had elevated levels of transcripts (group B and C, 19 (55.8% had a BCR-ABL/BCR ratio, 13 (38.2% patients had a 1% to 10% increase and 2 patients had a >10% increase of RT-qPCR. The last two patients had lost a CCyR, and none of them showed mutations in the ABL gene. Transient cytogenetic alterations in Ph-negative cells were observed in five (5.5% patients, and none of whom lost CCyR. Conclusions Despite an increase levels of BCR-ABL/BCR ratio variations by RT-qPCR, the majority of CML patients with MMR remained in CCyR. Thus, such single variations should neither be considered predictive of subsequent failure and nor an indication for altering imatinib dose or switching to second generation therapy. Changing of

  20. Structure of the β-Galactosidase Gene from Thermus sp. Strain T2: Expression in Escherichia coli and Purification in a Single Step of an Active Fusion Protein (United States)

    Vian, Alejandro; Carrascosa, Alfonso V.; García, José L.; Cortés, Estrella


    The nucleotide sequence of both the bgaA gene, coding for a thermostable β-galactosidase of Thermus sp. strain T2, and its flanking regions was determined. The deduced amino acid sequence of the enzyme predicts a polypeptide of 645 amino acids (Mr, 73,595). Comparative analysis of the open reading frames located in the flanking regions of the bgaA gene revealed that they might encode proteins involved in the transport and hydrolysis of sugars. The observed homology between the deduced amino acid sequences of BgaA and the β-galactosidase of Bacillus stearothermophilus allows us to classify the new enzyme within family 42 of glycosyl hydrolases. BgaA was overexpressed in its active form in Escherichia coli, but more interestingly, an active chimeric β-galactosidase was constructed by fusing the BgaA protein to the choline-binding domain of the major pneumococcal autolysin. This chimera illustrates a novel approach for producing an active and thermostable hybrid enzyme that can be purified in a single step by affinity chromatography on DEAE-cellulose, retaining the catalytic properties of the native enzyme. The chimeric enzyme showed a specific activity of 191,000 U/mg at 70°C and a Km value of 1.6 mM with o-nitrophenyl-β-d-galactopyranoside as a substrate, and it retained 50% of its initial activity after 1 h of incubation at 70°C. PMID:9603833

  1. Induction of specific humoral and cellular immune responses in a mouse model following gene fusion of HSP70C and Hantaan virus Gn and S0.7 in an adenoviral vector.

    Directory of Open Access Journals (Sweden)

    Linfeng Cheng

    Full Text Available Heat shock proteins (HSPs display adjuvant functions when given as fusion proteins to enhance vaccination efficiency. To evaluate enhanced potency of Hantaan virus (HTNV glycoprotein (GP and nucleocapsid protein (NP immunogenicity by heat shock protein 70 (HSP70, a recombinant adenovirus rAd-GnS0.7-pCAG-HSP70C expression vector was developed by genetically linking the HSP70 C-terminal gene (HSP70 359-610 aa, HSP70C to the Gn and 0.7 kb fragment of the NP (aa1-274-S0.7. C57BL/6 mice were immunized with these recombinant adenoviral vectors. A series of immunological assays determined the immunogenicity of the recombinant adenoviral vectors. The results showed that rAd-GnS0.7-pCAG-HSP70C induced a stronger humoral and cellular immune response than other recombinant adenoviruses (rAd-GnS0.7-pCAG and rAd-GnS0.7 and the HFRS vaccine control. Animal protection experiments showed that rAd-GnS0.7-pCAG-HSP70C was effective at protecting C57BL/6 mice from HTNV infection. The results of the immunological experiments showed that HSP70C lead to enhanced vaccine potency, and suggested significant potential in the development of genetically engineered vaccines against HTNV.

  2. Expression.purification and antigenicity of a recombinant respiratory syncyttal virus fusion gene encoded protein(amino acids 168-289)in the insect cells%呼吸道合胞病毒F蛋白第168~289氨基酸片段的昆虫细胞表达及其抗原性分析

    Institute of Scientific and Technical Information of China (English)

    单志娟; 项金忠


    DNA( Bsu36 Ⅰ -digested).The recombinant baculoviruses BacPAK F' was constructed and was able to express the recombinant protein in Sf9 insect cells.The recombinant protein was purified by Ni2+ NTA chromatography and its antigenicity was identified by Western blot(WB) analysis.Specimens including the nasopharyngeal aspirates(NPAs) and sera were collected from 33 infants and young children with acute lower respiration tract infection. Indirect immunofluorecenee assay (IFA) and WB were used to detect the RSV antigen in NPAs and the anti-RSV antibody in sera respectively.Results The recombinant protein (molecular weight 13 000) was expressed in Sf9 insect cells.WB analysis demonstrated that the purified recombinant protein had a specific RSV antibody-binding activity. The recombinant protein could be recognized by positive serum infected by RSV.The positive rate was 27.3% and 33.3% respectively.There was no significant difference between them(X2 = 0.29 ,P > 0.05).Conclusion The recombinant respiratory syncytial virus fusion gene (546-881 bp) encoding protein is expressed in Sf9 insect cells and it has strong antigenicity and could have clinical application value for detection of RSV infection.

  3. Cell fusions in mammals

    DEFF Research Database (Denmark)

    Larsson, Lars-Inge; Bjerregaard, Bolette; Talts, Jan Fredrik


    Cell fusions are important to fertilization, placentation, development of skeletal muscle and bone, calcium homeostasis and the immune defense system. Additionally, cell fusions participate in tissue repair and may be important to cancer development and progression. A large number of factors appear...... to regulate cell fusions, including receptors and ligands, membrane domain organizing proteins, proteases, signaling molecules and fusogenic proteins forming alpha-helical bundles that bring membranes close together. The syncytin family of proteins represent true fusogens and the founding member, syncytin-1......, has been documented to be involved in fusions between placental trophoblasts, between cancer cells and between cancer cells and host ells. We review the literature with emphasis on the syncytin family and propose that syncytins may represent universal fusogens in primates and rodents, which work...

  4. Cold nuclear fusion

    Directory of Open Access Journals (Sweden)

    Huang Zhenqiang Huang Yuxiang


    Full Text Available In normal temperature condition, the nuclear force constraint inertial guidance method, realize the combination of deuterium and tritium, helium and lithium... And with a magnetic moment of light nuclei controlled cold nuclear collide fusion, belongs to the nuclear energy research and development in the field of applied technology "cold nuclear collide fusion". According to the similarity of the nuclear force constraint inertial guidance system, the different velocity and energy of the ion beam mixing control, developed ion speed dc transformer, it is cold nuclear fusion collide, issue of motivation and the nuclear power plant start-up fusion and power transfer system of the important equipment, so the merger to apply for a patent

  5. Fusion Revisits CERN

    CERN Multimedia


    It's going to be a hot summer at CERN. At least in the Main Building, where from 13 July to 20 August an exhibition is being hosted on nuclear fusion, the energy of the Stars. Nuclear fusion is the engine driving the stars but also a potential source of energy for mankind. The exhibition shows the different nuclear fusion techniques and research carried out on the subject in Europe. Inaugurated at CERN in 1993, following collaboration between Lausanne's CRPP-EPFL and CERN, with input from Alessandro Pascolini of Italy's INFN, this exhibition has travelled round Europe before being revamped and returning to CERN. 'Fusion, Energy of the Stars', from 13 July onwards, Main Building

  6. Laser-Driven Fusion. (United States)

    Gibson, A. F.


    Discusses the present status and future prospects of laser-driven fusion. Current research (which is classified under three main headings: laser-matter interaction processes, compression, and laser development) is also presented. (HM)

  7. Fusion plasma physics

    CERN Document Server

    Stacey, Weston M


    This revised and enlarged second edition of the popular textbook and reference contains comprehensive treatments of both the established foundations of magnetic fusion plasma physics and of the newly developing areas of active research. It concludes with a look ahead to fusion power reactors of the future. The well-established topics of fusion plasma physics -- basic plasma phenomena, Coulomb scattering, drifts of charged particles in magnetic and electric fields, plasma confinement by magnetic fields, kinetic and fluid collective plasma theories, plasma equilibria and flux surface geometry, plasma waves and instabilities, classical and neoclassical transport, plasma-materials interactions, radiation, etc. -- are fully developed from first principles through to the computational models employed in modern plasma physics. The new and emerging topics of fusion plasma physics research -- fluctuation-driven plasma transport and gyrokinetic/gyrofluid computational methodology, the physics of the divertor, neutral ...

  8. Optical Fiber Fusion Splicing

    CERN Document Server

    Yablon, Andrew D


    This book is an up-to-date treatment of optical fiber fusion splicing incorporating all the recent innovations in the field. It provides a toolbox of general strategies and specific techniques that the reader can apply when optimizing fusion splices between novel fibers. It specifically addresses considerations important for fusion splicing of contemporary specialty fibers including dispersion compensating fiber, erbium-doped gain fiber, polarization maintaining fiber, and microstructured fiber. Finally, it discusses the future of optical fiber fusion splicing including silica and non-silica based optical fibers as well as the trend toward increasing automation. Whilst serving as a self-contained reference work, abundant citations from the technical literature will enable readers to readily locate primary sources.

  9. Sampling Based Average Classifier Fusion

    Directory of Open Access Journals (Sweden)

    Jian Hou


    fusion algorithms have been proposed in literature, average fusion is almost always selected as the baseline for comparison. Little is done on exploring the potential of average fusion and proposing a better baseline. In this paper we empirically investigate the behavior of soft labels and classifiers in average fusion. As a result, we find that; by proper sampling of soft labels and classifiers, the average fusion performance can be evidently improved. This result presents sampling based average fusion as a better baseline; that is, a newly proposed classifier fusion algorithm should at least perform better than this baseline in order to demonstrate its effectiveness.

  10. Recombination and Fusion Expression of Porcine Defensin Gene pBD-2 in E.Coli%猪防御素基因pBD-2在大肠杆菌中的重组和融合表达

    Institute of Scientific and Technical Information of China (English)

    徐海涛; 马立保; 何启盖; 廖冰麟; 任芬芬


    哺乳动物防御素是动物机体在抵御病原微生物的防御反应中产生的一类重要的抗菌肽物质,具有十分广泛的抗菌谱,在先天免疫上起重要作用.本研究根据已报道的pBD-2基因cDNA序列,合成了3条基因片段(1、2、3).片段1、2和片段2、3各有15个碱基互补配对,PCR扩增延伸得到pBD-2基因,将pBD-2基因克隆至原核表达载体pGEX-KG,成功地在大肠杆菌BL21(DE3)中表达出pBD-2融合蛋白.pBD-2基因的成功表达为进一步研究其抗菌活性打下了基础.%Defensin is an important antimicrobial peptide produced in the course of animals' defense against pathogens,which has a broad antibacterial spectrum and plays an important role in innate immunity. Based on cDNA sequence of porcine defensin βdefensin 2 gene reported in this study, three gene fragment (1,2,3) were synthesized in this experiment. Fragment 1,2 and fragment 2,3 had respectively 15 bases for complementary pair. pBD-2 gene was produced by PCR amplifing, and pBD-2 was inserted into expression vector (pGEX-KG) and recombinant vector transformed into E.coli, where recombinant vector produced fusion protein successfully. The expression of pBD-2 gene lays a foundation in research on antimicrobial activities of defensin.

  11. Glucose-ABL1-TOR Signaling Modulates Cell Cycle Tuning to Control Terminal Appressorial Cell Differentiation (United States)


    The conserved target of rapamycin (TOR) pathway integrates growth and development with available nutrients, but how cellular glucose controls TOR function and signaling is poorly understood. Here, we provide functional evidence from the devastating rice blast fungus Magnaporthe oryzae that glucose can mediate TOR activity via the product of a novel carbon-responsive gene, ABL1, in order to tune cell cycle progression during infection-related development. Under nutrient-free conditions, wild type (WT) M. oryzae strains form terminal plant-infecting cells (appressoria) at the tips of germ tubes emerging from three-celled spores (conidia). WT appressorial development is accompanied by one round of mitosis followed by autophagic cell death of the conidium. In contrast, Δabl1 mutant strains undergo multiple rounds of accelerated mitosis in elongated germ tubes, produce few appressoria, and are abolished for autophagy. Treating WT spores with glucose or 2-deoxyglucose phenocopied Δabl1. Inactivating TOR in Δabl1 mutants or glucose-treated WT strains restored appressorium formation by promoting mitotic arrest at G1/G0 via an appressorium- and autophagy-inducing cell cycle delay at G2/M. Collectively, this work uncovers a novel glucose-ABL1-TOR signaling axis and shows it engages two metabolic checkpoints in order to modulate cell cycle tuning and mediate terminal appressorial cell differentiation. We thus provide new molecular insights into TOR regulation and cell development in response to glucose. PMID:28072818

  12. Fusion ignition research experiment

    Energy Technology Data Exchange (ETDEWEB)

    Dale Meade


    Understanding the properties of high gain (alpha-dominated) fusion plasmas in an advanced toroidal configuration is the largest remaining open issue that must be addressed to provide the scientific foundation for an attractive magnetic fusion reactor. The critical parts of this science can be obtained in a compact high field tokamak which is also likely to provide the fastest and least expensive path to understanding alpha-dominated plasmas in advanced toroidal systems.

  13. Economically competitive fusion

    Directory of Open Access Journals (Sweden)

    David J. Ward


    Full Text Available Not since the oil crisis of the 1970s has the perception that energy is a crucial and precious resource been as strong as it is today. The need for a new approach to world energy supply, driven by concerns over resources, pollution, and security, is leading to a reappraisal of fusion. Fusion has enormous potential and major safety and environmental advantages, and hence could make a large difference to energy supplies.

  14. Fusion, cold fusion, and space policy

    Energy Technology Data Exchange (ETDEWEB)

    Rotegard, D. (CST Ltd. (United States))


    This paper critiques Americal science policy through a consideration of two examples-cold fusion and asteroid mining. It points out that the failure of central planning in science and technology policy is just as marked as in more mundane activities. It highlights the current low level of debate and points out some technical issues that need to be addressed. It concludes with evidence that the alliance of flawed policy options is further lowering the level of debate. (author).

  15. EML4-ALK与EGFR基因突变共存型非小细胞肺癌研究进展%Research Progress in Non-small Cell Lung Cancer with Concomitant EML4-ALK Fusion Gene and EGFR Gene Mutation

    Institute of Scientific and Technical Information of China (English)

    曾珠; 吴一龙


    Lung cancer is one of the most prevalence malignances, of which non-small cell lung cancer (NSCLC) account for 80%-85% of lung cancer. Molecular target therapy is one of the most popular and promising field of NSCLC treatment, and its hotspots includes EGFR (epidermal growth factor receptor), EML4-ALK (echinoderm microtubule associated protein like4-anaplastic lymphoma kinase), etc. Former researches indicated that EML4-ALK fusion and EGFR mutation were excluded mutually. However, cases of patients harbored concomitant EML4-ALK fusion gene and EGFR mutation have been reported continuously at recent. This review aims to summarize the incidence and molecular structure of EML4-ALK fusion gene and EGFR mutation, as well as clinical features of patients with the concomitant genes induced NSCLC.%肺癌是最常见的恶性肿瘤之一,其中非小细胞肺癌( non-small cell lung cancer,NSCLC)占肺癌的80%-85%.分子靶向治疗是目前NSCLC最热门也是最具前景的领域之一,其中的热点分子包括表皮生长因子受体(epidermal growth factor receptor,EGFR)、棘皮动物微管样蛋白4-间变淋巴瘤激酶(echinoderm microtubule associated protein like4-anaplastic lymphoma kinase,EML4-ALK)等.既往研究认为EML4-ALK融合基因与EGFR突变不能共存.近期陆续报道了EML4-ALK融合基因与EGFR突变共存的病例.本文就EML4-ALK融合基因及EGFR突变基因的分子结构、发生率和目前已报道双突变患者的临床特点等进行综述.

  16. Mitochondrial Fusion Proteins and Human Diseases

    Directory of Open Access Journals (Sweden)

    Michela Ranieri


    Full Text Available Mitochondria are highly dynamic, complex organelles that continuously alter their shape, ranging between two opposite processes, fission and fusion, in response to several stimuli and the metabolic demands of the cell. Alterations in mitochondrial dynamics due to mutations in proteins involved in the fusion-fission machinery represent an important pathogenic mechanism of human diseases. The most relevant proteins involved in the mitochondrial fusion process are three GTPase dynamin-like proteins: mitofusin 1 (MFN1 and 2 (MFN2, located in the outer mitochondrial membrane, and optic atrophy protein 1 (OPA1, in the inner membrane. An expanding number of degenerative disorders are associated with mutations in the genes encoding MFN2 and OPA1, including Charcot-Marie-Tooth disease type 2A and autosomal dominant optic atrophy. While these disorders can still be considered rare, defective mitochondrial dynamics seem to play a significant role in the molecular and cellular pathogenesis of more common neurodegenerative diseases, for example, Alzheimer’s and Parkinson’s diseases. This review provides an overview of the basic molecular mechanisms involved in mitochondrial fusion and focuses on the alteration in mitochondrial DNA amount resulting from impairment of mitochondrial dynamics. We also review the literature describing the main disorders associated with the disruption of mitochondrial fusion.

  17. Melissotarsus ants are likely able to digest plant polysaccharides. (United States)

    Mony, Ruth; Dejean, Alain; Bilong, Charles Félix Bilong; Kenne, Martin; Rouland-Lefèvre, Corinne


    Melissotarsus ants have an extremely specialized set of behaviours. Both workers and gynes tunnel galleries in their host tree bark. Workers walk with their mesothoracic legs pointing upwards and tend Diaspididae hemiptera for their flesh. The ants use their forelegs to plug the galleries with silk that they secrete themselves. We hypothesised that the ants' energetic needs for nearly constant gallery digging could be satisfied through the absorption of host tree tissues; so, using basic techniques, we examined the digestive capacities of workers from two species. We show that workers are able to degrade oligosaccharides and heterosides as well as, to a lesser degree, polysaccharides. This is one of the rare reports on ants able to digest plant polysaccharides other than starch.

  18. Foreign Investors Able to Establish Foreign- exclusively Exhibition Corporations

    Institute of Scientific and Technical Information of China (English)


    Foreign Investors Able to Establish Foreign-exclusively Exhibition Corporations In Feb, Ministry of Commerce issued its 1st decree on temporary regulation for foreign-investing corporations; the regulation allows foreign investors to set up foreign-investing convention & exhibition corporations exclusively or through cooperation with other Chinese corporations, enterprises or organizations. With legal protection on their regulatory management and legal rights, these foreign-investing corporations are in the charge of Department of Foreign Investment Administration, Ministry of Commerce.

  19. Isolation of Microorganisms Able To Metabolize Purified Natural Rubber


    Heisey, R. M.; Papadatos, S


    Bacteria able to grow on purified natural rubber in the absence of other organic carbon sources were isolated from soil. Ten isolates reduced the weight of vulcanized rubber from latex gloves by >10% in 6 weeks. Scanning electron microscopy clearly revealed the ability of the microorganisms to colonize, penetrate, and dramatically alter the physical structure of the rubber. The rubber-metabolizing bacteria were identified on the basis of fatty acid profiles and cell wall characteristics. Seve...

  20. Construction of recombinant plasmid of human FGFR4 gene and expression of fusion protein%人 FGFR4基因重组质粒的构建及融合蛋白的表达

    Institute of Scientific and Technical Information of China (English)

    刘彩红; 张婷婷; 纪书婷


    目的:探讨成纤维细胞生长因子4( FGFR4)重组质粒的构建方法,并检测FGFR4融合蛋白的表达。方法从HepG2细胞中提取总RNA,采用RT-PCR方法扩增FGFR4的全长编码序列,双酶切后与pcDNA3.1/Myc-HisA载体连接,构建pcDNA3.1/Myc-HisA-FGFR4重组质粒。重组质粒经双酶切和测序鉴定后瞬时转染人胚肾HEK293细胞,采用Western blot法检测FGFR4融合蛋白的表达。结果 FGFR4编码序列被成功克隆至pcDNA 3.1/Myc-HisA质粒中,成功构建了pcDNA3.1/Myc-HisA-FGFR4载体。转染HEK293细胞后检测到FGFR4融合蛋白的稳定表达,分子量约为89 kD。结论成功构建了FGFR4全长编码基因重组质粒,并检测到转染后FGFR4蛋白在HEK293细胞中的表达。%Objective To investigate the construction method of the recombinant plasmid of fibroblast growth factor re-ceptor 4(FGFR4) and to identify the expression of its fusion protein.Methods Total RNA was extracted from HepG2 cells.We obtained the full length of human FGFR4 coding sequence by RT-PCR, digested it by restriction enzyme and joined it with pcDNA3.1/Myc-HisA to construct the pcDNA3.1/Myc-HisA-FGFR4 recombinant plasmid.After the recom-binant plasmid was identified by enzyme digestion and sequencing, the plasmid was transfected into HEK293 cells.The ex-pression of fusion protein in HEK293 cells was detected by Western blotting.Results The coding sequence of human FG-FR4 was successfully cloned into pcDNA3.1/Myc-HisA.The pcDNA3.1/Myc-HisA-FGFR4 vector was successfully con-structed.After transfecting HEK293 cells, the stable expression of FGFR4 fusion protein with a molecular weight of 89 kD was detected.Conclusion The recombinant plasmid of FGFR4 full-length coding gene was successfully constructed and the FGFR4 protein was expressed in HEK293 cells.

  1. A novel bispecific peptide HIV-1 fusion inhibitor targeting the N-terminal heptad repeat and fusion peptide domains in gp41. (United States)

    Jiang, Xifeng; Jia, Qiyan; Lu, Lu; Yu, Fei; Zheng, Jishen; Shi, Weiguo; Cai, Lifeng; Jiang, Shibo; Liu, Keliang


    HIV-1 fusion with the target cell is initiated by the insertion of the gp41 fusion peptide (FP) into the target cell membrane and the interaction between the gp41 N- and C-terminal heptad repeats (NHR and CHR), followed by the formation of the six-helix bundle (6-HB) fusion core. Therefore, both FP and NHR are important targets for HIV-1 fusion inhibitors. Here, we designed and synthesized a dual-target peptidic HIV-1 fusion inhibitor, 4HR-LBD-VIRIP, in which 4HR-LBD is able to bind to the gp41 NHR domain, while VIRIP is able to interact with gp41 FP. We found that 4HR-LBD-VIRIP is about tenfold more potent than 4HR-LBD and VIRIP in inhibiting HIV-1IIIB infection and HIV-1 envelope glycoprotein (Env)-mediated cell-cell fusion, suggesting that this dual-target HIV-1 fusion inhibitor possesses a strong synergistic antiviral effect. A biophysical analysis indicates that 4HR-LBD-VIRIP can interact with N70 peptide that contains the gp41 NHR and FP domains and binds with lipid membrane. This study provides a new approach for designing novel viral fusion inhibitors against HIV and other enveloped viruses with class I membrane fusion proteins.

  2. Cell-type-specific expression and regulation of a c-fos-NGF fusion gene in neurons and astrocytes of transgenic mice. (United States)

    Onténiente, B; Horellou, P; Neveu, I; Makeh, I; Suzuki, F; Bourdet, C; Grimber, G; Colin, P; Brachet, P; Mallet, J


    A mouse line transgenic for nerve growth factor (NGF) was developed using the mouse prepro-NGF cDNA inserted within a plasmid containing the proximal region (-10 to -550 bp) of the c-fos promoter and the transcription termination and polyadenylation signals of the rabbit beta-globin gene. No significant modification of gross behavior or central nervous system anatomy was detected in adult animals as assessed by immunohistochemistry and in situ hybridization for NGF and choline acetyltransferase. The expression of the transgene and the possible regulation of its expression by agents acting on the promoter were investigated in vitro. Despite the presence of an additional pool of NGF mRNA specific to the transgene, basal levels of NGF in the supernatant of transgenic astrocytes were similar to normal ones. On the other hand, transgenic neurons spontaneously synthesized and released levels of NGF two to three times higher than normal neurons, while mRNA levels were barely detectable by conventional Northern blotting. The tissue-specificity of NGF expression was respected, with higher levels in hippocampal than neocortical neurons. Increases of NGF mRNA by agents acting on the promoter could be observed in normal and transgenic astrocytes only after inhibition of the protein synthesis by cycloheximide, suggesting a similar rapid turnover of normal and transgenic transcripts. Cyclic AMP agonists specifically increased the secretion of NGF protein by transgenic astrocytes and neurons, while activators of the protein kinase C had a similar effect on transgenic and normal cells. Differences between amounts of NGF secreted by neurons and astrocytes with regards to their respective content in mRNA suggest that transgenic transcripts are subject to normal cell- and tissue-specific post-transcriptional regulations. Agents acting on the c-fos promoter through the protein kinase C or cyclic AMP routes differentially increased the secretion of NGF by transgenic astrocytes or

  3. Gene (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,...

  4. Lateral Lumbar Interbody Fusion (United States)

    Hughes, Alexander; Girardi, Federico; Sama, Andrew; Lebl, Darren; Cammisa, Frank


    The lateral lumbar interbody fusion (LLIF) is a relatively new technique that allows the surgeon to access the intervertebral space from a direct lateral approach either anterior to or through the psoas muscle. This approach provides an alternative to anterior lumbar interbody fusion with instrumentation, posterior lumbar interbody fusion, and transforaminal lumbar interbody fusion for anterior column support. LLIF is minimally invasive, safe, better structural support from the apophyseal ring, potential for coronal plane deformity correction, and indirect decompression, which have has made this technique popular. LLIF is currently being utilized for a variety of pathologies including but not limited to adult de novo lumbar scoliosis, central and foraminal stenosis, spondylolisthesis, and adjacent segment degeneration. Although early clinical outcomes have been good, the potential for significant neurological and vascular vertebral endplate complications exists. Nevertheless, LLIF is a promising technique with the potential to more effectively treat complex adult de novo scoliosis and achieve predictable fusion while avoiding the complications of traditional anterior surgery and posterior interbody techniques. PMID:26713134

  5. The cytoplasmic domain of the gamete membrane fusion protein HAP2 targets the protein to the fusion site in Chlamydomonas and regulates the fusion reaction (United States)

    Liu, Yanjie; Pei, Jimin; Grishin, Nick; Snell, William J.


    Cell-cell fusion between gametes is a defining step during development of eukaryotes, yet we know little about the cellular and molecular mechanisms of the gamete membrane fusion reaction. HAP2 is the sole gamete-specific protein in any system that is broadly conserved and shown by gene disruption to be essential for gamete fusion. The wide evolutionary distribution of HAP2 (also known as GCS1) indicates it was present in the last eukaryotic common ancestor and, therefore, dissecting its molecular properties should provide new insights into fundamental features of fertilization. HAP2 acts at a step after membrane adhesion, presumably directly in the merger of the lipid bilayers. Here, we use the unicellular alga Chlamydomonas to characterize contributions of key regions of HAP2 to protein location and function. We report that mutation of three strongly conserved residues in the ectodomain has no effect on targeting or fusion, although short deletions that include those residues block surface expression and fusion. Furthermore, HAP2 lacking a 237-residue segment of the cytoplasmic region is expressed at the cell surface, but fails to localize at the apical membrane patch specialized for fusion and fails to rescue fusion. Finally, we provide evidence that the ancient HAP2 contained a juxta-membrane, multi-cysteine motif in its cytoplasmic region, and that mutation of a cysteine dyad in this motif preserves protein localization, but substantially impairs HAP2 fusion activity. Thus, the ectodomain of HAP2 is essential for its surface expression, and the cytoplasmic region targets HAP2 to the site of fusion and regulates the fusion reaction. PMID:25655701

  6. RNAi screen reveals an Abl kinase-dependent host cell pathway involved in Pseudomonas aeruginosa internalization.

    Directory of Open Access Journals (Sweden)

    Julia F Pielage


    Full Text Available Internalization of the pathogenic bacterium Pseudomonas aeruginosa by non-phagocytic cells is promoted by rearrangements of the actin cytoskeleton, but the host pathways usurped by this bacterium are not clearly understood. We used RNAi-mediated gene inactivation of approximately 80 genes known to regulate the actin cytoskeleton in Drosophila S2 cells to identify host molecules essential for entry of P. aeruginosa. This work revealed Abl tyrosine kinase, the adaptor protein Crk, the small GTPases Rac1 and Cdc42, and p21-activated kinase as components of a host signaling pathway that leads to internalization of P. aeruginosa. Using a variety of complementary approaches, we validated the role of this pathway in mammalian cells. Remarkably, ExoS and ExoT, type III secreted toxins of P. aeruginosa, target this pathway by interfering with GTPase function and, in the case of ExoT, by abrogating P. aeruginosa-induced Abl-dependent Crk phosphorylation. Altogether, this work reveals that P. aeruginosa utilizes the Abl pathway for entering host cells and reveals unexpected complexity by which the P. aeruginosa type III secretion system modulates this internalization pathway. Our results furthermore demonstrate the applicability of using RNAi screens to identify host signaling cascades usurped by microbial pathogens that may be potential targets for novel therapies directed against treatment of antibiotic-resistant infections.

  7. 纳米金放大计时库仑法的PML/RARα融合基因检测%Detection of PML/RARα Fusion Gene Using Gold Nanopaticles Modified Electrode by Chronocoulometry

    Institute of Scientific and Technical Information of China (English)

    王丽满; 林丽清; 翁少煌; 林新华; 陈元仲


    A gold nanoparticles modified electrode was prepared by electrodeposition at constant potential. The sulfydryl modified probe was immobilized onto surface of gold nanoparticles via Au--S covalent bond and hybrid- ized with complementary target DNA sequences, then the DNA sensor was fabricated for the detection of PML/ RARα fusion gene by chronocoulometry in acute promyelocytic leukemia (APL). Scanning electron microscopy (SEM) and electrochemical impedance spectroscopy (EIS) were used to characterize the surface morphology of gold nanoparticles and the constructing process of sensor. With hexaammineruthenium ( Ⅲ ) chloride (RuHex) as a novel electrochemical indicator, the chronocoulometric DNA biosensor was employed to monitor artificial APL PML/RARα fusion gene fragment. Experimental results showed that gold nanoparticles amplified the detection signal of RuHex. The linear relationship between the charge difference in the RuHex after and before hybridiza- tion and logarithmic concentrations(lgC) of target sequence was observed in the concentration range of 1.0 × 10-13 -1.0 × 10-9 mol L-1 ,and the detection limit had been estimated to be 3.7 ×10-14 mol L-1 (S/N = 3 ). This approach is simple, has good specificity, and is promising to apply for the detection of APL in the real sample for medical diagnostics.%应用恒电位在金基底表面电化学沉积纳米金,通过Au—S键将巯基修饰DNA探针固定在纳米金表面,与互补靶序列杂交,构建计时库仑电化学DNA传感器,并检测急性早幼粒细胞白血病(APL)PML/RARα融合基因.采用扫描电子显微术(SEM)与电化学交流阻抗技术(EIS)观察纳米金和表征DNA传感器的构筑过程.以氯化六氨合钌([Ru(NH3)6]Cl3,RuHex)作电化学杂交指示剂,由计时库仑法检测人工合成APL的PML/RARα融合基因.结果表明,纳米金能放大RuHex检测信号,杂交前后电量差值(ΔQ)与靶标链DNA

  8. Zebra finches are able to learn affixation-like patterns. (United States)

    Chen, Jiani; Jansen, Naomi; ten Cate, Carel


    Adding an affix to transform a word is common across the world languages, with the edges of words more likely to carry out such a function. However, detecting affixation patterns is also observed in learning tasks outside the domain of language, suggesting that the underlying mechanism from which affixation patterns have arisen may not be language or even human specific. We addressed whether a songbird, the zebra finch, is able to discriminate between, and generalize, affixation-like patterns. Zebra finches were trained and tested in a Go/Nogo paradigm to discriminate artificial song element sequences resembling prefixed and suffixed 'words.' The 'stems' of the 'words,' consisted of different combinations of a triplet of song elements, to which a fourth element was added as either a 'prefix' or a 'suffix.' After training, the birds were tested with novel stems, consisting of either rearranged familiar element types or novel element types. The birds were able to generalize the affixation patterns to novel stems with both familiar and novel element types. Hence, the discrimination resulting from the training was not based on memorization of individual stimuli, but on a shared property among Go or Nogo stimuli, i.e., affixation patterns. Remarkably, birds trained with suffixation as Go pattern showed clear evidence of using both prefix and suffix, while those trained with the prefix as the Go stimulus used primarily the prefix. This finding illustrates that an asymmetry in attending to different affixations is not restricted to human languages.

  9. Galectin-3: a novel substrate for c-Abl kinase (United States)

    Balan, Vitaly; Nangia-Makker, Pratima; Jung, Young Suk; Wang, Yi; Raz, Avraham


    Galectin-3, a ß-galactoside-binding lectin, is found in cellular and extracellular location of the cell and has pleiotropic biological functions such as cell growth, cell adhesion and cell-cell interaction. It may exhibit anti or pro-apoptotic activity depending on its localization and post-translational modifications. Two important post-translational modifications of galectin-3 have been reported: its cleavage and phosphorylation. Cleavage of galectin-3 was reported to be involved with angiogenic potential and apoptotic resistance. Phosphorylation of galectin-3 regulates its sugar-binding ability. In this report we have identified novel tyrosine phosphorylation sites in galectin-3 as well as the kinase responsible for its phosphorylation. Our results demonstrate that tyrosines at position 79, 107 and 118 can be phosphorylated in vitro and in vivo by c-Abl kinase. Tyrosine 107 is the main target of c-Abl. Expression of galectin-3 Y107F mutant in galectin-3 null SK-Br-3 cells leads to morphological changes and increased motility compared to wild type galectin-3. Further investigation is needed to better understand the functional significance of the novel tyrosine phosphorylated sites of galectin-3. PMID:20600357

  10. Mitochondrial fusion and inheritance of the mitochondrial genome. (United States)

    Takano, Hiroyoshi; Onoue, Kenta; Kawano, Shigeyuki


    Although maternal or uniparental inheritance of mitochondrial genomes is a general rule, biparental inheritance is sometimes observed in protists and fungi,including yeasts. In yeast, recombination occurs between the mitochondrial genomes inherited from both parents.Mitochondrial fusion observed in yeast zygotes is thought to set up a space for DNA recombination. In the last decade,a universal mitochondrial fusion mechanism has been uncovered, using yeast as a model. On the other hand, an alternative mitochondrial fusion mechanism has been identified in the true slime mold Physarum polycephalum.A specific mitochondrial plasmid, mF, has been detected as the genetic material that causes mitochondrial fusion in P. polycephalum. Without mF, fusion of the mitochondria is not observed throughout the life cycle, suggesting that Physarum has no constitutive mitochondrial fusion mechanism.Conversely, mitochondria fuse in zygotes and during sporulation with mF. The complete mF sequence suggests that one gene, ORF640, encodes a fusogen for Physarum mitochondria. Although in general, mitochondria are inherited uniparentally, biparental inheritance occurs with specific sexual crossing in P. polycephalum.An analysis of the transmission of mitochondrial genomes has shown that recombinations between two parental mitochondrial genomes require mitochondrial fusion,mediated by mF. Physarum is a unique organism for studying mitochondrial fusion.

  11. EML4-ALK在非小细胞肺癌中的表达及检测%Detection and Expression of the Transforming EML4-ALK Fusion Gene in Non-small Cell Lung Cancer

    Institute of Scientific and Technical Information of China (English)



    Non-small cell lung cancer ( NSCLC ) is one of the most common malignant tumors with rapidly increasing incidence and mortality, and has become the top cancer related death cause.Molecular target therapy has become one of the hot research points for treatments for late NSCLC, and the pertinent application of the target drug is the key of the treatment.Fusion genes of echinoderm microtubule associated protein like 4 ( EML4 ) and anaplastic lymphoma kinase ( ALK ) are mainly expressed in NSCLC ,which is probably the new target related to resistance of NSCLC epidermal growth factor receptor to tyrosine kinase inhibitors.%非小细胞肺癌(NSCLC)为当今最常见的恶性肿瘤之一,近年来我国肺癌的发病率和病死率增长迅速,已居癌症相关死亡首位.分子靶向治疗已成为晚期NSCLC治疗的研究热点之一,靶向药物如何针对性地用于患者的治疗是关键.棘皮动物微管结合蛋白4(EML4)与间变淋巴瘤激酶(ALK)形成的融合基因主要表达于NSCLC中,这可能是与NSCLC表皮生长因子受体酪氨酸激酶抑制剂耐药相关的新靶点.

  12. Sensor Data Fusion

    DEFF Research Database (Denmark)

    Plascencia, Alfredo; Stepán, Petr


    The main contribution of this paper is to present a sensor fusion approach to scene environment mapping as part of a Sensor Data Fusion (SDF) architecture. This approach involves combined sonar array with stereo vision readings.  Sonar readings are interpreted using probability density functions...... to the occupied and empty regions. Scale Invariant Feature Transform (SIFT) feature descriptors are interpreted using gaussian probabilistic error models. The use of occupancy grids is proposed for representing the sensor readings. The Bayesian estimation approach is applied to update the sonar array......  and the SIFT descriptors' uncertainty grids. The sensor fusion yields a significant reduction in the uncertainty of the occupancy grid compared to the individual sensor readings....

  13. Multibiometrics Belief Fusion

    CERN Document Server

    Kisku, Dakshina Ranjan; Gupta, Phalguni


    This paper proposes a multimodal biometric system through Gaussian Mixture Model (GMM) for face and ear biometrics with belief fusion of the estimated scores characterized by Gabor responses and the proposed fusion is accomplished by Dempster-Shafer (DS) decision theory. Face and ear images are convolved with Gabor wavelet filters to extracts spatially enhanced Gabor facial features and Gabor ear features. Further, GMM is applied to the high-dimensional Gabor face and Gabor ear responses separately for quantitive measurements. Expectation Maximization (EM) algorithm is used to estimate density parameters in GMM. This produces two sets of feature vectors which are then fused using Dempster-Shafer theory. Experiments are conducted on multimodal database containing face and ear images of 400 individuals. It is found that use of Gabor wavelet filters along with GMM and DS theory can provide robust and efficient multimodal fusion strategy.

  14. Fusion research at ORNL

    Energy Technology Data Exchange (ETDEWEB)


    The ORNL Fusion Program includes the experimental and theoretical study of two different classes of magnetic confinement schemes - systems with helical magnetic fields, such as the tokamak and stellarator, and the ELMO Bumpy Torus (EBT) class of toroidally linked mirror systems; the development of technologies, including superconducting magnets, neutral atomic beam and radio frequency (rf) heating systems, fueling systems, materials, and diagnostics; the development of databases for atomic physics and radiation effects; the assessment of the environmental impact of magnetic fusion; and the design of advanced demonstration fusion devices. The program involves wide collaboration, both within ORNL and with other institutions. The elements of this program are shown. This document illustrates the program's scope; and aims by reviewing recent progress.

  15. Peaceful Uses of Fusion (United States)

    Teller, E.


    Applications of thermonuclear energy for peaceful and constructive purposes are surveyed. Developments and problems in the release and control of fusion energy are reviewed. It is pointed out that the future of thermonuclear power reactors will depend upon the construction of a machine that produces more electric energy than it consumes. The fuel for thermonuclear reactors is cheap and practically inexhaustible. Thermonuclear reactors produce less dangerous radioactive materials than fission reactors and, when once brought under control, are not as likely to be subject to dangerous excursions. The interaction of the hot plasma with magnetic fields opens the way for the direct production of electricity. It is possible that explosive fusion energy released underground may be harnessed for the production of electricity before the same feat is accomplished in controlled fusion processes. Applications of underground detonations of fission devices in mining and for the enhancement of oil flow in large low-specific-yield formations are also suggested.

  16. Medical Image Fusion

    Directory of Open Access Journals (Sweden)

    Mitra Rafizadeh


    Full Text Available Technological advances in medical imaging in the past two decades have enable radiologists to create images of the human body with unprecedented resolution. MRI, PET,... imaging devices can quickly acquire 3D images. Image fusion establishes an anatomical correlation between corresponding images derived from different examination. This fusion is applied either to combine images of different modalities (CT, MRI or single modality (PET-PET."nImage fusion is performed in two steps:"n1 Registration: spatial modification (eg. translation of model image relative to reference image in order to arrive at an ideal matching of both images. Registration methods are feature-based and intensity-based approaches."n2 Visualization: the goal of it is to depict the spatial relationship between the model image and refer-ence image. We can point out its clinical application in nuclear medicine (PET/CT.

  17. Killing Effect of Fusion Suicide Genes on Nasopharyngeal Carcinoma Cells in vivo%融合自杀基因对鼻咽癌细胞的体内杀伤作用

    Institute of Scientific and Technical Information of China (English)

    卿菁; 赵素萍; 蒋卫红; 章华


    [目的]研究融合自杀基因对鼻咽癌细胞的体内杀伤作用.[方法]培养鼻咽癌CNE-2细胞,建立裸鼠荷瘤模型.随机分为6组,每组5只,A组:脂质体包裹质粒瘤内注射瘤内组;B组:脂质体包裹质粒瘤内注射+肿瘤局部照射γ射线(总剂量9Gy)组;C组:脂质体包裹质粒瘤内注射+腹腔注射5氟胞嘧啶(5-Fc)组;D组:脂质体包裹质粒瘤内注射+肿瘤局部照射γ射线(总剂量9Gy)+腹腔注射5-Fc组;E组:肿瘤局部照射γ射线(总剂量9Gy)+腹腔注射5-Fc,以及空白对照组.记录肿瘤体积变化;计算各组肿瘤抑制率,绘制肿瘤生长曲线;取肿瘤组织进行病理观察;鉴定肿瘤组织中CD/UPRT.UL49基因.肿瘤体积变化的比较采用单向方差分析,组间多重比较采用LSD法.[结果]成功建立鼻咽癌裸鼠荷瘤模型,生长曲线显示:5组瘤体积差异有统计学意义(F=720.684,P<0.01),测序结果证实,除E组外,其余各处理组均检测到目的基因,且Western blot检测结果示:B和D组肿瘤组织内自杀基因的表达高于其余各组.病理切片证实肿块为低分化鳞癌,其中D组肿瘤组织在干预作用下出现明显坏死.[结论] E6.BARF1p.CD/UPRT.UL49自杀基因载体联合前药5-Fc在放射线的作用下,可明显抑制鼻咽癌移植瘤的生长,转染该融合自杀基因可增强放疗敏感性,为鼻咽癌的基因—放射治疗开辟了新思路.%[Purpose] To explore the killing effect of fusion suicide genes in human nasopharyngeal carcinoma (NPC) in vivo.[Methods] NPC CNE-2 cell line was hosted to establish mice model with tumor which were randomly divided into 6 groups,where there were 5 mice with different interventions.The volumes of tumor were measured and compared,the tumor growth inhibition rates were calculated.The tumor tissue was taken out for analysis by Hematoxylin-Eosin staining,and the existence of CD/UPRT.UL49 was detected in tumor tissue.Analysis of variance was used for the comparison of

  18. Fusion Propulsion Study (United States)


    of propellant can be millions of times greater than the fuel, only a tiny fraction can completely push out the fuel. If the plasma is moving at a... push -plate for various explosive yields. It appears that the maximum specific impulse for such a system is -4000 to 5000 sec and increasing the base...Energy Agency, 1977, p. 507. Bourque, R.F., "OHTE as a Fusion Reactor," Proc. 4th Topl. Mt,. Tecnology of Controlled NV?4clear Fusion, King of Prussia

  19. Atomic data for fusion

    Energy Technology Data Exchange (ETDEWEB)

    Hunter, H.T.; Kirkpatrick, M.I.; Alvarez, I.; Cisneros, C.; Phaneuf, R.A. (eds.); Barnett, C.F.


    This report provides a handbook of recommended cross-section and rate-coefficient data for inelastic collisions between hydrogen, helium and lithium atoms, molecules and ions, and encompasses more than 400 different reactions of primary interest in fusion research. Published experimental and theoretical data have been collected and evaluated, and the recommended data are presented in tabular, graphical and parametrized form. Processes include excitation and spectral line emission, charge exchange, ionization, stripping, dissociation and particle interchange reactions. The range of collision energies is appropriate to applications in fusion-energy research.

  20. Fusion Welding Research. (United States)


    RD-AlSO 253 FUSION WELDING RESEARCH(U) MASSACHUSETTS INST OF TECH L/I CAMBRIDGE DEPT OF MATERIALS SCIENCE AND ENGINEERING T W EAGAR ET AL. 30 RPR 85...NUMBER 12. GOV’ ACCESSION NO. 3. RECICIE-S CATALOG NUMBER 4. T TL V nd Subtitle) S. P OFRPR PERIOD COVERED 5t h A~nnual Technical Report Fusion Welding ...research S on welding processes. Studies include metal vapors in the arc, development of a high speed infrared temperature monitor, digital signal

  1. 携SH2-Caspase8融合基因重组腺病毒的构建及对K562细胞增殖的影响%Construction of recombinant adenovirus carrying SH2-Caspase8 fusion gene and its effect on proliferation in K562 cells

    Institute of Scientific and Technical Information of China (English)

    刘双凤; 胡晶; 曹唯希; 史静; 彭智; 王海霞; 李亚娟; 冯文莉


    Objective To construct the recombinant adenovirus carrying SH2-Caspase8 fusion gene and its mutant to observe its inhibitory effect on proliferation of K562 cells. Methods SH2-Caspase8 fusion gene was amplified by RT-PCR and splicing PCR, and cloned into the shuttle plasmid pAdTrack-CMV of adenovirus. The shuttle plasmid pAdT-SC-EGFP was constructed and identified by double-enzyme digestion and DNA sequencing, and transformed into ultra-competent pAdEasy-BJ5183 after Pme Ⅰ digestion. Defective replication recombinant adenovirus plasmid pAdE-SC-EGFP and its mutant were generated by homologous recombination and further transfected into AD293 cells to package recombinant adenovirus after Pac Ⅰ digestion. Recombinant adenovirus plasmid AdE-SC-EGFP was identified by PCR and Western blotting, respectively, and amplified for the measurement of its titers. Results PCR, enzyme digestion and DNA sequencing showed that the shuttle and recombinant plasmids of adenovirus, pAdT-SC-EGFP and pAdE-SC-EGFP, were successfully constructed. PCR and EGFP expression confirmed that the recombinant plasmid of adenovirus, pAdE-SC-EGFP, was successfully packaged. After amplification, its titer was 1.5 ×l012pfu/ml. Western blot analysis displayed the expression of target protein. MTT and colony formation test could inhibit the proliferation of K562 cells. Conclusion Recombinant adenovirus plasmid, AdE-SC-EGFP, carrying SH2-Caspase8 fusion gene and its mutant, has been successfully constructed, and can significantly inhibit the proliferation of BCR-ABL positive K562 cells in vitro.%目的构建并鉴定携SH2 -Caspase8融合基因的重组腺病毒AdE-SC-EGFP及其突变体,观察其对K562细胞增殖的抑制作用。方法采用RT-PCR、重叠PCR扩增SH2-Caspase8融合基因,克隆至腺病毒穿梭载体pAdTrack-CMV中,构建穿梭质粒pAdT-SC-EGFP,进行酶切和测序鉴定。将PmeⅠ酶切后的穿梭质粒转化pAdEasy-BJ5183感受态,通过细菌内同源重组产

  2. To remember in order to be able to forget

    DEFF Research Database (Denmark)

    Jørholt, Eva


    affecting many Frenchmen's attitude towards North African immigrants and their descendants. Rather than addressing the war along Manichean national allegiance lines, however, the film emphasises similarities between the Algerian fight for freedom and the French Resistance during the Second World War. Its...... setting in Paris among Algerian immigrants highlights the war as a shared national heritage, and through its mainstream format it invites all Frenchmen, regardless of their backgrounds, to remember this largely occulted war, in order to be able to move on ... and forget. In a postcolonial understanding...... of Ernest Renan's claim that the establishment and consolidation of a nation requires its individual members to forget past controversies, Outside the Law can thus be said to contribute to the construction of a new and more inclusive understanding of the French nation....

  3. Hepatoma cell-specific ganciclovir-mediated toxicity of a lentivirally transduced HSV-TkEGFP fusion protein gene placed under the control of rat alpha-fetoprotein gene regulatory sequences. (United States)

    Uch, Rathviro; Gérolami, René; Faivre, Jamila; Hardwigsen, Jean; Mathieu, Sylvie; Mannoni, Patrice; Bagnis, Claude


    Suicide gene therapy combining herpes simplex virus thymidine kinase gene transfer and ganciclovir administration can be envisioned as a powerful therapeutical approach in the treatment of hepatocellular carcinoma; however, safety issues regarding transgene expression in parenchyma cells have to be addressed. In this study, we constructed LATKW, a lentiviral vector expressing the HSV-TkEGFP gene placed under the control of the promoter elements that control the expression of the rat alpha-fetoprotein, and assayed its specific expression in vitro in hepatocarcinoma and nonhepatocarcinoma human cell lines, and in epidermal growth factor stimulated human primary hepatocytes. Using LATKW, a strong expression of the transgene was found in transduced hepatocarcinoma cells compared to a very low expression in nonhepatocarcinoma human cell lines, as assessed by Northern blot, RT-PCR, FACS analysis and ganciclovir-mediated toxicity assay, and no expression was found in lentivirally transduced normal human hepatocytes. Altogether, these results demonstrate the possibility to use a lentivirally transduced expression unit containing the rat alpha-fetoprotein promoter to restrict the HSV-TK-mediated induced GCV sensitivity to human hepatocarcinoma cells.

  4. A fusion-inhibiting peptide against Rift Valley fever virus inhibits multiple, diverse viruses.

    Directory of Open Access Journals (Sweden)

    Jeffrey W Koehler

    Full Text Available For enveloped viruses, fusion of the viral envelope with a cellular membrane is critical for a productive infection to occur. This fusion process is mediated by at least three classes of fusion proteins (Class I, II, and III based on the protein sequence and structure. For Rift Valley fever virus (RVFV, the glycoprotein Gc (Class II fusion protein mediates this fusion event following entry into the endocytic pathway, allowing the viral genome access to the cell cytoplasm. Here, we show that peptides analogous to the RVFV Gc stem region inhibited RVFV infectivity in cell culture by inhibiting the fusion process. Further, we show that infectivity can be inhibited for diverse, unrelated RNA viruses that have Class I (Ebola virus, Class II (Andes virus, or Class III (vesicular stomatitis virus fusion proteins using this single peptide. Our findings are consistent with an inhibition mechanism similar to that proposed for stem peptide fusion inhibitors of dengue virus in which the RVFV inhibitory peptide first binds to both the virion and cell membranes, allowing it to traffic with the virus into the endocytic pathway. Upon acidification and rearrangement of Gc, the peptide is then able to specifically bind to Gc and prevent fusion of the viral and endocytic membranes, thus inhibiting viral infection. These results could provide novel insights into conserved features among the three classes of viral fusion proteins and offer direction for the future development of broadly active fusion inhibitors.

  5. Genetic characterization of fast-growing rhizobia able to nodulate Prosopis alba in North Spain. (United States)

    Iglesias, Olga; Rivas, Raúl; García-Fraile, Paula; Abril, Adriana; Mateos, Pedro F; Martinez-Molina, Eustoquio; Velázquez, Encarna


    Prosopis is a Mimosaceae legume tree indigenous to South America and not naturalized in Europe. In this work 18 rhizobial strains nodulating Prosopis alba roots were isolated from a soil in North Spain that belong to eight different randomly amplified polymorphic DNA groups phylogenetically related to Sinorhizobium medicae, Sinorhizobium meliloti and Rhizobium giardinii according to their intergenic spacer and 16S rRNA gene sequences. The nodC genes of isolates close to S. medicae and S. meliloti were identical to those of S. medicae USDA 1,037(T) and S. meliloti LMG 6,133(T) and accordingly all these strains were able to nodulate both alfalfa and Prosopis. These nodC genes were phylogenetically divergent from those of the isolates close to R. giardinii that were identical to that of R. giardinii H152(T) and therefore all these strains formed nodules in common beans and Prosopis. The nodC genes of the strains isolated in Spain were phylogenetically divergent from that carried by Mesorhizobium chacoense Pr-5(T) and Sinorhizobium arboris LMG 1,4919(T) nodulating Prosopis in America and Africa, respectively. Therefore, Prosopis is a promiscuous host which can establish symbiosis with strains carrying very divergent nodC genes and this promiscuity may be an important advantage for this legume tree to be used in reforestation.

  6. Fusion engineering device design description

    Energy Technology Data Exchange (ETDEWEB)

    Flanagan, C.A.; Steiner, D.; Smith, G.E.


    The US Magnetic Fusion Engineering Act of 1980 calls for the operation of a Fusion Engineering Device (FED) by 1990. It is the intent of the Act that the FED, in combination with other testing facilities, will establish the engineering feasibility of magnetic fusion energy. During 1981, the Fusion Engineering Design Center (FEDC), under the guidance of a Technical Management Board (TMB), developed a baseline design for the FED. This design is summarized herein.

  7. Fusion Engineering Device design description

    Energy Technology Data Exchange (ETDEWEB)

    Flanagan, C.A.; Steiner, D.; Smith, G.E.


    The US Magnetic Fusion Engineering Act of 1980 calls for the operation of a Fusion Engineering Device (FED) by 1990. It is the intent of the Act that the FED, in combination with other testing facilities, will establish the engineering feasibility of magnetic fusion energy. During 1981, the Fusion Engineering Design Center (FEDC), under the guidance of a Technical Management Board (TMB), developed a baseline design for the FED. This design is summarized herein.

  8. Synergetic Multisensor Fusion (United States)


    technology have led to increased interest in using DEMs for navigation and other applications. In particular, DEMs are attractive for use in aircraft...Multisensor Fusion for Computer Vision [67]. 30 6. POSI!IONAL zSTIM&TION TECEnIQUzs FOR AN OUTDOOR MOBLE ROBOT The autonomous navigation of mobile robots is

  9. Bouillabaisse sushi fusion power

    CERN Multimedia


    "If avant-garde cuisine is any guide, Japanese-French fusion does not work all that well. And the interminable discussions over the International Thermonuclear Experimental Reactor (ITER) suggest that what is true of cooking is true of physics" (1 page)

  10. International fusion og spaltning

    DEFF Research Database (Denmark)

    Hansen, Lone L.

    Bogen analyserer de nye muligheder fra 2007 i europæisk ret med hensyn til fusion eller spaltning mellem aktieselskaber og anpartsselskaber med hjemsted i forskellige europæiske lande. Bogen gennemgår de nye muligheder for strukturændringer, der herved er opstået mulighed for, og den sætter fokus...

  11. Fusion reactor materials

    Energy Technology Data Exchange (ETDEWEB)



    This paper discuses the following topics on fusion reactor materials: irradiation, facilities, test matrices, and experimental methods; dosimetry, damage parameters, and activation calculations; materials engineering and design requirements; fundamental mechanical behavior; radiation effects; development of structural alloys; solid breeding materials; and ceramics.

  12. Iterative guided image fusion

    Directory of Open Access Journals (Sweden)

    Alexander Toet


    Full Text Available We propose a multi-scale image fusion scheme based on guided filtering. Guided filtering can effectively reduce noise while preserving detail boundaries. When applied in an iterative mode, guided filtering selectively eliminates small scale details while restoring larger scale edges. The proposed multi-scale image fusion scheme achieves spatial consistency by using guided filtering both at the decomposition and at the recombination stage of the multi-scale fusion process. First, size-selective iterative guided filtering is applied to decompose the source images into approximation and residual layers at multiple spatial scales. Then, frequency-tuned filtering is used to compute saliency maps at successive spatial scales. Next, at each spatial scale binary weighting maps are obtained as the pixelwise maximum of corresponding source saliency maps. Guided filtering of the binary weighting maps with their corresponding source images as guidance images serves to reduce noise and to restore spatial consistency. The final fused image is obtained as the weighted recombination of the individual residual layers and the mean of the approximation layers at the coarsest spatial scale. Application to multiband visual (intensified and thermal infrared imagery demonstrates that the proposed method obtains state-of-the-art performance for the fusion of multispectral nightvision images. The method has a simple implementation and is computationally efficient.

  13. Muon catalyzed fusion

    Energy Technology Data Exchange (ETDEWEB)

    Ishida, K. [Advanced Meson Science Laboratory, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198 (Japan); Nagamine, K. [Muon Science Laboratory, IMSS-KEK, 1-1 Oho, Tsukuba, Ibaraki 305-0801 (Japan); Matsuzaki, T. [Advanced Meson Science Laboratory, RIKEN, 2-1 Hirosawa, Wako, Saitama 351-0198 (Japan); Kawamura, N. [Muon Science Laboratory, IMSS-KEK, 1-1 Oho, Tsukuba, Ibaraki 305-0801 (Japan)


    The latest progress of muon catalyzed fusion study at the RIKEN-RAL muon facility (and partly at TRIUMF) is reported. The topics covered are magnetic field effect, muon transfer to {sup 3}He in solid D/T and ortho-para effect in dd{mu} formation.

  14. Hugging fusion and related topics

    Energy Technology Data Exchange (ETDEWEB)

    Iwamoto, Akira [Japan Atomic Energy Research Inst., Tokai, Ibaraki (Japan). Tokai Research Establishment


    An important problem related to the synthesis of very heavy nuclides by fusion of two heavy-ions is the extra push effect. To avoid it, we propose a hugging fusion, which is the fusion of two well-deformed heavy-ions. (author)

  15. Prevalence of API2-MALT1 fusion gene in gastrointestinal mucosa-associated lymphoid tissue lymphomas and diffuse large B cell lymphomas%胃肠道黏膜相关边缘区B细胞淋巴瘤和弥漫性大B细胞淋巴瘤中API2-MALT1融合基因表达的差异

    Institute of Scientific and Technical Information of China (English)

    李百周; 陆洪芬; 盛伟琪; 施达仁


    Objective To investigate the difference of the prevalence of t(11;18) (q21;q21)/ AP12-MALT1 fusion gene between gastrointestinal mucosa-associated lymphoid tissue (MALT) lymphoma and diffuse large B cell lymphoma (DLBCL). Methods A total of 57 cases gastrointestinal MALT lymphomas (38 gastric and 19 intestinal lymphomas), 32 DLBCL (28 gastric and 4 intestinal lymphomas) and 7 cases gastric DLBCL accompanied MALT lymphoma were collected from the Cancer Hospital of Fudan University. API2-MALT1 fusion gene was detected by fluorescent in situ hybridization (FISH) using both dual fusion translocation and break apart probes. Results Among gastrointestinal MALT lymphomas, API2-MALT1 fusion gene was found in 12 of 57 cases (21.1% , 10 gastric and 2 intestinal lymphomas). In contrast, the fusion gene was not found in all 32 DLBCL and 7 gastric DLBCL with MALT lymphoma component. There was statistical significant difference between two groups (X~2=9.383, P=0.001). Conclusions API2-MALT1 fusion gene is a distinctive genetic aberration in MALT lymphomas, and is not present in DLBCL. The findings suggest that gastrointestinal tract MALT lymphomas with API2-MALT1 fusion geue may not transform into DLBCL, which may represent primary lymphoma or transformed API2-MALT1 negative MALT lymphomas.%目的 比较t(11;18)(q21;q21)/API2-MALX1融合基因在胃肠道黏膜相关边缘区B细胞淋巴瘤(MALN淋巴瘤)和弥漫性大B细胞淋巴瘤(DLBCL)中的发生情况,探讨t(11;18)(q21;q21)与胃肠道MALT淋巴瘤和DLBCL间演进的关系.方法 收集57例胃肠道MALT淋巴瘤(包括38例胃和19例肠),32例胃肠道DLBCL(包括28例胃和4例肠)和7例胃DLBCL同时合并MALT淋巴瘤成分,用荧光原位杂交(FISH)检测API2-MALT1融合基因.使用的探针包括API2-MALT1双色融合易位探针和MALT1双色分离重排探针.结果 在MALT淋巴瘤中有21.1%(12/57,包括10例胃和2例肠)发现API2-MALT1融合基因,而在32例DLBCL和7例DLBCL与MALT淋巴瘤混合

  16. Expression of SET-NUP214 Fusion Gene in Patients with T-cell Acute Lymphoblastic Leukemia and Its Clinical Significance%SET-NUP214融合基因在急性T淋巴细胞白血病患者中的表达及临床意义

    Institute of Scientific and Technical Information of China (English)

    戴海萍; 陈苏宁; 王谦; 吴丽丽; 平娜娜; 吴春晓; 解珺丹; 潘金兰; 薛永权; 吴德沛


    本研究旨在探讨SET-NUP214融合基因在急性T淋巴细胞白血病(T-ALL)中的表达,分析伴有SET-NUP214的T-ALL的临床及分子生物学特征.运用RT-PCR检测58例T-ALL患者骨髓标本中SET-NUP214的表达,以间期荧光原位杂交( FISH)及微阵列比较基因组杂交(Array-CGH)检测9q34缺失,基因测序法检测PHF6和NOTCH1基因突变.结果表明,仅在6例T-ALL患者中检测到SET-NUP214融合基因的表达.除T系抗原标记外,这些患者均表达髓系抗原CD13和(或)CD33,其中4例患者经FISH检测到9q34缺失,3例患者经Array-CGH检测到del(9)( q34.11q34.33).6例SET-NUP214阳性的T-ALL患者中有4例检测到PHF6基因突变,5例检测到NOTCH1基因突变.结论:SET-NUP214融合基因多由染色体9q34的缺失所致,SET-NUP214阳性的T-ALL常伴有PHF6及NOTCH基因突变.%This study was aimed to investigate the occurrence and clinical significance of the SET-NUP214 fusion gene in patients with T-cell acute lymphoblastic leukemia (T-ALL) , analyse clinical and biological characteristics in this disease. RT-PCR was used to detect the expression of SET-NUP214 fusion gene in 58 T-ALL cases. Interphase FISH and Array-CGH were used to detect the deletion of 9q34. Direct sequencing was applied to detect mutations of PHF6 and NOTCH1. The results showed that 6 out of 58 T-ALL cases (10. 3% ) were detected to have the SET-NUP214 fusion gene by RT-PCR. Besides T-lineage antigens, expression of CD13 and(or) CD33 were detected in all the 6 cases. Deletions of 9q34 were detected in 4 out of the 6 patients by FISH. Array-CGH results of 3 SET-NUP214 positive T-ALL patients confirmed that this fusion gene was resulted from a cryptic deletion of 9q34. Ilq34. 13. PHF6 and N0TCH1 gene mutations were found in 4 and 5 out of 6 SET-NUP214 positive T-ALL patients, respectively. It is concluded that SET-NUP214 fusion gene is often resulted from del(9) (q34). PHF6 and N0TCH1 mutations may be potential leukemo-genic event in SET

  17. Controlled fusion and plasma physics

    CERN Document Server

    Miyamoto, Kenro


    Resulting from ongoing, international research into fusion processes, the International Tokamak Experimental Reactor (ITER) is a major step in the quest for a new energy source.The first graduate-level text to cover the details of ITER, Controlled Fusion and Plasma Physics introduces various aspects and issues of recent fusion research activities through the shortest access path. The distinguished author breaks down the topic by first dealing with fusion and then concentrating on the more complex subject of plasma physics. The book begins with the basics of controlled fusion research, foll

  18. 来源于Alcaligenes A-6的D-氨基酰化酶基因的合成与融合表达%Gene Synthesis and Fusion Expression D-Aminoacylase Gene from Alcaligenes A-6

    Institute of Scientific and Technical Information of China (English)

    侯欣彤; 董媛; 林瑞东; 于梁; 任媛媛; 李剑光; 高朝辉; 滕利荣


    将来源于Alcaligenes A-6的D-氨基酰化酶基因用大肠杆菌中的丰沛密码子替换,利用化学和基于聚合酶链反应(PCR)技术的酶促方法进行基因全合成,利用pET-32a构建重组表达载体pET-dan,转化进E. coil BL21(DE3)中进行融合表达.经SDS-PAGE电泳、Western-blot检测和活性测定发现, D-ANase可在大肠杆菌中高效表达,目的蛋白可达到菌体总蛋白的69.2%,密码子优化后基因构建的工程菌发酵活性为96 U/mL,重组蛋白经超声细胞破碎及Ni2+柱亲和层析纯化,比活可达1692.3 U/mg,纯度可达95%以上.%The codons of D-ANase gene from Alcaligenes A-6 were substituted by the codons abundant in E. coli. , then the D-ANase gene was synthesized by the two-step method based on PCR technology. Synthetic gene and pET-32a vector were digested with BglII and XhoⅠ, ligated by T4 DNA ligase. The ligation mix-ture transformed into E. coli BL21(DE3) competent cell. Recombinant protein was detected by SDS-PAGE, Western-blot and activity assay. D-ANase can be expressed efficiently in E. coli and the expressed protein content can reach to 69.2% of the total bacterial protein content. In addition, the fermentation activity can achieve 96 U/mL. After the ultrasonic cell disruption, the recombinant protein was purified by Ni2+ affinity chromatography column. The specific activity of the purified recombinant enzyme was 1692.3 U/mg and the purity could be up to 95%. Furthermore a firm foundation was laid for the industrial use of the D-ANase.

  19. Novel linear polymers able to inhibit bacterial quorum sensing. (United States)

    Cavaleiro, Eliana; Duarte, Ana Sofia; Esteves, Ana Cristina; Correia, António; Whitcombe, Michael J; Piletska, Elena V; Piletsky, Sergey A; Chianella, Iva


    Bacterial phenotypes, such as biofilm formation, antibiotic resistance and virulence expression, are associated with quorum sensing. Quorum sensing is a density-dependent regulatory system of gene expression controlled by specific signal molecules, such as N-acyl homoserine lactones (AHLs), produced and released by bacteria. This study reports the development of linear polymers capable to attenuate quorum sensing by adsorption of AHLs. Linear polymers were synthesized using MMA as backbone monomer and methacrylic acid and itaconic acid as functional monomers. Two different quorum sensing-controlled phenotypes, Vibrio fischeri bioluminescence and Aeromonas hydrophila biofilm formation, were evaluated to test the polymers' efficiency. Results showed that both phenotypes were significantly affected by the polymers, with the itaconic acid-containing material being more effective than the methacrylic acid one. The polymer inhibitory effects were reverted by the addition of lactones, confirming attenuation of quorum sensing through sequestration of signal molecules. The polymers also showed no cytotoxicity when tested using a mammalian cell line.

  20. shRNA library screening identifies nucleocytoplasmic transport as a mediator of BCR-ABL1 kinase-independent resistance. (United States)

    Khorashad, Jamshid S; Eiring, Anna M; Mason, Clinton C; Gantz, Kevin C; Bowler, Amber D; Redwine, Hannah M; Yu, Fan; Kraft, Ira L; Pomicter, Anthony D; Reynolds, Kimberly R; Iovino, Anthony J; Zabriskie, Matthew S; Heaton, William L; Tantravahi, Srinivas K; Kauffman, Michael; Shacham, Sharon; Chenchik, Alex; Bonneau, Kyle; Ullman, Katharine S; O'Hare, Thomas; Deininger, Michael W


    The mechanisms underlying tyrosine kinase inhibitor (TKI) resistance in chronic myeloid leukemia (CML) patients lacking explanatory BCR-ABL1 kinase domain mutations are incompletely understood. To identify mechanisms of TKI resistance that are independent of BCR-ABL1 kinase activity, we introduced a lentiviral short hairpin RNA (shRNA) library targeting ∼5000 cell signaling genes into K562(R), a CML cell line with BCR-ABL1 kinase-independent TKI resistance expressing exclusively native BCR-ABL1. A customized algorithm identified genes whose shRNA-mediated knockdown markedly impaired growth of K562(R) cells compared with TKI-sensitive controls. Among the top candidates were 2 components of the nucleocytoplasmic transport complex, RAN and XPO1 (CRM1). shRNA-mediated RAN inhibition or treatment of cells with the XPO1 inhibitor, KPT-330 (Selinexor), increased the imatinib sensitivity of CML cell lines with kinase-independent TKI resistance. Inhibition of either RAN or XPO1 impaired colony formation of CD34(+) cells from newly diagnosed and TKI-resistant CML patients in the presence of imatinib, without effects on CD34(+) cells from normal cord blood or from a patient harboring the BCR-ABL1(T315I) mutant. These data implicate RAN in BCR-ABL1 kinase-independent imatinib resistance and show that shRNA library screens are useful to identify alternative pathways critical to drug resistance in CML.

  1. Alphavirus Entry and Membrane Fusion

    Directory of Open Access Journals (Sweden)

    Margaret Kielian


    Full Text Available The study of enveloped animal viruses has greatly advanced our understanding of the general properties of membrane fusion and of the specific pathways that viruses use to infect the host cell. The membrane fusion proteins of the alphaviruses and flaviviruses have many similarities in structure and function. As reviewed here, alphaviruses use receptor-mediated endocytic uptake and low pH-triggered membrane fusion to deliver their RNA genomes into the cytoplasm. Recent advances in understanding the biochemistry and structure of the alphavirus membrane fusion protein provide a clearer picture of this fusion reaction, including the protein’s conformational changes during fusion and the identification of key domains. These insights into the alphavirus fusion mechanism suggest new areas for experimental investigation and potential inhibitor strategies for anti-viral therapy.

  2. JET: Preparing the future in fusion (United States)

    Mlynár, J.; Ongena, J.; Duran, I.; Hron, M.; Pánek, R.; Petržílka, V.; Žáček, F.


    JET (Joint European Torus) is the largest tokamak in the world and the only fusion facility able to operate with Tritium, the fusion fuel, and Beryllium, the ITER first wall material. JET also features the most complete remote handling equipment for invessel maintenance. As a multinational research center, JET provides logistic experience in preparing for operation of the global facility, tokamak ITER. Experiments on JET are focused on ITER-relevant studies, in particular on detailing the operational scenarios (EL My H-modes and advanced regimes), on enhancing the heating systems, on developing diagnostics for burning plasmas etc. Pioneering real-time control techniques have been implemented that maximize performance and minimize internal disturbances of JET plasmas. In helium plasmas, ion cyclotron heating (ICRH) created fast α-particles, mimicking their populations in future burning plasmas. The recent successful Trace Tritium campaign provided important new data on fuel transport. Current enhancements on JET include a new ITER-like ELM-resilient high power ICRH antenna (7 MW) and over twenty new diagnostics that will further extend the JET scientific capabilities and push the facility even closer to the ITER parameters. A special mention is given to the involvement of the fusion experts from Association EURATOM-IPP.CR, who have been actively participating in the collective use of JET facility for more than three years.

  3. Enhanced Membrane Fusion in Sterol-enriched Vacuoles Bypasses the Vrp1p RequirementD⃞


    Tedrick, Kelly; Trischuk, Tim; Lehner, Richard; Eitzen, Gary


    Organization of lipids into membrane microdomains is a vital mechanism of protein processing. Here we show that overexpression of ERG6, a gene involved in ergosterol synthesis, elevates sterol levels 1.5-fold on the vacuole membrane and enhances their homotypic fusion. The mechanism of sterol-enhanced fusion is not via more efficient sorting, but instead promotes increased kinetics of fusion subreactions. We initially isolated ERG6 as a suppressor of a vrp1Δ growth defect selective for vacuol...

  4. Fusion Data Grid Service (United States)

    Shasharina, Svetlana; Wang, Nanbor


    Simulations and experiments in the fusion and plasma physics community generate large datasets at remote sites. Visualization and analysis of these datasets are difficult because of the incompatibility among the various data formats adopted by simulation, experiments, and analysis tools, and the large sizes of analyzed data. Grids and Web Services technologies are capable of providing solutions for such heterogeneous settings, but need to be customized to the field-specific needs and merged with distributed technologies currently used by the community. This paper describes how we are addressing these issues in the Fusion Grid Service under development. We also present performance results of relevant data transfer mechanisms including binary SOAP, DIME, GridFTP and MDSplus and CORBA. We will describe the status of data converters (between HDF5 and MDSplus data types), developed in collaboration with MIT (J. Stillerman). Finally, we will analyze bottlenecks of MDSplus data transfer mechanism (work performed in collaboration with General Atomics (D. Schissel and M. Qian).

  5. Alternate laser fusion drivers

    Energy Technology Data Exchange (ETDEWEB)

    Pleasance, L.D.


    Over the past few years, several laser systems have been considered as possible laser fusion drivers. Recently, there has been an increasing effort to evaluate these systems in terms of a reactor driver application. The specifications for such a system have become firmer and generally more restrictive. Several of the promising candidates such as the group VI laser, the metal vapor excimers and some solid state lasers can be eliminated on the basis of inefficiency. New solid state systems may impact the long range development of a fusion driver. Of the short wavelength gas lasers, the KrF laser used in conjunction with Raman compression and pulse stacking techniques is the most promising approach. Efficiencies approaching 10% may be possible with this system. While technically feasible, these approaches are complex and costly and are unsatisfying in an aethetic sense. A search for new lasers with more compelling features is still needed.

  6. Embryonic stem cell-somatic cell fusion and postfusion enucleation. (United States)

    Sumer, Huseyin; Verma, Paul J


    Embryonic stem (ES) cells are able to reprogram somatic cells following cell fusion. The resulting cell hybrids have been shown to have similar properties to pluripotent cells. It has also been shown that transcriptional changes can occur in a heterokaryon, without nuclear hybridization. However it is unclear whether these changes can be sustained following removal of the dominant ES nucleus. In this chapter, methods are described for the cell fusion of mouse tetraploid ES cells with somatic cells and enrichment of the resulting heterokaryons. We next describe the conditions for the differential removal of the ES cell nucleus, allowing for the recovery of somatic cells.

  7. Spatial Diversity Aware Data Fusion for Cooperative Spectrum Sensing

    DEFF Research Database (Denmark)

    Kiilerich Pratas, Nuno; Prasad, Neeli R.; Rodrigues, António;


    similar signal conditions are grouped by the fusion center in clusters and where the data fusion is then done separately at each cluster. The proposed algorithm uses the correlation between the binary decisions of the local detectors over an observation window to select the cluster where each local...... detector should go. It was observed that in the case where there is only one signal source, that the proposed algorithm is able to achieve the same level of performance when compared to the perfect clustering algorithm where full information about the signal conditions at each local detector is available....

  8. Novel 5′ Fusion Partners of ETV1 and ETV4 in Prostate Cancer

    Directory of Open Access Journals (Sweden)

    Joao D. Barros-Silva


    Full Text Available Gene fusions involving the erythroblast transformation-specific (ETS transcription factors ERG, ETV1, ETV4, ETV5, and FLI1 are a common feature of prostate carcinomas (PCas. Themost common upstream fusion partner described is the androgenregulated prostate-specific gene TMPRSS2, most frequently with ERG, but additional 5′fusion partners have been described. We performed 5′ rapid amplification of cDNA ends in 18 PCas with ETV1, ETV4, or ETV5 outlier expression to identify the 5′ fusion partners. We also evaluated the exon-level expression profile of these ETS genes in 14 cases. We identified and confirmed by fluorescent in situ hybridization (FISH and reverse transcription-polymerase chain reaction the two novel chimeric genes OR51E2-ETV1 and UBTF-ETV4 in two PCas. OR51E2 encodes a G-protein.coupled receptor that is overexpressed in PCas, whereas UBTF is a ubiquitously expressed gene encoding an HMG-box DNA-binding protein involved in ribosome biogenesis. We additionally describe two novel gene fusion combinations of previously described genes, namely, SLC45A3- ETV4 and HERVK17-ETV4. Finally, we found one PCa with TMPRSS2-ETV1, one with C15orf21-ETV1, one with EST14-ETV1, and two with 14q133-q21.1-ETV1. In nine PCas (eight ETV1 and one ETV5, exhibiting ETS outlier expression and genomic rearrangement detected by FISH, no 5′ fusion partner was found. Our findings contribute significantly to characterize the heterogeneous group of ETS gene fusions and indicate that all genes described as 5′ fusion partners with one ETS gene can most likely be rearranged with any of the other ETS genes involved in prostate carcinogenesis.

  9. (Fusion energy research)

    Energy Technology Data Exchange (ETDEWEB)

    Phillips, C.A. (ed.)


    This report discusses the following topics: principal parameters achieved in experimental devices (FY88); tokamak fusion test reactor; Princeton beta Experiment-Modification; S-1 Spheromak; current drive experiment; x-ray laser studies; spacecraft glow experiment; plasma deposition and etching of thin films; theoretical plasma; tokamak modeling; compact ignition tokamak; international thermonuclear experimental reactor; Engineering Department; Project Planning and Safety Office; quality assurance and reliability; and technology transfer.

  10. Materials for Fusion Applications

    Directory of Open Access Journals (Sweden)

    Jiří Matějíček


    Full Text Available An overview of materials foreseen for use or already used in fusion devices is given. The operating conditions, material requirements and characteristics of candidate materials in several specific application segments are briefly reviewed. These include: construction materials, electrical insulation, permeation barriers and plasma facing components. Special attention will be paid to the latter and to the issues of plasma-material interaction, materials joining and fuctionally graded interlayers.

  11. Fc-fusion mimetics



    The Fc-fusion mimetic RpR 2 was prepared by disulfide bridging conjugation using a PEG in the place of the Fc. RpR 2 displayed higher affinity for VEGF than aflibercept caused primarily by a slower dissociation rate, which can prolong a drug at its site of action. RpRs have considerable potential for development as stable, organ specific therapeutics.

  12. Fusion development and technology

    Energy Technology Data Exchange (ETDEWEB)

    Montgomery, D.B.


    This report discusses the following: superconducting magnet technology; high field superconductors; advanced magnetic system and divertor development; poloidal field coils; gyrotron development; commercial reactor studies--aries; ITER physics: alpha physics and alcator R D for ITER; lower hybrid current drive and heating in the ITER device; ITER superconducting PF scenario and magnet analysis; ITER systems studies; and safety, environmental and economic factors in fusion development.

  13. Modular Aneutronic Fusion Engine

    Energy Technology Data Exchange (ETDEWEB)

    Gary Pajer, Yosef Razin, Michael Paluszek, A.H. Glasser and Samuel Cohen


    NASA's JUNO mission will arrive at Jupiter in July 2016, after nearly five years in space. Since operational costs tend to rise with mission time, minimizing such times becomes a top priority. We present the conceptual design for a 10MW aneutronic fusion engine with high exhaust velocities that would reduce transit time for a Jupiter mission to eighteen months and enable more challenging exploration missions in the solar system and beyond. __________________________________________________

  14. RNA interference screen identifies Abl kinase and PDGFR signaling in Chlamydia trachomatis entry.

    Directory of Open Access Journals (Sweden)

    Cherilyn A Elwell


    Full Text Available The strain designated Chlamydia trachomatis serovar L2 that was used for experiments in this paper is Chlamydia muridarum, a species closely related to C. trachomatis (and formerly termed the Mouse Pneumonitis strain of C. trachomatis. This conclusion was verified by deep sequencing and by PCR using species-specific primers. All data presented in the results section that refer to C. trachomatis should be interpreted as referring to C. muridarum. Since C. muridarum TARP lacks the consensus tyrosine repeats present in C. trachomatis TARP, we cannot make any conclusions about the role of TARP phosphorylation and C. muridarum entry. However, the conclusion that C. trachomatis L2 TARP is a target of Abl kinase is still valid as these experiments were performed with C. trachomatis L2 TARP [corrected]. To elucidate the mechanisms involved in early events in Chlamydia trachomatis infection, we conducted a large scale unbiased RNA interference screen in Drosophila melanogaster S2 cells. This allowed identification of candidate host factors in a simple non-redundant, genetically tractable system. From a library of 7,216 double stranded RNAs (dsRNA, we identified approximately 226 host genes, including two tyrosine kinases, Abelson (Abl kinase and PDGF- and VEGF-receptor related (Pvr, a homolog of the Platelet-derived growth factor receptor (PDGFR. We further examined the role of these two kinases in C. trachomatis binding and internalization into mammalian cells. Both kinases are phosphorylated upon infection and recruited to the site of bacterial attachment, but their roles in the infectious process are distinct. We provide evidence that PDGFRbeta may function as a receptor, as inhibition of PDGFRbeta by RNA interference or by PDGFRbeta neutralizing antibodies significantly reduces bacterial binding, whereas depletion of Abl kinase has no effect on binding. Bacterial internalization can occur through activation of PDGFRbeta or through independent

  15. Menin-MLL inhibitors reverse oncogenic activity of MLL fusion proteins in leukemia. (United States)

    Grembecka, Jolanta; He, Shihan; Shi, Aibin; Purohit, Trupta; Muntean, Andrew G; Sorenson, Roderick J; Showalter, Hollis D; Murai, Marcelo J; Belcher, Amalia M; Hartley, Thomas; Hess, Jay L; Cierpicki, Tomasz


    Translocations involving the mixed lineage leukemia (MLL) gene result in human acute leukemias with very poor prognosis. The leukemogenic activity of MLL fusion proteins is critically dependent on their direct interaction with menin, a product of the multiple endocrine neoplasia (MEN1) gene. Here we present what are to our knowledge the first small-molecule inhibitors of the menin-MLL fusion protein interaction that specifically bind menin with nanomolar affinities. These compounds effectively reverse MLL fusion protein-mediated leukemic transformation by downregulating the expression of target genes required for MLL fusion protein oncogenic activity. They also selectively block proliferation and induce both apoptosis and differentiation of leukemia cells harboring MLL translocations. Identification of these compounds provides a new tool for better understanding MLL-mediated leukemogenesis and represents a new approach for studying the role of menin as an oncogenic cofactor of MLL fusion proteins. Our findings also highlight a new therapeutic strategy for aggressive leukemias with MLL rearrangements.

  16. Antileukemia effects of xanthohumol in Bcr/Abl-transformed cells involve nuclear factor-kappaB and p53 modulation. (United States)

    Monteghirfo, Stefano; Tosetti, Francesca; Ambrosini, Claudia; Stigliani, Sara; Pozzi, Sarah; Frassoni, Francesco; Fassina, Gianfranco; Soverini, Simona; Albini, Adriana; Ferrari, Nicoletta


    The oncogenic Bcr-Abl tyrosine kinase activates various signaling pathways including phosphoinositide 3-kinase/Akt and nuclear factor-kappaB that mediate proliferation, transformation, and apoptosis resistance in Bcr-Abl+ myeloid leukemia cells. The hop flavonoid xanthohumol inhibits tumor growth by targeting the nuclear factor-kappaB and Akt pathways and angiogenesis. Here, we show that xanthohumol has in vitro activity against Bcr-Abl+ cells and clinical samples and retained its cytotoxicity when imatinib mesylate-resistant K562 cells were examined. Xanthohumol inhibition of K562 cell viability was associated with induction of apoptosis, increased p21 and p53 expression, and decreased survivin levels. We show that xanthohumol strongly inhibited Bcr-Abl expression at both mRNA and protein levels and show that xanthohumol caused elevation of intracellular reactive oxygen species and that the antioxidant N-acetylcysteine blunted xanthohumol-induced events. Further, we observed that xanthohumol inhibits leukemia cell invasion, metalloprotease production, and adhesion to endothelial cells, potentially preventing in vivo life-threatening complications of leukostasis and tissue infiltration by leukemic cells. As structural mutations and/or gene amplification in Bcr-Abl can circumvent an otherwise potent anticancer drug such as imatinib, targeting Bcr-Abl expression as well as its kinase activity could be a novel additional therapeutic approach for the treatment of Bcr-Abl+ myeloid leukemia.

  17. MicroRNA-320a acts as a tumor suppressor by targeting BCR/ABL oncogene in chronic myeloid leukemia. (United States)

    Xishan, Zhu; Ziying, Lin; Jing, Du; Gang, Liu


    Accumulating evidences demonstrated that the induction of epithelial-mesenchymal transition (EMT) and aberrant expression of microRNAs (miRNAs) are associated with tumorigenesis, tumor progression, metastasis and relapse in cancers, including chronic myeloid leukemia (CML). We found that miR-320a expression was reduced in K562 and in CML cancer stem cells. Moreover, we found that miR-320a inhibited K562 cell migration, invasion, proliferation and promoted apoptosis by targeting BCR/ABL oncogene. As an upstream regulator of BCR/ABL, miR-320a directly targets BCR/ABL. The enhanced expression of miR-320a inhibited the phosphorylation of PI3K, AKT and NF-κB; however, the expression of phosphorylated PI3K, AKT and NF-κB were restored by the overexpression of BCR/ABL. In K562, infected with miR-320a or transfected with SiBCR/ABL, the protein levels of fibronectin, vimentin, and N-cadherin were decreased, but the expression of E-cadherin was increased. The expression of mesenchymal markers in miR-320a-expressing cells was restored to normal levels by the restoration of BCR/ABL expression. Generally speaking, miR-320a acts as a novel tumor suppressor gene in CML and miR-320a can decrease migratory, invasive, proliferative and apoptotic behaviors, as well as CML EMT, by attenuating the expression of BCR/ABL oncogene.

  18. Color image fusion for concealed weapon detection (United States)

    Toet, Alexander


    Recent advances in passive and active imaging sensor technology offer the potential to detect weapons that are concealed underneath a person's clothing or carried along in bags. Although the concealed weapons can sometimes easily be detected, it can be difficult to perceive their context, due to the non-literal nature of these images. Especially for dynamic crowd surveillance purposes it may be impossible to rapidly asses with certainty which individual in the crowd is the one carrying the observed weapon. Sensor fusion is an enabling technology that may be used to solve this problem. Through fusion the signal of the sensor that depicts the weapon can be displayed in the context provided by a sensor of a different modality. We propose an image fusion scheme in which non-literal imagery can be fused with standard color images such that the result clearly displays the observed weapons in the context of the original color image. The procedure is such that the relevant contrast details from the non-literal image are transferred to the color image without altering the original color distribution of this image. The result is a natural looking color image that fluently combines all details from both input sources. When an observer who performs a dynamic crowd surveillance task, detects a weapon in the scene, he will also be able to quickly determine which person in the crowd is actually carrying the observed weapon (e.g. "the man with the red T-shirt and blue jeans"). The method is illustrated by the fusion of thermal 8-12 μm imagery with standard RGB color images.

  19. Transient Gene Expression and Purification of a Mouse-Derived Anti-Allergy Fusion Protein%鼠源抗过敏融合蛋白的瞬时表达及纯化

    Institute of Scientific and Technical Information of China (English)

    程露; 易小萍; 孙祥明; 张元兴


    To improve the transient gene expression of a mouse-derived anti-allergy fusion protein(mAAFP) in 5 L bioreactor,feeding and mild hypothermia were adopted together to provide sufficient nutrition for cell growth and extend the production duration.Firstly,two different expression vectors of pCI-mAAFP and pID-mAAFP were transiently transfected in CHO-S cells with an optimal transient transfection protocol and a better level of mAAFP expression was reached when using pID-mAAFP as an expression vector.Transient gene expression(TGE) was conducted in 1.3 L bioreactor,with combined use of fed-batch and mild hypothermic culture,so the expression level of mAAFP was increased 8-fold compared with batch culture.Finally,the TGE process was carried out in a 5 L bioreactor and a high expression level of 25 mg/L was achieved.mAAFP was subsequently purified by rProteinA Sepharose Fast Flow.The obtained mAAFP was identified by SDS-PAGE and Western blot.This work provided the necessary basis to assay mAAFP efficacy in mouse later on.%采用流加和降温培养工艺,实现在5 L搅拌式生物反应器规模的瞬时基因表达。首先比较了两种表达载体在CHO-S细胞中的瞬时基因表达,发现pID-mAAFP载体介导的瞬时转染表达可以获得mAAFP的高水平表达。采用pID-mAAFP载体在1.3 L搅拌式生物反应器中进行瞬时基因表达,与分批培养相比,通过流加和降温培养工艺对转染过程进行控制,培养时间延长1倍,蛋白表达浓度提高8倍。在5 L搅拌式生物反应器中对上述工艺进行成功放大,mAAFP质量浓度达到25 mg/L。之后,利用rProtein A Sepharose Fast Flow介质纯化mAAFP,并通过SDS-PAGE、Western blot验证其纯度,为下一步利用小鼠模型研究蛋白功效和作用机理做好准备。

  20. 非小细胞肺癌间变性淋巴瘤激酶融合基因的荧光原位杂交检测规范化流程探讨%Standard protocol of ALK fusion gene assessment by fluorescent in situ hybridization in non-small cell lung cancer

    Institute of Scientific and Technical Information of China (English)

    郭蕾; 郑闪; 谢永强


    Objective To investigate the standard protocol for anaplastic lymphoma kinase (ALK) fusion gene assessment by fluorescent in situ hybridization (FISH) in non-small cell lung cancer (NSCLC).Methods Tissue specimens of NSCLC cases were retrospectively collected from Jan.2011 to July 2012.ALK fusion gene was examined by FISH using break-apart ALK gene probes (Vysis company).The identification of ALK fusion gene was determined by fluorescent signals under a fluorescence microscope.Results One hundred and forty-six eligible NSCLC tumor specimens were tested for ALK fusion gene by FISH.The specimens included 110 cases (75.4%) of surgically-removed tissues,11 cases (7.5%) of biopsy,19 cases (13.0%) of lymph node and 6 cases (4.1%) of other metastatic tissues.The positivity of ALK fusion gene was 8.9% (13/146).Conclusions The assessment of ALK fusing gene by FISH using standard protocol for formalin-fixed,paraffin-embedded (FFPE) tissue is feasible.The protocol can used to test in surgically-removed tissues,biopsies,metastatic lymph nodes and other metastastic specimens.%目的 探讨非小细胞肺癌(NSCLC)患者荧光原位杂交(FISH)方法检测间变性淋巴瘤激酶(ALK)融合基因的操作规范.方法 收集中国医学科学肿瘤医院病理科2011年1月至2012年7月共146例NSCLC患者肿瘤病理组织学标本,应用Vysis公司的ALK基因断裂探针、采用FISH方法对ALK融合基因进行检测,荧光显微镜下根据荧光信号进行结果判读.结果 146例合格NSCLC患者病理组织学标本,其中手术切除标本110例(75.4%),肺活检标本11例(7.5%),淋巴结转移标本19例(13.0%),其他转移灶6例(4.1%).ALK融合基因阳性率为8.9% (13/146).结论 采用标准化的流程进行病理组织标本切片、切片预处理、FISH方法检测ALK融合基因是可行的.该流程适用于检测包括手术切除和支气管镜活检标本、淋巴结及其他转移灶标本在内的常见标本ALK融合基因的检测.

  1. J亚群禽白血病病毒 SUJ及兔 IgGFc基因在腺病毒表达系统中的融合表达%Fusion expression of subgroup J avian leukosis virus gp85 gene with rabbit IgGFc gene in adenovirus expression system and its application

    Institute of Scientific and Technical Information of China (English)

    梅梅; 钱科; 唐应华; 秦爱建; 侯继波


    To obtain the fusion protein of subgroup J avian leukosis virus ( ALV-J) envelope protein gp85 and rabbit IgGFc, ALV-J gp85 gene fused with rabbit IgGFc was expressed by adenovirus expression system. The ALV-J gp85 ( SU) and rabbit IgGFc ( rIgGFc) gene were digested from plasmid pcDNA3. 1-SUJ-rIgGFc with XhoⅠand KpnⅠand cloned into pShuttle-CMV to construct the recombinant plasmid pShuttle-CMV-SUJ-rIgGFc which was transformed into BJ5183-AD-1 competent cells containing pAdeasy-1 to construct the recombinant adenovirus plasmid pAd-SUJ-rIgGFc. The recombinant adenovirus was obtained by transfecting the recom-binant plasmid pAd-SUJ-rIgGFc into 293T cells. The fusion pro-tein can be recognized by mAb JE9 specific to gp85 of ALV-J and antibodies against rabbit IgG with the molecular weight about 9. 5× 104. The result of Co-IP showed that the fusion protein SUJ-rIgGFc reacted with the membrane proteins from DF1 cells and produced differential proteins, indicative of the specific re-action between fused protein and the protein from host cells of ALV-J.%为获得J亚群禽白血病病毒( ALV-J)囊膜蛋白gp85和兔IgGFc的融合蛋白,通过腺病毒表达系统融合表达ALV-J SUJ和兔IgGFc( rIgGFc)基因。将pcDNA3.1-SUJ-rIgGFc用XhoⅠ、KpnⅠ进行双酶切,获得SUJ-rIgGFc基因,将其克隆至 pShuttle-CMV 质粒,构建穿梭载体 pShuttle-CMV-SUJ-rIgGFc,重组质粒转化含 pAdeasy-1的BJ5183-AD-1感受态细胞,获得重组腺病毒质粒 pAd-SUJ-rIgGFc,最后转染人293T细胞。表达的融合蛋白可被ALV-J单克隆抗体JE9以及羊抗兔IgG识别,重组蛋白的分子量大小约9.5×104,且与JE9及羊抗兔IgG都有很好的反应性;免疫共沉淀试验结果显示融合蛋白可与DF1细胞膜蛋白混合物反应并获得差异蛋白。该融合蛋白可与ALV-J宿主细胞蛋白发生特异性反应。

  2. Inertial fusion energy; L'energie de fusion inertielle

    Energy Technology Data Exchange (ETDEWEB)

    Decroisette, M.; Andre, M.; Bayer, C.; Juraszek, D. [CEA Bruyeres-le-Chatel, Dir. des Systemes d' Information (CEA/DIF), 91 (France); Le Garrec, B. [CEA Centre d' Etudes Scientifiques et Techniques d' Aquitaine, 33 - Le Barp (France); Deutsch, C. [Paris-11 Univ., 91 - Orsay (France); Migus, A. [Institut d' Optique Centre scientifique, 91 - Orsay (France)


    We first recall the scientific basis of inertial fusion and then describe a generic fusion reactor with the different components: the driver, the fusion chamber, the material treatment unit, the target factory and the turbines. We analyse the options proposed at the present time for the driver and for target irradiation scheme giving the state of art for each approach. We conclude by the presentation of LMJ (laser Megajoule) and NIF (national ignition facility) projects. These facilities aim to demonstrate the feasibility of laboratory DT ignition, first step toward Inertial Fusion Energy. (authors)

  3. A universal assay for detection of oncogenic fusion transcripts by oligo microarray analysis

    Directory of Open Access Journals (Sweden)

    Ribeiro Franclim R


    Full Text Available Abstract Background The ability to detect neoplasia-specific fusion genes is important not only in cancer research, but also increasingly in clinical settings to ensure that correct diagnosis is made and the optimal treatment is chosen. However, the available methodologies to detect such fusions all have their distinct short-comings. Results We describe a novel oligonucleotide microarray strategy whereby one can screen for all known oncogenic fusion transcripts in a single experiment. To accomplish this, we combine measurements of chimeric transcript junctions with exon-wise measurements of individual fusion partners. To demonstrate the usefulness of the approach, we designed a DNA microarray containing 68,861 oligonucleotide probes that includes oligos covering all combinations of chimeric exon-exon junctions from 275 pairs of fusion genes, as well as sets of oligos internal to all the exons of the fusion partners. Using this array, proof of principle was demonstrated by detection of known fusion genes (such as TCF3:PBX1, ETV6:RUNX1, and TMPRSS2:ERG from all six positive controls consisting of leukemia cell lines and prostate cancer biopsies. Conclusion This new method bears promise of an important complement to currently used diagnostic and research tools for the detection of fusion genes in neoplastic diseases.

  4. Computational Plasma Physics at the Bleeding Edge: Simulating Kinetic Turbulence Dynamics in Fusion Energy Sciences (United States)

    Tang, William


    Advanced computing is generally recognized to be an increasingly vital tool for accelerating progress in scientific research in the 21st Century. The imperative is to translate the combination of the rapid advances in super-computing power together with the emergence of effective new algorithms and computational methodologies to help enable corresponding increases in the physics fidelity and the performance of the scientific codes used to model complex physical systems. If properly validated against experimental measurements and verified with mathematical tests and computational benchmarks, these codes can provide more reliable predictive capability for the behavior of complex systems, including fusion energy relevant high temperature plasmas. The magnetic fusion energy research community has made excellent progress in developing advanced codes for which computer run-time and problem size scale very well with the number of processors on massively parallel supercomputers. A good example is the effective usage of the full power of modern leadership class computational platforms from the terascale to the petascale and beyond to produce nonlinear particle-in-cell simulations which have accelerated progress in understanding the nature of plasma turbulence in magnetically-confined high temperature plasmas. Illustrative results provide great encouragement for being able to include increasingly realistic dynamics in extreme-scale computing campaigns to enable predictive simulations with unprecedented physics fidelity. Some illustrative examples will be presented of the algorithmic progress from the magnetic fusion energy sciences area in dealing with low memory per core extreme scale computing challenges for the current top 3 supercomputers worldwide. These include advanced CPU systems (such as the IBM-Blue-Gene-Q system and the Fujitsu K Machine) as well as the GPU-CPU hybrid system (Titan).

  5. Transferred BCR/ABL DNA from K562 extracellular vesicles causes chronic myeloid leukemia in immunodeficient mice.

    Directory of Open Access Journals (Sweden)

    Jin Cai

    Full Text Available Our previous study showed that besides mRNAs and microRNAs, there are DNA fragments within extracellular vesicles (EVs. The BCR/ABL hybrid gene, involved in the pathogenesis of chronic myeloid leukemia (CML, could be transferred from K562 EVs to neutrophils and decrease their phagocytic activity in vitro. Our present study provides evidence that BCR/ABL DNAs transferred from EVs have pathophysiological significance in vivo. Two months after injection of K562 EVs into the tail vein of Sprague-Dawley (SD rats, they showed some characteristics of CML, e.g., feeble, febrile, and thin, with splenomegaly and neutrophilia but with reduced neutrophil phagocytic activity. These findings were also observed in immunodeficient NOD/SCID mice treated with K562 EVs; BCR/ABL mRNA and protein were found in their neutrophils. The administration of actinomycin D, an inhibitor of de novo mRNA synthesis, prevented the abnormalities caused by K562 EVs in NOD/SCID mice related to CML, including neutrophilia and bone marrow hyperplasia. As a specific inhibitor of tyrosine kinases, imatinib blocked the activity of tyrosine kinases and the expression of phospho-Crkl, induced by the de novo BCR/ABL protein caused by K562 EVs bearing BCR/ABL DNA. Our current study shows the pathophysiological significance of transferred tumor gene from EVs in vivo, which may represent an important mechanism for tumorigenesis, tumor progression, and metastasis.

  6. BCR-ABL isoforms associated with intrinsic or acquired resistance to imatinib : more heterogeneous than just ABL kinase domain point mutations?

    NARCIS (Netherlands)

    Gruber, Franz X.; Lundan, Tuija; Goll, Rasmus; Silye, Aleksandra; Mikkola, Ingvild; Rekvig, Ole Petter; Knuutila, Sakari; Remes, Kari; Gedde-Dahl, Tobias; Porkka, Kimmo; Hjorth-Hansen, Henrik


    Imatinib, a small molecule inhibitor of ABL, PDGFR and C-KIT, has revolutionized treatment of chronic myeloid leukaemia (CML). However, resistance to treatment is of increasing importance and often is due to point mutations in the Abl kinase domain (Abl KD). Here, we analysed clinical outcome and mu

  7. Adenovirus mediated fusion gene system driven by KDR promoter kills selectively pancreatic cancer cells%双自杀基因重组腺病毒对胰腺癌细胞特异性杀伤作用

    Institute of Scientific and Technical Information of China (English)

    闫振宇; 陈旭; 孔恒; 黄宗海; 俞金龙; 厉周


    were analyzed by MTT. The pathological character of ceils was observed with electron microscopy and distribution of cell cycle was detected by flow cytometry. An animal model of human pancreatic cancer in nude mice was repro-duced. Ad-KDR-CD/TK was injected directly into the tumor. Then,the nude mice were given intrap-inhibition rates for tumor growth were also calculated. Results The infection rate of the resultant re-combinant Ad to Capan-2 and HepG2 cells was not apparently different,and it increased gradually with the addition of multiple of infection (MOI) of Ads. RT-PCR demonstrated that there existed the prod-uet of CDTK gene in Capan-2 cell infected by Ad-KDR-CDTK. Prodrug could inhibit proliferation of Capan-2 and the effect was dose-dependent,while the infected HepG2 cells were not. There was con-siderable bystander effect by MTT. Apoptotic peak was also shown by flow cytometry. Morphologic features of apoptosis in Capan-2 cells were displayed via electron microscopy. The analysis of cell cycle revealed that the rate of cells at the G0-G1 phase was increased and the rate at the G2-M and S phase was decreased by treatment with pro-drug. Furthermore, the tumors decreased in size during treatment of double suicide gene system by establish subcutaneous transplanted model. Conelusion The CDTK fusion gene system controlled by KDR promoter have selectively killing effects on the KDR-expressing Capan-2 cells and inducing the cell apoptosis. It can significantly inhibit the growth of xenografted pancreatic cancer in nude mice.

  8. Fc fusion as a platform technology: potential for modulating immunogenicity. (United States)

    Levin, Ditza; Golding, Basil; Strome, Scott E; Sauna, Zuben E


    The platform technology of fragment crystallizable (Fc) fusion, in which the Fc region of an antibody is genetically linked to an active protein drug, is among the most successful of a new generation of bioengineering strategies. Immunogenicity is a critical safety concern in the development of any protein therapeutic. While the therapeutic goal of generating Fc-fusion proteins has been to extend half-life, there is a critical mass of literature from immunology indicating that appropriate design of the Fc component has the potential to engage the immune system for product-specific outcomes. In the context of Fc-fusion therapeutics, a review of progress in understanding Fc biology suggests the prospect of engineering products that have an extended half-life and are able to modulate the immune system.

  9. Fusion Rings for Quantum Groups

    DEFF Research Database (Denmark)

    Andersen, Henning Haahr; Stroppel, Catharina


    We study the fusion rings of tilting modules for a quantum group at a root of unity modulo the tensor ideal of negligible tilting modules. We identify them in type A with the combinatorial rings from Korff, C., Stroppel, C.: The sl(ˆn)k-WZNW fusion ring: a combinato-rial construction and a realis......We study the fusion rings of tilting modules for a quantum group at a root of unity modulo the tensor ideal of negligible tilting modules. We identify them in type A with the combinatorial rings from Korff, C., Stroppel, C.: The sl(ˆn)k-WZNW fusion ring: a combinato-rial construction...

  10. Cold nuclear fusion

    Energy Technology Data Exchange (ETDEWEB)

    Tsyganov, E.N., E-mail: [Cold Fusion Power, International (United States); Bavizhev, M.D. [LLC “Radium”, Moscow (Russian Federation); Buryakov, M.G. [Joint Institute for Nuclear Research (JINR), Dubna (Russian Federation); Dabagov, S.B. [RAS P.N. Lebedev Physical Institute, Leninsky pr. 53, 119991 Moscow (Russian Federation); National Research Nuclear University MEPhI, Kashirskoe shosse 31, 115409 Moscow (Russian Federation); Golovatyuk, V.M.; Lobastov, S.P. [Joint Institute for Nuclear Research (JINR), Dubna (Russian Federation)


    If target deuterium atoms were implanted in a metal crystal in accelerator experiments, a sharp increase in the probability of DD-fusion reaction was clearly observed when compared with the reaction’s theoretical value. The electronic screening potential, which for a collision of free deuterium atoms is about 27 eV, reached 300–700 eV in the case of the DD-fusion in metallic crystals. These data leads to the conclusion that a ban must exist for deuterium atoms to be in the ground state 1s in a niche filled with free conduction electrons. At the same time, the state 2p whose energy level is only 10 eV above that of state 1s is allowed in these conditions. With anisotropy of 2p, 3p or above orbitals, their spatial positions are strictly determined in the lattice coordinate system. When filling out the same potential niches with two deuterium atoms in the states 2p, 3p or higher, the nuclei of these atoms can be permanently positioned without creating much Coulomb repulsion at a very short distance from each other. In this case, the transparency of the potential barrier increases dramatically compared to the ground state 1s for these atoms. The probability of the deuterium nuclei penetrating the Coulomb barrier by zero quantum vibration of the DD-system also increases dramatically. The so-called cold nuclear DD-fusion for a number of years was registered in many experiments, however, was still rejected by mainstream science for allegedly having no consistent scientific explanation. Finally, it received the validation. Below, we outline the concept of this explanation and give the necessary calculations. This paper also considers the further destiny of the formed intermediate state of {sup 4}He{sup ∗}.

  11. Fusion Advanced Design Studies

    Energy Technology Data Exchange (ETDEWEB)

    El-Guebaly, Laila; Henderson, Douglass; Wilson, Paul; Blanchard, Jake


    During the January 1, 2013 – December 31, 2015 contract period, the UW Fusion Technology Institute personnel have actively participated in the ARIES-ACT and FESS-FNSF projects, led the nuclear and thermostructural tasks, attended several project meetings, and participated in all conference calls. The main areas of effort and technical achievements include updating and documenting the nuclear analysis for ARIES-ACT1, performing nuclear analysis for ARIES-ACT2, performing thermostructural analysis for ARIES divertor, performing disruption analysis for ARIES vacuum vessel, and developing blanket testing strategy and Materials Test Module for FNSF.

  12. miR-29b suppresses CML cell proliferation and induces apoptosis via regulation of BCR/ABL1 protein

    Energy Technology Data Exchange (ETDEWEB)

    Li, Yajuan; Wang, Haixia; Tao, Kun [Department of Clinical Hematology, Key Laboratory of Laboratory Medical Diagnostics Designated of Ministry of Education, Chongqing Medical University, 1 Yixueyuan Road, Yuzhong District, Chongqing 400016 (China); Xiao, Qing [Department of Hematology, The First Affiliated Hospital, Chongqing Medical University, 1 Yixueyuan Road, Yuzhong District, Chongqing 400016 (China); Huang, Zhenglan; Zhong, Liang; Cao, Weixi [Department of Clinical Hematology, Key Laboratory of Laboratory Medical Diagnostics Designated of Ministry of Education, Chongqing Medical University, 1 Yixueyuan Road, Yuzhong District, Chongqing 400016 (China); Wen, Jianping [Department of Pathology and Molecular Medicine, McMaster University, Hamilton, Ontario, Canada L8N 3Z5 (Canada); Feng, Wenli, E-mail: [Department of Clinical Hematology, Key Laboratory of Laboratory Medical Diagnostics Designated of Ministry of Education, Chongqing Medical University, 1 Yixueyuan Road, Yuzhong District, Chongqing 400016 (China)


    MicroRNAs (miRNAs) are small RNAs that regulate gene expression posttranscriptionally and are critical for many cellular pathways. Recent evidence has shown that aberrant miRNA expression profiles and unique miRNA signaling pathways are present in many cancers. Here, we demonstrate that miR-29b is markedly lower expressed in CML patient samples. Bioinformatics analysis reveals a conserved target site for miR-29b in the 3′-untranslated region (UTR) of ABL1. miR-29b significantly suppresses the activity of a luciferase reporter containing ABL1-3′UTR and this activity is not observed in cells transfected with mutated ABL1-3′UTR. Enforced expression of miR-29b in K562 cells inhibits cell growth and colony formation ability thereby inducing apoptosis through cleavage of procaspase 3 and PARP. Furthermore, K562 cells transfected with a siRNA targeting ABL1 show similar growth and apoptosis phenotypes as cells overexpression of miR-29b. Collectively, our results suggest that miR-29b may function as a tumor suppressor by targeting ABL1 and BCR/ABL1. - Highlights: ► miR-29b expression was downregulated in CML patients. ► ABL1 was identified as a direct target gene of miR-29b. ► Enforced expression of miR-29b inhibits cell proliferation and induces apoptosis. ► miR-29b might be a therapeutic target to CML.

  13. MicroRNA-125a-5p modulates human cervical carcinoma proliferation and migration by targeting ABL2

    Directory of Open Access Journals (Sweden)

    Qin X


    Full Text Available Xian Qin,1 Yajun Wan,1 Saiying Wang,2 Min Xue1 1Department of Obstetrics and Gynecology, 2Department of Anesthesiology, Third Xiangya Hospital, Central South University, Changsha, People’s Republic of China Background: In this study, we intended to understand the regulatory mechanisms of microRNA-125a-5p (miR-125a-5p in human cervical carcinoma.Methods: The gene expressions of miR-125a-5p in seven cervical carcinoma cell lines and 12 human cervical carcinoma samples were evaluated by quantitative real-time reverse transcription polymerase chain reaction. Ca-Ski and HeLa cells were transduced with lentivirus carrying miR-125a-5p mimics, and the effects of lentivirus-induced miR-125a-5p upregulation on cervical carcinoma proliferation and migration were examined by 3-(4,5-dimethylthiazol-2-yl-2,5-diphenyltetrazolium bromide and transwell assays, respectively. In additional, HeLa cells were inoculated into null mice to evaluate the effect of miR-125a-5p upregulation on in vivo cervical carcinoma growth. The direct regulation of miR-125a-5p on its target gene, ABL proto-oncogene 2 (ABL2, in cervical carcinoma was evaluated by quantitative real-time reverse transcription polymerase chain reaction, Western blotting and luciferase reporter assays, respectively. ABL2 was then downregulated by small interfering RNA to examine its effect on cervical carcinoma proliferation and migration.Results: miR-125a-5p was downregulated in both cervical carcinoma cell lines and human cervical carcinomas. In Ca-Ski and HeLa cells, lentivirus-mediated miR-125a-5p upregulation inhibited cancer proliferation and migration in vitro and cervical carcinoma transplantation in vivo. ABL2 was shown to be directly targeted by miR-125a-5p. In cervical carcinoma, ABL2 gene and protein levels were both downregulated by miR-125a-5p. Small interfering RNA-mediated ABL2 downregulation also had tumor-suppressive effects on cervical carcinoma proliferation and migration

  14. EML4-ALK阳性表达非小细胞肺癌患者的临床特征及治疗现状%Clinicopathologic characteristics and treatment of non-small cell lung cancer with EML4-ALK fusion gene

    Institute of Scientific and Technical Information of China (English)

    孙霞; 魏嘉; 刘宝瑞


    棘皮动物微管相关类蛋白4(EML4)与间变性淋巴瘤激酶(ALK)融合基因首次被发现存在于部分非小细胞肺癌中.该融合基因是由2号染色体上2区1带和2区3带易位形成,可以诱导肿瘤生成,而ALK抑制剂能够拮抗其促肿瘤生成活性.本文旨在介绍EML4-ALK基因阳性表达肺癌患者的临床特征及该基因在肺癌诊断、治疗中的意义.%The echinoderm microtubule-associated protein-like 4-anaplastic lymphoma kinase(EML4-ALK) fusion gene resulting from the chromosome inversion inv(2) (p21 ;p23) recently was identified in non-small cell lung cancer. EML4-ALK fusion gene is oncogenic, which could be suppressed by ALK-inhibitor through blocking the downstream signaling pass-way of EML4-ALK. This review will focus on the clinicopathologic characteristics and targeted therapy of EML4-ALK in lung cancer.

  15. Ten-a affects the fusion of central complex primordia in Drosophila.

    Directory of Open Access Journals (Sweden)

    Xuebo Cheng

    Full Text Available The central complex of Drosophila melanogaster plays important functions in various behaviors, such as visual and olfactory memory, visual orientation, sleep, and movement control. However little is known about the genes regulating the development of the central complex. Here we report that a mutant gene affecting central complex morphology, cbd (central brain defect, was mapped to ten-a, a type II trans-membrane protein coding gene. Down-regulation of ten-a in pan-neural cells contributed to abnormal morphology of central complex. Over-expression of ten-a by C767-Gal4 was able to partially restore the abnormal central complex morphology in the cbd mutant. Tracking the development of FB primordia revealed that C767-Gal4 labeled interhemispheric junction that separated fan-shaped body precursors at larval stage withdrew to allow the fusion of the precursors. While the C767-Gal4 labeled structure did not withdraw properly and detached from FB primordia, the two fan-shaped body precursors failed to fuse in the cbd mutant. We propose that the withdrawal of C767-Gal4 labeled structure is related to the formation of the fan-shaped body. Our result revealed the function of ten-a in central brain development, and possible cellular mechanism underlying Drosophila fan-shaped body formation.

  16. Prospective molecular monitoring of BCR/ABL transcript in children with Ph+ acute lymphoblastic leukaemia unravels differences in treatment response. (United States)

    Cazzaniga, Giovanni; Lanciotti, Marina; Rossi, Vincenzo; Di Martino, Daniela; Aricò, Maurizio; Valsecchi, Maria Grazia; Basso, Giuseppe; Masera, Giuseppe; Micalizzi, Concetta; Biondi, Andrea


    Children with Philadelphia-chromosome-positive (Ph+) acute lymphoblastic leukaemia (ALL) represent a subgroup at very high risk for treatment failure, despite intensive chemotherapy. However, recent retrospective studies showed that Ph+ childhood ALL is a heterogeneous disease with regard to treatment response. We have prospectively monitored, by reverse transcription polymerase chain reaction (RT-PCR) during follow-up, the presence of the BCR/ABL fusion transcript in Ph+ ALL children diagnosed in the Italian multicentre Associazione Italiana Ematologia Oncologia Pediatrica ALL-AIEOP-95 therapy protocol. To our knowledge, this is the first report on the evaluation of minimal residual disease (MRD) in childhood Ph+ ALL prospectively enrolled in an intensive, Berlin-Frankfurt-Munster (BFM)-type treatment protocol. Twenty-seven of 36 (75.0%) Ph+ patients consecutively enrolled into the high-risk group of the AIEOP-ALL protocol between May 1995 and October 1999 were successfully analysed. Twenty were good responders to the pre-phase of prednisone/intrathecal methotrexate treatment (PGR) and seven were poor responders (PPR). Within the PPR group, the RT-PCR monitoring constantly showed positivity for the BCR/ABL fusion transcript and all the patients died of disease progression. In contrast, highly sensitive qualitative RT-PCR monitoring revealed heterogeneity within the PGR group of Ph+ childhood ALL patients. Three different subgroups could be defined, according to the clearance of Ph+ cells within the first 5 months of treatment. This provides useful information on the capability of chemotherapy to reduce the leukaemic clone, with prognostic implications.


    Institute of Scientific and Technical Information of China (English)

    于翠娟; 孟艳玲; 桂俊豪; 赵晶; 金明; 王智; 王成济; 杨安钢


    Objective: To obtain the recombinant fusion AIF genes inserted into the eukaryotic expression vector Pires2-EGFP, to observe the expression and location of the fusion AIF genes (3NE: PE(280-358)-AIFΔ1-120, and 4NE: PE(280-364)-AIFΔ1-120), and to detect and compare their apoptosis inducing effects on the transfected HeLa cells. Methods: Full-length human AIF gene was cloned by RT-PCR, and its N-terminal mitochondrial localization sequence (MLS) was replaced by part sequence of Psuedomonas exotoxin A (PE) translocation domain (PEII(280-358/364)), then the recombinant fusion genes were inserted into the Pires2-EGFP eukaryotic expression vector. After these genes were transiently transfected into HeLa cells with LipofectAmine, the expression of the recombinant fusion AIF genes and their effects on HeLa cells were detected by fluorescent microscopy, laser confocal microscopy and electron microscopy. Results: The eukaryotic expression vectors containing the recombinant fusion AIF genes (Pires2-EGFP-PEII(280-358/364)- AIFΔ1- 120) were constructed successfully. It was demonstrated that the fusion AIF protein genes were expressed effectively in the transfected cells, with the GFP comco-expressed in cells by indirect immunofluorescence staining analysis. After transfection, expression of the genes could induce HeLa cells to exhibit the typical apoptosis features: such as plasma membrane blebbing and peripheral chromatin condensation. As compared with control groups, the untreated cells and the void vector transfected cells, the living cell number of the AIF gene transfected cells reduced distinctly. Conclusion: Our data prove that the expression of the recombinant human AIF fusion genes could induce apoptosis in transfected HeLa cells, which provides new strategy for cancer killing.

  18. 抗Ⅰ型糖尿病融合基因HSP65-6×P277在烟草中的表达%Expression of anti-type I diabetes fusion gene HSP65-6×P277 in tobacco

    Institute of Scientific and Technical Information of China (English)

    郑卿; 郭书巧; 杨洁; 吴洁; 刘景晶; 倪万潮


    With the purpose of studying the expression of reeombinant gene HSP65-6×P277 in plant, the plant constitutive expression vector pCAMBlA2301-HSP65-6×5xre77 containing fusion gene HSP65-6×P277 was constructed and transformed into tobacco plants via Agrobacterium tumefaciens. Sixteen resistant plants containing anti-type I diabetes gene were obtained by screening with kanamysin. The resistant plants were analyzed by PCR, RT-PCR and SDS-PAGE. The results showed that the fusion gene HSP65-6×P277 was expressed in the resistant tobacco plants on DNA, mRNA and protein levels.%为了研究抗Ⅰ型糖尿病融合蛋白质基因HSP65-6×P277在植物中的表达,构建了含有融合基因HSP65-6×P277的植物表达载体pCAM BIA 2301-HSP65-6×P277,通过农杆菌介导法将重组载体转入烟草,利用卡那霉素作为植物筛选标记,获得含有抗Ⅰ型糖尿病基因的16株转基因烟草植株.通过PCR、半定量RT-PCR及SDSPAGE蛋白质电泳等方法检测,初步验证在抗性烟草植株中融合基因HSP65-6×P277在mRNA和蛋白质水平上进行了表达.

  19. Inverse regulation of bridging integrator 1 and BCR-ABL1 in chronic myeloid leukemia. (United States)

    Trino, Stefania; De Luca, Luciana; Simeon, Vittorio; Laurenzana, Ilaria; Morano, Annalisa; Caivano, Antonella; La Rocca, Francesco; Pietrantuono, Giuseppe; Bianchino, Gabriella; Grieco, Vitina; Signorino, Elisabetta; Fragasso, Alberto; Bochicchio, Maria Teresa; Venturi, Claudia; Rosti, Gianantonio; Martinelli, Giovanni; Del Vecchio, Luigi; Cilloni, Daniela; Musto, Pellegrino


    Endocytosis is the major regulator process of tyrosine kinase receptor (RTK) functional activities. Bridging integrator 1 (BIN1) is a key protein involved in RTK intracellular trafficking. Here, we report, by studying 34 patients with chronic myeloid leukemia (CML) at diagnosis, that BIN1 gene is downregulated in CML as compared to healthy controls, suggesting an altered endocytosis of RTKs. Rab interactor 1 (RIN1), an activator of BIN1, displayed a similar behavior. Treatment of 57 patients by tyrosine kinase inhibitors caused, along with BCR-ABL1 inactivation, an increase of BIN1 and RIN1 expression, potentially restoring endocytosis. There was a significant inverse correlation between BIN1-RIN1 and BCR-ABL1 expression. In vitro experiments on both CML and nontumorigenic cell lines treated with Imatinib confirmed these results. In order to provide another proof in favor of BIN1 and RIN1 endocytosis function in CML, we demonstrated that Imatinib induced, in K562 cell line, BIN1-RIN1 upregulation accompanied by a parallel AXL receptor internalization into cytoplasmic compartment. This study shows a novel deregulated mechanism in CML patients, indicating BIN1 and RIN1 as players in the maintenance of the abnormal RTK signaling in this hematological disease.

  20. Utility requirements for fusion

    Energy Technology Data Exchange (ETDEWEB)

    Vondrasek, R.J.


    This report describes work done and results obtained during performance of Task 1 of a study of Utility Requirements and Criteria for Fusion Options. The work consisted of developing a list of utility requirements for fusion optics containing definition of the requirements and showing their relative importance to the utility industry. The project team members developed a preliminary list which was refined by discussions and literature searches. The refined list was recast as a questionnaire which was sent to a substantial portion of the utility industry in this country. Forty-three questionnaire recipients responded including thirty-two utilities. A workshop was held to develop a revised requirements list using the survey responses as a major input. The list prepared by the workshop was further refined by a panel consisting of vice presidents of the three project team firms. The results of the study indicate that in addition to considering the cost of energy for a power plant, utilities consider twenty-three other requirements. Four of the requirements were judged to be vital to plant acceptability: Plant Capital Cost, Financial Liability, Plant Safety and Licensability.

  1. JENDL fusion file 99

    Energy Technology Data Exchange (ETDEWEB)

    Chiba, Satoshi; Fukahori, Tokio; Shibata, Keiichi [Japan Atomic Energy Research Inst., Tokai, Ibaraki (Japan). Tokai Research Establishment; Yu Baosheng [China Institute of Atomic Energy, Beijing (China); Kosako, Kazuaki [Sumitomo Atomic Industries, Tokyo (Japan); Yamamuro, Nobuhiro [Data Engineering Co. Ltd., Yokohama, Kanagawa (Japan)


    The double-differential cross sections (DDXs) of secondary neutrons have been evaluated for 79 isotopes and 13 natural elements ranging from H to Bi to improve the accuracy of predictions for the neutronics calculations in the D-T thermonuclear fusion applications. The data given in JENDL-3.1, which was the newest version of JENDL general purpose file when this project was initiated, was combined with new calculations based on the optical model, DWBA, pre-equilibrium and multi-step statistical models, and the DDX data were generated based on various kinds of systematics for medium-mass nuclei. Different methods were employed for light nuclei to which the above method could not be applied. In addition, the DDXs for emission of charged particles (p, d, t, {sup 3}He and {alpha}-particle) were given for {sup 2}H, {sup 9}Be and elements heavier or equal to F. The present results give an overall good description of the measured DDX data of both the neutron and charged particles emission channels. The data were compiled in ENDF-6 format, and released in 1999 as a special purpose file of JENDL family, namely, JENDL Fusion File 99. (author)

  2. Clean steels for fusion

    Energy Technology Data Exchange (ETDEWEB)

    Gelles, D.S.


    Fusion energy production has an inherent advantage over fission: a fuel supply with reduced long term radioactivity. One of the leading candidate materials for structural applications in a fusion reactor is a tungsten stabilized 9% chromium Martensitic steel. This alloy class is being considered because it offers the opportunity to maintain that advantage in the reactor structure as well as provide good high temperature strength and radiation induced swelling and embrittlement resistance. However, calculations indicate that to obtain acceptable radioactivity levels within 500 years after service, clean steel will be required because the niobium impurity levels must be kept below about 2 appm and nickel, molybdenum, nitrogen, copper, and aluminum must be intentionally restricted. International efforts are addressing the problems of clean steel production. Recently, a 5,000 kg heat was vacuum induction melted in Japan using high purity commercial raw materials giving niobium levels less than 0.7 appm. This paper reviews the need for reduced long term radioactivity, defines the advantageous properties of the tungsten stabilized Martensitic steel class, and describes the international efforts to produce acceptable clean steels.

  3. Homeland security application of the Army Soft Target Exploitation and Fusion (STEF) system (United States)

    Antony, Richard T.; Karakowski, Joseph A.


    A fusion system that accommodates both text-based extracted information along with more conventional sensor-derived input has been developed and demonstrated in a terrorist attack scenario as part of the Empire Challenge (EC) 09 Exercise. Although the fusion system was developed to support Army military analysts, the system, based on a set of foundational fusion principles, has direct applicability to department of homeland security (DHS) & defense, law enforcement, and other applications. Several novel fusion technologies and applications were demonstrated in EC09. One such technology is location normalization that accommodates both fuzzy semantic expressions such as behind Library A, across the street from the market place, as well as traditional spatial representations. Additionally, the fusion system provides a range of fusion products not supported by traditional fusion algorithms. Many of these additional capabilities have direct applicability to DHS. A formal test of the fusion system was performed during the EC09 exercise. The system demonstrated that it was able to (1) automatically form tracks, (2) help analysts visualize behavior of individuals over time, (3) link key individuals based on both explicit message-based information as well as discovered (fusion-derived) implicit relationships, and (4) suggest possible individuals of interest based on their association with High Value Individuals (HVI) and user-defined key locations.

  4. Fusion of NUP98 and the SET binding protein 1 (SETBP1) gene in a paediatric acute T cell lymphoblastic leukaemia with t(11;18)(p15;q12)

    DEFF Research Database (Denmark)

    Panagopoulos, Ioannis; Kerndrup, Gitte; Carlsen, Niels


    Three NUP98 chimaeras have previously been reported in T cell acute lymphoblastic leukaemia (T-ALL): NUP98/ADD3, NUP98/CCDC28A, and NUP98/RAP1GDS1. We report a T-ALL with t(11;18)(p15;q12) resulting in a novel NUP98 fusion. Fluorescent in situ hybridisation showed NUP98 and SET binding protein 1......(SETBP1) fusion signals; other analyses showed that exon 12 of NUP98 was fused in-frame with exon 5 of SETBP1. Nested polymerase chain reaction did not amplify the reciprocal SETBP1/NUP98, suggesting that NUP98/SETBP1 transcript is pathogenetically important. SETBP1 has previously not been implicated...

  5. Gene fusions with human carbonic anhydrase II for efficient expression and rapid single-step recovery of recombinant proteins: expression of the Escherichia coli F1-ATPase epsilon subunit. (United States)

    Van Heeke, G; Shaw, R; Schnizer, R; Couton, J M; Schuster, S M; Wagner, F W


    A new expression vector was constructed which allows the overproduction in Escherichia coli of tripartite proteins consisting of human carbonic anhydrase isozyme II (hCAII), a peptide linker containing an enterokinase cleavage site, and a target protein of interest. Carbonic anhydrase is soluble and stable in E. coli and serves as a highly specific purification tag in the recovery of the fusion protein by a single affinity chromatography step. The enterokinase cleavage site was engineered into the construct to allow accurate and efficient release of the target protein. To demonstrate the practical value of this vector, the E. coli F1-ATPase epsilon subunit was expressed as a fusion with hCAII. After a single purification step, biologically active recombinant E. coli F1-ATPase epsilon subunit was recovered following proteolytic removal of the hCAII moiety.

  6. Application of optical tweezers and excimer laser to study protoplast fusion (United States)

    Kantawang, Titirat; Samipak, Sompid; Limtrakul, Jumras; Chattham, Nattaporn


    Protoplast fusion is a physical phenomenon that two protoplasts come in contact and fuse together. Doing so, it is possible to combine specific genes from one protoplast to another during fusion such as drought resistance and disease resistance. There are a few possible methods to induce protoplast fusion, for example, electrofusion and chemical fusion. In this study, chemical fusion was performed with laser applied as an external force to enhance rate of fusion and observed under a microscope. Optical tweezers (1064 nm with 100X objective N.A. 1.3) and excimer laser (308 nm LMU-40X-UVB objective) were set with a Nikon Ti-U inverted microscope. Samples were prepared by soaking in hypertonic solution in order to induce cell plasmolysis. Elodea Canadensis and Allium cepa plasmolysed leaves were cut and observed under microscope. Concentration of solution was varied to induce difference turgor pressures on protoplasts pushing at cell wall. Free protoplasts in solution were trapped by optical tweezers to study the effect of Polyethylene glycol (PEG) solution. PEG was diluted by Ca+ solution during the process to induced protoplast cell contact and fusion. Possibility of protoplast fusion by excimer laser was investigated and found possible. Here we report a novel tool for plant cell fusion using excimer laser. Plant growth after cell fusion is currently conducted.

  7. The status of cold fusion (United States)

    Storms, E.

    This report attempts to update the status of the phenomenon of cold fusion. The new field is continuing to grow as a variety of nuclear reactions are discovered to occur in a variety of chemical environments at modest temperatures. However, it must be cautioned that most scientists consider cold fusion as something akin to UFO's, ESP, and numerology.

  8. The quest for fusion power (United States)

    Cowley, Steven C.


    Fusion power is one of a very few sustainable options to replace fossil fuels as the world's primary energy source. Although the conditions for fusion have been reached, much remains to be done to turn scientific success into commercial electrical power.

  9. Fusion Policy Advisory Committee (FPAC)

    Energy Technology Data Exchange (ETDEWEB)


    This document is the final report of the Fusion Policy Advisory Committee. The report conveys the Committee's views on the matters specified by the Secretary in his charge and subsequent letters to the Committee, and also satisfies the provisions of Section 7 of the Magnetic Fusion Energy Engineering Act of 1980, Public Law 96-386, which require a triennial review of the conduct of the national Magnetic Fusion Energy program. Three sub-Committee's were established to address the large number of topics associated with fusion research and development. One considered magnetic fusion energy, a second considered inertial fusion energy, and the third considered issues common to both. For many reasons, the promise of nuclear fusion as a safe, environmentally benign, and affordable source of energy is bright. At the present state of knowledge, however, it is uncertain that this promise will become reality. Only a vigorous, well planned and well executed program of research and development will yield the needed information. The Committee recommends that the US commit to a plan that will resolve this critically important issue. It also outlines the first steps in a development process that will lead to a fusion Demonstration Power Plant by 2025. The recommended program is aggressive, but we believe the goal is reasonable and attainable. International collaboration at a significant level is an important element in the plan.

  10. Fusion research programme in India

    Indian Academy of Sciences (India)

    Shishir Deshpande; Predhiman Kaw


    The fusion energy research program of India is summarized in the context of energy needs and scenario of tokamak advancements on domestic and international fronts. In particular, the various technologies that will lead us to ultimately build a fusion power reactor are identified along with the steps being taken for their indigenous development.