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Sample records for c-myc affects mrna

  1. c-Myc affects mRNA translation, cell proliferation and progenitor cell function in the mammary gland

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    Trumpp Andreas

    2009-09-01

    Full Text Available Abstract Background The oncoprotein c-Myc has been intensely studied in breast cancer and mouse mammary tumor models, but relatively little is known about the normal physiological role of c-Myc in the mammary gland. Here we investigated functions of c-Myc during mouse mammary gland development using a conditional knockout approach. Results Generation of c-mycfl/fl mice carrying the mammary gland-specific WAPiCre transgene resulted in c-Myc loss in alveolar epithelial cells starting in mid-pregnancy. Three major phenotypes were observed in glands of mutant mice. First, c-Myc-deficient alveolar cells had a slower proliferative response at the start of pregnancy, causing a delay but not a block of alveolar development. Second, while milk composition was comparable between wild type and mutant animals, milk production was reduced in mutant glands, leading to slower pup weight-gain. Electron microscopy and polysome fractionation revealed a general decrease in translational efficiency. Furthermore, analysis of mRNA distribution along the polysome gradient demonstrated that this effect was specific for mRNAs whose protein products are involved in milk synthesis. Moreover, quantitative reverse transcription-polymerase chain reaction analysis revealed decreased levels of ribosomal RNAs and ribosomal protein-encoding mRNAs in mutant glands. Third, using the mammary transplantation technique to functionally identify alveolar progenitor cells, we observed that the mutant epithelium has a reduced ability to repopulate the gland when transplanted into NOD/SCID recipients. Conclusion We have demonstrated that c-Myc plays multiple roles in the mouse mammary gland during pregnancy and lactation. c-Myc loss delayed, but did not block proliferation and differentiation in pregnancy. During lactation, lower levels of ribosomal RNAs and proteins were present and translation was generally decreased in mutant glands. Finally, the transplantation studies suggest a role

  2. Systematic analysis of the contribution of c-myc mRNA constituents upon cap and IRES mediated translation.

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    Meristoudis, Christos; Trangas, Theoni; Lambrianidou, Andromachi; Papadopoulos, Vasilios; Dimitriadis, Euthymios; Courtis, Nelly; Ioannidis, Panayotis

    2015-12-01

    Fine tuning of c-MYC expression is critical for its action and is achieved by several regulatory mechanisms. The contribution of c-myc mRNA regulatory sequences on its translational control has been investigated individually. However, putative interactions have not been addressed so far. The effect of these interactions upon the translatability of monocistronic and bicistronic chimaeric mRNAs, carrying combinations of the c-myc mRNA 5'-untranlated region (UTR), 3'-UTR, and coding region instability element (CRD) was investigated on this study. The presence of the 5'-UTR induced an increase in translatability of 50%. The presence of the CRD element, when in frame, reduced translatability by approximately 50%, regardless of the expression levels of the wild type CRD- binding protein (CRD-BP/IMP1). Conversely, overexpression of a mutated CRD-BP/IMP1 (Y396F) further impeded translation of the chimaeric mRNAs carrying its cognate sequences. The presence of the c-myc 3'-UTR increased translatability by approximately 300% affecting both cap and c-myc internal ribosome entry site (IRES) mediated translation. In addition, 3'-UTR rescued the cap mediated translation in the presence of the polyadenylation inhibitor cordycepin. Furthermore, the 3'-UTR rescued cap mediated translation under metabolic stress conditions and this was enhanced in the absence of a long poly (A) tail.

  3. Regulation of c-Myc mRNA by L11 in Response to UV and Gamma Irradiation

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    2014-12-01

    L11 mutations are associated with cleft palate and abnormal thumbs in Diamond-Blackfan anemia patients. Am. J. Hum. Genet. 83:769–780. 23. Gomez-Roman...24 (Figure 1). We also tested an array of miRNAs possessing tumor suppressor functions for L11 binding miR-16 miR-1248 miR-3944 (-) miR-191 miR...32), and tristetraprolin (TTP) (39), have been found to bind to c-myc AREs and regulate c-myc mRNA sta- bility. To test whether L11 binds to c-myc

  4. Sensitive electrochemiluminescence detection of c-Myc mRNA in breast cancer cells on a wireless bipolar electrode.

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    Wu, Mei-Sheng; Qian, Guang-sheng; Xu, Jing-Juan; Chen, Hong-Yuan

    2012-06-19

    We report an ultrasensitive wireless electrochemiluminescence (ECL) protocol for the detection of a nucleic acid target in tumor cells on an indium tin oxide bipolar electrode (BPE) in a poly(dimethylsiloxane) microchannel. The approach is based on the modification of the anodic pole of the BPE with antisense DNA as the recognition element, Ru(bpy)(3)(2+)-conjugated silica nanoparticles (RuSi@Ru(bpy)(3)(2+)) as the signal amplification tag, and reporter DNA as a reference standard. It employs the hybridization-induced changes of RuSi@Ru(bpy)(3)(2+) ECL efficiency for the specific detection of reporter DNA released from tumor cells. Prior to ECL detection, tumor cells are transfected with CdSe@ZnS quantum dot (QD)-antisense DNA/reporter DNA conjugates. Upon the selective binding of antisense DNA probes to intracellular target mRNA, reporter DNA will be released from the QDs, which indicates the amount of the target mRNA. The proof of concept is demonstrated using a proto-oncogene c-Myc mRNA in MCF-7 cells (breast cancer cell line) as a model target. The wireless ECL biosensor exhibited excellent ECL signals which showed a good linear range over 2 × 10(-16) to 1 × 10(-11) M toward the reporter DNA detection and could accurately quantify c-Myc mRNA copy numbers in living cells. C-Myc mRNA in each MCF-7 cell and LO2 cell was estimated to be 2203 and 13 copies, respectively. This wireless ECL strategy provides great promise in a miniaturized device and may facilitate the achievement of point of care testing.

  5. Specific regulation of c-myc oncogene expression in a murine B-cell lymphoma.

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    McCormack, J E; Pepe, V H; Kent, R B; Dean, M; Marshak-Rothstein, A; Sonenshein, G E

    1984-01-01

    The c-myc oncogene has been implicated in a wide spectrum of B-cell neoplasias. In normal cells, the level of expression of the c-myc gene correlates with growth status. In the present study, we examined the effect of receptor-mediated inhibition of growth on c-myc expression in a B-cell lymphoma. The murine lymphoma line WEHI 231 has been characterized as an early B cell; it bears surface-bound IgM and has unrearranged c-myc genes. Following treatment of a WEHI 231 culture with anti-mouse Ig antiserum, the cells undergo one round of division and further proliferation is inhibited. We observed that this treatment specifically affected cytoplasmic levels of c-myc mRNA. An initial early increase is followed by a precipitous drop such that by 4 hr (after exposure) the amount of c-myc mRNA is below control values by a factor of approximately equal to 10. The drop in c-myc precedes cessation of DNA synthesis. During the 2- to 4-hr period, c-myc mRNA had a maximal half-life of between 20 and 30 min. In contrast, even 24 hr after anti-Ig exposure, the amounts of most major mRNAs, including mu heavy chain and actin, were not significantly altered. These results indicate that expression of an unrearranged c-myc gene can be selectively responsive to receptor-mediated regulatory events. Images PMID:6206500

  6. Electrochemiluminescence detection of c-Myc mRNA in breast cancer cells on a wireless bipolar electrode.

    Science.gov (United States)

    Wu, Mei-Sheng; Qian, Guang-sheng; Xu, Jing-Juan; Chen, Hong-Yuan

    2013-01-01

    Electrochemiluminescence (ECL) on bipolar electrode (BPE) is a sensitive, portable, and low-cost approach which has been employed to detect DNA and proteins. Here, we develop an ultrasensitive method for intracellular mRNA assay based on mRNA-mediated reporter DNA liberation and Ru(bpy)3(2+)-conjugated silica nanoparticles (RuSi@Ru(bpy)3(2+)) tag-based signal amplification.

  7. "Retroposon" insertion into the cellular oncogene c-myc in canine transmissible venereal tumor.

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    Katzir, N; Rechavi, G; Cohen, J B; Unger, T; Simoni, F; Segal, S; Cohen, D; Givol, D

    1985-01-01

    We examined by Southern blotting the state of the cellular oncogene c-myc in the dog transmissible venereal tumor. The tumor DNA contains a 16.8-kilobase pair (kbp) rearranged c-myc fragment in addition to the normal 15-kbp and 7.5-kbp fragments. We compared the structure of the cloned rearranged c-myc (re-myc) with that of a cloned normal c-myc and found that the rearrangement was due to the insertion of a 1.8-kbp DNA upstream to the first exon of c-myc. The inserted DNA is flanked by 10-base-pair direct repeats and contains a dA-rich tail, suggesting its origin from mRNA. Partial sequence of the inserted element showed 62% homology with the primate interdispersed Kpn I repetitive element. These results provide an example for the behavior of repetitive DNA sequences like the Kpn I family, as movable elements that can transpose nearby to oncogenes or other structural genes and perhaps affect their activity. Images PMID:2983328

  8. Humanized c-Myc mouse.

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    Frank M Lehmann

    Full Text Available BACKGROUND: A given tumor is usually dependent on the oncogene that is activated in the respective tumor entity. This phenomenon called oncogene addiction provides the rationale for attempts to target oncogene products in a therapeutic manner, be it by small molecules, by small interfering RNAs (siRNA or by antigen-specific T cells. As the proto-oncogene product is required also for the function of normal cells, this raises the question whether there is a therapeutic window between the adverse effects of specific inhibitors or T cells to normal tissue that may limit their application, and their beneficial tumor-specific therapeutic action. To address this crucial question, suitable mouse strains need to be developed, that enable expression of the human proto-oncogene not only in tumor but also in normal cells. The aim of this work is to provide such a mouse strain for the human proto-oncogene product c-MYC. PRINCIPAL FINDINGS: We generated C57BL/6-derived embryonic stem cells that are transgenic for a humanized c-Myc gene and established a mouse strain (hc-Myc that expresses human c-MYC instead of the murine ortholog. These transgenic animals harbor the humanized c-Myc gene integrated into the endogenous murine c-Myc locus. Despite the lack of the endogenous murine c-Myc gene, homozygous mice show a normal phenotype indicating that human c-MYC can replace its murine ortholog. CONCLUSIONS: The newly established hc-Myc mouse strain provides a model system to study in detail the adverse effects of therapies that target the human c-MYC protein. To mimic the clinical situation, hc-Myc mice may be cross-bred to mice that develop tumors due to overexpression of human c-MYC. With these double transgenic mice it will be possible to study simultaneously the therapeutic efficiency and adverse side effects of MYC-specific therapies in the same mouse.

  9. [The expression of oncogene c-myc and its role on human laryngeal cancer].

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    Long, Xiaobo; Hu, Shuang; Cao, Pingping; Liu, Zheng; Zhen, Hongtao; Cui, Yonghua

    2009-12-01

    To detect the expression of oncogene c-myc and explore its role on human laryngeal cancer. The mRNA and of oncogene c-myc levels were detected in human laryngeal cancer tissues and laryngeal normal tissues by the means of real-time PCR and immunohistochemistry, respectively. Compared with normal laryngeal tissues, the mRNA and protein levels of oncogene c-myc were both upregulated in laryngeal cancer tissues of all differentiation degree (P<0.05). Moreover, the level of c-myc was inverse correlated with the differentiation degree in human laryngeal cancer tissues. The oncogene c-myc is overexpressed in human laryngeal cancer tissues, and c-myc may play an important role in carcinogenesis and progression of human laryngeal cancer tissues. It may provide a new target for gene therapy of human laryngeal cancer.

  10. Constitutive gray hair in mice induced by melanocyte-specific deletion of c-Myc.

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    Pshenichnaya, Irina; Schouwey, Karine; Armaro, Marzia; Larue, Lionel; Knoepfler, Paul S; Eisenman, Robert N; Trumpp, Andreas; Delmas, Véronique; Beermann, Friedrich

    2012-05-01

    c-Myc is involved in the control of diverse cellular processes and implicated in the maintenance of different tissues including the neural crest. Here, we report that c-Myc is particularly important for pigment cell development and homeostasis. Targeting c-Myc specifically in the melanocyte lineage using the floxed allele of c-Myc and Tyr::Cre transgenic mice results in a congenital gray hair phenotype. The gray coat color is associated with a reduced number of functional melanocytes in the hair bulb and melanocyte stem cells in the hair bulge. Importantly, the gray phenotype does not progress with time, suggesting that maintenance of the melanocyte through the hair cycle does not involve c-Myc function. In embryos, at E13.5, c-Myc-deficient melanocyte precursors are affected in proliferation in concordance with a reduction in numbers, showing that c-Myc is required for the proper melanocyte development. Interestingly, melanocytes from c-Myc-deficient mice display elevated levels of the c-Myc paralog N-Myc. Double deletion of c-Myc and N-Myc results in nearly complete loss of the residual pigmentation, indicating that N-Myc is capable of compensating for c-Myc loss of function in melanocytes. © 2012 John Wiley & Sons A/S.

  11. Overexpression of C-MYC oncogene in prostate cancer predicts biochemical recurrence.

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    Hawksworth, D; Ravindranath, L; Chen, Y; Furusato, B; Sesterhenn, I A; McLeod, D G; Srivastava, S; Petrovics, G

    2010-12-01

    Alterations of chromosome 8, including amplification at 8q24 harboring the C-MYC oncogene, have been noted as one of the most common chromosomal abnormalities in prostate cancer (CaP) progression. However, the frequency of C-MYC alterations in CaP has remained uncertain. A recent study, using a new anti-MYC antibody, described prevalent upregulation of nuclear C-MYC protein expression as an early oncogenic alteration in CaP. Further, we have recently reported regulation of C-MYC expression by ERG and a significant correlation between C-MYC overexpression and TMPRSS2-ERG fusion in early stage CaP. These emerging data suggest that increased C-MYC expression may be a critical and early oncogenic event driving CaP progression. In this study, we assessed whether C-MYC mRNA overexpression in primary prostate tumors was predictive of more aggressive tumor or disease progression. Our approach was to quantitatively determine C-MYC mRNA expression levels in laser capture micro-dissected tumor cells and matched benign epithelial cells in a radical prostatectomy cohort with long follow-up data available. On the basis of our results, we conclude that elevated C-MYC expression in primary prostate tumor is biologically relevant and may be a predictor of future biochemical recurrence.

  12. Thrombopoietin (TPO) induces c-myc expression through a PI3K- and MAPK-dependent pathway that is not mediated by Akt, PKCzeta or mTOR in TPO-dependent cell lines and primary megakaryocytes.

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    Chanprasert, Supantitra; Geddis, Amy E; Barroga, Charlene; Fox, Norma E; Kaushansky, Kenneth

    2006-08-01

    Thrombopoietin (TPO) and its receptor (c-Mpl) are the major regulators of megakaryocyte and platelet production and serve a critical and non-redundant role in hematopoietic stem cell (HSC) biology. TPO signals through the Jak-STAT, Ras-Raf-MAPK, and PI3K pathways, and promotes survival, proliferation, and polyploidization in megakaryocytes. The proto-oncogene c-myc also plays an important role in many of these same processes. In this work we studied the regulated expression of c-myc in megakaryocytic cell lines and primary cells by quantitative real-time RT-PCR. We found that TPO induced expression of c-myc in 1 h in both hematopoietic cell lines (UT-7 and BaF3/Mpl) and mature murine megakaryocytes. The TPO-induced expression of c-myc was blocked by a phosphatidylinositol 3-kinase (PI3K) inhibitor, suggesting that TPO stimulated c-myc expression through a PI3K-dependent pathway. Of interest, our study showed that overexpression of active Akt did not rescue the effect of PI3K blockade on c-myc expression, rather, enhanced it. In addition, inhibitors of protein kinase C (PKC)zeta and the target of rapamycin (mTOR) also failed to affect c-myc mRNA expression, while c-myc mRNA expression was reduced by inhibition of the mitogen activated protein kinase (MAPK) pathway. Therefore, we conclude that TPO stimulates c-myc expression in primary megakaryocytes through a PI3K- and MAPK-dependent pathway that is not mediated by Akt, PKCzeta or mTOR.

  13. Metformin targets c-MYC oncogene to prevent prostate cancer.

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    Akinyeke, Tunde; Matsumura, Satoko; Wang, Xinying; Wu, Yingjie; Schalfer, Eric D; Saxena, Anjana; Yan, Wenbo; Logan, Susan K; Li, Xin

    2013-12-01

    Prostate cancer (PCa) is the second leading cause of cancer-related death in American men and many PCa patients develop skeletal metastasis. Current treatment modalities for metastatic PCa are mostly palliative with poor prognosis. Epidemiological studies indicated that patients receiving the diabetic drug metformin have lower PCa risk and better prognosis, suggesting that metformin may have antineoplastic effects. The mechanism by which metformin acts as chemopreventive agent to impede PCa initiation and progression is unknown. The amplification of c-MYC oncogene plays a key role in early prostate epithelia cell transformation and PCa growth. The purpose of this study is to investigate the effect of metformin on c-myc expression and PCa progression. Our results demonstrated that (i) in Hi-Myc mice that display murine prostate neoplasia and highly resemble the progression of human prostate tumors, metformin attenuated the development of prostate intraepithelial neoplasia (PIN, the precancerous lesion of prostate) and PCa lesions. (ii) Metformin reduced c-myc protein levels in vivo and in vitro. In Myc-CaP mouse PCa cells, metformin decreased c-myc protein levels by at least 50%. (iii) Metformin selectively inhibited the growth of PCa cells by stimulating cell cycle arrest and apoptosis without affecting the growth of normal prostatic epithelial cells (RWPE-1). (iv) Reduced PIN formation by metformin was associated with reduced levels of androgen receptor and proliferation marker Ki-67 in Hi-Myc mouse prostate glands. Our novel findings suggest that by downregulating c-myc, metformin can act as a chemopreventive agent to restrict prostatic neoplasia initiation and transformation. Metformin, an old antidiabetes drug, may inhibit prostate intraepithelial neoplasia transforming to cancer lesion via reducing c-MYC, an 'old' overexpressed oncogene. This study explores chemopreventive efficacy of metformin in prostate cancer and its link to cMYC in vitro and in vivo.

  14. Elevated c-myc protooncogene expression in autosomal recessive polycystic kidney disease

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    Cowley, B.D. Jr.; Smardo, F.L. Jr.; Grantham, J.J.; Calvet, J.P.

    1987-12-01

    The polycystic kidney diseases (PKDs) are a group of disorders characterized by the growth of epithelial cysts from the nephrons and collecting ducts of kidney tubules. The diseases can be inherited or can be provoked by environmental factors. To investigate the molecular basis of the abnormal cell growth associated with PKD, c-myc protooncogene expression was studied in a mouse model for autosomal recessive PKD. Homozygous recessive C57BL/6J (cpk/cpk) mice develop massively enlarged cystic kidneys and die from renal failure shortly after 3 weeks of age. Quantitative dot blot and RNA blot hybridization experiments in which whole kidney poly(A)/sup +/ RNA was hybridized with a c-myc RNA probe showed a 2- to 6-fold increase in c-myc mRNA at 2 weeks, and a 25- to 30-fold increase in c-myc mRNA at 3 weeks of age in polycystic mice, as compared to normal littermates. c-myc expression was also examined under two conditions in which kidney cell growth was experimentally induced in normal adult mice: compensatory renal hypertrophy and tubule regeneration following folic acid-induced renal cell injury. While compensatory hypertrophy resulted in only a small increase in c-myc, folic acid treatment gave rise after 24 hr to a 12-fold increase in c-myc RNA. The induction of c-myc by folic acid is consistent with increased cellular proliferation regenerating tubules. In contrast, polycystic kidneys show only a minimal increase in cellular proliferation over that seen in normal kidneys, while c-myc levels were found to be markedly elevated. Thus, the level of c-myc expression in cystic kidneys appears to be out of proportion to the rate of cell division, suggesting that elevated and potentially abnormal c-myc expression may be involved in the pathogenesis of PKD.

  15. p53 inhibits the upregulation of sirtuin 1 expression induced by c-Myc.

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    Yuan, Fang; Liu, Lu; Lei, Yonghong; Tang, Peifu

    2017-10-01

    Sirtuin 1 (Sirt1), a conserved NAD(+) dependent deacetylase, is a mediator of life span by calorie restriction. However, Sirt1 may paradoxically increase the risk of cancer. Accordingly, the expression level of Sirt1 is selectively elevated in numerous types of cancer cell; however, the mechanisms underlying the differential regulation remain largely unknown. The present study demonstrated that oncoprotein c-Myc was a direct regulator of Sirt1, which accounts for the upregulation of Sirt1 expression only in the cells without functional p53. In p53 deficient cells, the overexpression of c-Myc increased Sirt1 mRNA and protein expression levels as well as its promoter activity, whereas the inhibitor of c-Myc, 10058-F4, induced decreased Sirt1 basal mRNA and protein expression levels. Deletion/mutation mapping analyses revealed that c-Myc bound to the conserved E-box[-189 to -183 base pair (bp)] of the Sirt1 promoter. In addition, p53 and c-Myc shared at least response element and the presence of p53 may block the binding of c-Myc to the Sirt1 promoter, thus inhibit the c-Myc mediated upregulation of Sirt1 promoter activity. The present study indicated that the expression level of Sirt1 was tightly regulated by oncoprotein c-Myc and tumor suppressor p53, which aids an improved understanding of its expression regulation and tumor promoter role in certain conditions.

  16. c-myc, c-fos, and c-jun regulation in the regenerating livers of normal and H-2K/c-myc transgenic mice.

    OpenAIRE

    Morello, D; Fitzgerald, M J; Babinet, C; Fausto, N

    1990-01-01

    We investigated the mechanisms of regulation of c-myc, c-fos, and c-jun at the early stages of liver regeneration in mice. We show that the transient increase in steady-state levels of c-myc mRNA at the start of liver regeneration is most probably regulated by posttranscriptional mechanisms. Although there was a marked increase in c-myc transcriptional initiation shortly after partial hepatectomy, a block in elongation prevented the completion of most transcripts. To gain further information ...

  17. [Inhibition of proliferation in MCF-7 breast cancer cells by plasmid-based siRNA targeting to oncogene c-myc].

    Science.gov (United States)

    Zhou, Chang-Hua; Peng, Xiao-Dong; Wu, Jing; Zhang, Ping; Zhao, Zong-Rong; Wei, Da-Peng; Zhang, Chong-Jie

    2008-05-01

    To investigate the effects of plasmid-based siRNA targeting to oncogene c-myc on c-myc/ c-Myc expressions and cells proliferation in MCF-7 breast cancer cells. siRNA eukaryotic expression plasmid p-Mat01-1 targeting to the sequence 589-609 of oncogene c-myc and its mismatch plasmid p-Mis09-1 were constructed, and transiently transfected MCF-7 cells using Lipo2000. Semi-quantitative RT-PCR and Western blot were used to analyze the expressions of c-myc/c-Myc in MCF-7 cells, and cells proliferation was detected by MTT assay. p-Mat01-1 inhibited the expressions of c-myc mRNA (24 h: P < 0.01) and c-Myc protein (5 d. P < 0.01) in MCF-7 cells as compared with pEGFP-C1 and p-Mis09-1 controls, and suppressed the proliferation of MCF-7 cells significantly (3 d: P < 0.05, 5, 7 d: P < 0. 01). Plasmid-based siRNA targeting to oncogene c-myc could inhibit the expressions of c-myc/c-Myc in MCF-7 breast cancer cells efficiently, suggesting that the downregulation of c-myc/c-Myc could suppress the proliferation of MCF-7 cells in vitro.

  18. [Role of microRNA-223 and its target gene oncogene c-myc in hepatocellular carcinoma pathogenesis].

    Science.gov (United States)

    Zhao, Wen-Yue; Wang, Dong-Dong; Song, Meng-Qi; Yang, Ling; Ye, Jin; Chen, Li-Bo

    2011-02-01

    To investigate the regulatory role of microRNA-223 (miR-223) on c-myc and its role in hepatocarcinogenesis. miR-223 and c-myc mRNA expressions in normal tissue, paraneoplastic tissue, liver cancer tissue and liver cancer cells were tested with microRNA microarray and quantitative real-time PCR (qRT-PCR). C-myc protein expression was detected by Western blot. MiR-223 mimic was transfected into HepG2 cells and the expression changes of c-myc mRNA and protein were tested with qRT-PCR and Western blot respectively. MiR-223 was down-regulated by 61.53% and 30.77% respectively in hepatocellular carcinoma and adjacent tissues as compared to normal liver tissues and the expression of miR-223 was also decreased in HepG2 cell as compared to fetal liver cells L02, whereas the expressions of c-myc mRNA and protein increased in paraneoplastic and HCC tissues compared with normal liver tissues. It prompts that the expressions of miR-223 and c-myc are negatively correlated. No obvious difference found among c-myc mRNA expressions after miR-223 mimics transfection. The c-myc abnormal high-expression may play a dynamic role in hepatocarcinogenesis due to the miR-223 down-regulation.

  19. Upregulation of the oncogene c-myc in Barrett’s adenocarcinoma: induction of c-myc by acidified bile acid in vitro

    Science.gov (United States)

    Tselepis, C; Morris, C D; Wakelin, D; Hardy, R; Perry, I; Luong, Q T; Harper, E; Harrison, R; Attwood, S E A; Jankowski, J A Z

    2003-01-01

    Background and aims: C-myc over expression is implicated in malignancy although to date this has not been studied in Barrett’s metaplasia. We sought to determine c-myc expression in the malignant progression of Barrett’s metaplasia and whether it may be induced by bile acids seen in gastro-oesophageal refluxate. Methods: C-myc protein and mRNA levels were assessed in 20 Barrett’s metaplasia and 20 oesophageal adenocarcinoma samples by western blotting and real time polymerase chain reaction. Levels of c-myc and proliferation were also assessed in cell lines OE21, OE33, SW-480, and TE-7 stimulated with pulses or continuous exposure to the bile acids deoxycholic acid and chenodeoxycholic acid. Results: C-myc protein was upregulated in 50% of Barrett’s metaplasia and 90% of oesophageal adenocarcinoma samples compared with squamous, gastric, and duodenal controls. C-myc immunolocalisation in Barrett’s metaplasia revealed discrete nuclear localisation, becoming more diffuse with progression from low to high grade dysplasia to adenocarcinoma. Both continual and pulsed bile acid induced c-myc at pH 4, with no effect at pH 7 or with acidified media alone. Pulsed bile acid treatment induced proliferation (p<0.05); in contrast, continuous exposure led to suppression of proliferation (p<0.05). Conclusions: We have shown upregulation of c-myc with malignant progression of Barrett’s metaplasia and suggest that acidified bile may be a novel agent responsible for induction of this oncogene. PMID:12524396

  20. Increased transcription of the c-myc oncogene in two methylcholanthrene-induced quail fibroblastic cell lines

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    Saule, S.; Martin, P.; Gegonne, A.; Begue, A.; Lagrou, C.; Stehelin, D.

    1984-12-01

    The expression of three c-onc genes (c-erb, c-myc, c-myb) was investigated in five cell lines established from fibrosarcomas induced with 20-methylcholanthrene (MCA) of Japanese quails. These cell lines showed low levels of the three c-onc genes, with the exception of two cell lines that accumulated moderate (MCAQ 1-4) and large amounts (MCAQ 3-5) of c-myc RNA. Molecular cloning and restriction endonuclease analyses indicated that expression of c-myc in these two cell lines were not associated with detectable rearrangements in the c-myc locus, that the size of the c-myc transcript (2.7 kb) in MCAQ 3-5 was similar to that of the normal c-myc messenger RNAs (mRNA) and that the transcriptional activatin observed in MCAQ 3-5 was not mediated by the LTR (long terminal repeat) of a proximate ALV (avian leukosis virus) provirus. Finally, when analyzed with the restriction enzymes Msp I and Hpa II, the c-myc locus of MCAQ 3-5 and MCAQ 1-4 was found hypomethylated as compared with that of the other cell lines tested that show low levels of c-myc transcripts. Results suggest that one of the ways methylcholanthrene could mediate transformation is by inducing an abnormal regulation of the c-myc gene.

  1. Antiproliferative effects of antisense oligonucleotides directed to the RNA of c-myc oncogene.

    Science.gov (United States)

    Degols, G; Leonetti, J P; Mechti, N; Lebleu, B

    1991-01-01

    Several groups have reported the use of antisense oligonucleotides to inhibit c-myc gene expression and study its biological role. However high concentrations of free oligonucleotides were generally needed. To lower their concentration and stabilize the antisense effect against c-myc, oligonucleotides were covalently linked to poly(L-lysine) and administered in ternary complexes formed with heparin (100 micrograms/ml). A sequence specific growth inhibition was observed at concentrations lower than 1 microM, while oligonucleotide-poly(L-lysine) conjugates alone were inefficient. Similar results occurred with other polyanionic compounds. Inhibition of proliferation was correlated to a reduction of c-myc protein and to a transient decrease in c-myc mRNA level. However, implication of RNase H in this process could not be demonstrated. Images PMID:1708128

  2. Posttranscriptional regulation of cellular gene expression by the c-myc oncogene

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    Prendergast, G.C.; Cole, M.D. (Princeton Univ., NJ (USA). Dept. of Biology)

    1989-01-01

    The c-myc oncogene has been implicated in the development of many different cancers, yet the mechanism by which the c-myc protein alters cellular growth control has proven elusive. The authors used a cDNA hybridization difference assay to isolate two genes, mr1 and mr2, that were constitutively expressed (i.e., deregulated) in rodent fibroblast cell lines immortalized by transfection of a viral promoter-linked c-myc gene. Both cDNAs were serum inducible in quiescent G/sub o/ fibroblasts, suggesting that they are functionally related to cellular proliferative processes. Although there were significant differences in cytoplasmic mRNA levels between myc-immortalized and control cells, the rates of transcription and mRNA turnover of both genes were similar, suggesting that c-myc regulates mr1 and mr2 expression by some nuclear posttranscriptional mechanism. Their results provide evidence that c-myc can rapidly modulate cellular gene expression and suggest that c-myc may function in gene regulation at the level of RNA export, splicing, or nuclear RNA turnover.

  3. Antiproliferative protein Tob directly regulates c-myc proto-oncogene expression through cytoplasmic polyadenylation element-binding protein CPEB.

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    Ogami, K; Hosoda, N; Funakoshi, Y; Hoshino, S

    2014-01-02

    The regulation of mRNA deadenylation constitutes a pivotal mechanism of the post-transcriptional control of gene expression. Here we show that the antiproliferative protein Tob, a component of the Caf1-Ccr4 deadenylase complex, is involved in regulating the expression of the proto-oncogene c-myc. The c-myc mRNA contains cis elements (CPEs) in its 3'-untranslated region (3'-UTR), which are recognized by the cytoplasmic polyadenylation element-binding protein (CPEB). CPEB recruits Caf1 deadenylase through interaction with Tob to form a ternary complex, CPEB-Tob-Caf1, and negatively regulates the expression of c-myc by accelerating the deadenylation and decay of its mRNA. In quiescent cells, c-myc mRNA is destabilized by the trans-acting complex (CPEB-Tob-Caf1), while in cells stimulated by the serum, both Tob and Caf1 are released from CPEB, and c-Myc expression is induced early after stimulation by the stabilization of its mRNA as an 'immediate-early gene'. Collectively, these results indicate that Tob is a key factor in the regulation of c-myc gene expression, which is essential for cell growth. Thus, Tob appears to function in the control of cell growth at least, in part, by regulating the expression of c-myc.

  4. Domain-specific c-Myc ubiquitylation controls c-Myc transcriptional and apoptotic activity

    Science.gov (United States)

    Zhang, Qin; Spears, Erick; Boone, David N.; Li, Zhaoliang; Gregory, Mark A.; Hann, Stephen R.

    2013-01-01

    The oncogenic transcription factor c-Myc causes transformation and tumorigenesis, but it can also induce apoptotic cell death. Although tumor suppressors are necessary for c-Myc to induce apoptosis, the pathways and mechanisms are unclear. To further understand how c-Myc switches from an oncogenic protein to an apoptotic protein, we examined the mechanism of p53-independent c-Myc–induced apoptosis. We show that the tumor suppressor protein ARF mediates this switch by inhibiting ubiquitylation of the c-Myc transcriptional domain (TD). Whereas TD ubiquitylation is critical for c-Myc canonical transcriptional activity and transformation, inhibition of ubiquitylation leads to the induction of the noncanonical c-Myc target gene, Egr1, which is essential for efficient c-Myc–induced p53-independent apoptosis. ARF inhibits the interaction of c-Myc with the E3 ubiquitin ligase Skp2. Overexpression of Skp2, which occurs in many human tumors, inhibits the recruitment of ARF to the Egr1 promoter, leading to inhibition of c-Myc–induced apoptosis. Therapeutic strategies could be developed to activate this intrinsic apoptotic activity of c-Myc to inhibit tumorigenesis. PMID:23277542

  5. Linc-RoR promotes c-Myc expression through hnRNP I and AUF1.

    Science.gov (United States)

    Huang, Jianguo; Zhang, Ali; Ho, Tsui-Ting; Zhang, Ziqiang; Zhou, Nanjiang; Ding, Xianfeng; Zhang, Xu; Xu, Min; Mo, Yin-Yuan

    2016-04-20

    Linc-RoR was originally identified to be a regulator for induced pluripotent stem cells in humans and it has also been implicated in tumorigenesis. However, the underlying mechanism of Linc-RoR-mediated gene expression in cancer is poorly understood. The present study demonstrates that Linc-RoR plays an oncogenic role in part through regulation of c-Myc expression. Linc-RoR knockout (KO) suppresses cell proliferation and tumor growth. In particular, Linc-RoR KO causes a significant decrease in c-Myc whereas re-expression of Linc-RoR in the KO cells restores the level of c-Myc. Mechanistically, Linc-RoR interacts with heterogeneous nuclear ribonucleoprotein (hnRNP) I and AU-rich element RNA-binding protein 1 (AUF1), respectively, with an opposite consequence to their interaction with c-Myc mRNA. While Linc-RoR is required for hnRNP I to bind to c-Myc mRNA, interaction of Linc-RoR with AUF1 inhibits AUF1 to bind to c-Myc mRNA. As a result, Linc-RoR may contribute to the increased stability of c-Myc mRNA. Although hnRNP I and AUF1 can interact with many RNA species and regulate their functions, with involvement of Linc-RoR they would be able to selectively regulate mRNA stability of specific genes such as c-Myc. Together, these results support a role for Linc-RoR in c-Myc expression in part by specifically enhancing its mRNA stability, leading to cell proliferation and tumorigenesis. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

  6. Cooverexpression of EpCAM and c-myc genes in malignant breast ...

    Indian Academy of Sciences (India)

    oncogene, affects progression, treatment, and diagnosis of many adenocarcinomas. C-myc has been shown to be a downstream target of EpCAM and is also one of the most important proto-oncogenes routinely overexpressed in breast cancer.

  7. Detection of the c-myc oncogene product in colonic polyps and carcinomas.

    Science.gov (United States)

    Stewart, J.; Evan, G.; Watson, J.; Sikora, K.

    1986-01-01

    The c-myc oncogene has been implicated in the processes of normal cell proliferation and differentiation. Elevated levels of c-myc mRNA and its gene product (p62c-myc), have been detected in a variety of solid tumours and cultured cel lines. Its precise role in normal cell function and in neoplastic transformation and progression has yet to be elucidated. We have used a monoclonal antibody, raised by peptide immunisation, to determine the distribution by immunoperoxidase staining of the c-myc oncogene product in archival specimens of colonic polyps and carcinomas. Samples from 42 patients with colon carcinoma, 24 with benign polyps and 15 normal colon biopsies were examined. Normal colon revealed maximum staining in the mid-zone of the crypts, corresponding to the zone of differentiation and maturation. The staining was predominantly cytoplasmic. Adenomatous polyps revealed the most intense pattern of staining in areas of dysplastic change. Colonic tumours showed a wide range of staining. Well differentiated tumours contained more cytoplasmic p62c-myc than poorly differentiated tumours. These findings suggest that the c-myc oncogene product may play an important role in the evolution of colonic neoplasia. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:3511934

  8. ADA3 regulates normal and tumor mammary epithelial cell proliferation through c-MYC.

    Science.gov (United States)

    Griffin, Nicolas I; Sharma, Gayatri; Zhao, Xiangshan; Mirza, Sameer; Srivastava, Shashank; Dave, Bhavana J; Aleskandarany, Mohammed; Rakha, Emad; Mohibi, Shakur; Band, Hamid; Band, Vimla

    2016-11-16

    We have established the critical role of ADA3 as a coactivator of estrogen receptor (ER), as well as its role in cell cycle progression. Furthermore, we showed that ADA3 is predominantly nuclear in mammary epithelium, and in ER+, but is cytoplasmic in ER- breast cancers, the latter correlating with poor survival. However, the role of nuclear ADA3 in human mammary epithelial cells (hMECs), and in ER+ breast cancer cells, as well as the importance of ADA3 expression in relation to patient prognosis and survival in ER+ breast cancer have remained uncharacterized. We overexpressed ADA3 in hMECs or in ER+ breast cancer cells and assessed the effect on cell proliferation. The expression of ADA3 was analyzed then correlated with the expression of various prognostic markers, as well as survival of breast cancer patients. Overexpression of ADA3 in ER- hMECs as well as in ER+ breast cancer cell lines enhanced cell proliferation. These cells showed increased cyclin B and c-MYC, decreased p27 and increased SKP2 levels. This was accompanied by increased mRNA levels of early response genes c-FOS, EGR1, and c-MYC. Analysis of breast cancer tissue specimens showed a significant correlation of ADA3 nuclear expression with c-MYC expression. Furthermore, nuclear ADA3 and c-MYC expression together showed significant correlation with tumor grade, mitosis, pleomorphism, NPI, ER/PR status, Ki67 and p27 expression. Importantly, within ER+ cases, expression of nuclear ADA3 and c-MYC also significantly correlated with Ki67 and p27 expression. Univariate Kaplan Meier analysis of four groups in the whole, as well as the ER+ patients showed that c-MYC and ADA3 combinatorial phenotypes showed significantly different breast cancer specific survival with c-MYC-high and ADA3-Low subgroup had the worst outcome. Using multivariate analyses within the whole cohort and the ER+ subgroups, the significant association of ADA3 and c-MYC expression with patients' outcome was independent of tumor grade

  9. The structure and nucleotide sequence of the 5' end of the human c-myc oncogene.

    Science.gov (United States)

    Watt, R; Nishikura, K; Sorrentino, J; ar-Rushdi, A; Croce, C M; Rovera, G

    1983-01-01

    We have established the structure and nucleotide sequence of the 5' end of the human c-myc oncogene, using a cloned genomic fragment isolated from a fetal liver library (clone lambda MC41) and cloned cDNA from the human leukemic cell line K562. The human c-myc oncogene consists of three exons and two introns. Primer extension of the human c-myc mRNA of three different cell lines and S1 nuclease protection experiments served to establish the position of two transcription initiation sites. The splicing site of the first exon-intron boundary was determined by comparative analysis of the sequences of the genomic and cDNA clones. The first exon contains termination codons in all three reading frames and no translation initiation signals, confirming our previous observation that the c-myc mRNA has a long 5' noncoding sequence. This first exon also was found to be utilized in the formation of c-myc mRNAs in a variety of human cell lines. Images PMID:6578511

  10. Shikonin regulates C-MYC and GLUT1 expression through the MST1-YAP1-TEAD1 axis

    Energy Technology Data Exchange (ETDEWEB)

    Vališ, Karel, E-mail: karel.valis@biomed.cas.cz [Laboratory of Structural Biology and Cell Signaling, Institute of Microbiology, v.v.i., The Czech Academy of Sciences, Prague (Czech Republic); Faculty of Science, Charles University, Prague (Czech Republic); Talacko, Pavel; Grobárová, Valéria [Laboratory of Structural Biology and Cell Signaling, Institute of Microbiology, v.v.i., The Czech Academy of Sciences, Prague (Czech Republic); Faculty of Science, Charles University, Prague (Czech Republic); Černý, Jan [Faculty of Science, Charles University, Prague (Czech Republic); Novák, Petr, E-mail: pnovak@biomed.cas.cz [Laboratory of Structural Biology and Cell Signaling, Institute of Microbiology, v.v.i., The Czech Academy of Sciences, Prague (Czech Republic); Faculty of Science, Charles University, Prague (Czech Republic)

    2016-12-10

    The general mechanism underlying the tumor suppressor activity of the Hippo signaling pathway remains unclear. In this study, we explore the molecular mechanisms connecting the Hippo signaling pathway with glucose metabolism. We have found that two key regulators of glycolysis, C-MYC and GLUT1, are targets of the Hippo signaling pathway in human leukemia cells. Our results revealed that activation of MST1 by the natural compound shikonin inhibited the expression of GLUT1 and C-MYC. Furthermore, RNAi experiments confirmed the regulation of GLUT1 and C-MYC expression via the MST1-YAP1-TEAD1 axis. Surprisingly, YAP1 was found to positively regulate C-MYC mRNA levels in complex with TEAD1, while it negatively regulates C-MYC levels in cooperation with MST1. Hence, YAP1 serves as a rheostat for C-MYC, which is regulated by MST1. In addition, depletion of MST1 stimulates lactate production, whereas the specific depletion of TEAD1 has an opposite effect. The inhibition of lactate production and cellular proliferation induced by shikonin also depends on the Hippo pathway activity. Finally, a bioinformatic analysis revealed conserved TEAD-binding motifs in the C-MYC and GLUT1 promoters providing another molecular data supporting our observations. In summary, regulation of glucose metabolism could serve as a new tumor suppressor mechanism orchestrated by the Hippo signaling pathway. - Highlights: • Shikonin inhibits C-MYC and GLUT1 expression in MST1 and YAP1 dependent manner. • YAP1-TEAD1 interaction activates C-MYC and GLUT1 expression. • MST1 in cooperation with YAP1 inhibits C-MYC and GLUT1 expression. • MST1-YAP1-TEAD1 axis regulates lactate production by leukemic cells. • MST1 and YAP1 proteins block proliferation of leukemic cells.

  11. C-MYC aberrations as prognostic factors in diffuse large B-cell lymphoma: a meta-analysis of epidemiological studies.

    Directory of Open Access Journals (Sweden)

    Kuangguo Zhou

    Full Text Available OBJECTIVES: Various studies have investigated the prognostic value of C-MYC aberrations in diffuse large B-cell lymphoma (DLBCL. However, the role of C-MYC as an independent prognostic factor in clinical practice remains controversial. A systematic review and meta-analysis were performed to clarify the clinical significance of C-MYC aberrations in DLBCL patients. METHODS: The pooled hazard ratios (HRs for overall survival (OS and event-free survival (EFS were calculated as the main effect size estimates. The procedure was conducted according to the Cochrane handbook and PRISMA guidelines, including the use of a heterogeneity test, publication bias assessment, and meta-regression, as well as subgroup analyses. RESULTS: Twenty-four eligible studies enrolling 4662 patients were included in this meta-analysis. According to the nature of C-MYC aberrations (gene, protein, and mRNA, studies were divided into several subgroups. For DLBCL patients with C-MYC gene abnormalities, the combined HR was 2.22 (95% confidence interval, 1.89 to 2.61 for OS and 2.29 (95% confidence interval, 1.81 to 2.90 for EFS, compared to patients without C-MYC gene abnormalities. For DLBCL patients with overexpression of C-MYC protein and C-MYC mRNA, pooled HRs for OS were 2.13 and 1.62, respectively. C-MYC aberrations appeared to play an independent role among other well-known prognostic factors in DLBCL. Addition of rituximab could not overcome the inferior prognosis conferred by C-MYC. CONCLUSION: The present systematic review and meta-analysis confirm the prognostic value of C-MYC aberrations. Screening of C-MYC should have definite prognostic meaning for DLBCL stratification, thus guaranteeing a more tailored therapy.

  12. c-myc down-regulates class I HLA expression in human melanomas

    NARCIS (Netherlands)

    Versteeg, R.; NOORDERMEER, I. A.; Krüse-Wolters, M.; Ruiter, D. J.; Schrier, P. I.

    1988-01-01

    Expression of class I HLA antigen has been shown to be reduced in a number of human tumours. Here we show that in a panel of 11 melanoma cell lines with variable class I HLA expression an inverse correlation exists between the mRNA levels of c-myc and class I HLA. This suggests that high expression

  13. Deubiquitinating enzyme USP22 positively regulates c-Myc stability and tumorigenic activity in mammalian and breast cancer cells.

    Science.gov (United States)

    Kim, Dongyeon; Hong, Ahyoung; Park, Hye In; Shin, Woo Hyun; Yoo, Lang; Jeon, Seo Jeong; Chung, Kwang Chul

    2017-12-01

    The proto-oncogene c-Myc has a pivotal function in growth control, differentiation, and apoptosis and is frequently affected in human cancer, including breast cancer. Ubiquitin-specific protease 22 (USP22), a member of the USP family of deubiquitinating enzymes (DUBs), mediates deubiquitination of target proteins, including histone H2B and H2A, telomeric repeat binding factor 1, and cyclin B1. USP22 is also a component of the mammalian SAGA transcriptional co-activating complex. In this study, we explored the functional role of USP22 in modulating c-Myc stability and its physiological relevance in breast cancer progression. We found that USP22 promotes deubiquitination of c-Myc in several breast cancer cell lines, resulting in increased levels of c-Myc. Consistent with this, USP22 knockdown reduces c-Myc levels. Furthermore, overexpression of USP22 stimulates breast cancer cell growth and colony formation, and increases c-Myc tumorigenic activity. In conclusion, the present study reveals that USP22 in breast cancer cell lines increases c-Myc stability through c-Myc deubiquitination, which is closely correlated with breast cancer progression. © 2017 Wiley Periodicals, Inc.

  14. Acidosis decreases c-Myc oncogene expression in human lymphoma cells: a role for the proton-sensing G protein-coupled receptor TDAG8.

    Science.gov (United States)

    Li, Zhigang; Dong, Lixue; Dean, Eric; Yang, Li V

    2013-10-11

    Acidosis is a biochemical hallmark of the tumor microenvironment. Here, we report that acute acidosis decreases c-Myc oncogene expression in U937 human lymphoma cells. The level of c-Myc transcripts, but not mRNA or protein stability, contributes to c-Myc protein reduction under acidosis. The pH-sensing receptor TDAG8 (GPR65) is involved in acidosis-induced c-Myc downregulation. TDAG8 is expressed in U937 lymphoma cells, and the overexpression or knockdown of TDAG8 further decreases or partially rescues c-Myc expression, respectively. Acidic pH alone is insufficient to reduce c-Myc expression, as it does not decrease c-Myc in H1299 lung cancer cells expressing very low levels of pH-sensing G protein-coupled receptors (GPCRs). Instead, c-Myc is slightly increased by acidosis in H1299 cells, but this increase is completely inhibited by ectopic overexpression of TDAG8. Interestingly, TDAG8 expression is decreased by more than 50% in human lymphoma samples in comparison to non-tumorous lymph nodes and spleens, suggesting a potential tumor suppressor function of TDAG8 in lymphoma. Collectively, our results identify a novel mechanism of c-Myc regulation by acidosis in the tumor microenvironment and indicate that modulation of TDAG8 and related pH-sensing receptor pathways may be exploited as a new approach to inhibit Myc expression.

  15. Acidosis Decreases c-Myc Oncogene Expression in Human Lymphoma Cells: A Role for the Proton-Sensing G Protein-Coupled Receptor TDAG8

    Directory of Open Access Journals (Sweden)

    Zhigang Li

    2013-10-01

    Full Text Available Acidosis is a biochemical hallmark of the tumor microenvironment. Here, we report that acute acidosis decreases c-Myc oncogene expression in U937 human lymphoma cells. The level of c-Myc transcripts, but not mRNA or protein stability, contributes to c-Myc protein reduction under acidosis. The pH-sensing receptor TDAG8 (GPR65 is involved in acidosis-induced c-Myc downregulation. TDAG8 is expressed in U937 lymphoma cells, and the overexpression or knockdown of TDAG8 further decreases or partially rescues c-Myc expression, respectively. Acidic pH alone is insufficient to reduce c-Myc expression, as it does not decrease c-Myc in H1299 lung cancer cells expressing very low levels of pH-sensing G protein-coupled receptors (GPCRs. Instead, c-Myc is slightly increased by acidosis in H1299 cells, but this increase is completely inhibited by ectopic overexpression of TDAG8. Interestingly, TDAG8 expression is decreased by more than 50% in human lymphoma samples in comparison to non-tumorous lymph nodes and spleens, suggesting a potential tumor suppressor function of TDAG8 in lymphoma. Collectively, our results identify a novel mechanism of c-Myc regulation by acidosis in the tumor microenvironment and indicate that modulation of TDAG8 and related pH-sensing receptor pathways may be exploited as a new approach to inhibit Myc expression.

  16. hTERT protein expression is independent of clinicopathological parameters and c-Myc protein expression in human breast cancer

    Directory of Open Access Journals (Sweden)

    Meligonis G

    2005-01-01

    Full Text Available Abstract Background Telomerase is a ribonucleoprotein enzyme that synthesises telomeres after cell division and maintains chromosomal length and stability thus leading to cellular immortalisation. The hTERT (human telomerase reverse transcriptase subunit seems to be the rate-limiting determinant of telomerase and knowledge of factors controlling hTERT transcription may be useful in therapeutic strategies. The hTERT promoter contains binding sites for c-Myc and there is some experimental and in vitro evidence that c-Myc may increase hTERT expression. We previously reported no correlation between c-Myc mRNA expression and hTERT mRNA or telomerase activity in human breast cancer. This study aims to examine the correlation between hTERT expression as determined by immunohistochemistry and c-Myc expression, lymph node status, and tumour size and grade in human breast cancer. Materials and methods The immunohistochemical expression of hTERT and c-Myc was investigated in 38 malignant breast tumours. The expression of hTERT was then correlated with the lymph node status, c-Myc expression and other clinicopathological parameters of the tumours. Results hTERT expression was positive in 27 (71% of the 38 tumours. 15 (79% of 19 node positive tumours were hTERT positive compared with 11 (63% of 19 node negative tumours. The expression was higher in node positive tumours but this failed to reach statistical significance (p = 0.388. There was no significant association with tumour size, tumour grade or c-Myc expression. However, hTERT expression correlated positively with patients' age (correlation coefficient = 0.415, p = 0.0097. Conclusion hTERT protein expression is independent of lymph node status, tumour size and grade and c-Myc protein expression in human breast cancer

  17. A proteomic study of cMyc improvement of CHO culture

    Directory of Open Access Journals (Sweden)

    Dunn Michael J

    2010-03-01

    Full Text Available Abstract Background The biopharmaceutical industry requires cell lines to have an optimal proliferation rate and a high integral viable cell number resulting in a maximum volumetric recombinant protein product titre. Nutrient feeding has been shown to boost cell number and productivity in fed-batch culture, but cell line engineering is another route one may take to increase these parameters in the bioreactor. The use of CHO-K1 cells with a c-myc plasmid allowing for over-expressing c-Myc (designated cMycCHO gives a higher integral viable cell number. In this study the differential protein expression in cMycCHO is investigated using two-dimensional gel electrophoresis (2-DE followed by image analysis to determine the extent of the effect c-Myc has on the cell and the proteins involved to give the new phenotype. Results Over 100 proteins that were differentially expressed in cMycCHO cells were detected with high statistical confidence, of which 41 were subsequently identified by tandem mass spectrometry (LC-MS/MS. Further analysis revealed proteins involved in a variety of pathways. Some examples of changes in protein expression include: an increase in nucleolin, involved in proliferation and known to aid in stabilising anti-apoptotic protein mRNA levels, the cytoskeleton and mitochondrial morphology (vimentin, protein biosysnthesis (eIF6 and energy metabolism (ATP synthetase, and a decreased regulation of all proteins, indentified, involved in matrix and cell to cell adhesion. Conclusion These results indicate several proteins involved in proliferation and adhesion that could be useful for future approaches to improve proliferation and decrease adhesion of CHO cell lines which are difficult to adapt to suspension culture.

  18. Transactivation of proto-oncogene c-Myc by hepatitis B virus transactivator MHBst167.

    Science.gov (United States)

    Lun, Yong-Zhi; Cheng, Jun; Chi, Qing; Wang, Xue-Lei; Gao, Meng; Sun, Li-DA

    2014-08-01

    C-terminally truncated hepatitis B virus (HBV) middle size surface proteins (MHBst) has been shown to be a transcriptional activator and may be relevant to hepatocarcinogenesis by transactivating gene expression. In the present study, a pcDNA3.1(-)-MHBst167 vector coding for MHBst truncated at amino acid 167 (MHBst167) was constructed and transfected into the HepG2 hepatoma cell line. mRNA and protein expression of MHBst167 in the cells was detected by reverse transcription-polymerase chain reaction (RT-PCR) and western blot analysis. A cDNA library of genes transactivated by the truncated protein in HepG2 cells was made in pGEM-T Easy using suppression subtractive hybridization. The cDNAs were sequenced and analyzed with BLAST searching against the sequences in GenBank. The results showed that certain sequences, such as that of human proto-oncogene c-Myc, may be involved in tumor development. An expression vector pCAT3/c-Myc containing the chloramphenicol acetyltransferase (CAT) gene under the control of a c-Myc promoter was generated, and the transcriptional transactivating effect of MHBst167 on the c-Myc promoter was investigated by RT-PCR and western blotting. MHBst167 was found to upregulate the transcriptional activity of the promoter, as well as transcription and translation of c-Myc. MHBst167 was also shown to transactivate SV40 immediate early promoter, and transcriptionally transactivate the expression of human c-Myc. These findings provide new directions for studying the biological functions of MHBst167, and for a better understanding of the tumor development mechanisms of HBV infection.

  19. Recessive genetic deregulation abrogates c-myc suppression by interferon and is implicated in oncogenesis

    Energy Technology Data Exchange (ETDEWEB)

    Kimchi, A.; Resnitzky, D.; Ber, R.; Gat, G.

    1988-07-01

    Previously the authors demonstrated that many hematopoietic tumor cells are resistant to the inhibitory effects that interferon exerts on c-myc mRNA expression without losing other receptor-mediated intracellular responses. They report here that this partial resistance was overridden in two independent stable somatic cell hybrids prepared by fusion between sensitive and resistant cells. The c-myc mRNA transcribed from the active allele of the resistant parent cell was reduced by interferon within the context of the cell hybrid. It was therefore concluded that changes in the cis-acting sequences of c-myc were not involved in this type of relaxed regulation and that resistance resulted rather from inactivation or loss of postreceptor elements which operate in trans. The growth-stimulating effect that this genetic deregulation might have on cells was tested in experimental systems of cell differentiation in which an autocrine interferon is produced. For that purpose the authors isolated variant clones of M1 myeloid cells which were partially resistant to alpha and beta interferons and tested their growth behaviour during in vitro-induced differentiation. The resistant clones displayed higher proliferative activity on days 2 and 3 of differentiation than did the sensitive clones, which stopped proliferating. The loss of c-myc responses to the self-produced interferon disrupted the normal cessation of growth during differentiation and therefore might lead cells along the pathway of neoplasia.

  20. Insertion of the LINE-1 element in the C-MYC gene and immunoreactivity of C-MYC, p53, p21 and p27 proteins in different morphological patterns of the canine TVT

    OpenAIRE

    C.R.O. Lima; Faleiro,M.B.R.; Rabelo,R.E.; V.A.S. Vulcani; Rubini,M.R.; Torres,F.A.G.; Moura,V.M.B.D.

    2016-01-01

    ABSTRACT The canine transmissible venereal tumor (TVT) affects the external genitalia of dogs by the natural transplant of viable tumor cells. Thus, this research aimed to diagnose and characterize TVT morphological patterns, identify the insertion of the LINE-1 element in C-MYC gene, by means of the polymerase chain reaction (PCR), and evaluate the immunohistochemical expression of C-MYC, p53, p21 and p27 proteins. The relationship between C-MYC and p53 proteins and their interference on the...

  1. The enhancement of c-myc expression in cultured epithelial cells by some cytotoxic metals.

    Science.gov (United States)

    Skilleter, D N; Price, R J; McNerney, R

    1991-01-01

    The toxic or carcinogenic metal ions Cd(2+), Hg(2+), Co(2+), Ni(2+) and Be(2+) are known to be potent inhibitors of cell division in cultured cells. The effects of these metal ions on the biphasic expression of the cell proliferation-associated proto-oncogene c-myc, have now been examined in epithelial cells (BL9L) derived from rat liver, using mRNA hybridization analysis following serum stimulation of synchronized (G(0)/G(1) cell cycle phase) confluent (quiescent) and non-confluent (proliferating) monolayer cultures. Exposure of the cells under these conditions to antiproliferative concentrations of BeSO(4) (50-100 mum), NiCl(2) (50 mum), CoCl(2) (50 mum), HgCl(2) (20-50 mum) or CdCl(2) (5-10 mum) showed that whereas Be(2+), Cd(2+) and Hg(2+) further increased steady-state c-myc mRNA levels throughout the treatment period, particularly in non-confluent cultures (two- to eight-fold enhancement), Co(2+) and Ni(2+) did not have a significant effect. In contrast, mRNA transcripts for constantly expressed cytoskeletal actin were essentially unchanged by all the metal ion treatments of the cells. The extent of the enhanced c-myc expression maintained in the cells by Be(2+), Cd(2+) or Hg(2+) treatment could also be broadly correlated with the degree of cell detachment from the culture dishes, which was ultimately produced within 20-24 hr. RNA and protein synthesis inhibitor studies indicate that the cytotoxic metal ions either directly or indirectly modify the normal control mechanisms for c-myc expression. It is concluded that an enhanced c-myc expression is a feature of the cells' response to certain cytotoxic metal ions, the magnitude of which may also be a potential index of pending cell damage.

  2. Equol, an Isoflavone Metabolite, Regulates Cancer Cell Viability and Protein Synthesis Initiation via c-Myc and eIF4G*

    Science.gov (United States)

    de la Parra, Columba; Borrero-Garcia, Luis D.; Cruz-Collazo, Ailed; Schneider, Robert J.; Dharmawardhane, Suranganie

    2015-01-01

    Epidemiological studies implicate dietary soy isoflavones as breast cancer preventives, especially due to their anti-estrogenic properties. However, soy isoflavones may also have a role in promoting breast cancer, which has yet to be clarified. We previously reported that equol, a metabolite of the soy isoflavone daidzein, may advance breast cancer potential via up-regulation of the eukaryotic initiation factor 4GI (eIF4GI). In estrogen receptor negative (ER−) metastatic breast cancer cells, equol induced elevated levels of eIF4G, which were associated with increased cell viability and the selective translation of mRNAs that use non-canonical means of initiation, including internal ribosome entry site (IRES), ribosome shunting, and eIF4G enhancers. These mRNAs typically code for oncogenic, survival, and cell stress molecules. Among those mRNAs translationally increased by equol was the oncogene and eIF4G enhancer, c-Myc. Here we report that siRNA-mediated knockdown of c-Myc abrogates the increase in cancer cell viability and mammosphere formation by equol, and results in a significant down-regulation of eIF4GI (the major eIF4G isoform), as well as reduces levels of some, but not all, proteins encoded by mRNAs that are translationally stimulated by equol treatment. Knockdown of eIF4GI also markedly reduces an equol-mediated increase in IRES-dependent mRNA translation and the expression of specific oncogenic proteins. However, eIF4GI knockdown did not reciprocally affect c-Myc levels or cell viability. This study therefore implicates c-Myc as a potential regulator of the cancer-promoting effects of equol via up-regulation of eIF4GI and selective initiation of translation on mRNAs that utilize non-canonical initiation, including certain oncogenes. PMID:25593313

  3. Cooverexpression of EpCAM and c-myc genes in malignant breast tumours.

    Science.gov (United States)

    Sadeghi, Samira; Hojati, Zohreh; Tabatabaeian, Hossein

    2017-03-01

    The overexpression of epithelial cell adhesion molecule (EpCAM), a proto-oncogene, affects progression, treatment, and diagnosis of many adenocarcinomas. C-myc has been shown to be a downstream target of EpCAM and is also one of the most important proto-oncogenes routinely overexpressed in breast cancer. However, cooverexpression of EpCAM and c-myc genes has not been investigated in breast cancer tissues, particularly in Iranian population. The aim of this study was to assess the expression of EpCAM and c-myc genes in malignant breast cancer tissues using reverse transcriptase-quantitative polymerase chain reaction (RT-qPCR) followed by analyses of the association between the outcomes. In this study, 122 fresh tissues, including 104 malignant and 18 benign samples, were disrupted by mortar and pestle, and then the RNA was isolated from the samples and converted to cDNA. The relative expression levels of EpCAM and c-myc genes were measured by 2 -ΔΔCt method using RT-qPCR. EpCAM protein level was also assessed in 66 cases using Western blot technique. Using RT-qPCR method, our results showed that EpCAM was overexpressed in 48% of malignant and 11.1% of benign samples. Evaluating EpCAM protein overexpression in a portion of samples depicted the fully concordance rate between Western blot and RT-qPCR techniques. C-myc expression was first evaluated by RT-qPCR method, showing the overexpression rate of 39% and 28% in malignant and benign samples, respectively. These data were also quite concordant with the clinically available immunohistochemistry reports of the same samples studied in this study. Importantly, overexpression of EpCAM and c-myc was significantly associated and showed an agreement of 57.3%. This study demonstrated the cooverexpression of EpCAM and c-myc in breast tumours collected from breast cancer patients of the Iranian population. EpCAM and c-myc positive cases were significantly associated with reduced and enhanced risk of ER/PR positivity

  4. c-Myc-Dependent Cell Competition in Human Cancer Cells.

    Science.gov (United States)

    Patel, Manish S; Shah, Heta S; Shrivastava, Neeta

    2017-07-01

    Cell Competition is an interaction between cells for existence in heterogeneous cell populations of multicellular organisms. This phenomenon is involved in initiation and progression of cancer where heterogeneous cell populations compete directly or indirectly for the survival of the fittest based on differential gene expression. In Drosophila, cells having lower dMyc expression are eliminated by cell competition through apoptosis when present in the milieu of cells having higher dMyc expression. Thus, we designed a study to develop c-Myc (human homolog) dependent in vitro cell competition model of human cancer cells. Cells with higher c-Myc were transfected with c-myc shRNA to prepare cells with lower c-Myc and then co-cultured with the same type of cells having a higher c-Myc in equal ratio. Cells with lower c-Myc showed a significant decrease in numbers when compared with higher c-Myc cells, suggesting "loser" and "winner" status of cells, respectively. During microscopy, engulfment of loser cells by winner cells was observed with higher expression of JNK in loser cells. Furthermore, elimination of loser cells was prevented significantly, when co-cultured cells were treated with the JNK (apoptosis) inhibitor. Above results indicate elimination of loser cells in the presence of winner cells by c-Myc-dependent mechanisms of cell competition in human cancer cells. This could be an important mechanism in human tumors where normal cells are eliminated by c-Myc-overexpressed tumor cells. J. Cell. Biochem. 118: 1782-1791, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  5. Acidified bile acids enhance tumor progression and telomerase activity of gastric cancer in mice dependent on c-Myc expression.

    Science.gov (United States)

    Wang, Xiaolong; Sun, Lei; Wang, Xijing; Kang, Huafeng; Ma, Xiaobin; Wang, Meng; Lin, Shuai; Liu, Meng; Dai, Cong; Dai, Zhijun

    2017-04-01

    c-Myc overexpression has been implicated in several malignancies including gastric cancer. Here, we report that acidified bile acids enhance tumor progression and telomerase activity in gastric cancer via c-Myc activation both in vivo and in vitro. c-Myc mRNA and protein levels were assessed in ten primary and five local recurrent gastric cancer samples by quantitative real-time polymerase chain reaction and western blotting analysis. The gastric cancer cell line MGC803 was exposed to bile salts (100 μmol/L glycochenodeoxycholic acid and deoxycholic acid) in an acid medium (pH 5.5) for 10 min daily for 60 weeks to develop an MGC803-resistant cell line. Control MGC803 cells were grown without acids or bile salts for 60 weeks as a control. Cell morphology, proliferation, colony formation and apoptosis of MGC803-resistant cells were analyzed after 60 weeks. To determine the involvement of c-Myc in tumor progression and telomere aging in MGC803-resistant cells, we generated xenografts in nude mice and measured xenograft volume and in vivo telomerase activity. The c-Myc and hTERT protein and mRNA levels were significantly higher in local recurrent gastric cancer samples than in primary gastric cancer samples. MGC803-resistant cells showed a marked phenotypic change under normal growth conditions with more clusters and acini, and exhibited increased cell viability and colony formation and decreased apoptosis in vitro. These phenotypic changes were found to be dependent on c-Myc activation using the c-Myc inhibitor 10058-F4. MGC803-resistant cells also showed a c-Myc-dependent increase in xenograft growth and telomerase activity in vivo. In conclusion, these observations support the hypothesis that acidified bile acids enhance tumor progression and telomerase activity in gastric cancer and that these effects are dependent on c-Myc activity. These findings suggest that acidified bile acids play an important role in the malignant progression of local recurrent

  6. Telomerase in relation to expression of p53, c-Myc and estrogen receptor in ovarian tumours

    NARCIS (Netherlands)

    Wisman, GBA; Hollema, H; Helder, MN; Knol, AJ; Van Der Meer, GT; Krans, M; De Jong, S; De Vries, EGE; Van Der Zee, AGJ

    2003-01-01

    Telomerase activity and its subunits (hTERC, hTERT mRNA) were evaluated in ovarian tumours in relation to the expression of p53, c-Myc and estrogen receptor (ER). Furthermore, relations between telomerase activity, hTERC and hTERT with known clinicopathologic prognostic factors and survival in

  7. [Progress of research on Proto-oncogene c-myc, c-myb in platelet diseases].

    Science.gov (United States)

    Zhang, Ying; Chen, Rui; Zhao, Li

    2011-02-01

    The Proto-oncogene c-myc and c-myb has been shown to be crucial in the development of the hematopoietic system. The changes in the expression of c-myc are concerned the cell proliferation and differentiation, the expression products of which play an important regulatory role in cell growth, differentiation or malignant transformation. The c-myb involves in transcription and affects cell proliferation, differentiation, apoptosis. More recently, the researches on proto-oncogene c-myc, c-myb in hematopoietic regulation have gradually increased along with development of molecular biology, molecular immunology and cell biology. Scientists point out that the directive differentiation of erythroid and megakaryocytic progenitors, and platelet abnormalities all relate to the level of their expressions. The most common thrombocytopathy includes thrombocytopenia, thrombocytosis and so on. The etiology and the mechanism of these diseases are unknown. This article reviews the structure, function and the expression of c-myc and c-myb in platelet diseases and their significance.

  8. Expression of c-myc oncogene in coeliac disease.

    OpenAIRE

    P. J. Ciclitira; Stewart, J.(DESY, 15738, Zeuthen, Germany); Evan, G; Wight, D G; Sikora, K

    1987-01-01

    A monoclonal antibody, produced by peptide immunisation was used to detect the distribution of p62c/myc by immunohistology in normal and coeliac small intestinal mucosa. The effect of gluten in four treated coeliac patients was investigated by taking serial jejunal biopsy specimens for six hours after a 10 g oral gluten challenge. There was a progressive increase in p62c/myc staining intensity in the villus enterocytes extending to the crypts, which accompanied the classical morphological cha...

  9. c-Myc-Induced Extrachromosomal Elements Carry Active Chromatin

    Directory of Open Access Journals (Sweden)

    Greg Smith

    2003-03-01

    Full Text Available Murine Pre-13 lymphocytes with experimentally activated MycER show both chromosomal and extrachromosomal gene amplification. In this report, we have elucidated the size, structure, functional components of c-Myc-induced extrachromosomal elements (EEs. Scanning electron microscopy revealed that EEs isolated from MycER-activated Pre-B+ cells are an average of 10 times larger than EEs isolated from non-MycER-activated control Pre-B- cells. We demonstrate that these large c-Myc-induced EEs are associated with histone proteins, whereas EEs of non-MycER-activated Pre B- cells are not. Immunohistochemistry and Western blot analyses using pan -histone-specific, histone H3 phosphorylation-specific, histone H4 acetylation-specific antibodies indicate that a significant proportion of EEs analyzed from MycER-activated cells harbors transcriptionally competent and/or active chromatin. Moreover, these large, c-Myc-induced EEs carry genes. Whereas the total genetic make-up of these c-Myc-induced EEs is unknown, we found that 30.2% of them contain the dihydrofolate reductase (DHFR gene, whereas cyclin C (CCNC was absent. In addition, 50% of these c-Myc-activated Pre-B+ EEs incorporated bromodeoxyuridine (BrdU, identifying them as genetic structures that self-propagate. In contrast, EEs isolated from non-Myc-activated cells neither carry the DHFR gene nor incorporate BrdU, suggesting that c-Myc deregulation generates a new class of EEs.

  10. Analysis of polyadenylation site usage of the c-myc oncogene.

    OpenAIRE

    Laird-Offringa, I A; Elfferich, P; Knaken, H J; Ruiter, J.; van der Eb, A J

    1989-01-01

    The c-myc gene contains 2 well conserved polyadenylation (pA) sites. In all human and rat cell lines from various differentiation stages and tissue types the amount of mRNA terminating at the second pA site is 6-fold higher than the amount ending at the upstream site. This is not due to a difference in stability of the two mRNA types and therefore must be due to preferential usage of the downstream site. The usage of the pA sites is not altered during growth factor induction of quiescent cell...

  11. Dynamic regulation of c-Myc proto-oncogene expression during lymphocyte development revealed by a GFP-c-Myc knock-in mouse.

    Science.gov (United States)

    Huang, Ching-Yu; Bredemeyer, Andrea L; Walker, Laura M; Bassing, Craig H; Sleckman, Barry P

    2008-02-01

    c-Myc induces widely varying cellular effects, including cell proliferation and cell death. These different cellular effects are determined, in part, by c-Myc protein expression levels, which are regulated through several transcriptional and post-transcriptional pathways. c-Myc transcripts can be detected in cells at all stages of B and T lymphocyte development. However, little is known about c-Myc protein expression, and how it varies, in developing lymphocytes. Here mice have been generated in which the endogenous c-Myc locus has been modified (c-Myc(G)) so that it encodes a GFP-c-Myc fusion protein. c-Myc(G/G) mice are viable, appear normal and exhibit grossly normal lymphocyte development. Flow cytometric analyses revealed significant heterogeneity in c-Myc protein expression levels in developing c-Myc(G/G) B and T lymphocytes. GFP-c-Myc expression levels were highest in proliferating lymphocytes, suggesting that c-Myc up-regulation is important for promoting lymphocyte cell division, and demonstrating that GFP-c-Myc expression is a marker of proliferating lymphocytes in vivo.

  12. TCEAL7 Inhibition of c-Myc Activity in Alternative Lengthening of Telomeres Regulates hTERT Expression

    Directory of Open Access Journals (Sweden)

    Kyle Lafferty-Whyte

    2010-05-01

    Full Text Available Replicative senescence forms a major barrier to tumor progression. Cancer cells bypass this by using one of the two known telomere maintenance mechanisms: telomerase or the recombination-based alternative lengthening of telomeres (ALT mechanism. The molecular details of ALT are currently poorly understood. We have previously shown that telomerase is actively repressed through complex networks of kinase, gene expression, and chromatin regulation. In this study, we aimed to gain further understanding of the role of kinases in the regulation of telomerase expression in ALT cells. Using a whole human kinome small interfering RNA (siRNA screen, we highlighted 106 kinases whose expression is linked to human telomerase reverse transcriptase (hTERT promoter activity. Network modeling of transcriptional regulation implicated c-Myc as a key regulator of the 106 kinase hits. Given our previous observations of lower c-Myc activity in ALT cells, we further explored its potential to regulate telomerase expression in ALT. We found increased c-Myc binding at the hTERT promoter in telomerase-positive compared with ALT cells, although no expression differences in c-Myc, Mad, or Max were observed between ALT and telomerase-positive cells that could explain decreased c-Myc activity in ALT. Instead, we found increased expression of the c-Myc competitive inhibitor TCEAL7 in ALT cells and tumors and that alteration of TCEAL7 expression levels in ALT and telomerase-positive cells affects hTERT expression. Lower c-Myc activity in ALT may therefore be obtained through TCEAL7 regulation. Thus, TCEAL7 may present an interesting novel target for cancer therapy, which warrants further investigation.

  13. [Influence of RNA interference targeting against human telomerase reverse transcriptase on expression of C-myc protein].

    Science.gov (United States)

    Chi, Huaming; Tao, Zezhang; Chen, Shiming; Xiao, Bokui; Zhan, Hanzhang

    2005-11-01

    To investigate the effect of inhibiting human telomerase reverse transcriptase (hTERT) on expression of C-myc protein by RNA interference (RNAi) in the larynx cancer cell line, Hep-2. The primary structures of hTERT cDNA were found in GeneBank. Then the structure analyses were done according to the strategy of RNAi, which determined the specific base sequences to design shRNA plasmid. One type of plasmid, pshRNA1, involved in fluorescein gene was synthesized based on the specific base sequence. Control pshRNA2-a random sequence-were also constructed. METAFECTENE was used as the transfect ion reagent. Cells were treated daily with pshRNA1-2 or normal culture medium respectively. After administration of pshRNA1-2, hTERT mRNA was detected by RT-PCR, hTERT protein and C-myc protein were examined by Western Blot. The expression of hTERT mRNA and protein were both significantly decreased after treated by pshRNA1 (P < 0.05). The expression of C-myc protein was significantly increased after treated by pshRNA1 (P < 0.01). The inhibition of hTERT expression could increase the expression of C-myc protein in Hep-2 cells.

  14. Control of c-fos and c-myc proto-oncogene induction in rat thyroid cells in culture

    Energy Technology Data Exchange (ETDEWEB)

    Isozaki, O.; Kohn, L.D. (National Institute of Diabetes, Digestive and Kidney Diseases, Bethesda, MD (USA))

    1987-11-01

    Removal of TSH, insulin, and cortisol from the medium, and decreasing the serum content to 0.2%, abolishes both the proliferate and differentiated state of FRTL-5 rat thyroid cells in culture. In these basal conditions, the individual addition of TSH, insulin, insulin-like growth factor-I (IGF-I), phorbol 12-myristate 13-acetate (TPA), alpha 1-adrenergic agents, or A23187, increase c-myc and/or c-fos proto-oncogene expression. Under the same conditions, only the addition of TSH increased cAMP levels; 8-bromo-cAMP can increase c-myc or c-fos mRNA levels. Pretreatment of cells with phorbol 12,13-dibutyrate, an agent which down regulates the C-kinase, completely inhibits the effect of TPA on proto-oncogene expression but has no affect on the A23187 induced-increase. The sum of these results indicate that at least four separate signal systems independently increase c-myc or c-fos gene expression in FRTL-5 cells cAMP (TSH), C-kinase (TPA), Ca++/phosphoinositide (A23187), and that influenced by insulin/IGF-I. None of the ligands, when individually returned to cells in basal medium (no TSH, insulin, or cortisol and only 0.2% serum), increases cell number; norepinephrine, and A23187 do not increase (3H)thymidine incorporation into DNA under these conditions; and combinations of the ligands can be more than additive in effecting (3H)thymidine incorporation into DNA but are only additive in effecting proto-oncogene expression. Insulin/IGF-I plus TSH or insulin/IGF-I plus norepinephrine can increase both proto-oncogene expression and (3H)thymidine incorporation into DNA to the same extent; however, the former combination can increase cell number whereas the latter cannot. There is therefore no simple correlation between the ability of the above ligands to increase proto-oncogene expression and their ability to increase cell number or induce DNA synthesis.

  15. Interaction of c-Myc with the pRb-related protein p107 results in inhibition of c-Myc-mediated transactivation

    NARCIS (Netherlands)

    Beijersbergen, R.L.; Hijmans, E.M.; Zhu, L.; Bernards, R.A.

    1994-01-01

    The product of the c-myc proto-oncogene, c-Myc, is a sequence-specific DNA binding protein with an Nterminal transactivation domain and a C-terminal DNA binding domain. Several lines of evidence indicate that c-Myc activity is essential for normal cell cycle progression. Since the abundance of

  16. Differential control of proto-oncogene c-myc and c-fos expression in lymphocytes and fibroblasts.

    Science.gov (United States)

    McNerney, R; Darling, D; Johnstone, A

    1987-01-01

    In lymphocytes stimulated with the mitogen phytohaemagglutinin, an inhibitor of the enzyme ADP-ribosyltransferase (ADPRT) completely blocks the proliferative response and the increase in expression of the proto-oncogene c-myc without affecting c-fos significantly. Conversely, in fibroblasts the serum-induced growth is not affected by the ADPRT inhibitor, and both oncogenes are dramatically super-induced. Hence there are differences between lymphocyte and fibroblast early responses to mitogenic stimulation and also between regulation of c-fos and c-myc gene expression. Images Fig. 2. Fig. 3. PMID:3117047

  17. Expression of c-myc oncogene in coeliac disease.

    Science.gov (United States)

    Ciclitira, P J; Stewart, J; Evan, G; Wight, D G; Sikora, K

    1987-01-01

    A monoclonal antibody, produced by peptide immunisation was used to detect the distribution of p62c/myc by immunohistology in normal and coeliac small intestinal mucosa. The effect of gluten in four treated coeliac patients was investigated by taking serial jejunal biopsy specimens for six hours after a 10 g oral gluten challenge. There was a progressive increase in p62c/myc staining intensity in the villus enterocytes extending to the crypts, which accompanied the classical morphological changes occurring in the mucosa. Images Fig 1 Fig 2 PMID:3549790

  18. Alteraciones del gen c-Myc en la oncogénesis = c-Myc gene alterations in oncogenesis

    Directory of Open Access Journals (Sweden)

    Ospina Pérez, Mariano

    2011-12-01

    Full Text Available La familia de protooncogenes MYC (c-Myc, N-Myc y L-Myc se relaciona con el origen de diversas neoplasias en seres humanos. Estos genes actúan como factores de transcripción y participan en la regulación del ciclo celular, la proliferación y diferenciación celulares, la apoptosis y la inmortalización. Los genes MYC se expresan en diferentes tejidos y responden a diversas señales internas y externas; codifican para la síntesis de factores de transcripción que se unen al ADN para regular la expresión de múltiples genes. El gen más ampliamente estudiado de esta familia es c-Myc, que se expresa en las células con mayor tasa de proli­feración. C-Myc se encuentra alterado en un gran número de tumores sólidos, leucemias y linfomas. Las alteraciones de c-Myc encontradas con mayor frecuencia en células cancero­sas son las amplificaciones, translocaciones, mutaciones y reordenamientos cromosómicos que involucran el locus de este gen y conducen a que se desregule su expresión en diversas neoplasias humanas. La amplificación de c-Myc es una alteración común en los cánceres de mama, pulmón, ovario y próstata, así como en leucemias y linfomas, mientras que la pérdida de su regulación es común en el cáncer de colon, en tumores ginecológicos y melanoma. En neoplasias con defectos de c-Myc los estudios actuales están dirigidos al desarrollo de nuevas estrategias terapéuticas.

  19. Effects of microRNA-24 targeting C-myc on apoptosis, proliferation and cytokine expressions in chondrocytes of rats with osteoarthritis via MAPK signaling pathway.

    Science.gov (United States)

    Wu, Yuan-Hao; Liu, Wei; Zhang, Lei; Liu, Xiao-Ya; Wang, Yi; Xue, Bin; Liu, Bin; Duan, Ran; Zhang, Bo; Ji, Yang

    2017-11-16

    To investigate whether microRNA-24 (miR-24) targeting C-myc affects chondrocytes of rats with osteoarthritis (OA) via the MAPK signaling pathway. Thirty rats were assigned as a sham group and an OA group (established as OA rat models by cutting the anterior cruciate ligaments and removing 1/3 medial meniscus). TUNEL staining and immunohistochemistry were conducted for cell apoptosis index (AI) and positive expression rate of C-myc protein. Enzyme-linked immuno sorbent assay (ELISA) was carried out for serum level of IL-1β and TNF-α. Primary chondrocytes were assigned into the blank, negative control (NC), miR-24 mimics, miR-24 inhibitors, siRNA-C-myc, and miR-24 inhibitors + siRNA-C-myc groups. The expressions of miR-24, C-myc, p38, ERK, JNK, IL-1β, and TNF-α in tissues and cells were detected using reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR) and Western blotting. CCK8 assay and flow cytometry were performed for cell proliferation and apoptosis. The OA group showed higher IL-1β, TNF-α, AI, and C-myc than the sham group. C-myc is a target gene of miR-24. Compared with the blank group, the miR-24 mimics and siRNA-C-myc groups showed reduced expression of C-myc, IL-1β, TNF-α, p38, p-p38, ERK, p-ERK, JNK, and p-JNK, apoptosis rate yet increased cell proliferation; however, the miR-24 inhibitors group exhibited an opposite trend. The miR-24 inhibitors + siRNA-C-myc group presented a same tendency compared to the siRNA-C-myc group. Upregulated miR-24 downregulates C-myc could suppress apoptosis and promote proliferation of chondrocytes to prevent the occurrence and subsequent progression of OA via inactivating the MAPK signaling pathway. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.

  20. Equol, an isoflavone metabolite, regulates cancer cell viability and protein synthesis initiation via c-Myc and eIF4G.

    Science.gov (United States)

    de la Parra, Columba; Borrero-Garcia, Luis D; Cruz-Collazo, Ailed; Schneider, Robert J; Dharmawardhane, Suranganie

    2015-03-06

    Epidemiological studies implicate dietary soy isoflavones as breast cancer preventives, especially due to their anti-estrogenic properties. However, soy isoflavones may also have a role in promoting breast cancer, which has yet to be clarified. We previously reported that equol, a metabolite of the soy isoflavone daidzein, may advance breast cancer potential via up-regulation of the eukaryotic initiation factor 4GI (eIF4GI). In estrogen receptor negative (ER-) metastatic breast cancer cells, equol induced elevated levels of eIF4G, which were associated with increased cell viability and the selective translation of mRNAs that use non-canonical means of initiation, including internal ribosome entry site (IRES), ribosome shunting, and eIF4G enhancers. These mRNAs typically code for oncogenic, survival, and cell stress molecules. Among those mRNAs translationally increased by equol was the oncogene and eIF4G enhancer, c-Myc. Here we report that siRNA-mediated knockdown of c-Myc abrogates the increase in cancer cell viability and mammosphere formation by equol, and results in a significant down-regulation of eIF4GI (the major eIF4G isoform), as well as reduces levels of some, but not all, proteins encoded by mRNAs that are translationally stimulated by equol treatment. Knockdown of eIF4GI also markedly reduces an equol-mediated increase in IRES-dependent mRNA translation and the expression of specific oncogenic proteins. However, eIF4GI knockdown did not reciprocally affect c-Myc levels or cell viability. This study therefore implicates c-Myc as a potential regulator of the cancer-promoting effects of equol via up-regulation of eIF4GI and selective initiation of translation on mRNAs that utilize non-canonical initiation, including certain oncogenes. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.

  1. C-myc oncogene product P62c-myc in ovarian mucinous neoplasms: immunohistochemical study correlated with malignancy.

    Science.gov (United States)

    Polacarz, S V; Hey, N A; Stephenson, T J; Hill, A S

    1989-01-01

    The monoclonal antibody Myc 1-6E10 was used to determine the cellular distribution of the c-myc oncogene product p62c-myc in 60 mucinous ovarian tumours. Three patterns of immunostaining were apparent: (i) nuclear staining alone; (ii) staining of the nucleus and basal cytoplasm; and (iii) staining of the entire cell. Of the 21 cases of mucinous cystadenoma, 11 showed nuclear staining alone, and a further case showed additional weak staining of the basal cytoplasm. Nuclear staining alone was not present in any of the 17 borderline mucinous tumours examined. Strong staining of the nucleus and basal cytoplasm was seen in 16 of these borderline cases, six of which also showed focal staining of the apical cytoplasm. All 22 cases of mucinous cystadenocarcinoma showed staining of the cell nucleus and entire cell cytoplasm. Focal staining of the apical cytoplasm in six of 17 borderline mucinous tumours produced a pattern of c-myc immunostaining similar to that of cystadenocarcinoma. Retrospective analysis of the clinical data showed that no significant differences between patients with borderline tumours of these two categories could be defined. Although immunostaining with Myc 1-6E10 can be used in the categorisation of mucinous ovarian tumours, it is concluded that standard histological criteria are more accurate indicators of tumour behaviour than is an assessment of c-myc expression. Images Fig 1 Fig 2 Fig 3 Fig 4 PMID:2921356

  2. Postnatal liver growth and regeneration are independent of c-myc in a mouse model of conditional hepatic c-myc deletion

    Directory of Open Access Journals (Sweden)

    Sanders Jennifer A

    2012-03-01

    Full Text Available Abstract Background The transcription factor c-myc regulates genes involved in hepatocyte growth, proliferation, metabolism, and differentiation. It has also been assigned roles in liver development and regeneration. In previous studies, we made the unexpected observation that c-Myc protein levels were similar in proliferating fetal liver and quiescent adult liver with c-Myc displaying nucleolar localization in the latter. In order to investigate the functional role of c-Myc in adult liver, we have developed a hepatocyte-specific c-myc knockout mouse, c-mycfl/fl;Alb-Cre. Results Liver weight to body weight ratios were similar in control and c-myc deficient mice. Liver architecture was unaffected. Conditional c-myc deletion did not result in compensatory induction of other myc family members or in c-Myc's binding partner Max. Floxed c-myc did have a negative effect on Alb-Cre expression at 4 weeks of age. To explore this relationship further, we used the Rosa26 reporter line to assay Cre activity in the c-myc floxed mice. No significant difference in Alb-Cre activity was found between control and c-mycfl/fl mice. c-myc deficient mice were studied in a nonproliferative model of liver growth, fasting for 48 hr followed by a 24 hr refeeding period. Fasting resulted in a decrease in liver mass and liver protein, both of which recovered upon 24 h of refeeding in the c-mycfl/fl;Alb-Cre animals. There was also no effect of reducing c-myc on recovery of liver mass following 2/3 partial hepatectomy. Conclusions c-Myc appears to be dispensable for normal liver growth during the postnatal period, restoration of liver mass following partial hepatectomy and recovery from fasting.

  3. Expression of c-myc, bcl-2 and survivin in cutaneous and oral ...

    African Journals Online (AJOL)

    Expression of c-myc, bcl-2 and survivin in cutaneous and oral squamous cell carcinoma, basal cell carcinoma and actinic keratosis. ... Conclusion: c-myc expressions correspond to the survivin expressions. c-myc expression was stronger in OSCC than in CSCC and BCC, and weaker in AK than in other malignant tumors.

  4. Ezrin mediates c-Myc actions in prostate cancer cell invasion

    DEFF Research Database (Denmark)

    Chuan, Yin Choy; Iglesias Gato, Diego; Fernandez-Perez, L

    2010-01-01

    The forced overexpression of c-Myc in mouse prostate and in normal human prostate epithelial cells results in tumor transformation with an invasive phenotype. How c-Myc regulates cell invasion is poorly understood. In this study, we have investigated the interplay of c-Myc and androgens in the re...

  5. Insertion of the LINE-1 element in the C-MYC gene and immunoreactivity of C-MYC, p53, p21 and p27 proteins in different morphological patterns of the canine TVT

    Directory of Open Access Journals (Sweden)

    C.R.O. Lima

    2016-06-01

    Full Text Available ABSTRACT The canine transmissible venereal tumor (TVT affects the external genitalia of dogs by the natural transplant of viable tumor cells. Thus, this research aimed to diagnose and characterize TVT morphological patterns, identify the insertion of the LINE-1 element in C-MYC gene, by means of the polymerase chain reaction (PCR, and evaluate the immunohistochemical expression of C-MYC, p53, p21 and p27 proteins. The relationship between C-MYC and p53 proteins and their interference on the expression of p21 and p27 were also studied. For that, 20 samples of naturally occurring TVT were used, subjected to cytopathological, histopathological and immunohistochemical analysis, and to molecular diagnosis of neoplasia. The increased tissue expression and the correlation among C-MYC, p53, p21 and p27 proteins indicate reduction and/or loss of their functionality in the TVT microenvironment, with consequent apoptotic suppression, maintenance of cell growth and progression of neoplasia.

  6. C-myc oncogene expression in anal squamous neoplasia.

    Science.gov (United States)

    Ogunbiyi, O A; Scholefield, J H; Rogers, K; Sharp, F; Smith, J H; Polacarz, S V

    1993-01-01

    AIMS: To determine the pattern of c-myc oncogene expression in anal squamous neoplasia and to determine if this could be used as a marker of disease progression. METHODS: The presence and localisation of the c-myc gene product p62 in archival specimens of anal squamous epithelium, normal and neoplastic, was examined using immunohistochemical staining with the monoclonal antibody Myc1-6E10. Ten normal and epithelia, 10 anal intraepithelial neoplasia (AIN) III, and 31 anal squamous cancers were examined. RESULTS: There was a noticeable difference between the staining characteristics of invasive tumours, normal anal epithelium, and AIN III. Intense, diffuse, mixed nuclear and cytoplasmic (n = 14) and exclusively nuclear (n = 8) staining in 22 of 31 (71%) of invasive anal tumours was observed. All positively staining tumours were well differentiated histologically, while the negatively staining nine of 31 (29%) were poorly differentiated (n = 7) and moderately well differentiated (n = 2). In six positively staining tumour sections adjacent areas of AIN III and non-dysplastic anal epithelium had staining characteristics similar to those of the invasive component. Staining in both normal anal epithelium (4/10) and AIN III specimens obtained from patients without a history of invasive disease (8/10) was less intense, focal in distribution, and exclusively nuclear. No difference in staining characteristics could be detected in these two groups. CONCLUSIONS: The results of this study suggest that c-myc oncogene expression is implicated in the pathogenesis of anal squamous neoplasia, and that immunohistochemical staining for c-myc protein may be helpful in identifying those AIN III lesions most likely to progress to invasive tumours. Images PMID:7679417

  7. A critical appraisal of the immunohistochemical detection of the c-myc oncogene product in colorectal cancer.

    Science.gov (United States)

    Jones, D. J.; Ghosh, A. K.; Moore, M.; Schofield, P. F.

    1987-01-01

    Expression of c-myc was studied immunohistochemically in 100 colorectal carcinomas, using a monoclonal antibody, Myc 1-6E10, which is purported to recognize the oncoprotein (p62c-myc) in paraffin-embedded material. In normal epithelium, maturing crypt cells and terminally differentiated surface cells were positive, and proliferating basal crypt cells negative. All carcinomas stained positively, but intensity was independent of histological differentiation, Dukes' stage, DNA ploidy and survival. Staining was predominantly cytoplasmic despite the suspected nuclear location of p62c-myc and there was considerable staining of fibroblasts. When staining was compared in frozen and paraffin-embedded sections fixed in different ways, different patterns were observed. Acetone-fixed frozen sections exhibited weak nuclear and cytoplasmic staining or were negative. In formol-saline fixed frozen sections, there was stronger predominantly nuclear staining. In paraffin-embedded sections staining was predominantly cytoplasmic. This study suggests that c-myc expression is enhanced in the majority of colorectal carcinomas and although independent of clinical behaviour, may be a common event in malignant transformation. However, since staining is affected by fixation and processing, data obtained using Myc 1-6E10 on routinely processed specimens should be interpreted with caution. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:3325094

  8. C-myc proto-oncogene amplification detected by polymerase chain reaction in archival human ovarian carcinomas.

    Science.gov (United States)

    Schreiber, G.; Dubeau, L.

    1990-01-01

    Polymerase chain reaction (PCR) technology was used to examine the state of amplification of the proto-oncogene c-myc in archival ovarian carcinomas. Sequences from the c-myc gene and from a control gene were amplified simultaneously by PCR and the ratios of the two products measured. The results provided an accurate measurement of the relative number of copies of the two genes in each tumor genome if the control and test sequences amplified by PCR were of equal lengths. The results were not affected by the number of PCR cycles used. This technique should facilitate gene amplification studies in clinical medicine. Increased c-myc copy number was found in 17% of the 30 cases examined when a control from the same chromosome as c-myc was used, but in 37% of cases if a control from another chromosome was used. This underlines the importance of the genetic location of the selected control genes for such studies. Images Figure 2 PMID:2205100

  9. [Expression of c-myc protein on rats' brains after brain concussion].

    Science.gov (United States)

    Fang, Wei-Hua; Wang, Dong-Liang; Wang, Feng

    2006-10-15

    To study the changes of expression of c-myc protein on rats' brains after brain concussion. sixty rats were randomly divided into brain concussion groups and control group. The expression of c-myc protein was microscopically observed by immunohistochemical method. No expression of c-myc protein in control group were observed. However, positive expression of c-myc protein in some neurons was seen at 20 min after brain concussion, and reach to the peak at 8h after brain concussion and then decreased gradually. These findings suggest that the detection of c-myc protein could be an index of diagnosis of brain concussion.

  10. Detection of the c-myc oncogene product in testicular cancer.

    OpenAIRE

    Sikora, K; Evan, G; Stewart, J.(DESY, 15738, Zeuthen, Germany); Watson, J. V.

    1985-01-01

    A set of monoclonal antibodies was constructed by immunising mice with peptide fragments of the c-myc oncogene product. One such antibody, Myc 1-6E10 was shown to bind to a 62,000 dalton protein identifiable with the c-myc product (p62c-myc). The antigen recognised was not destroyed by paraffin wax embedding. Myc 1-6E10 was used to characterise the distribution of p62c-myc in archival testicular tumour material. Normal testes expressed only small amounts of p62c-myc. Seminomas showed increase...

  11. MicroRNA-184 Deregulated by the MicroRNA-21 Promotes Tumor Malignancy and Poor Outcomes in Non-small Cell Lung Cancer via Targeting CDC25A and c-Myc.

    Science.gov (United States)

    Lin, Tsang-Chi; Lin, Po-Lin; Cheng, Ya-Wen; Wu, Tzu-Chin; Chou, Ming-Chih; Chen, Chih-Yi; Lee, Huei

    2015-12-01

    MicroRNA (miR)-184 has been reported to have a dual role in human cancers. However, the role of miR-184 in non-small cell lung cancer (NSCLC) remains unclear. Wild-type or mutant CDC25A promoters were constructed by PCR and site-directed mutagenesis to verify whether miR-184 could inhibit CDC25A expression at post-transcription level. Boyden chamber assay was used to assess whether miR-184 could modulate cell invasiveness via targeting CDC25A and c-Myc. We utilized 124 tumors from NSCLC patients to determine miR-184, miR-21, PDCD4 mRNA, c-Myc mRNA, and CDC25A mRNA expression levels by means of real-time PCR analysis. The prognostic value of CDC25A, c-Myc, and miR-184 on overall survival (OS) and relapse-free survival (RFS) was evaluated by Kaplan-Meier and Cox regression analysis. MiR-184 suppressed CDC25A expression by enhancing the instability of its mRNA as a result of miR-184 binding to its coding region. An increase in CDC25A expression by means of a reduction in miR-184 promotes cell invasiveness. Moreover, a concomitant increase in CDC25A and c-Myc expression as a result of decreased miR-184 via the miR-21-mediated PDCD4 reduction is responsible for cell invasiveness. Among patients, miR-184 expression in lung tumors was found to correlate negatively with CDC25A mRNA, c-Myc mRNA, and miR-21 expression, but was positively related to PDCD4 mRNA expression. High-miR-184, High-CDC25A, or high-c-Myc mRNA tumors exhibited shorter OS and RFS periods than their counterparts. The worst OS and RFS were observed in low-miR-184/high-CDC25A/high-c-Myc tumors, followed by low-miR-184 /high-CDC25A, low-miR-184/high-c-Myc, high-c-Myc, and high-CDC25A tumors. MiR-184 as a tumor suppressor miR inhibits cell proliferation and invasion capability via targeting CDC25A and c-Myc. Low miR-184 level may predict worse prognosis in NSCLC patients.

  12. Post-transcriptional control of c-myc proto-oncogene expression by glucocorticoid hormones in human T lymphoblastic leukemia cells

    Energy Technology Data Exchange (ETDEWEB)

    Maroder, M.; Vacca, A.; Screpanti, I.; Petrangeli, E.; Frati, L. (Univ. La Sapienza, Roma (Italy)); Martinotti, S.; Gulino, A. (Univ. of L' Aquila (Italy))

    1990-03-11

    The authors have studied the regulation of the human c-myc proto-oncogene by glucocorticoid hormones in T lymphoblastic leukemic cells. A significant decrease (50%) of the steady state levels of c-myc mRNA was observed as early as 3 hours after dexamethasone treatment of CEM-1.3 human lymphoma cells, reaching less than 5% values, with respect to untreated cells, 24 hours after hormone administration. Nuclear run-on experiments showed no modifications of the transcriptional rate from the first exon. However, a slight decrease (15%) of the transcript elongation from the first exon/first intron boundary was observed in the dexamethasone-treated cells. Using actinomycin D to block gene transcription, they observed a significant increase in the rate of c-myc RNA specific decay after dexamethasone treatment. The data suggest that dexamethasone is able to inhibit human c-myc gene expression primarily at the post-transcriptional level, through the synthesis of hormone-transcriptional level, through the synthesis of hormone-induced regulatory protein(s) controlling c-myc transcript stability.

  13. Far upstream element-binding protein-1, a novel caspase substrate, acts as a cross-talker between apoptosis and the c-myc oncogene.

    Science.gov (United States)

    Jang, M; Park, B C; Kang, S; Chi, S-W; Cho, S; Chung, S J; Lee, S C; Bae, K-H; Park, S G

    2009-03-26

    Far upstream element-binding protein-1 (FBP-1) binds to an upstream element of the c-myc promoter and regulates the c-myc mRNA level. Earlier, FBP-1 was identified as a candidate substrate of caspase-7. Here, we report that FBP-1 is cleaved by executor caspases, both in vitro and during apoptosis. Cleavage occurs at the caspase consensus site (DQPD(74)) located within the classical bipartite nuclear localization signal sequence. In cells subjected to apoptotic stimuli, the caspase-mediated cleavage of FBP-1 leads to its decreased presence in the nucleus, concomitant with the marked downregulation of c-Myc and its various target proteins. By contrast, cells transfected with a non-cleavable mutant of FBP-1 (D74A) maintain higher levels of c-Myc and are protected from apoptosis. On the basis of these results, we suggest that the oncogenic potential of c-Myc is 'switched off' after apoptosis induction as a consequence of the caspase-mediated cleavage of FBP-1.

  14. Endogenous c-Myc is essential for p53-induced apoptosis in response to DNA damage in vivo

    NARCIS (Netherlands)

    Phesse, T. J.; Myant, K. B.; Cole, A. M.; Ridgway, R. A.; Pearson, H.; Muncan, V.; van den Brink, G. R.; Vousden, K. H.; Sears, R.; Vassilev, L. T.; Clarke, A. R.; Sansom, O. J.

    2014-01-01

    Recent studies have suggested that C-MYC may be an excellent therapeutic cancer target and a number of new agents targeting C-MYC are in preclinical development. Given most therapeutic regimes would combine C-MYC inhibition with genotoxic damage, it is important to assess the importance of C-MYC

  15. The nucleolar ubiquitin-specific protease USP36 deubiquitinates and stabilizes c-Myc

    Science.gov (United States)

    Sun, Xiao-Xin; He, Xia; Yin, Li; Komada, Masayuki; Sears, Rosalie C.; Dai, Mu-Shui

    2015-01-01

    c-Myc protein stability and activity are tightly regulated by the ubiquitin-proteasome system. Aberrant stabilization of c-Myc contributes to many human cancers. c-Myc is ubiquitinated by SCFFbw7 (a SKP1-cullin-1-F-box complex that contains the F-box and WD repeat domain-containing 7, Fbw7, as the F-box protein) and several other ubiquitin ligases, whereas it is deubiquitinated and stabilized by ubiquitin-specific protease (USP) 28. The bulk of c-Myc degradation appears to occur in the nucleolus. However, whether c-Myc is regulated by deubiquitination in the nucleolus is not known. Here, we report that the nucleolar deubiquitinating enzyme USP36 is a novel c-Myc deubiquitinase. USP36 interacts with and deubiquitinates c-Myc in cells and in vitro, leading to the stabilization of c-Myc. This USP36 regulation of c-Myc occurs in the nucleolus. Interestingly, USP36 interacts with the nucleolar Fbw7γ but not the nucleoplasmic Fbw7α. However, it abolished c-Myc degradation mediated both by Fbw7γ and by Fbw7α. Consistently, knockdown of USP36 reduces the levels of c-Myc and suppresses cell proliferation. We further show that USP36 itself is a c-Myc target gene, suggesting that USP36 and c-Myc form a positive feedback regulatory loop. High expression levels of USP36 are found in a subset of human breast and lung cancers. Altogether, these results identified USP36 as a crucial and bono fide deubiquitinating enzyme controlling c-Myc’s nucleolar degradation pathway. PMID:25775507

  16. Celastrol inhibits chondrosarcoma proliferation, migration and invasion through suppression CIP2A/c-MYC signaling pathway

    Directory of Open Access Journals (Sweden)

    Jinhui Wu

    2017-05-01

    Full Text Available Chondrosarcomas (CS is the second most frequent tumors of cartilage origin. A small compound extracted from Thunder God Vine (Tripterygium wilfordii Hook. F. called celastrol can directly bound CIP2A protein and effectively inhibit cell proliferation and induce apoptosis in several cancer cells. However, little knowledge is concern about the important role of CIP2A in CS patients and the therapeutic value of celastrol on CS. Our results showed that CIP2A and c-MYC were verified to be oncoproteins by detecting their mRNA and protein expression in 10 human CS tissues by qRT-PCR and Western blots. After treatment of celastrol, the proliferation, migration and invasion were significantly inhibited; whereas the apoptosis was largely induced in human CS cell lines. In addition, celastrol inhibited the expression of CIP2A, c-MYC, and suppressed apoptotic proteins BAX and caspase-8 in human CS cells, on the other hand, it induced the expression of antiapoptotic protein Bcl-2. Finally, knockdown of CIP2A also inhibited the migration and invasion and induced apoptosis of human CS cells. To sum up, we found that celastrol had effects on inhibiting proliferation, migration, invasion and inducing apoptosis through suppression CIP2A/c-MYC signaling pathway in vitro, which may provide a new therapeutic regimen for CS.

  17. O-GlcNAc transferase integrates metabolic pathways to regulate the stability of c-MYC in human prostate cancer cells.

    Science.gov (United States)

    Itkonen, Harri M; Minner, Sarah; Guldvik, Ingrid J; Sandmann, Mareike Julia; Tsourlakis, Maria Christina; Berge, Viktor; Svindland, Aud; Schlomm, Thorsten; Mills, Ian G

    2013-08-15

    Metabolic disruptions that occur widely in cancers offer an attractive focus for generalized treatment strategies. The hexosamine biosynthetic pathway (HBP) senses metabolic status and produces an essential substrate for O-linked β-N-acetylglucosamine transferase (OGT), which glycosylates and thereby modulates the function of its target proteins. Here, we report that the HBP is activated in prostate cancer cells and that OGT is a central regulator of c-Myc stability in this setting. HBP genes were overexpressed in human prostate cancers and androgen regulated in cultured human cancer cell lines. Immunohistochemical analysis of human specimens (n = 1987) established that OGT is upregulated at the protein level and that its expression correlates with high Gleason score, pT and pN stages, and biochemical recurrence. RNA interference-mediated siliencing or pharmacologic inhibition of OGT was sufficient to decrease prostate cancer cell growth. Microarray profiling showed that the principal effects of OGT inhibition in prostate cancer cells were related to cell-cycle progression and DNA replication. In particular, c-MYC was identified as a candidate upstream regulator of OGT target genes and OGT inhibition elicited a dose-dependent decrease in the levels of c-MYC protein but not c-MYC mRNA in cell lines. Supporting this relationship, expression of c-MYC and OGT was tightly correlated in human prostate cancer samples (n = 1306). Our findings identify HBP as a modulator of prostate cancer growth and c-MYC as a key target of OGT function in prostate cancer cells.

  18. [Expression of c-myc and HER-1 genes in the development of human esophageal cancer].

    Science.gov (United States)

    Liang, Y Y

    1991-05-01

    To understand the possible role of oncogenes in the development of human esophageal cancer, the expression of c-myc and HER-1 genes was studied by in situ hybridization. The results showed that: (a) The c-myc and HER-1 protooncogenes were transcriptionally activated (b) Activation of c-myc gene was observed in hyperplastic cells and carcinoma cells. (c) The degree of pathological changes of the esophageal epithelium was related to the level of c-myc transcription, the highest level of c-myc expression was seen in invasive carcinoma cells. (d) Expression of HER-1 gene in carcinoma cells is higher than that in normal and adjacent non-tumor cells, but its frequency is lower than that of c-myc gene.

  19. c-myc oncogene product expression and prognosis in operable breast cancer.

    Science.gov (United States)

    Locker, A. P.; Dowle, C. S.; Ellis, I. O.; Elston, C. W.; Blamey, R. W.; Sikora, K.; Evan, G.; Robins, R. A.

    1989-01-01

    The 62 kDa protein product of the c-myc oncogene (p62 c-myc) is thought to be involved in the control of normal cellular proliferation and differentiation. We have measured oncoprotein levels using a flow cytometric assay in 141 operable breast cancers and have correlated levels with prognostic variables, patient survival and disease free intervals. High levels of p62 c-myc were associated with well differentiated tumours. There was no correlation with tumour DNA index, lymph node or oestrogen receptor status. C-myc oncoprotein levels were not predictive of patient survival or disease free interval. This relationship of oncoprotein levels with tumour histological grade is in keeping with the suggestion that the c-myc oncogene is important in the control of cellular differentiation. The other findings imply that measurement of c-myc oncoprotein levels does not yield useful prognostic information. PMID:2679850

  20. Small Molecules Targeting c-Myc Oncogene: Promising Anti-Cancer Therapeutics

    OpenAIRE

    Chen, Bing-Jia; Wu, Yan-Ling; Tanaka, Yoshimasa; ZHANG, Wen

    2014-01-01

    The nuclear transcription factor c-Myc is a member of the Myc gene family with multiple functions and located on band q24.1 of chromosome 8. The c-Myc gene is activated by chromosomal translocation, rearrangement, and amplification. Its encoded protein transduces intracellular signals to the nucleus, resulting in the regulation of cell proliferation, differentiation, and apoptosis, and has the ability to transform cells and bind chromosomal DNA. c-Myc also plays a critical role in malignant t...

  1. Amplification and expression of a cellular oncogene (c-myc) in human gastric adenocarcinoma cells.

    Science.gov (United States)

    Shibuya, M; Yokota, J; Ueyama, Y

    1985-01-01

    Three of 16 human gastric adenocarcinoma samples, maintained as solid tumors in nude mice, were found to carry amplified c-myc genes. In two samples with a high degree of c-myc DNA amplification (15- to 30-fold), double minute chromosomes were observed in karyotype analysis. The level of c-myc RNA was markedly elevated in a rapidly growing and poorly differentiated tumor, whereas it was only slightly elevated in a slowly growing and more differentiated tumor. Images PMID:2579323

  2. c-Myc Oncoprotein: A Dual Pathogenic Role in Neoplasia and Cardiovascular Diseases?

    Science.gov (United States)

    Napoli, Claudio; Lerman, Lilach O; de Nigris, Filomena; Sica, Vincenzo

    2002-01-01

    Abstract A growing body of evidence indicates that c-Myc can play a pivotal role both in neoplasia and cardiovascular diseases. Indeed, alterations of the basal machinery of the cell and perturbations of c-Myc-dependent signaling network are involved in the pathogenesis of certain cardiovascular disorders. Down-regulation of c-Myc induced by intervention with antioxidants or by antisense technology may protect the integrity of the arterial wall as well as neoplastic tissues. Further intervention studies are necessary to investigate the effects of tissue-specific block of c-Myc overexpression in the development of cardiovascular diseases. PMID:11988837

  3. Growth-promoting and tumourigenic activity of c-Myc is suppressed by Hhex.

    Science.gov (United States)

    Marfil, V; Blazquez, M; Serrano, F; Castell, J V; Bort, R

    2015-06-04

    c-Myc transcription factor is a key protein involved in cellular growth, proliferation and metabolism. c-Myc is one of the most frequently activated oncogenes, highlighting the need to identify intracellular molecules that interact directly with c-Myc to suppress its function. Here we show that Hhex is able to interact with the basic region/helix-loop-helix/leucine zipper of c-Myc. Knockdown of Hhex increases proliferation rate in hepatocellular carcinoma cells, whereas Hhex expression cell-autonomously reduces cell proliferation rate in multiple cell lines by increasing G1 phase length through a c-Myc-dependent mechanism. Global transcriptomic analysis shows that Hhex counter-regulates multiple c-Myc targets involved in cell proliferation and metabolism. Concomitantly, Hhex expression leads to reduced cell size, lower levels of cellular RNA, downregulation of metabolism-related genes, decreased sensitivity to methotrexate and severe reduction in the ability to form tumours in nude mouse xenografts, all indicative of decreased c-Myc activity. Our data suggest that Hhex is a novel regulator of c-Myc function that limits c-Myc activity in transformed cells.

  4. Small molecules targeting c-Myc oncogene: promising anti-cancer therapeutics.

    Science.gov (United States)

    Chen, Bing-Jia; Wu, Yan-Ling; Tanaka, Yoshimasa; Zhang, Wen

    2014-01-01

    The nuclear transcription factor c-Myc is a member of the Myc gene family with multiple functions and located on band q24.1 of chromosome 8. The c-Myc gene is activated by chromosomal translocation, rearrangement, and amplification. Its encoded protein transduces intracellular signals to the nucleus, resulting in the regulation of cell proliferation, differentiation, and apoptosis, and has the ability to transform cells and bind chromosomal DNA. c-Myc also plays a critical role in malignant transformation. The abnormal over-expression of c-Myc is frequently observed in some tumors, including carcinomas of the breast, colon, and cervix, as well as small-cell lung cancer, osteosarcomas, glioblastomas, and myeloid leukemias, therefore making it a possible target for anticancer therapy. In this minireview, we summarize unique characteristics of c-Myc and therapeutic strategies against cancer using small molecules targeting the oncogene, and discuss the prospects in the development of agents targeting c-Myc, in particular G-quadruplexes formed in c-Myc promoter and c-Myc/Max dimerization. Such information will be of importance for the research and development of c-Myc-targeted drugs.

  5. Involvement of SIRT1 in hypoxic down-regulation of c-Myc and β-catenin and hypoxic preconditioning effect of polyphenols

    Energy Technology Data Exchange (ETDEWEB)

    Hong, Kyung-Soo [Department of Biochemistry, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Research Center for Ischemic Tissue regeneration, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Park, Jun-Ik [Department of Biochemistry, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Kim, Mi-Ju; Kim, Hak-Bong; Lee, Jae-Won [Department of Biochemistry, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Research Center for Ischemic Tissue regeneration, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Dao, Trong Tuan; Oh, Won Keun [BK21 Project Team, College of Pharmacy, Chosun University, Gwangju (Korea, Republic of); Kang, Chi-Dug, E-mail: kcdshbw@pusan.ac.kr [Department of Biochemistry, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Kim, Sun-Hee, E-mail: ksh7738@pusan.ac.kr [Department of Biochemistry, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Research Center for Ischemic Tissue regeneration, Pusan National University School of Medicine, Yangsan (Korea, Republic of)

    2012-03-01

    SIRT1 has been found to function as a Class III deacetylase that affects the acetylation status of histones and other important cellular nonhistone proteins involved in various cellular pathways including stress responses and apoptosis. In this study, we investigated the role of SIRT1 signaling in the hypoxic down-regulations of c-Myc and β-catenin and hypoxic preconditioning effect of the red wine polyphenols such as piceatannol, myricetin, quercetin and resveratrol. We found that the expression of SIRT1 was significantly increased in hypoxia-exposed or hypoxic preconditioned HepG2 cells, which was closely associated with the up-regulation of HIF-1α and down-regulation of c-Myc and β-catenin expression via deacetylation of these proteins. In addition, blockade of SIRT1 activation using siRNA or amurensin G, a new potent SIRT1 inhibitor, abolished hypoxia-induced HIF-1α expression but increased c-Myc and β-catenin expression. SIRT1 was also found to stabilize HIF-1α protein and destabilize c-Myc, β-catenin and PHD2 under hypoxia. We also found that myricetin, quercetin, piceatannol and resveratrol up-regulated HIF-1α and down-regulated c-Myc, PHD2 and β-catenin expressions via SIRT1 activation, in a manner that mimics hypoxic preconditioning. This study provides new insights of the molecular mechanisms of hypoxic preconditioning and suggests that polyphenolic SIRT1 activators could be used to mimic hypoxic/ischemic preconditioning. -- Graphical abstract: Polyphenols mimicked hypoxic preconditioning by up-regulating HIF-1α and SIRT1 and down-regulating c-Myc, PHD2, and β-catenin. HepG2 cells were pretreated with the indicated doses of myricetin (MYR; A), quercetin (QUR; B), or piceatannol (PIC; C) for 4 h and then exposed to hypoxia for 4 h. Levels of HIF-1α, SIRT1, c-Myc, β-catenin, and PHD2 were determined by western blot analysis. The data are representative of three individual experiments. Highlights: ► SIRT1 expression is increased in hypoxia

  6. A balance between self-renewal and commitment in the murine erythroleukemia cells with the transferred c-myc gene; an in vitro stochastic model.

    Science.gov (United States)

    Yamamoto, T; Masuko, K; Takada, S; Kume, T U; Obinata, M

    1989-11-01

    When murine erythroleukemia (MEL) cells, having the transferred rat c-myc gene under the control of human metallothionein II gene promoter, are induced to differentiate with dimethyl sulfoxide (DMSO), the level of differentiation is dependent on the c-myc levels which are modulated by the Zn++ ion. The clonal transformant cell line (38-2) can continuously grow in the presence of both DMSO and Zn++ ion. The proportion of differentiated cells in a population of the continuous culture is strongly affected by the concentration of Zn++ ions. These results suggested that a balance between self-renewal and commitment to differentiation of MEL cells is determined by the c-myc level, and that this cell line may be suitable for studying the stochastic process of growth and differentiation of hemopoietic stem cells.

  7. Tumor suppressor NDRG2 inhibits glycolysis and glutaminolysis in colorectal cancer cells by repressing c-Myc expression.

    Science.gov (United States)

    Xu, Xinyuan; Li, Jianying; Sun, Xiang; Guo, Yan; Chu, Dake; Wei, Li; Li, Xia; Yang, Guodong; Liu, Xinping; Yao, Libo; Zhang, Jian; Shen, Lan

    2015-09-22

    Cancer cells use glucose and glutamine as the major sources of energy and precursor intermediates, and enhanced glycolysis and glutamimolysis are the major hallmarks of metabolic reprogramming in cancer. Oncogene activation and tumor suppressor gene inactivation alter multiple intracellular signaling pathways that affect glycolysis and glutaminolysis. N-Myc downstream regulated gene 2 (NDRG2) is a tumor suppressor gene inhibiting cancer growth, metastasis and invasion. However, the role and molecular mechanism of NDRG2 in cancer metabolism remains unclear. In this study, we discovered the role of the tumor suppressor gene NDRG2 in aerobic glycolysis and glutaminolysis of cancer cells. NDRG2 inhibited glucose consumption and lactate production, glutamine consumption and glutamate production in colorectal cancer cells. Analysis of glucose transporters and the catalytic enzymes involved in glycolysis revealed that glucose transporter 1 (GLUT1), hexokinase 2 (HK2), pyruvate kinase M2 isoform (PKM2) and lactate dehydrogenase A (LDHA) was significantly suppressed by NDRG2. Analysis of glutamine transporter and the catalytic enzymes involved in glutaminolysis revealed that glutamine transporter ASC amino-acid transporter 2 (ASCT2) and glutaminase 1 (GLS1) was also significantly suppressed by NDRG2. Transcription factor c-Myc mediated inhibition of glycolysis and glutaminolysis by NDRG2. More importantly, NDRG2 inhibited the expression of c-Myc by suppressing the expression of β-catenin, which can transcriptionally activate C-MYC gene in nucleus. In addition, the growth and proliferation of colorectal cancer cells were suppressed significantly by NDRG2 through inhibition of glycolysis and glutaminolysis. Taken together, these findings indicate that NDRG2 functions as an essential regulator in glycolysis and glutaminolysis via repression of c-Myc, and acts as a suppressor of carcinogenesis through coordinately targeting glucose and glutamine transporter, multiple catalytic

  8. Acetylation of the c-MYC oncoprotein is required for cooperation with the HTLV-1 p30{sup II} accessory protein and the induction of oncogenic cellular transformation by p30{sup II}/c-MYC

    Energy Technology Data Exchange (ETDEWEB)

    Romeo, Megan M.; Ko, Bookyung; Kim, Janice; Brady, Rebecca; Heatley, Hayley C.; He, Jeffrey; Harrod, Carolyn K.; Barnett, Braden [Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design, and Delivery, Southern Methodist University, Dallas, TX 75275-0376 (United States); Ratner, Lee [Departments of Medicine and Molecular Microbiology, Washington University School of Medicine, St. Louis, MO 63110 (United States); Lairmore, Michael D. [University of California-Davis, School of Veterinary Medicine, One Shields Avenue, Davis, CA 95618 (United States); Martinez, Ernest [Department of Biochemistry, University of California, Riverside, CA 92521 (United States); Lüscher, Bernhard [Institute of Biochemistry, Klinikum, RWTH Aachen University, Pauwelsstrasse 30, 52057 Aachen (Germany); Robson, Craig N. [Northern Institute for Cancer Research, Newcastle University, The Medical School, Newcastle upon Tyne, NE2 4HH (United Kingdom); Henriksson, Marie [Department of Microbiology, Cell and Tumor Biology, Karolinska Institutet, Stockholm (Sweden); Harrod, Robert, E-mail: rharrod@smu.edu [Laboratory of Molecular Virology, Department of Biological Sciences, and The Dedman College Center for Drug Discovery, Design, and Delivery, Southern Methodist University, Dallas, TX 75275-0376 (United States)

    2015-02-15

    The human T-cell leukemia retrovirus type-1 (HTLV-1) p30{sup II} protein is a multifunctional latency-maintenance factor that negatively regulates viral gene expression and deregulates host signaling pathways involved in aberrant T-cell growth and proliferation. We have previously demonstrated that p30{sup II} interacts with the c-MYC oncoprotein and enhances c-MYC-dependent transcriptional and oncogenic functions. However, the molecular and biochemical events that mediate the cooperation between p30{sup II} and c-MYC remain to be completely understood. Herein we demonstrate that p30{sup II} induces lysine-acetylation of the c-MYC oncoprotein. Acetylation-defective c-MYC Lys→Arg substitution mutants are impaired for oncogenic transformation with p30{sup II} in c-myc{sup −/−} HO15.19 fibroblasts. Using dual-chromatin-immunoprecipitations (dual-ChIPs), we further demonstrate that p30{sup II} is present in c-MYC-containing nucleoprotein complexes in HTLV-1-transformed HuT-102 T-lymphocytes. Moreover, p30{sup II} inhibits apoptosis in proliferating cells expressing c-MYC under conditions of genotoxic stress. These findings suggest that c-MYC-acetylation is required for the cooperation between p30{sup II}/c-MYC which could promote proviral replication and contribute to HTLV-1-induced carcinogenesis. - Highlights: • Acetylation of c-MYC is required for oncogenic transformation by HTLV-1 p30{sup II}/c-MYC. • Acetylation-defective c-MYC mutants are impaired for foci-formation by p30{sup II}/c-MYC. • The HTLV-1 p30{sup II} protein induces lysine-acetylation of c-MYC. • p30{sup II} is present in c-MYC nucleoprotein complexes in HTLV-1-transformed T-cells. • HTLV-1 p30{sup II} inhibits apoptosis in c-MYC-expressing proliferating cells.

  9. Cooverexpression of EpCAM and c-myc genes in malignant breast ...

    Indian Academy of Sciences (India)

    most important proto-oncogenes routinely overexpressed in breast cancer. However, cooverexpression of EpCAM and c-myc genes has not been investigated in breast cancer tissues, particularly in Iranian population. The aim of this study was to assess the expression of EpCAM and c-myc genes in malignant breast cancer ...

  10. B-cell activating factor and v-Myc myelocytomatosis viral oncogene homolog (c-Myc) influence progression of chronic lymphocytic leukemia.

    Science.gov (United States)

    Zhang, Weizhou; Kater, Arnon P; Widhopf, George F; Chuang, Han-Yu; Enzler, Thomas; James, Danelle F; Poustovoitov, Maxim; Tseng, Ping-Hui; Janz, Siegfried; Hoh, Carl; Herschman, Harvey; Karin, Michael; Kipps, Thomas J

    2010-11-02

    Mice bearing a v-Myc myelocytomatosis viral oncogene homolog (c-Myc) transgene controlled by an Ig-alpha heavy-chain enhancer (iMyc(Cα) mice) rarely develop lymphomas but instead have increased rates of memory B-cell turnover and impaired antibody responses to antigen. We found that male progeny of iMyc(Cα) mice mated with mice transgenic (Tg) for CD257 (B-cell activating factor, BAFF) developed CD5(+) B-cell leukemia resembling human chronic lymphocytic leukemia (CLL), which also displays a male gender bias. Surprisingly, leukemic cells of Myc/Baff Tg mice expressed higher levels of c-Myc than did B cells of iMyc(Cα) mice. We found that CLL cells of many patients with progressive disease also expressed high amounts of c-MYC, particularly CLL cells whose survival depends on nurse-like cells (NLC), which express high-levels of BAFF. We find that BAFF could enhance CLL-cell expression of c-MYC via activation the canonical IκB kinase (IKK)/NF-κB pathway. Inhibition of the IKK/NF-κB pathway in mouse or human leukemia cells blocked the capacity of BAFF to induce c-MYC or promote leukemia-cell survival and significantly impaired disease progression in Myc/Baff Tg mice. This study reveals an important relationship between BAFF and c-MYC in CLL which may affect disease development and progression, and suggests that inhibitors of the canonical NF-κB pathway may be effective in treatment of patients with this disease.

  11. B Lymphocyte commitment program is driven by the proto-oncogene c-Myc.

    Science.gov (United States)

    Vallespinós, Mireia; Fernández, David; Rodríguez, Lorena; Alvaro-Blanco, Josué; Baena, Esther; Ortiz, Maitane; Dukovska, Daniela; Martínez, Dolores; Rojas, Ana; Campanero, Miguel R; Moreno de Alborán, Ignacio

    2011-06-15

    c-Myc, a member of the Myc family of transcription factors, is involved in numerous biological functions including the regulation of cell proliferation, differentiation, and apoptosis in various cell types. Of all of its functions, the role of c-Myc in cell differentiation is one of the least understood. We addressed the role of c-Myc in B lymphocyte differentiation. We found that c-Myc is essential from early stages of B lymphocyte differentiation in vivo and regulates this process by providing B cell identity via direct transcriptional regulation of the ebf-1 gene. Our data show that c-Myc influences early B lymphocyte differentiation by promoting activation of B cell identity genes, thus linking this transcription factor to the EBF-1/Pax-5 pathway.

  12. The translocated c-myc oncogene of Raji Burkitt lymphoma cells is not expressed in human lymphoblastoid cells.

    Science.gov (United States)

    Nishikura, K; Erikson, J; ar-Rushdi, A; Huebner, K; Croce, C M

    1985-01-01

    We hybridized Raji Burkitt lymphoma cells, which carry a t(8;14) chromosome translocation, with human lymphoblastoid cells to study the expression of the translocated cellular myc oncogene (c-myc) in the hybrid cells. In Raji cells the c-myc oncogene is translocated to a switch region of the gamma heavy chain locus (S gamma). Because of sequence alterations in the 5' exon of the translocated c-myc oncogene in this cell line, it is possible to distinguish the transcripts of the translocated c-myc gene and of the normal c-myc gene. S1 nuclease protection experiments with a c-myc first exon probe indicate that Raji cells express predominantly the translocated c-myc gene, while the level of expression of the normal c-myc gene is less than 2% of that of the translocated c-myc gene. Somatic cell hybrids between Raji and human lymphoblastoid cells retain the lymphoblastoid phenotype and express only the normal c-myc oncogene. This result indicates that the activation of a c-myc oncogene translocated to a S region depends on the stage of B-cell differentiation of the cells harboring the translocated c-myc gene and not on alterations in the structure of the translocated c-myc oncogene. Images PMID:3857623

  13. Glutathione Depletion Induced by c-Myc Downregulation Triggers Apoptosis on Treatment with Alkylating Agents1

    Science.gov (United States)

    Biroccio, Annamaria; Benassi, Barbara; Fiorentino, Francesco; Zupi, Gabriella

    2004-01-01

    Abstract Here we investigate the mechanism(s) involved in the c-Myc-dependent drug response of melanoma cells. By using three M14-derived c-Myc low-expressing clones, we demonstrate that alkylating agents, cisplatin and melphalan, trigger apoptosis in the c-Myc antisense transfectants, but not in the parental line. On the contrary, topoisomerase inhibitors, adriamycin and camptothecin, induce apoptosis to the same extent regardless of c-Myc expression. Because we previously demonstrated that c-Myc downregulation decreases glutathione (GSH) content, we evaluated the role of GSH in the apoptosis induced by the different drugs. In control cells treated with one of the alkylating agents or the others, GSH depletion achieved by l-buthionine-sulfoximine preincubation opens the apoptotic pathway. The apoptosis proceeded through early Bax relocalization, cytochrome c release, and concomitant caspase-9 activation, whereas reactive oxygen species production and alteration of mitochondria membrane potential were late events. That GSH was determining in the c-Myc-dependent drug-induced apoptosis was demonstrated by altering the intracellular GSH content of the c-Myc low-expressing cells up to the level of controls. Indeed, GSH ethyl ester-mediated increase of GSH abrogated apoptosis induced by cisplatin and melphalan by inhibition of Bax/cytochrome c redistribution. The relationship among c-Myc, GSH content, and the response to alkylating agent has been also evaluated in the M14 Myc overexpressing clones as well as in the melanoma JR8 c-Myc antisense transfectants. All together, these results demonstrate that GSH plays a key role in governing c-Myc-dependent drug-induced apoptosis. PMID:15153331

  14. Detection of the c-myc oncogene product in testicular cancer.

    Science.gov (United States)

    Sikora, K.; Evan, G.; Stewart, J.; Watson, J. V.

    1985-01-01

    A set of monoclonal antibodies was constructed by immunising mice with peptide fragments of the c-myc oncogene product. One such antibody, Myc 1-6E10 was shown to bind to a 62,000 dalton protein identifiable with the c-myc product (p62c-myc). The antigen recognised was not destroyed by paraffin wax embedding. Myc 1-6E10 was used to characterise the distribution of p62c-myc in archival testicular tumour material. Normal testes expressed only small amounts of p62c-myc. Seminomas showed increased nuclear and cytoplasmic staining. Undifferentiated teratoma showed little activity, whereas p62c-myc was abundant in the nuclei of differentiating epithelial structures, yolk sacs and embryoid bodies. Only small amounts of p62c-myc were seen in the tumours of 5 patients who subsequently died from their disease. Images Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:4027160

  15. The proto-oncogene c-myc regulates antibody secretion and Ig class switch recombination.

    Science.gov (United States)

    Fernández, David; Ortiz, Maitane; Rodríguez, Lorena; García, Arancha; Martinez, Dolores; Moreno de Alborán, Ignacio

    2013-06-15

    The immune response involves the generation of Ab-secreting cells and memory B cells through a process called terminal B lymphocyte differentiation. This program requires the transcriptional repressor Blimp-1, which inhibits c-myc expression and terminates proliferation. Although the role of c-Myc in cell proliferation is well characterized, it is not known whether it has other functions in terminal differentiation. In this study, we show that c-Myc not only regulates cell proliferation, but it is also essential for Ab-secreting cell function and differentiation in vivo. c-Myc-deficient B lymphocytes hypersecrete IgM and do not undergo Ig class switch recombination (CSR). CSR has been previously linked to proliferation, and in this study we mechanistically link class switching and proliferation via c-Myc. We observed that c-Myc regulates CSR by transcriptionally activating the B cell-specific factor activation-induced cytidine deaminase. By linking cell proliferation and CSR, c-Myc is thus a critical component for a potent immune response.

  16. Translocation of an immunoglobulin kappa locus to a region 3' of an unrearranged c-myc oncogene enhances c-myc transcription.

    Science.gov (United States)

    Erikson, J; Nishikura, K; ar-Rushdi, A; Finan, J; Emanuel, B; Lenoir, G; Nowell, P C; Croce, C M

    1983-01-01

    We have studied somatic cell hybrids between mouse myeloma and JI Burkitt lymphoma cells carrying a t(2;8) chromosome translocation for the expression of human kappa chains. and for the presence and rearrangements of the human c-myc oncogene and kappa chain genes. Our results indicate that the c-myc oncogene is unrearranged and remains on the 8q+ chromosome of JI cells. Two rearranged C kappa genes were detected: the expressed allele on normal chromosome 2 and the excluded kappa allele that was translocated from chromosome 2 to the involved chromosome 8 (8q+). The distribution of V kappa and C kappa genes in hybrid clones retaining different human chromosomes indicated that C kappa is distal to V kappa on 2p and that the breakpoint in this Burkitt lymphoma is within the region carrying V kappa genes. High levels of transcripts of the c-myc gene were found when it resided on the 8q+ chromosome but not on the normal chromosome 8, demonstrating that translocation of a kappa locus to region distal to the c-myc oncogene enhances c-myc transcription. Images PMID:6424112

  17. A genetic screen to identify genes that rescue the slow growth phenotype of c-myc null fibroblasts

    NARCIS (Netherlands)

    Berns, K.; Hijmans, E.M.; Koh, E.; Daley, G.Q.; Bernards, R.A.

    2000-01-01

    The c-myc gene is frequently over-expressed in human cancers and is involved in regulation of proliferation, differentiation and apoptosis. c-Myc is a transcription factor that acts primarily by regulating the expression of other genes. However, it has been very difficult to identify bona fide c-Myc

  18. c-Myc is essential to prevent endothelial pro-inflammatory senescent phenotype.

    Directory of Open Access Journals (Sweden)

    Victoria Florea

    Full Text Available The proto-oncogene c-Myc is vital for vascular development and promotes tumor angiogenesis, but the mechanisms by which it controls blood vessel growth remain unclear. In the present work we investigated the effects of c-Myc knockdown in endothelial cell functions essential for angiogenesis to define its role in the vasculature. We provide the first evidence that reduction in c-Myc expression in endothelial cells leads to a pro-inflammatory senescent phenotype, features typically observed during vascular aging and pathologies associated with endothelial dysfunction. c-Myc knockdown in human umbilical vein endothelial cells using lentivirus expressing specific anti-c-Myc shRNA reduced proliferation and tube formation. These functional defects were associated with morphological changes, increase in senescence-associated-β-galactosidase activity, upregulation of cell cycle inhibitors and accumulation of c-Myc-deficient cells in G1-phase, indicating that c-Myc knockdown in endothelial cells induces senescence. Gene expression analysis of c-Myc-deficient endothelial cells showed that senescent phenotype was accompanied by significant upregulation of growth factors, adhesion molecules, extracellular-matrix components and remodeling proteins, and a cluster of pro-inflammatory mediators, which include Angptl4, Cxcl12, Mdk, Tgfb2 and Tnfsf15. At the peak of expression of these cytokines, transcription factors known to be involved in growth control (E2f1, Id1 and Myb were downregulated, while those involved in inflammatory responses (RelB, Stat1, Stat2 and Stat4 were upregulated. Our results demonstrate a novel role for c-Myc in the prevention of vascular pro-inflammatory phenotype, supporting an important physiological function as a central regulator of inflammation and endothelial dysfunction.

  19. A cyclometallated iridium(III) complex as a c-myc G-quadruplex stabilizer and down-regulator of c-myc oncogene expression.

    Science.gov (United States)

    Yang, H; Ma, V P-Y; Chan, D S-H; He, H-Z; Leung, C-H; Ma, D-L

    2013-01-01

    A new cyclometallated iridium(III) complex with the 2,2'-biquinoline N-donor ligand has been synthesized and characterized. The interaction and affinity of the complex towards c-myc G-quadruplex and duplex DNA have been investigated using UV/Vis spectroscopy and gel mobility shift assay. These studies revealed that complex 1 binds to c-myc G-quadruplexes (Pu22 and Pu27) with high affinity but does not interact with duplex DNA either by intercalation or groove binding. The ability of 1 to stabilize c-myc G-quadruplex DNA in vitro has also been examined through a PCR stop assay and a cell-based luciferase reporter assay. Complex 1 displays promising cytotoxic activity against the HeLa cell line with sub-micromolar potency.

  20. UV damage and repair in the domain of the human c-myc oncogene

    Energy Technology Data Exchange (ETDEWEB)

    Bianchi, N.O.; Bianchi, M.S.; Alitalo, K.; De La Chapelle, A. (IMBICE, La Plata (Argentina))

    1991-03-01

    We have used a modification of the Southern hybridization method to analyze the removal of UV-induced pyrimidine cyclobutane dimers from the domain of the c-myc oncogene. The study was performed in human COLO320HSR cells, which exhibit a 30- to 40-fold amplification of c-myc that is maintained in a marker chromosome as a homogeneously staining region. Intron 2 and the region upstream from the gene showed better dimer removal than intron 1 or the region downstream from the c-myc gene. Regions showing less repair coincide with regions that are hotspots for mutations and chromosome translocations. Therefore, it is proposed that the inefficiency of DNA repair may play an important role in the origin of c-myc rearrangements.

  1. Cooverexpression of EpCAM and c-myc genes in malignant breast ...

    Indian Academy of Sciences (India)

    SAMIRA SADEGHI

    and c-myc in breast tumours collected from breast cancer patients of the Iranian population. EpCAM .... Type. Fibroadenoma. 1. 9. 10. 2. 8. 10. Ductal or lobular hyperplasia. 0. 3. 3. 1. 2. 3. Lobular carcinoma in situ. 1. 4. 5. 2. 3. 5. Total. 2. 16. 18. 5. 13. 18. Age. 50<. 2. 12 ..... Distribution of EpCAM/c-myc positive and negative.

  2. Arf tumor suppressor disrupts the oncogenic positive feedback loop including c-Myc and DDX5.

    Science.gov (United States)

    Tago, K; Funakoshi-Tago, M; Itoh, H; Furukawa, Y; Kikuchi, J; Kato, T; Suzuki, K; Yanagisawa, K

    2015-01-15

    Tumor suppressor protein p19(ARF) (Arf; p14(ARF) in humans) functions in both p53-dependent and -independent modes to counteract hyper-proliferative signals caused by proto-oncogene activation, but its p53-independent activities remain poorly understood. Using the tandem affinity purification-tag technique, we purified Arf-containing protein complexes and identified p68 DEAD-box protein (DDX5) as a novel interacting protein of Arf. In this study, we found that DDX5 interacts with c-Myc, and harbors essential roles for c-Myc-mediated transcription and its transforming activity. Furthermore, when c-Myc was forcibly expressed, the expression level of DDX5 protein was drastically increased through the acceleration of protein synthesis of DDX5, suggesting the presence of an oncogenic positive feedback loop including c-Myc and DDX5. Strikingly, Arf blocked the physical interaction between DDX5 and c-Myc, and drove away DDX5 from the promoter of c-Myc target genes. These observations most likely indicate the mechanism by which Arf causes p53-independent tumor-suppressive activity.

  3. Structure-based design of flavone derivatives as c-myc oncogene down-regulators.

    Science.gov (United States)

    Yang, Hui; Zhong, Hai-Jing; Leung, Ka-Ho; Chan, Daniel Shiu-Hin; Ma, Victor Pui-Yan; Fu, Wai-Chung; Nanjunda, Rupesh; Wilson, W David; Ma, Dik-Lung; Leung, Chung-Hang

    2013-01-23

    Based on molecular docking analysis of complexes between flavone and the c-myc G-quadruplex, we designed and screened 30 flavone derivatives containing various side chains that could potentially form interactions with the G-quadruplex grooves. As a proof-of-concept, the highest-scoring flavone derivatives containing cationic pyridinium side chains were synthesized and their interactions with the c-myc G-quadruplex were examined using a PCR-stop assay. The stabilizing effects of the flavone derivatives were found to be selective towards the c-myc G-quadruplex over other biologically relevant G-quadruplex structures, such as the human telomeric sequence (HTS). The interaction between the most potent compound of the series and the c-myc G-quadruplex was examined in depth using UV-Vis titration, molecular modeling and CD spectroscopy. Our results suggest that in addition to stabilizing the c-myc G-quadruplex, the flavone derivatives were capable of inducing the formation of the G-quadruplex structure even in the absence of monovalent cations. The flavone derivatives were found to be potent inhibitors of c-myc promoters within the cellular environment and displayed promising cytotoxic behavior against human cancer cell lines. Copyright © 2012 Elsevier B.V. All rights reserved.

  4. The c-myc oncogene is translocated to the involved chromosome 12 in mouse plasmacytoma.

    Science.gov (United States)

    Erikson, J; Miller, D A; Miller, O J; Abcarian, P W; Skurla, R M; Mushinski, J F; Croce, C M

    1985-01-01

    Although it is known that the c-myc oncogene is rearranged in a head-to-head fashion with the immunoglobulin heavy chain locus in mouse plasmacytomas, it has not been clear whether the c-myc oncogene is translocated to the heavy chain locus on mouse chromosome 12 or whether the heavy chain locus is translocated to the c-myc locus on mouse chromosome 15. To determine which of these two possibilities is correct, we hybridized Chinese hamster fibroblasts with J558 mouse plasmacytoma cells that carry a reciprocal chromosome translocation between chromosomes 12 and 15, and we examined the segregating hybrids for the presence of the normal and rearranged mouse c-myc genes, for the presence of different regions of the mouse heavy chain locus, and for the presence of genes located on mouse chromosomes 12 and 15. The results of this analysis indicate that, as in human Burkitt lymphomas with the 8;14 chromosome translocation, the c-myc gene is translocated to the heavy chain locus in mouse plasmacytomas. Thus the orientation of the heavy chain locus on mouse chromosome 12 and of the c-myc gene on mouse chromosome 15 is the same as the orientation of the homologous loci in man. Images PMID:3923490

  5. Hfq affects mRNA levels independently of degradation

    Directory of Open Access Journals (Sweden)

    Hajnsdorf Eliane

    2010-02-01

    Full Text Available Abstract Background The bacterial Lsm protein, Hfq, is an RNA chaperone involved in many reactions related to RNA metabolism, such as replication and stability, control of small RNA activity and polyadenylation. Despite this wide spectrum of known functions, the global role of Hfq is almost certainly undervalued; its capacity to bind DNA and to interact with many other proteins are only now beginning to be taken into account. Results The role of Hfq in the maturation and degradation of the rpsO mRNA of E. coli was investigated in vivo. The data revealed a decrease in rpsO mRNA abundance concomitant to an increase in its stability when Hfq is absent. This indicates that the change in mRNA levels in hfq mutants does not result from its modification of RNA stability. Moreover, a series of independent experiments have revealed that the decrease in mRNA level is not a consequence of a reduction of translation efficiency and that Hfq is not directly implicated in translational control of rpsO expression. Reduced steady-state mRNA levels in the absence of Hfq were also shown for rpsT, rpsB and rpsB-tsf, but not for lpp, pnp or tRNA transcripts. The abundance of chimeric transcripts rpsO-lacZ and rpsB-lacZ, whose expression was driven by rpsO and rpsB promoters, respectively, was also lower in the hfq null-mutants, while the β-galactosidase yield remained about the same as in the parent wild-type strain. Conclusions The data obtained suggest that alteration of rpsO, rpsT and rpsB-tsf transcript levels observed under conditions of Hfq deficiency is not caused by the post-transcriptional events, such as mRNA destabilization or changes in translation control, and may rather result from changes in transcriptional activity. So far, how Hfq affects transcription remains unclear. We propose that one of the likely mechanisms of Hfq-mediated modulation of transcription might operate early in the elongation step, when interaction of Hfq with a nascent transcript

  6. Tumor suppressor PDCD4 modulates miR-184-mediated direct suppression of C-MYC and BCL2 blocking cell growth and survival in nasopharyngeal carcinoma.

    Science.gov (United States)

    Zhen, Yan; Liu, Zhen; Yang, Huiling; Yu, Xiaoli; Wu, Qiangyun; Hua, Shengni; Long, Xiaobin; Jiang, Qingping; Song, Ye; Cheng, Chao; Wang, Hao; Zhao, Menyang; Fu, Qiaofen; Lyu, Xiaoming; Chen, Yiyu; Fan, Yue; Liu, Yan; Li, Xin; Fang, Weiyi

    2013-10-24

    Programmed cell death 4 (PDCD4), a novel tumor suppressor, inhibits cell proliferation, migration and invasion as well as promotes cell apoptosis in tumors. However, the molecular mechanism of its tumor-suppressive function remains largely unknown in tumors including nasopharyngeal carcinoma (NPC). In this study, downregulated PDCD4 expression was significantly associated with the status of NPC progression and poor prognosis. PDCD4 markedly suppressed the ability of cell proliferation and cell survival by modulating C-MYC-controlled cell cycle and BCL-2-mediated mitochondrion apoptosis resistance signals, and oncogenic transcription factor C-JUN in NPC. Furthermore, miR-184, a tumor-suppressive miRNA modulated by PDCD4 directly targeting BCL2 and C-MYC, participated in PDCD4-mediated suppression of cell proliferation and survival in NPC. Further, we found that PDCD4 decreased the binding of C-Jun to the AP-1 element on the miR-184 promoter regions by PI3K/AKT/JNK/C-Jun pathway and stimulated miR-184 expression. In clinical fresh specimens, reduced PDCD4 mRNA level was positively correlated with miR-184 expression in NPC. Our studies are the first to demonstrate that PDCD4 as tumor suppressor regulated miR-184-mediated direct targeting of BCL2 and C-MYC via PI3K/AKT and JNK/C-Jun pathway attenuating cell proliferation and survival in NPC.

  7. Chromosome translocation activates heterogeneously initiated, bipolar transcription of a mouse c-myc gene.

    Science.gov (United States)

    Calabi, F; Neuberger, M S

    1985-01-01

    In many mouse plasmacytomas, the active c-myc gene has been truncated by chromosome translocation with the resultant severance of the protein-coding sequence from the normal promoter. Transcripts of such truncated c-myc genes were analyzed by Northern blotting, nuclease S1 mapping, primer extension assays and cDNA cloning. We conclude that transcription originates from multiple initiation sites on both c-myc coding and non-coding strands with the two-sets of transcripts derived from adjacent but essentially non-overlapping regions located greater than 1 kb from the translocation junction. In X63Ag8, where c-myc is translocated to the immunoglobulin C gamma 2b gene, the c-myc non-coding strand transcripts include the translocation junction and then splice directly into the gamma 2b CH1 exon. We propose that chromosome translocation activates a cryptic promoter in the first intron and that the heterogeneously initiated, bipolar transcription reflects the absence of a suitably placed TATA box element. Images Fig. 1. Fig. 2. Fig. 3. Fig. 4. Fig. 7. Fig. 8. PMID:3924591

  8. Induction of c-myc and c-jun proto-oncogene expression in rat L6 myoblasts by cadmium is inhibited by zinc preinduction of the metallothionein gene

    Energy Technology Data Exchange (ETDEWEB)

    Abshire, M.K.; Buzard, G.S.; Shiraishi, Noriyuki; Waalkers, M.P. [National Cancer Institute, Fredrick, MD (United States)

    1996-07-01

    Certain proto-oncogenes transfer growth regulatory signals from the cell surface to the nucleus. These genes often show activation soon after cells are exposed to mitogenic stimulation but can also be activated as a nonmitogenic stress response. Cadmium (Cd) is a carcinogenic metal in humans and rodents and, though its mechanism of action is unknown, it could involve activation of such proto-oncogenes. Metallothionein (MT), a metal-inducible protein that binds Cd, can protect against many aspects of Cd toxicity, including genotoxicity and possibly carcinogenesis. Thus, the effects of Cd on expression of c-myc and c-jun in rat L6 myoblasts, and the effect of preactivation of the MT gene by Zn treatment on such oncogene expression, were studied. MT protein levels were measured using oligonucleotide hybridization and standardized to {beta}-actin levels. Cd (5 {mu}M CdCl{sub 2}, 0-30 h) stimulated both c-myc and c-jun mRNA expression. An initial peak of activation of c-myc expression occurred 2 h after initiation of Cd exposure, and levels remained elevated throughout the assessment period. Zn pretreatment markedly reduced the activation of c-myc expression by Cd compared to cells not receiving Zn pretreatment. Cd treatment increased c-jun mRNA levels by up to 3.5-fold. Again, Zn pretreatment markedly reduced. 10 refs., 8 figs.

  9. c-MYC—Making Liver Sick: Role of c-MYC in Hepatic Cell Function, Homeostasis and Disease

    Science.gov (United States)

    Zheng, Kang; Cubero, Francisco Javier; Nevzorova, Yulia A.

    2017-01-01

    Over 35 years ago, c-MYC, a highly pleiotropic transcription factor that regulates hepatic cell function, was identified. In recent years, a considerable increment in the number of publications has significantly shifted the way that the c-MYC function is perceived. Overexpression of c-MYC alters a wide range of roles including cell proliferation, growth, metabolism, DNA replication, cell cycle progression, cell adhesion and differentiation. The purpose of this review is to broaden the understanding of the general functions of c-MYC, to focus on c-MYC-driven pathogenesis in the liver, explain its mode of action under basal conditions and during disease, and discuss efforts to target c-MYC as a plausible therapy for liver disease. PMID:28422055

  10. Discovery of a natural product-like c-myc G-quadruplex DNA groove-binder by molecular docking.

    Directory of Open Access Journals (Sweden)

    Dik-Lung Ma

    Full Text Available The natural product-like carbamide (1 has been identified as a stabilizer of the c-myc G-quadruplex through high-throughput virtual screening. NMR and molecular modeling experiments revealed a groove-binding mode for 1. The biological activity of 1 against the c-myc G-quadruplex was confirmed by its ability to inhibit Taq polymerase-mediated DNA extension and c-myc expression in vitro, demonstrating that 1 is able to control c-myc gene expression at the transcriptional level presumably through the stabilization of the c-myc promoter G-quadruplex. Furthermore, the interaction between carbamide analogues and the c-myc G-quadruplex was also investigated by in vitro experiments in order to generate a brief structure-activity relationship (SAR for the observed potency of carbamide 1.

  11. Acetylation of the c-MYC oncoprotein is required for cooperation with the HTLV-1 p30II accessory protein and the induction of oncogenic cellular transformation by p30II/c-MYC

    Science.gov (United States)

    Romeo, Megan M.; Ko, Bookyung; Kim, Janice; Brady, Rebecca; Heatley, Hayley C.; He, Jeffrey; Harrod, Carolyn K.; Barnett, Braden; Ratner, Lee; Lairmore, Michael D.; Martinez, Ernest; Lüscher, Bernhard; Robson, Craig N.; Henriksson, Marie; Harrod, Robert

    2014-01-01

    The human T-cell leukemia retrovirus type-1 (HTLV-1) p30II protein is a multifunctional latency-maintenance factor that negatively regulates viral gene expression and deregulates host signaling pathways involved in aberrant T-cell growth and proliferation. We have previously demonstrated that p30II interacts with the c-MYC oncoprotein and enhances c-MYC-dependent transcriptional and oncogenic functions. However, the molecular and biochemical events that mediate the cooperation between p30II and c-MYC remain to be completely understood. Herein we demonstrate that p30II induces lysine-acetylation of the c-MYC oncoprotein. Acetylation-defective c-MYC Lys→Arg substitution mutants are impaired for oncogenic transformation with p30II in c-myc−/− HO15.19 fibroblasts. Using dual-chromatin-immunoprecipitations (dual-ChIPs), we further demonstrate that p30II is present in c-MYC-containing nucleoprotein complexes in HTLV-1-transformed HuT-102 T-lymphocytes. Moreover, p30II inhibits apoptosis in proliferating cells expressing c-MYC under conditions of genotoxic stress. These findings suggest that c-MYC-acetylation is required for the cooperation between p30II/c-MYC which could promote proviral replication and contribute to HTLV-1-induced carcinogenesis. PMID:25569455

  12. Endogenous c-Myc is essential for p53-induced apoptosis in response to DNA damage in vivo.

    Science.gov (United States)

    Phesse, T J; Myant, K B; Cole, A M; Ridgway, R A; Pearson, H; Muncan, V; van den Brink, G R; Vousden, K H; Sears, R; Vassilev, L T; Clarke, A R; Sansom, O J

    2014-06-01

    Recent studies have suggested that C-MYC may be an excellent therapeutic cancer target and a number of new agents targeting C-MYC are in preclinical development. Given most therapeutic regimes would combine C-MYC inhibition with genotoxic damage, it is important to assess the importance of C-MYC function for DNA damage signalling in vivo. In this study, we have conditionally deleted the c-Myc gene in the adult murine intestine and investigated the apoptotic response of intestinal enterocytes to DNA damage. Remarkably, c-Myc deletion completely abrogated the immediate wave of apoptosis following both ionizing irradiation and cisplatin treatment, recapitulating the phenotype of p53 deficiency in the intestine. Consistent with this, c-Myc-deficient intestinal enterocytes did not upregulate p53. Mechanistically, this was linked to an upregulation of the E3 Ubiquitin ligase Mdm2, which targets p53 for degradation in c-Myc-deficient intestinal enterocytes. Further, low level overexpression of c-Myc, which does not impact on basal levels of apoptosis, elicited sustained apoptosis in response to DNA damage, suggesting c-Myc activity acts as a crucial cell survival rheostat following DNA damage. We also identify the importance of MYC during DNA damage-induced apoptosis in several other tissues, including the thymus and spleen, using systemic deletion of c-Myc throughout the adult mouse. Together, we have elucidated for the first time in vivo an essential role for endogenous c-Myc in signalling DNA damage-induced apoptosis through the control of the p53 tumour suppressor protein.

  13. Mitochondrial structure, function and dynamics are temporally controlled by c-Myc.

    Directory of Open Access Journals (Sweden)

    J Anthony Graves

    Full Text Available Although the c-Myc (Myc oncoprotein controls mitochondrial biogenesis and multiple enzymes involved in oxidative phosphorylation (OXPHOS, the coordination of these events and the mechanistic underpinnings of their regulation remain largely unexplored. We show here that re-expression of Myc in myc-/- fibroblasts is accompanied by a gradual accumulation of mitochondrial biomass and by increases in membrane polarization and mitochondrial fusion. A correction of OXPHOS deficiency is also seen, although structural abnormalities in electron transport chain complexes (ETC are not entirely normalized. Conversely, the down-regulation of Myc leads to a gradual decrease in mitochondrial mass and a more rapid loss of fusion and membrane potential. Increases in the levels of proteins specifically involved in mitochondrial fission and fusion support the idea that Myc affects mitochondrial mass by influencing both of these processes, albeit favoring the latter. The ETC defects that persist following Myc restoration may represent metabolic adaptations, as mitochondrial function is re-directed away from producing ATP to providing a source of metabolic precursors demanded by the transformed cell.

  14. c-Myc-induced transcription factor AP4 is required for CD8+ T cell-mediated host protection

    OpenAIRE

    Chou, Chun; Pinto, Amelia K.; Curtis, Jonathan D.; Persaud, Stephen P.; Cella, Marina; Lin, Chih-Chung; Edelson, Brian T.; Allen, Paul M.; Colonna, Marco; Pearce, Erika L; Diamond, Michael S.; Egawa, Takeshi

    2014-01-01

    Although c-Myc is essential to establish a metabolically active and proliferative state in T cells after priming, its expression is transient. It remains unknown how T cell activation is maintained after c-Myc down-regulation. Here, we identify AP4 as the transcription factor that is induced by c-Myc and sustains activation of antigen-specific CD8+ T cells. Despite normal priming, AP4-deficient CD8+ T cells fail to continue transcription of a broad range of c-Myc-dependent targets. Mice lacki...

  15. [Advances Research on C-MYC Proto-oncogene in Multiple Myeloma -Review].

    Science.gov (United States)

    Huang, He; Guo, Wen-Jian; Yao, Ron-Xin

    2016-08-01

    Multiple myeloma(MM) as one of the most common tumors of hmatologic system, is characterized by malignant proliferation of plasma cells, and the chemotherapy is the main therapeutic method. MM is an incurable disease because of drug-resistance of MM cells. Although the pathogenesis of MM remains unknown, the chromosome abnormalities exit in half of the patients, particularly the highly expressed gene C-MYC. Furthermore, plenty of clinical researches indicated a high expression level of C-MYC implied worse progression and/or poor prognosis of MM. Recently, the work exploiting the compounds targeting MYC has made substantial progress, even in the MM therapy. In this article, briefly the recent advances of the research on C-MYC proto-oncogene in multiple myeloma are reviewed.

  16. Balance of Yin and Yang: Ubiquitylation-Mediated Regulation of p53 and c-Myc

    Directory of Open Access Journals (Sweden)

    Mu-Shui Dai

    2006-08-01

    Full Text Available Protein ubiquitylation has been demonstrated to play a vital role not only in mediating protein turnover but also in modulating protein activity. The stability and activity of the tumor suppressor p53 and of the oncoprotein c-Myc are no exception. Both are regulated through independent ubiquitylation by several E3 ubiquitin ligases. Interestingly, p53 and c-Myc are functionally connected by some of these E3 enzymes and their regulator ARF, although these proteins play opposite roles in controlling cell growth and proliferation. The balance of this complex ubiquitylation network and its disruption during oncogenesis will be the topics of this review.

  17. What real influence does the proto-oncogene c-myc have in OSCC behavior?

    Science.gov (United States)

    Pérez-Sayáns, Mario; Suárez-Peñaranda, José Manuel; Pilar, Gayoso-Diz; Barros-Angueira, Francisco; Gándara-Rey, José Manuel; García-García, Abel

    2011-08-01

    The influence of c-myc in the carcinogenic process has been previously described although in the specific case of oral tumors it has been poorly tested. Myc proteins are a family of proto-oncogenes involved in the cell proliferation regulation, differentiation and apoptosis. The goal of this paper is to describe the functions of c-myc and its role as oncogene, assessing its expression by immunohistochemistry and genetic amplification studies, and studying its relationship with tumoral clinical and pathological variables, and describing genetic and molecular interactions in OSCC. Copyright © 2011 Elsevier Ltd. All rights reserved.

  18. Alterations in TP53, cyclin D2, c-Myc, p21WAF1/CIP1 and p27KIP1 expression associated with progression in B-CLL

    Directory of Open Access Journals (Sweden)

    Antosz Halina

    2010-04-01

    Full Text Available B-cell chronic lymphocytic leukaemia (B-CLL originates from B lymphocytes that may differ in the activationlevel, maturation state or cellular subgroups in peripheral blood. Tumour progression in CLL B cells seems to result in gradualaccumulation of the clone of resting B lymphocytes in the early phases (G0/G1 of the cell cycle. The G1 phase isimpaired in B-CLL. We investigated the gene expression of five key cell cycle regulators: TP 53, c-Myc, cyclin D2,p21WAF1/CIP1 and p27KIP1, which primarily regulate the G1 phase of the cell cycle, or S-phase entry and ultimately controlthe proliferation and cell growth as well as their role in B-CLL progression. The study was conducted in peripheral bloodCLL lymphocytes of 40 previously untreated patients. Statistical analysis of correlations of TP53, cyclin D2, c-Myc,p21WAF1/CIP1 and p27KIP1 expressions in B-CLL patients with different Rai stages demonstrated that the progression of diseasewas accompanied by increases in p53, cyclin D2 and c-Myc mRNA expression. The expression of p27KIP1 was nearlystatistically significant whereas that of p21 WAF1/CIP1 showed no such correlation. Moreover, high expression levels of TP53and c-Myc genes were found to be closely associated with more aggressive forms of the disease requiring earlier therapy.

  19. Lymphocytes DNA Content, P53, C-Myc And Bcl-2 As Predictive ...

    African Journals Online (AJOL)

    Cell cycle parameters as well as apoptotic and tumor markers directly control cell growth. DNA ploidy and S phase fraction, apoptosis fraction in addition to apoptotic inducer (p53, c-myc) and antiapoptotic marker (Bcl-2) were investigated in childhood with acute lymphoblastic leukemia (ALL) leukemia as a predictive ...

  20. Astroglial c-Myc overexpression predisposes mice to primary malignant gliomas

    DEFF Research Database (Denmark)

    Jensen, Niels Aagaard; Pedersen, Karen-Marie; Lihme, Frederikke

    2003-01-01

    Malignant astrocytomas are common human primary brain tumors that result from neoplastic transformation of astroglia or their progenitors. Here we show that deregulation of the c-Myc pathway in developing astroglia predisposes mice to malignant astrocytomas within 2-3 weeks of age. The genetically...

  1. RESEARCH ARTICLE Co-overexpression of EpCAM and c-myc ...

    Indian Academy of Sciences (India)

    Methods: To this purpose, 122 fresh tissues including 104 malignant and 18 benign samples were disrupted by mortar and pestle and RNA was then isolated from the samples and converted to cDNA. The relative expression levels of EpCAM and c-myc genes were measured by2−ΔΔCt method using RT-. qPCR method.

  2. Diet-induced obesity promotes murine gastric cancer growth through a nampt/sirt1/c-myc positive feedback loop.

    Science.gov (United States)

    Li, Hai-Jun; Che, Xiang-Ming; Zhao, Wei; He, Shi-Cai; Zhang, Zheng-Liang; Chen, Rui; Fan, Lin; Jia, Zong-Liang

    2013-11-01

    Obesity increases the risk of gastric cancer and may promote its growth, as was recently demonstrated by our novel in vivo mouse model. However, the underlying mechanisms of this correlation remain unclear. The purpose of this study was to investigate the precise effects of obesity on gastric cancer growth and to elucidate the potential molecular mechanisms. Diet-induced obese mice were insulin-resistant, glucose-intolerant and had high serum visfatin concentration. In the subcutaneous mouse model, tumors were more aggressive in diet-induced obese mice compared with lean mice. Tumor weights showed a significant positive correlation with mouse body weights, as well as serum insulin and visfatin concentrations. Immunohistochemical staining showed that the expression levels of iNampt, Sirt1 and c-MYC proteins were upregulated in the subcutaneous tumors from obese mice compared to those from lean animals. Furthermore, obesity not only prompted significantly murine forestomach carcinoma cell migration, proliferation, but also affected cellular apoptosis and cell cycle by endocrine mechanisms. These were associated with increased expression of the pro-survival nampt/sirt1/c-myc positive feedback loop confirmed by RT-PCR and western blotting. These results suggested that diet-induced obesity could promote murine gastric cancer growth by upregulating the expression of the nampt, sirt1 and c-myc genes.

  3. Inhibition of c-Myc overcomes cytotoxic drug resistance in acute myeloid leukemia cells by promoting differentiation.

    Directory of Open Access Journals (Sweden)

    Xiao-Na Pan

    Full Text Available Nowadays, drug resistance still represents a major obstacle to successful acute myeloid leukemia (AML treatment and the underlying mechanism is not fully elucidated. Here, we found that high expression of c-Myc was one of the cytogenetic characteristics in the drug-resistant leukemic cells. c-Myc over-expression in leukemic cells induced resistance to chemotherapeutic drugs, enhanced colony formation capacity and inhibited cell differentiation induced by all-trans retinoic acid (ATRA. Meanwhile, inhibition of c-Myc by shRNA or specific c-Myc inhibitor 10058-F4 rescued the sensitivity to cytotoxic drugs, restrained the colony formation ability and promoted differentiation. RT-PCR and western blotting analysis showed that down-regulation of C/EBPβ contributed to the poor differentiation state of leukemic cells induced by c-Myc over-expression. Importantly, over-expression of C/EBPβ could reverse c-Myc induced drug resistance. In primary AML cells, the c-Myc expression was negatively correlated with C/EBPβ. 10058-F4, displayed anti-proliferative activity and increased cellular differentiation with up-regulation of C/EBPβ in primary AML cells. Thus, our study indicated that c-Myc could be a novel target to overcome drug resistance, providing a new approach in AML therapy.

  4. c-MYC Copy-Number Gain Is an Independent Prognostic Factor in Patients with Colorectal Cancer.

    Science.gov (United States)

    Lee, Kyu Sang; Kwak, Yoonjin; Nam, Kyung Han; Kim, Duck-Woo; Kang, Sung-Bum; Choe, Gheeyoung; Kim, Woo Ho; Lee, Hye Seung

    2015-01-01

    The aim of this study was to determine the incidence and clinicopathological significance of c-MYC gene copy-number (GCN) gain in patients with primary colorectal cancer (CRC). The c-MYC GCN was investigated in 367 consecutive CRC patients (cohort 1) by using dual-color silver in situ hybridization. Additionally, to evaluate regional heterogeneity, we examined CRC tissue from 3 sites including the primary cancer, distant metastasis, and lymph-node metastasis in 152 advanced CRC patients (cohort 2). KRAS exons 2 and 3 were investigated for mutations. In cohort 1, c-MYC gene amplification, defined by a c-MYC:centromere of chromosome 8 ratio ≥ 2.0, was detected in 31 (8.4%) of 367 patients. A c-MYC GCN gain, defined by ≥ 4.0 c-MYC copies/nucleus, was found in 63 (17.2%) patients and was associated with poor prognosis (P = 0.015). Multivariate Cox regression analysis showed that the hazard ratio for c-MYC GCN gain was 2.35 (95% confidence interval, 1.453-3.802; P patients, c-MYC GCN gain was significantly associated with poor prognosis by univariate (P = 0.034) and multivariate (P = 0.040) analyses. c-MYC protein overexpression was observed in 201 (54.8%) out of 367 patients and weakly correlated with c-MYC GCN gain (ρ, 0.211). In cohort 2, the c-MYC genetic status was heterogenous in advanced CRC patients. Discordance between GCN gain in the primary tumor and either distant or lymph-node metastasis was 25.7% and 30.4%, respectively. A similar frequency for c-MYC GCN gain and amplification was observed in CRC patients with both wild-type and mutated KRAS. c-MYC GCN gain was an independent factor for poor prognosis in consecutive CRC patients and in the stage II-III subgroup. Our findings indicate that the status of c-MYC may be helpful in predicting the patients' outcome and for managing CRC patients.

  5. Effects of Klf4 and c-Myc Knockdown on Pluripotency Maintenance in Porcine Induced Pluripotent Stem Cell.

    Science.gov (United States)

    Liao, Yu-Jing; Chen, Yi-Shiou; Lee, Ja-Xin; Chen, Lih-Ren; Yang, Jenn-Rong

    2018-01-01

    The importance of Oct4 and Sox2 in maintaining pluripotency and self-renewal is well-understood, but the functions of Klf4 and c-Myc has not been fully investigated. In the present study, we attempted to determine the roles of Klf4 and c-Myc on pluripotency maintenance of porcine induced pluripotent stem (piPS) cells. In this experimental study, we performed short hairpin RNA (shRNA) to knock down the Klf4 and c-Myc functions of piPS cells and examined pluripotency markers and teratoma formation to evaluate piPS cell pluripotency. The shRNA-Klf4 and shRNA-c-Myc vectors containing a reporter gene, TagFP635, were transfected into piPS cells by lentivirus infection. The piPS cells fully expressing infrared fluorescence were selected to confirm gene knockdown of Klf4 and c-Myc reverse transcription-polymerase chain reaction (RT-PCR). Next, for pluripotency evaluation, expression of pluripotency markers was detected by immunocytochemical staining, and capability of teratoma formation was investigated by piPS cell transplantation into nonobese diabetic-severe combined immunodeficiency (NOD-SCID) mice. Our findings indicated that Klf4 and c-Myc functions of piPS cells were knocked down by shRNA transfection, and knockdown of Klf4 and c-Myc functions impaired expression of pluripotency markers such as Oct4, AP, SSEA-3, SSEA-4, TRA-1-6, and TRA-1-81. Furthermore, piPS cells without Klf4 and c-Myc expression failed to form teratomas. The pluripotency of piPS cells are crucially dependent upon Klf4 and c-Myc expression. These findings, suggesting potential mechanisms of Klf4 and c-Myc contribution to piPS cell formation, have important implications for application, regulation, and tumorigenesis of piPS cells.

  6. Oncogene amplification detected by in situ hybridization in radiation induced rat skin tumors. [C-myc:a3

    Energy Technology Data Exchange (ETDEWEB)

    Yi Jin.

    1991-02-01

    Oncogene activation may play an important role in radiation induced carcinogenesis. C-myc oncogene amplification was detected by in situ hybridization in radiation-induced rat skin tumors, including squamous and basal cell carcinomas. In situ hybridization was performed with a biotinylated human c-myc third exon probe, visualized with an avidin-biotinylated alkaline phosphate detection system. No c-myc oncogene amplification was detected in normal rat skin at very early times after exposure to ionizing radiation, which is consistent with the view that c-myc amplification is more likely to be related to carcinogenesis than to normal cell proliferation. The incorporation of tritiated thymidine into the DNA of rat skin cells showed that the proliferation of epidermal cells reached a peak on the seventh day after exposure to ionizing radiation and then decreased. No connection between the proliferation of epidermal cell and c-myc oncogene amplification in normal or irradiated rat skin was found. The results indicated that c-myc amplification as measured by in situ hybridization was correlated with the Southern bolt results, but only some of the cancer cells were amplified. The c-myc positive cells were distributed randomly within regions of the tumor and exhibited a more uniform nuclear structure in comparison to the more vacuolated c-myc negative cells. No c-myc signal was detected in unirradiated normal skin or in irradiated skin cells near the tumors. C-myc amplification appears to be cell or cell cycle specific within radiation-induced carcinomas. 28 refs., 3 figs., 3 tabs.

  7. The Histone Acetyltransferase GCN5 Expression Is Elevated and Regulated by c-Myc and E2F1 Transcription Factors in Human Colon Cancer

    Science.gov (United States)

    Yin, Yan-Wei; Jin, Hong-Jian; Zhao, Wenjing; Gao, Beixue; Fang, Jiangao; Wei, Junmin; Zhang, Donna D.; Zhang, Jianing; Fang, Deyu

    2017-01-01

    The histone acetyltransferase GCN5 has been suggested to be involved in promoting cancer cell growth. But its role in human colon cancer development remains unknown. Herein we discovered that GCN5 expression is significantly upregulated in human colon adenocarcinoma tissues. We further demonstrate that GCN5 is upregulated in human colon cancer at the mRNA level. Surprisingly, two transcription factors, the oncogenic c-Myc and the proapoptotic E2F1, are responsible for GCN5 mRNA transcription. Knockdown of c-Myc inhibited colon cancer cell proliferation largely through downregulating GCN5 transcription, which can be fully rescued by the ectopic GCN5 expression. In contrast, E2F1 expression induced human colon cancer cell death, and suppression of GCN5 expression in cells with E2F1 overexpression further facilitated cell apoptosis, suggesting that GCN5 expression is induced by E2F1 as a possible negative feedback in suppressing E2F1-mediated cell apoptosis. In addition, suppression of GCN5 with its specific inhibitor CPTH2 inhibited human colon cancer cell growth. Our studies reveal that GCN5 plays a positive role in human colon cancer development, and its suppression holds a great therapeutic potential in antitumor therapy. PMID:26637399

  8. Conversion of androgen receptor signaling from a growth suppressor in normal prostate epithelial cells to an oncogene in prostate cancer cells involves a gain of function in c-Myc regulation.

    Science.gov (United States)

    Vander Griend, Donald J; Litvinov, Ivan V; Isaacs, John T

    2014-01-01

    In normal prostate, androgen-dependent androgen receptor (AR) signaling within prostate stromal cells induces their secretion of paracrine factors, termed "andromedins" which stimulate growth of the epithelial cells. The present studies demonstrate that androgen-dependent andromedin-driven growth stimulation is counter-balanced by androgen-induced AR signaling within normal adult prostate epithelial cells resulting in terminal G0 growth arrest coupled with terminal differentiation into ΔNp63-negative, PSA-expressing secretory luminal cells. This cell autonomous AR-driven terminal differentiation requires DNA-binding of the AR protein, is associated with decreases in c-Myc m-RNA and protein, are coupled with increases in p21, p27, and SKP-2 protein expression, and does not require functional p53. These changes result in down-regulation of Cyclin D1 protein and RB phosphoryation. shRNA knockdown documents that neither RB, p21, p27 alone or in combination are required for such AR-induced G0 growth arrest. Transgenic expression of a constitutive vector to prevent c-Myc down-regulation overrides AR-mediated growth arrest in normal prostate epithelial cells, which documents that AR-induced c-Myc down-regulation is critical in terminal growth arrest of normal prostate epithelial cells. In contrast, in prostate cancer cells, androgen-induced AR signaling paradoxically up-regulates c-Myc expression and stimulates growth as documented by inhibition of both of these responses following exposure to the AR antagonist, bicalutamide. These data document that AR signaling is converted from a growth suppressor in normal prostate epithelial cells to an oncogene in prostate cancer cells during prostatic carcinogenesis and that this conversion involves a gain of function for regulation of c-Myc expression.

  9. Alterations of c-Myc and c-erbB-2 genes in ovarian tumours

    Directory of Open Access Journals (Sweden)

    Pastor Tibor

    2009-01-01

    Full Text Available Introduction. According to clinical and epidemiological studies, ovarian cancer ranks fifth in cancer deaths among women. The causes of ovarian cancer remain largely unknown but various factors may increase the risk of developing it, such as age, family history of cancer, childbearing status etc. This cancer results from a succession of genetic alterations involving oncogenes and tumour suppressor genes, which have a critical role in normal cell growth regulation. Mutations and/or overexpression of three oncogenes, c-erbB-2, c-Myc and K-ras, and of the tumour suppressor gene p53, have been frequently observed in a sporadic ovarian cancer. Objective. The aim of the present study was to analyze c-Myc and c-erbB-2 oncogene alterations, specifically amplification, as one of main mechanisms of their activation in ovarian cancers and to establish a possible association with the pathogenic process. Methods. DNA was isolated from 15 samples of malignant and 5 benign ovarian tumours, using proteinase K digestion, followed by phenol-chloroform isoamyl extraction and ethanol precipitation. C-Myc and c-erbB-2 amplification were detected by differential PCR. The level of gene copy increase was measured using the Scion image software. Results. The amplification of both c-Myc and c-erbB-2 was detected in 26.7% of ovarian epithelial carcinoma specimens. Only one tumour specimen concomitantly showed increased gene copy number for both studied genes. Interestingly, besides amplification, gene deletion was also detected (26.7% for c-erbB-2. Most of the ovarian carcinomas with alterations in c-Myc and c-erbB-2 belonged to advanced FIGO stages. Conclusion. The amplification of c-Myc and c-erbB-2 oncogenes in ovarian epithelial carcinomas is most probably a late event in the pathogenesis conferring these tumours a more aggressive biological behaviour. Similarly, gene deletions point to genomic instability in epithelial carcinomas in higher clinical stages as the

  10. Driving gradual endogenous c-myc overexpression by flow-sorting: intracellular signaling and tumor cell phenotype correlate with oncogene expression

    DEFF Research Database (Denmark)

    Knudsen, Kasper Jermiin; Holm, G.M.N.; Krabbe, J.S.

    2009-01-01

    Insulin-exposed rat mammary cancer cells were flow sorted based on a c-myc reporter plasmid encoding a destabilized green fluorescent protein. Sorted cells exhibited gradual increases in c-myc levels. Cells overexpressing c-myc by only 10% exhibited phenotypic changes attributable to c-myc overex...... an alternative modeling of the receptor-mediated carcinogenic process, compared to the currently used approaches of recombinant constitutive or conditional overexpression of oncogenic transmembrane receptor tyrosine kinases or oncogenic transcription factors....

  11. The parafibromin tumor suppressor protein inhibits cell proliferation by repression of the c-myc proto-oncogene.

    Science.gov (United States)

    Lin, Ling; Zhang, Jian-Hua; Panicker, Leelamma M; Simonds, William F

    2008-11-11

    Parafibromin is a tumor suppressor protein encoded by HRPT2, a gene recently implicated in the hereditary hyperparathyroidism-jaw tumor syndrome, parathyroid cancer, and a subset of kindreds with familial isolated hyperparathyroidism. Human parafibromin binds to RNA polymerase II as part of a PAF1 transcriptional regulatory complex. The physiologic targets of parafibromin and the mechanism by which its loss of function can lead to neoplastic transformation are poorly understood. We show here that RNA interference with the expression of parafibromin or Paf1 stimulates cell proliferation and increases levels of the c-myc proto-oncogene product, a DNA-binding protein and established regulator of cell growth. This effect results from both c-myc protein stabilization and activation of the c-myc promoter, without alleviation of the c-myc transcriptional pause. Chromatin immunoprecipitation demonstrates the occupancy of the c-myc promoter by parafibromin and other PAF1 complex subunits in native cells. Knockdown of c-myc blocks the proliferative effect of RNA interference with parafibromin or Paf1 expression. These experiments provide a previously uncharacterized mechanism for the anti-proliferative action of the parafibromin tumor suppressor protein resulting from PAF1 complex-mediated inhibition of the c-myc proto-oncogene.

  12. Repression of c-Myc responsive genes in cycling cells causes G1 arrest through reduction of cyclin E/CDK2 kinase activity

    NARCIS (Netherlands)

    Berns, K.; Hijmans, E.M.; Bernards, R.A.

    1997-01-01

    The c-myc gene encodes a sequence-specific DNA binding protein involved in proliferation and oncogenesis. Activation of c-myc expression in quiescent cells is sufficient to mediate cell cycle entry, whereas inhibition of c-myc expression causes cycling cells to withdraw from the cell cycle. To

  13. Induced pluripotent stem cells without c-Myc reduce airway responsiveness and allergic reaction in sensitized mice.

    Science.gov (United States)

    Wang, Chien-Ying; Chiou, Guang-Yuh; Chien, Yueh; Wu, Chia-Chao; Wu, Tzee-Chung; Lo, Wen-Tsung; Chen, Shyi-Jou; Chiou, Shih-Hwa; Peng, Ho-Jen; Huang, Ching-Feng

    2013-12-15

    Allergic disorders have increased substantially in recent years. Asthma is characterized by airway damage and remodeling. Reprogramming induced pluripotent stem cells (iPSCs) from adult somatic cells transfected by Oct-4/Sox-2/Klf-4, but not c-Myc, has shown the potential of embryonic-like cells. These cells have potential for multilineage differentiation and provide a resource for stem cell-based utility. However, the therapeutic potential of iPSCs without c-Myc (iPSC-w/o-c-Myc) in allergic diseases and airway hyperresponsiveness has not been investigated. The aim of this study was to evaluate the therapeutic effect of iPSC-w/o-c-Myc transplantation in a murine asthma model. BALB/c mice were sensitized with alum-adsorbed ovalbumin (OVA) and then challenged with aerosolized OVA. Phosphate-buffered saline or iPSC-w/o-c-Myc was then intravenously injected after inhalation. Serum allergen-specific antibody levels, airway hyperresponsiveness, cytokine levels in spleen cells and bronchoalveolar lavage fluid (BALF), and cellular distribution in BALF were then examined. Treatment with iPSC-w/o-c-Myc effectively suppressed both Th1 and Th2 antibody responses, which was characterized by reduction in serum allergen-specific IgE, IgG, IgG1, and IgG2a levels as well as in interleukin-5 and interferon-γ levels in BALF and in OVA-incubated splenocytes. Meanwhile, regulatory cytokine, interleukin-10, was enhanced. Transplantation of iPSC-w/o-c-Myc also significantly attenuated cellular infiltration in BALF and allergic airway hyperresponsiveness. However, no tumor formation was observed 6 months after transplantation. Administration of iPSC-w/o-c-Myc not only inhibited Th1 inflammatory responses but also had therapeutic effects on systemic allergic responses and airway hyperresponsiveness. iPSC-w/o-c-Myc transplantation may be a potential modality for treating allergic reactions and bronchial asthma.

  14. Bioinformatic analysis of c-Myc target from laryngeal cancer cell gene of laryngeal cancer.

    Science.gov (United States)

    Zhang, Wei-Dong; Chen, He-Xin; Wang, Yun-Xia; Chen, Zhi-Ping; Shan, Zhong-Jie; Xu, Guang

    2016-01-01

    The aim was to explore the structure and functions of new target spot c-Myc target from laryngeal cancer cell. (MTLC) of c-Myc gene. This study adopted bioinformatic methods to analyze the physicochemical property, secondary structure, hydrophobic region, a transmembrane region, and prediction of functions. The results showed that the whole length of the open reading frames was 708 bp, coding was 235 amino acids. This protein was a basic protein possessed two transmembrane structures and weight was about 26592.5 Da. The main elements of secondary structure were alpha-helix and random coil. MTLC was a membrane constitutive protein that possessed signal transduction and regulation may locate on karyotheca as results of subcellular localization and function prediction. This study has provided the theoretical basis for the further discussion of the effect and mechanism of action of MTLC in the occurrence of laryngeal cancer.

  15. c-Myc activates multiple metabolic networks to generate substrates for cell-cycle entry.

    Science.gov (United States)

    Morrish, F; Isern, N; Sadilek, M; Jeffrey, M; Hockenbery, D M

    2009-07-09

    Cell proliferation requires the coordinated activity of cytosolic and mitochondrial metabolic pathways to provide ATP and building blocks for DNA, RNA and protein synthesis. Many metabolic pathway genes are targets of the c-myc oncogene and cell-cycle regulator. However, the contribution of c-Myc to the activation of cytosolic and mitochondrial metabolic networks during cell-cycle entry is unknown. Here, we report the metabolic fates of [U-(13)C] glucose in serum-stimulated myc(-/-) and myc(+/+) fibroblasts by (13)C isotopomer NMR analysis. We demonstrate that endogenous c-myc increased (13)C labeling of ribose sugars, purines and amino acids, indicating partitioning of glucose carbons into C1/folate and pentose phosphate pathways, and increased tricarboxylic acid cycle turnover at the expense of anaplerotic flux. Myc expression also increased global O-linked N-acetylglucosamine protein modification, and inhibition of hexosamine biosynthesis selectively reduced growth of Myc-expressing cells, suggesting its importance in Myc-induced proliferation. These data reveal a central organizing function for the Myc oncogene in the metabolism of cycling cells. The pervasive deregulation of this oncogene in human cancers may be explained by its function in directing metabolic networks required for cell proliferation.

  16. Quantitative characterization of the interactions among c-myc transcriptional regulators FUSE, FBP, and FIR.

    Science.gov (United States)

    Hsiao, Hsin-Hao; Nath, Abhinav; Lin, Chi-Yen; Folta-Stogniew, Ewa J; Rhoades, Elizabeth; Braddock, Demetrios T

    2010-06-08

    Human c-myc is critical for cell homeostasis and growth but is a potent oncogenic factor if improperly regulated. The c-myc far-upstream element (FUSE) melts into single-stranded DNA upon active transcription, and the noncoding strand FUSE recruits an activator [the FUSE-binding protein (FBP)] and a repressor [the FBP-interacting repressor (FIR)] to fine-tune c-myc transcription in a real-time manner. Despite detailed biological experiments describing this unique mode of transcriptional regulation, quantitative measurements of the physical constants regulating the protein-DNA interactions remain lacking. Here, we first demonstrate that the two FUSE strands adopt different conformations upon melting, with the noncoding strand DNA in an extended, linear form. FBP binds to the linear noncoding FUSE with a dissociation constant in the nanomolar range. FIR binds to FUSE more weakly, having its modest dissociation constants in the low micromolar range. FIR is monomeric under near-physiological conditions but upon binding of FUSE dimerizes into a 2:1 FIR(2)-FUSE complex mediated by the RRMs. In the tripartite interaction, our analysis suggests a stepwise addition of FIR onto an activating FBP-FUSE complex to form a quaternary FIR(2)-FBP-FUSE inhibitory complex. Our quantitative characterization enhances understanding of DNA strand preference and the mechanism of the stepwise complex formation in the FUSE-FBP-FIR regulatory system.

  17. CUDR promotes liver cancer stem cell growth through upregulating TERT and C-Myc

    Science.gov (United States)

    Pu, Hu; Zheng, Qidi; Li, Haiyan; Wu, Mengying; An, Jiahui; Gui, Xin; Li, Tianming; Lu, Dongdong

    2015-01-01

    Cancer up-regulated drug resistant (CUDR) is a novel non-coding RNA gene. Herein, we demonstrate excessive CUDR cooperates with excessive CyclinD1 or PTEN depletion to accelerate liver cancer stem cells growth and liver stem cell malignant transformation in vitro and in vivo. Mechanistically, we reveal the decrease of PTEN in cells may lead to increase binding capacity of CUDR to CyclinD1. Therefore, CUDR-CyclinD1 complex loads onto the long noncoding RNA H19 promoter region that may lead to reduce the DNA methylation on H19 promoter region and then to enhance the H19 expression. Strikingly, the overexpression of H19 increases the binding of TERT to TERC and reduces the interplay between TERT with TERRA, thus enhancing the cell telomerase activity and extending the telomere length. On the other hand, insulator CTCF recruits the CUDR-CyclinD1 complx to form the composite CUDR-CyclinD1-insulator CTCF complex which occupancied on the C-myc gene promoter region, increasing the outcome of oncogene C-myc. Ultimately, excessive TERT and C-myc lead to liver cancer stem cell and hepatocyte-like stem cell malignant proliferation. To understand the novel functions of long noncoding RNA CUDR will help in the development of new liver cancer therapeutic and diagnostic approaches. PMID:26513297

  18. c-Myc oncogene expression in selected odontogenic cysts and tumors: An immunohistochemical study.

    Science.gov (United States)

    Moosvi, Zama; Rekha, K

    2013-01-01

    To investigate the role of c-Myc oncogene in selected odontogenic cysts and tumors. Ten cases each of ameloblastoma, adenomatoid odontogenic tumor (AOT), odontogenic keratocyst (OKC), dentigerous cyst, and radicular cyst were selected and primary monoclonal mouse anti-human c-Myc antibody was used in a dilution of 1: 50. Statistical Analysis was performed using Mann Whitney U test. 80% positivity was observed in ameloblastoma, AOT and OKC; 50% positivity in radicular cyst and 20% positivity in dentigerous cyst. Comparison of c-Myc expression between ameloblastoma and AOT did not reveal significant results. Similarly, no statistical significance was observed when results of OKC were compared with ameloblastoma and AOT. In contrast, significant differences were seen on comparison of dentigerous cyst with ameloblastoma and AOT and radicular cyst with AOT. From the above data we conclude that (1) Ameloblastoma and AOT have similar proliferative potential and their biologic behavior cannot possibly be attributed to it. (2) OKC has an intrinsic growth potential which is absent in other cysts and reinforces its classification as keratocystic odontogenic tumor.

  19. Angiotensin II Reduces Cardiac AdipoR1 Expression through AT1 Receptor/ROS/ERK1/2/c-Myc Pathway

    Science.gov (United States)

    Lei, Hong; Wang, Cheng; Wu, Li-Peng; Wang, Jin-Yu; Fu, Feng-Ying; Zhu, Wei-Guo; Wu, Li-Ling

    2013-01-01

    Adiponectin, an abundant adipose tissue-derived protein, exerts protective effect against cardiovascular disease. Adiponectin receptors (AdipoR1 and AdipoR2) mediate the beneficial effects of adiponectin on the cardiovascular system. However, the alteration of AdipoRs in cardiac remodeling is not fully elucidated. Here, we investigated the effect of angiotensin II (AngII) on cardiac AdipoRs expression and explored the possible molecular mechanism. AngII infusion into rats induced cardiac hypertrophy, reduced AdipoR1 but not AdipoR2 expression, and attenuated the phosphorylations of adenosine monophosphate-activated protein kinase and acetyl coenzyme A carboxylase, and those effects were all reversed by losartan, an AngII type 1 (AT1) receptor blocker. AngII reduced expression of AdipoR1 mRNA and protein in cultured neonatal rat cardiomyocytes, which was abolished by losartan, but not by PD123319, an AT2 receptor antagonist. The antioxidants including reactive oxygen species (ROS) scavenger NAC, NADPH oxidase inhibitor apocynin, Nox2 inhibitor peptide gp91 ds-tat, and mitochondrial electron transport chain complex I inhibitor rotenone attenuated AngII-induced production of ROS and phosphorylation of extracellular signal-regulated kinase (ERK) 1/2. AngII-reduced AdipoR1 expression was reversed by pretreatment with NAC, apocynin, gp91 ds-tat, rotenone, and an ERK1/2 inhibitor PD98059. Chromatin immunoprecipitation assay demonstrated that AngII provoked the recruitment of c-Myc onto the promoter region of AdipoR1, which was attenuated by PD98059. Moreover, AngII-induced DNA binding activity of c-Myc was inhibited by losartan, NAC, apocynin, gp91 ds-tat, rotenone, and PD98059. c-Myc small interfering RNA abolished the inhibitory effect of AngII on AdipoR1 expression. Our results suggest that AngII inhibits cardiac AdipoR1 expression in vivo and in vitro and AT1 receptor/ROS/ERK1/2/c-Myc pathway is required for the downregulation of AdipoR1 induced by AngII. PMID

  20. Knocking-down of CREPT prohibits the progression of oral squamous cell carcinoma and suppresses cyclin D1 and c-Myc expression.

    Directory of Open Access Journals (Sweden)

    Juntao Ma

    Full Text Available As a regulator essential for many cell cycle-related proteins, the robust expression of Cell cycle-Related and Expression-elevated Protein in Tumor (CREPT implicates a poor diagnosis of endoderm and mesoderm-derived tumors. Whether CREPT plays the same role in the tumorigenesis derived from ectodermal tissues remains elusive.To explore the role of CREPT in ectoderm-derived tumors, cells from 7oral squamous cell carcinoma (OSCC lines and 84clinical OSCC samples were exploited in this study. Quantitative PCR, Western blot assay and immunohistochemistry were applied in the evaluation of CREPT, cyclin D1 and c-Myc expression. Knocking-down of CREPT was performed by lentivirus delivering specific shRNA of CREPT. The effects of CREPT on OSCC were examined by cell proliferation, colony formation, apoptosis, cell migration and xenograft implantation experiments.Compared with human normal oral keratinocytes, OSCC cell lines showed a significantly elevated expression of CREPT in both mRNA and protein levels. Consistently, samples from OSCC patients also exhibited a noticeably stronger CREPT expression than the noncancerous samples. In contrast, knocking down of CREPT in OSCC cell lines significantly reduced proliferation, colony formation and migration as well as the expression of cyclin D1 and c-Myc, but promoted apoptosis. Statistical analysis also suggested that CREPT expression was significantly correlated with the T and N classification of OSCC. Furthermore, CAL27 mouse xenograft model confirmed that down-regulation of CREPT prohibited cyclin D1 and c-Myc expression, through which decreased the in vivo tumor growth, but increased the survival ratio of hosts.In OSCC cell lines, up-regulated CREPT expression enhanced cell proliferation, migration and cell cycle as well as promoted cyclin D1 and c-Myc expression as it did in endoderm and mesoderm-origin tumors. Our study strongly suggests that CREPT could be used as a marker for the OSCC prognosis and

  1. Angiotensin II reduces cardiac AdipoR1 expression through AT1 receptor/ROS/ERK1/2/c-Myc pathway.

    Directory of Open Access Journals (Sweden)

    Li Li

    Full Text Available Adiponectin, an abundant adipose tissue-derived protein, exerts protective effect against cardiovascular disease. Adiponectin receptors (AdipoR1 and AdipoR2 mediate the beneficial effects of adiponectin on the cardiovascular system. However, the alteration of AdipoRs in cardiac remodeling is not fully elucidated. Here, we investigated the effect of angiotensin II (AngII on cardiac AdipoRs expression and explored the possible molecular mechanism. AngII infusion into rats induced cardiac hypertrophy, reduced AdipoR1 but not AdipoR2 expression, and attenuated the phosphorylations of adenosine monophosphate-activated protein kinase and acetyl coenzyme A carboxylase, and those effects were all reversed by losartan, an AngII type 1 (AT1 receptor blocker. AngII reduced expression of AdipoR1 mRNA and protein in cultured neonatal rat cardiomyocytes, which was abolished by losartan, but not by PD123319, an AT2 receptor antagonist. The antioxidants including reactive oxygen species (ROS scavenger NAC, NADPH oxidase inhibitor apocynin, Nox2 inhibitor peptide gp91 ds-tat, and mitochondrial electron transport chain complex I inhibitor rotenone attenuated AngII-induced production of ROS and phosphorylation of extracellular signal-regulated kinase (ERK 1/2. AngII-reduced AdipoR1 expression was reversed by pretreatment with NAC, apocynin, gp91 ds-tat, rotenone, and an ERK1/2 inhibitor PD98059. Chromatin immunoprecipitation assay demonstrated that AngII provoked the recruitment of c-Myc onto the promoter region of AdipoR1, which was attenuated by PD98059. Moreover, AngII-induced DNA binding activity of c-Myc was inhibited by losartan, NAC, apocynin, gp91 ds-tat, rotenone, and PD98059. c-Myc small interfering RNA abolished the inhibitory effect of AngII on AdipoR1 expression. Our results suggest that AngII inhibits cardiac AdipoR1 expression in vivo and in vitro and AT1 receptor/ROS/ERK1/2/c-Myc pathway is required for the downregulation of AdipoR1 induced by AngII.

  2. Proteomic Characterization of the World Trade Center dust-activated mdig and c-myc signaling circuit linked to multiple myeloma.

    Science.gov (United States)

    Wu, Kai; Li, Lingzhi; Thakur, Chitra; Lu, Yongju; Zhang, Xiangmin; Yi, Zhengping; Chen, Fei

    2016-11-11

    Several epidemiological studies suggested an increased incidence rate of multiple myeloma (MM) among first responders and other individuals who exposed to World Trade Center (WTC) dust. In this report, we provided evidence showing that WTC dust is potent in inducing mdig protein and/or mRNA in bronchial epithelial cells, B cells and MM cell lines. An increased mdig expression in MM bone marrow was observed, which is associated with the disease progression and prognosis of the MM patients. Through integrative genomics and proteomics approaches, we further demonstrated that mdig directly interacts with c-myc and JAK1 in MM cell lines, which contributes to hyperactivation of the IL-6-JAK-STAT3 signaling important for the pathogenesis of MM. Genetic silencing of mdig reduced activity of the major downstream effectors in the IL-6-JAK-STAT3 pathway. Taken together, these data suggest that WTC dust may be one of the key etiological factors for those who had been exposed for the development of MM by activating mdig and c-myc signaling circuit linked to the IL-6-JAK-STAT3 pathway essential for the tumorigenesis of the malignant plasma cells.

  3. The 5T mouse multiple myeloma model: Absence of c-myc oncogene rearrangement in early transplant generations

    NARCIS (Netherlands)

    Radl, J.; Punt, Y.A.; Enden-Vieveen, M.H.M. van den; Bentvelzen, P.A.J.; Bakkus, M.H.C.; Akker T., W. van den; Benner, R.

    1990-01-01

    Consistent chromosomal translocations involving the c-myc cellular oncogene and one of the three immunoglobulin loci are typical for human Burkitt's lymphoma, induced mouse plasmacytoma (MPC) and spontaneously arising rat immunocytoma (RIC). Another plasma cell malignancy, multiple myeloma (MM),

  4. Not All G-Quadruplexes are Created Equally: An Investigation of the Structural Polymorphism of the c-Myc G-Quadruplex-Forming Sequence and its Interaction with the Porphyrin meso-Tetra(N-methyl-4-pyridyl)porphine

    Science.gov (United States)

    Le, Huy T.; Miller, M. Clarke; Buscaglia, Robert; Dean, William L.; Holt, Patrick A.; Chaires, Jonathan B.; Trent, John O.

    2012-01-01

    G-quadruplexes, DNA tertiary structures highly localized to functionally important sites within the human genome, have emerged as important new drug targets. The putative G-quadruplex-forming sequence (Pu27) in the NHE-III1 promoter region of the c-Myc gene is of particular interest as stabilization of this G-quadruplex with TMPyP4 has been shown to repress c-Myc transcription. In this study, we examine the Pu27 G-quadruplex-forming sequence and its interaction with TMPyP4. We report that the Pu27 sequence exists as a heterogeneous mixture of monomeric and higher-order G-quadruplex species in vitro and that this mixture can be partially resolved by size exclusion chromatography (SEC) separation. Within this ensemble of configurations, the equilibrium can be altered by modifying the buffer composition, annealing procedure, and dialysis protocol thereby affecting the distribution of G-quadruplex species formed. TMPyP4 was found to bind preferentially to higher-order G-quadruplex species suggesting the possibility of stabilization of the junctions of the c-Myc G-quadruplex multimers by porphyrin end-stacking. We also examined four modified c-Myc sequences that have been previously reported and found a narrower distribution of quadruplex configurations compared to the parent Pu27 sequence. We could not definitively conclude whether these G-quadruplex structures were selected from the original ensemble or if they are new G-quadruplex structures. Since these sequences differ considerably from the wild-type promoter sequence, it is unclear whether their structures have any actual biological relevance. Additional studies are needed to examine how the polymorphic nature of G-quadruplexes affects the interpretation of in vitro data for c-Myc and other G-quadruplexes. The findings reported here demonstrate that experimental conditions contribute significantly to G-quadruplex formation and should be carefully considered, controlled, and reported in detail. PMID:23108607

  5. Prognostic Value of Beta-Tubulin-3 and c-Myc in Muscle Invasive Urothelial Carcinoma of the Bladder

    Science.gov (United States)

    Massari, Francesco; Bria, Emilio; Ciccarese, Chiara; Munari, Enrico; Modena, Alessandra; Zambonin, Valentina; Sperduti, Isabella; Artibani, Walter; Cheng, Liang; Martignoni, Guido; Tortora, Giampaolo; Brunelli, Matteo

    2015-01-01

    Background To date, putative prognostic biomarkers have shown limited utility from the clinical perspective for bladder urothelial carcinoma. Herein, the expression of beta-tubulin-3 and c-Myc was evaluated to determine their prognostic potential. Methods In formalin fixed-paraffin embedded blocks, immunohistochemical expression of c-Myc and beta-tubulin-3 was evaluated. H score ranging from 0 to 300 was obtained by multiplying the percentage of positive cells by intensity (0–3); c-Myc and beta-tubulin-3 expression was defined: 0: negative, 1: weakly positive, 2: strongly positive. Results beta-tubulin-3 and c-Myc immunoexpression was available for 46 cases. At the univariate analysis, node-involvement, beta-tubulin-3 and c-Myc overexpression discriminate shorter DFS (HR 2.19, p = 0.043; HR 3.10, p = 0.24 and HR 3.05, p = 0.011, respectively); 2-yrs DFS log-rank analysis according to low versus high level of immunoexpression were statistically significant; beta-tubulin-3, 53% low vs 12.7% high (p = value 0.02) and c-Myc 28 low vs 8 high (p-value 0.007). Patients displaying negative beta-tubulin-3/c-Myc had statistically significant better 2-yrs DFS than those with mixed expression or double positivity (54.5% versus 18.7% versus 0%, log-rank p = 0.006). Conclusions c-Myc and beta-tubulin-3 show improvement for prognostic risk stratification in patients with muscle invasive bladder urothelial carcinoma. These molecular pathways may also be candidate to improve predictiveness to targeted therapies. PMID:26046361

  6. Prognostic Value of Beta-Tubulin-3 and c-Myc in Muscle Invasive Urothelial Carcinoma of the Bladder.

    Directory of Open Access Journals (Sweden)

    Francesco Massari

    Full Text Available To date, putative prognostic biomarkers have shown limited utility from the clinical perspective for bladder urothelial carcinoma. Herein, the expression of beta-tubulin-3 and c-Myc was evaluated to determine their prognostic potential.In formalin fixed-paraffin embedded blocks, immunohistochemical expression of c-Myc and beta-tubulin-3 was evaluated. H score ranging from 0 to 300 was obtained by multiplying the percentage of positive cells by intensity (0-3; c-Myc and beta-tubulin-3 expression was defined: 0: negative, 1: weakly positive, 2: strongly positive.beta-tubulin-3 and c-Myc immunoexpression was available for 46 cases. At the univariate analysis, node-involvement, beta-tubulin-3 and c-Myc overexpression discriminate shorter DFS (HR 2.19, p = 0.043; HR 3.10, p = 0.24 and HR 3.05, p = 0.011, respectively; 2-yrs DFS log-rank analysis according to low versus high level of immunoexpression were statistically significant; beta-tubulin-3, 53% low vs 12.7% high (p = value 0.02 and c-Myc 28 low vs 8 high (p-value 0.007. Patients displaying negative beta-tubulin-3/c-Myc had statistically significant better 2-yrs DFS than those with mixed expression or double positivity (54.5% versus 18.7% versus 0%, log-rank p = 0.006.c-Myc and beta-tubulin-3 show improvement for prognostic risk stratification in patients with muscle invasive bladder urothelial carcinoma. These molecular pathways may also be candidate to improve predictiveness to targeted therapies.

  7. C-Myc negatively controls the tumor suppressor PTEN by upregulating miR-26a in glioblastoma multiforme cells

    Energy Technology Data Exchange (ETDEWEB)

    Guo, Pin; Nie, Quanmin; Lan, Jin; Ge, Jianwei [Department of Neurosurgery, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200127 (China); Qiu, Yongming, E-mail: qiuzhoub@hotmail.com [Department of Neurosurgery, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200127 (China); Shanghai Institute of Head Trauma, Shanghai 200127 (China); Mao, Qing, E-mail: maoq@netease.com [Department of Neurosurgery, Ren Ji Hospital, School of Medicine, Shanghai Jiao Tong University, Shanghai 200127 (China); Shanghai Institute of Head Trauma, Shanghai 200127 (China)

    2013-11-08

    Highlights: •The c-Myc oncogene directly upregulates miR-26a expression in GBM cells. •ChIP assays demonstrate that c-Myc interacts with the miR-26a promoter. •Luciferase reporter assays show that PTEN is a specific target of miR-26a. •C-Myc–miR-26a suppression of PTEN may regulate the PTEN/AKT pathway. •Overexpression of c-Myc enhances the proliferative capacity of GBM cells. -- Abstract: The c-Myc oncogene is amplified in many tumor types. It is an important regulator of cell proliferation and has been linked to altered miRNA expression, suggesting that c-Myc-regulated miRNAs might contribute to tumor progression. Although miR-26a has been reported to be upregulated in glioblastoma multiforme (GBM), the mechanism has not been established. We have shown that ectopic expression of miR-26a influenced cell proliferation by targeting PTEN, a tumor suppressor gene that is inactivated in many common malignancies, including GBM. Our findings suggest that c-Myc modulates genes associated with oncogenesis in GBM through deregulation of miRNAs via the c-Myc–miR-26a–PTEN signaling pathway. This may be of clinical relevance.

  8. The distribution of the c-myc oncogene product in malignant lymphomas and various normal tissues as demonstrated by immunocytochemistry.

    Science.gov (United States)

    Jack, A. S.; Kerr, I. B.; Evan, G.; Lee, F. D.

    1986-01-01

    The expression of c-myc was studied in 51 malignant lymphomas and in a variety of normal tissues by immunocytochemistry using monoclonal antibodies raised to different synthetic peptides and reacting monospecifically with the c-myc product (p62c-myc). The c-myc product was detected in only a minority of malignant lymphomas principally those containing cells with immunoblastic characteristics, and was predominantly localised to the cytoplasm. In normal lymphoid tissues only plasma cells and histiocytes were found to have immunoreactivity. In non-lymphoid normal tissues, however, the c-myc product was distributed widely. Marked differences in its intracellular distribution were apparent in different tissues. These findings suggest that the relationship of p62c-myc expression to cell division may be more complex than previously suggested by in vitro studies, and raise the possibility that it may have other functions within the cell. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:3521694

  9. Significance of HER2 and C-MYC oncogene amplifications in breast cancer in atomic bomb survivors: associations with radiation exposure and histologic grade.

    Science.gov (United States)

    Miura, Shiro; Nakashima, Masahiro; Ito, Masahiro; Kondo, Hisayoshi; Meirmanov, Serik; Hayashi, Tomayoshi; Soda, Midori; Matsuo, Takeshi; Sekine, Ichiro

    2008-05-15

    It has been postulated that radiation induces breast cancers in atomic bomb (A-bomb) survivors. Oncogene amplification is an important mechanism during breast carcinogenesis and also serves as an indicator of genomic instability (GIN). The objective of this study was to clarify the association of oncogene amplification in breast cancer in A-bomb survivors with radiation exposure. In total, 593 breast cancers were identified in A-bomb survivors from 1968 to 1999, and the association between breast cancer incidence and A-bomb radiation exposure was evaluated. Invasive ductal cancers from 67 survivors and 30 nonsurvivors were analyzed for amplification of the HER2 and C-MYC genes by fluorescence in situ hybridization, and expression levels of hormone receptors were analyzed by immunostaining. The incidence rate increased significantly as exposure distance decreased from the hypocenter (hazard ratio per 1-km decrement, 1.47; 95% confidence interval [95% CI], 1.30-1.66). The incidence of HER2 and C-MYC amplification was increased significantly in the order of the control group, the distal group (P = .0238), and the proximal group (P = .0128). Multivariate analyses revealed that distance was a risk factor for the coamplification of C-MYC and HER2 in breast cancer in survivors (odds ratio per 1-km increment, 0.17; 95% CI, 0.01-0.63). The histologic grade of breast cancers became significantly higher in the order of the control group, the distal group, and the proximal group and was associated with oncogene amplifications. The current results suggested that A-bomb radiation may affect the development of oncogene amplification by inducing GIN and may be associated with a higher histologic grade in breast cancer among A-bomb survivors. (c) 2008 American Cancer Society.

  10. Localisation of lung cancer by a radiolabelled monoclonal antibody against the c-myc oncogene product

    Energy Technology Data Exchange (ETDEWEB)

    Chan, S.Y.T.; Evan, G.I.; Ritson, A.; Watson, J.; Wraight, P.; Sikora, K.

    1986-11-01

    A set of mouse monoclonal antibodies against the c-myc oncogene product, a 62,000 dalton nuclear binding protein involved in cell cycle control, has been constructed by immunisation with synthetic peptide fragments. One such antibody, CT14, was radiolabelled with /sup 131/I and administered to 20 patients with different malignant diseases. Good tumour localisation was observed in 12 out of 14 patients with primary bronchial carcinoma but not in patients with pulmonary metastases from primary tumours elsewhere. Successfully localised tumours were all 3 cm or more in diameter. Monoclonal antibodies against oncogene products may provide novel selective tools for the diagnosis and therapy of cancer.

  11. The role of the proto-oncogene c-myc in B lymphocyte differentiation.

    Science.gov (United States)

    Fernandez, David; Sanchez-Arevalo, Victor Javier; de Alboran, Ignacio Moreno

    2012-01-01

    Since the discovery of the myc gene, few genes are likely to have such influence on biomedical research. The diversity of the biological functions regulated by this transcription factor and its impact in human health have attracted investigators from many different fields. The development of conditional knockout mouse models has allowed for the characterization of Myc-driven molecular mechanisms in primary cells in physiological and pathological conditions. In this review, we discuss some of the main functions and recent findings regarding c-Myc in in vivo B lymphocyte differentiation from early progenitors to terminally differentiated cells.

  12. Impact of C-Myc gene-related aberrations in newly diagnosed myeloma with bortezomib/dexamethasone therapy.

    Science.gov (United States)

    Sekiguchi, Naohiro; Ootsubo, Kaori; Wagatsuma, Miyuki; Midorikawa, Kiyoe; Nagata, Akihisa; Noto, Satoshi; Yamada, Kazuaki; Takezako, Naoki

    2014-03-01

    Recent studies have suggested that c-Myc over-expression may be a factor indicating poor prognosis in multiple myeloma (MM), although c-Myc gene-related abnormalities, including translocation and gene amplification, have not been fully investigated in the novel agent era. Additional chromosome 8 may be considered as aggressive disease in the 1990s. To clarify the impact of these aberrations, we retrospectively analyzed newly diagnosed MM (NDMM) and relapsed/refractory MM (RRMM) with bortezomib and dexamethasone induction therapy. In the present study, the high-risk group was defined as having at least one of the following present: non-hyperdiploidy, IgH/FGFR3, and del p53. Forty NDMM cases were analyzed. At the median follow-up duration of 14.1 months, 14 RRMM were recognized. The proportions of patients in the high-risk, c-Myc gene-related aberrations, and additional chromosome 8 groups at diagnosis were 45.5, 22.5, and 10 %, respectively. The proportions of patients who developed RRMM in the high-risk, c-Myc gene-related aberrations, and additional chromosome 8 groups were 41.7, 77.7, and 50 %, respectively. Furthermore, patients with c-Myc gene-related abnormalities tended to exhibit inferior progression-free survival (PFS), and those with c-Myc gene-related abnormalities and/or additional chromosome 8 showed statistically shorter PFS. Therefore, c-Myc gene-related abnormalities and additional chromosome 8 may be related to a poorer prognosis.

  13. C-MYC overexpression predicts aggressive transformation and a poor outcome in mucosa-associated lymphoid tissue lymphomas.

    Science.gov (United States)

    Huang, Wenting; Guo, Lei; Liu, Hongyan; Zheng, Bo; Ying, Jianming; Lv, Ning

    2014-01-01

    Mucosa-associated lymphoid tissue (MALT) lymphoma is a relatively common, indolent B-cell lymphoma. MALT lymphoma with large tumor cells (LTCs) is believed to have the potential to transform to aggressive diffuse large B-cell lymphoma (DLBCL) which may have a poor prognosis. C-MYC is a transcription factor. Its translocation and overexpression predicts an inferior prognosis and poor response to therapy in cases of DLBCL. In the current study, C-MYC expression was detected in MALT lymphomas, and its relationship to the occurrence of LTCs, clinicopathological parameters and prognosis was assessed. A total of 69 cases were enrolled in the study, including 42 cases of MALT lymphoma without LTCs, 20 cases of MALT lymphoma with LTCs and 7 cases of DLBCL with a MALT lymphoma component (DLBCL+MALT). Immunohistochemistry and fluorescent in situ hybridization analyses were performed. In total, 15/42 (35.7%) cases were nuclear positive for C-MYC expression in the group without LTCs, whereas 15/20 (75.0%) and 4/7 (57.1%) cases were positive in the group with LTCs and in the group with DLBCL+MALT, respectively (P=0.004). Univariate and multivariate analysis were used to determine the correlations of C-MYC expression and clinicopathological parameters with overall survival (OS). C-MYC expression, Ann Arbor stage, LDH level and IPI were considerably associated with OS according to the univariate analysis. However, only C-MYC expression ≥ 20% showed a statistical significance in the multivariate analysis (HR=20.604, 95% CI: 1.909-222.412, P=0.013). Therefore, C-MYC overexpression may play an important role in aggressive transformation and is an independent prognostic factor in MALT lymphoma.

  14. miR-494 acts as an anti-oncogene in gastric carcinoma by targeting c-myc.

    Science.gov (United States)

    He, Weiling; Li, Yuhuang; Chen, Xinlin; Lu, Liya; Tang, Bin; Wang, Zhixiong; Pan, Yunbao; Cai, Shirong; He, Yulong; Ke, Zunfu

    2014-01-01

    We recently showed that miR-494 was downregulated in gastric carcinoma (GC). The objectives of this study were to determine the role of miR-494 in GC malignancy and to identify its target genes. Real-time polymerase chain reaction was employed to quantify the expression level of miR-494 and c-myc in gastric cancer tissues. Bioinformatics was used to predict the downstream target genes of miR-494, which were confirmed by luciferase and RNA immunoprecipitation assays. Cell functional analyses and a xenograft mouse model were used to evaluate the role of miR-494 in malignancy. miR-494 was downregulated in human GC tissues and in GC cells and was negatively correlated with c-myc expression. High level of c-myc or low level of miR-494 correlated with poor prognosis. The miR-494-binding site in the c-myc 3' untranslated region was predicted using TargetScan and was confirmed by the luciferase assay. Additionally, c-myc and miR-494 were enriched in coimmunoprecipitates with tagged Argonaute2 proteins in cells overexpressing miR-494. Furthermore, a miR-494 mimic significantly downregulated endogenous c-myc expression, which may contribute to the delayed G1/S transition, decreased synthesis phase bromodeoxyuridine incorporation, and impaired cell growth and colony formation; on the other hand, treatment with a miR-494 inhibitor displayed the opposite effects. Reduced tumor burden and decreased cell proliferation were observed following the delivery of miR-494 into xenograft mice. miR-494 is downregulated in human GC and acts as an anti-oncogene by targeting c-myc. miR-494 plays a role in the pathogenesis of gastric cancer in a recessive fashion. © 2014 Journal of Gastroenterology and Hepatology Foundation and Wiley Publishing Asia Pty Ltd.

  15. Alternative DNA structure formation in the mutagenic human c-MYC promoter.

    Science.gov (United States)

    Del Mundo, Imee Marie A; Zewail-Foote, Maha; Kerwin, Sean M; Vasquez, Karen M

    2017-05-05

    Mutation 'hotspot' regions in the genome are susceptible to genetic instability, implicating them in diseases. These hotspots are not random and often co-localize with DNA sequences potentially capable of adopting alternative DNA structures (non-B DNA, e.g. H-DNA and G4-DNA), which have been identified as endogenous sources of genomic instability. There are regions that contain overlapping sequences that may form more than one non-B DNA structure. The extent to which one structure impacts the formation/stability of another, within the sequence, is not fully understood. To address this issue, we investigated the folding preferences of oligonucleotides from a chromosomal breakpoint hotspot in the human c-MYC oncogene containing both potential G4-forming and H-DNA-forming elements. We characterized the structures formed in the presence of G4-DNA-stabilizing K+ ions or H-DNA-stabilizing Mg2+ ions using multiple techniques. We found that under conditions favorable for H-DNA formation, a stable intramolecular triplex DNA structure predominated; whereas, under K+-rich, G4-DNA-forming conditions, a plurality of unfolded and folded species were present. Thus, within a limited region containing sequences with the potential to adopt multiple structures, only one structure predominates under a given condition. The predominance of H-DNA implicates this structure in the instability associated with the human c-MYC oncogene. © The Author(s) 2017. Published by Oxford University Press on behalf of Nucleic Acids Research.

  16. DSC Deconvolution of the Structural Complexity of c-MYC P1 Promoter G-Quadruplexes

    Science.gov (United States)

    Dettler, Jamie M.; Buscaglia, Robert; Le, Vu H.; Lewis, Edwin A.

    2011-01-01

    We completed a biophysical characterization of the c-MYC proto-oncogene P1 promoter quadruplex and its interaction with a cationic porphyrin, 5,10,15,20-tetra(N-methyl-4-pyridyl)porphyrin (TMPyP4), using differential scanning calorimetry, isothermal titration calorimetry, and circular dichroism spectroscopy. We examined three different 24-mer oligonucleotides, including the wild-type (WT) sequence found in the c-MYC P1 promoter and two mutant G→T sequences that are known to fold into single 1:2:1 and 1:6:1 loop isomer quadruplexes. Biophysical experiments were performed on all three oligonucleotide sequences at two different ionic strengths (30 mM [K+] and 130 mM [K+]). Differential scanning calorimetry experiments demonstrated that the WT quadruplex consists of a mixture of at least two different folded conformers at both ionic strengths, whereas both mutant sequences exhibit a single two-state melting transition at both ionic strengths. Isothermal titration calorimetry experiments demonstrated that both mutant sequences bind 4 mols of TMPyP4 to 1 mol of DNA, in similarity to the WT sequence. The circular dichroism spectroscopy signatures for all three oligonucleotides at both ionic strengths are consistent with an intramolecular parallel stranded G-quadruplex structure, and no change in quadruplex structure is observed upon addition of saturating amounts of TMPyP4 (i.e., 4:1 TMPyP4/DNA). PMID:21402034

  17. The oncogene c-Myc coordinates regulation of metabolic networks to enable rapid cell cycle entry.

    Science.gov (United States)

    Morrish, Fionnuala; Neretti, Nicola; Sedivy, John M; Hockenbery, David M

    2008-04-15

    The c-myc proto-oncogene is rapidly activated by serum and regulates genes involved in metabolism and cell cycle progression. This gene is thereby uniquely poised to coordinate both the metabolic and cell cycle regulatory events required for cell cycle entry. However, this function of Myc has not been evaluated. Using a rat fibroblast model of isogenic cell lines, myc(-/-), myc(+/-), myc(+/+) and myc(-/-) cells with an inducible c-myc transgene (mycER), we show that the Myc protein programs cells to utilize both oxidative phosphorylation and glycolysis to drive cell cycle progression. We demonstrate this coordinate regulation of metabolic networks is essential, as specific inhibitors of these pathways block Myc-induced proliferation. Metabolic events temporally correlated with cell cycle entry include increased oxygen consumption, mitochondrial function, pyruvate and lactate production, and ATP generation. Treatment of normal cells with inhibitors of oxidative phosphorylation recapitulates the myc(-/-) phenotype, resulting in impaired cell cycle entry and reduced metabolism. Combined with a kinetic expression profiling analysis of genes linked to mitochondrial function, our study indicates that Myc's ability to coordinately regulate the mitochondrial metabolic network transcriptome is required for rapid cell cycle entry. This function of Myc may underlie the pervasive presence of Myc in many human cancers.

  18. Effect of c-myc on the ultrastructural structure of cochleae in guinea pigs with noise induced hearing loss

    Energy Technology Data Exchange (ETDEWEB)

    Han, Yu; Zhong, Cuiping [Department of Otolaryngology, Xijing Hospital, Fourth Military Medical University, Xi' an, 710032 Shaanxi Province (China); Hong, Liu [First Division of Digestive Diseases, Xijing Hospital, Fourth Military Medical University, Xi' an, 710032 Shaanxi Province (China); Wang, Ye; Qiao, Li [Department of Otolaryngology, Xijing Hospital, Fourth Military Medical University, Xi' an, 710032 Shaanxi Province (China); Qiu, Jianhua, E-mail: qiujh@fmmu.edu.cn [Department of Otolaryngology, Xijing Hospital, Fourth Military Medical University, Xi' an, 710032 Shaanxi Province (China)

    2009-12-18

    Noise over-stimulation may induce hair cells loss and hearing deficit. The c-myc oncogene is a major regulator for cell proliferation, growth, and apoptosis. However, the role of this gene in the mammalian cochlea is still unclear. The study was designed to firstly investigate its function under noise condition, from the aspect of cochlear ultrastructural changes. We had established the adenoviral vector of c-myc gene and delivered the adenovirus suspension into the scala tympani of guinea pigs 4 days before noise exposure. The empty adenoviral vectors were injected as control. Then, all subjects were exposed to 4-kHz octave-band noise at 110 dB SPL for 8 h/day, 3 days consecutively. Auditory thresholds were assessed by auditory brainstem response, prior to and 7 days following noise exposure. On the seventh days after noise exposure, the cochlear sensory epithelia surface was observed microscopically and the cochleae were taken to study the ultrastructural changes. The results indicated that auditory threshold shift after noise exposure was higher in the ears treated with Ad.EGFP than that treated with Ad.c-myc-EGFP. Stereocilia loss and the disarrangement of outer hair cells were observed, with greater changes found in the Ad.EGFP group. Also, the ultrastructure changes were severe in the Ad.EGFP group, but not obvious in the Ad.c-myc-EGFP group. Therefore, c-myc gene might play an unexpected role in hearing functional and morphological protection from acoustic trauma.

  19. Infection by Toxoplasma gondii Specifically Induces Host c-Myc and the Genes This Pivotal Transcription Factor Regulates

    Science.gov (United States)

    Franco, Magdalena; Shastri, Anjali J.

    2014-01-01

    Toxoplasma gondii infection has previously been described to cause dramatic changes in the host transcriptome by manipulating key regulators, including STATs, NF-κB, and microRNAs. Here, we report that Toxoplasma tachyzoites also mediate rapid and sustained induction of another pivotal regulator of host cell transcription, c-Myc. This induction is seen in cells infected with all three canonical types of Toxoplasma but not the closely related apicomplexan parasite Neospora caninum. Coinfection of cells with both Toxoplasma and Neospora still results in an increase in the level of host c-Myc, showing that c-Myc is actively upregulated by Toxoplasma infection (rather than repressed by Neospora). We further demonstrate that this upregulation may be mediated through c-Jun N-terminal protein kinase (JNK) and is unlikely to be a nonspecific host response, as heat-killed Toxoplasma parasites do not induce this increase and neither do nonviable parasites inside the host cell. Finally, we show that the induced c-Myc is active and that transcripts dependent on its function are upregulated, as predicted. Hence, c-Myc represents an additional way in which Toxoplasma tachyzoites have evolved to specifically alter host cell functions during intracellular growth. PMID:24532536

  20. The expression of p53, Rb, and c-myc protein in cervical cancer by immunohistochemistry stain

    Directory of Open Access Journals (Sweden)

    AMBAR MUDIGDO

    2005-07-01

    Full Text Available The pathogenesis of cancer as whole (50% is caused by gene mutation. Pathogenesis of cervical cancer has focusing on Human Papilloma Virus (HPV. Early-7 (E7 proteins of HPV shell bind the Rb tumor suppressor gene, so pRb (Rb protein can’t express. Because of the E2F transcription factor gene can’t bound with pRb, so E2F gene are going active and help c-myc for DNA replication and to stimuli the cell cycle. E6 protein of HPV is bind to and facilitates the degradation of the p53 tumor suppressor gene product. The objective of this experiment is to known the expression of p53, Rb and c-myc proteins in cancer of uterine cervix. Nineteen blocks paraffin tissue of cervical cancer are cut in thoroughly cleaned cryotome and place in glass plate that covered with poly-elysine. The immunohistochemistry is done with monoclonal antibody anti p53, Rb and c-myc proteins. The Result of this experiment is shown that the expression of proteins of p53 protein is 40%, Rb protein is 30.8% and c-myc protein is 50.1%. The conclusion from this experiment is that the expression of p53, Rb and c-myc proteins in cervical cancer are in mild category (30-70%. The experiment about cervical cancer is suggested.

  1. Deficiency in the DNA glycosylases UNG1 and OGG1 does not potentiate c-Myc-induced B-cell lymphomagenesis

    DEFF Research Database (Denmark)

    Green, Blerta; Martin, Alberto; Belcheva, Antoaneta

    2017-01-01

    C-Myc overexpression mediates lymphomagenesis, however, secondary genetic lesions are required for its full oncogenic potential. The origin and the mechanism of formation of these mutations are unclear. Here we investigated the role of Ogg1 and UNG glycosylases in c-Myc driven lymphomagenesis...... but found that their deficiencies did not influence the disease outcome in the Eµ c-Myc model. We also found that Rag proteins do not contribute to this process. Instead, our work suggests that secondary mutations that potentiate C-Myc lymphomagenesis arise early in B-cell ontogeny, occur at C:G basepairs...

  2. Human adipose tissue-derived stem cells cultured in xeno-free culture condition enhance c-MYC expression increasing proliferation but bypassing spontaneous cell transformation.

    Science.gov (United States)

    Paula, Ana C C; Martins, Thaís M M; Zonari, Alessandra; Frade, Soraia P P J; Angelo, Patrícia C; Gomes, Dawidson A; Goes, Alfredo M

    2015-04-14

    Human adipose tissue-derived stem cells (hASCs) are attractive cells for therapeutic applications and are currently being evaluated in multiple clinical trials. Prior to their clinical application, hASCs must be expanded ex vivo to obtain the required number of cells for transplantation. Fetal bovine serum is the supplement most widely used for cell culture, but it has disadvantages and it is not safe for cell therapy due to the risks of pathogen transmission and immune reaction. Furthermore, the cell expansion poses a risk of accumulating genetic abnormalities that could lead to malignant cell transformation. In this study, our aim was to evaluate the proliferation pattern as well as the resistance to spontaneous transformation of hASCs during expansion in a xeno-free culture condition. hASCs were expanded in Dulbecco's modified Eagle's medium supplemented with pooled allogeneic human serum or fetal bovine serum to enable a side-by-side comparison. Cell viability and differentiation capacity toward the mesenchymal lineages were assessed, along with immunophenotype. Ki-67 expression and the proliferation kinetics were investigated. The expression of the transcription factors c-FOS and c-MYC was examined with Western blot, and MYC, CDKN2A, ERBB2 and TERT gene expression was assessed with quantitative PCR. Senescence was evaluated by β-gal staining. Karyotype analysis was performed and tumorigenesis assay in vivo was also evaluated. The hASCs expanded in medium with pooled allogeneic human serum did not show remarkable differences in morphology, viability, differentiation capacity or immunophenotype. The main difference observed was a significantly higher proliferative effect on hASCs cultured in pooled allogeneic human serum. There was no significant difference in C-FOS expression; however, C-MYC protein expression was enhanced in pooled allogeneic human serum cultures compared to fetal bovine serum cultures. No difference was observed in MYC and TERT mRNA levels

  3. Mechanism of action of EBV, Bcl-2, p53, c-Myc and Rb in non-Hodgkin's lymphoma.

    Science.gov (United States)

    Song, W; Liu, M-G; Zhang, J-B; Zhang, J-J; Sun, M-M; Yu, Q-K

    2016-01-01

    The aim of the present study is to explore the mechanism of action of several proteins, including Epstein-Barr virus (EBV), B-cell lymphoma (Bcl)-2, p53, c-Myc and retinoblastoma (Rb), in Non-Hodgkin's lymphoma (NHL). Between July 2010 and July 2015, samples of 142 patients with pathologically confirmed NHL which presented at our institution were included in the observation group. In addition, samples from 55 patients with hyperplastic lymphadenitis presented during the same period were enrolled as control group. The expressions of EBV (+), p53(+), Bcl-2(+), Rb(-) and c-Myc(+) were determined and compared Between July 2010 and July 2015, samples of 142 patients with pathologically confirmed NHL which presented at our institution were included in the observation group. In addition, samples from 55 patients with hyperplastic lymphadenitis presented during the same period were enrolled as control group. The expressions of EBV (+), p53(+), Bcl-2(+), Rb(-) and c-Myc(+) were determined and compared among different subtypes and stages of NHLs of observation group. Besides, the correlation of EBV with p53, Bcl-2, Rb and c-Myc were investigated in NHLs of observation group. In the observation group, the expression rates of EBV(+), p53(+), Bcl-2(+), Rb(-), and c-Myc(+) were significantly higher than those, respectively, in the control group (p EBV expression and the expressions of p53, Bcl-2, Rb and c-Myc in the observation group (p > 0.05). The expression rates of p53(+) and Bcl-2(+) were significantly higher in aggressive and highly-aggressive NHLs than in indolent NHLs of the observation group (p EBV(+), p53(+), Bcl-2(+), Rb(-), and c-Myc(+) were significantly higher in stage III-IV NHLs than in stage I-II NHLs (p EBV(+), p53(+), Bcl-2(+), Rb(-), and c-Myc(+) are closely associated with NHL pathogenesis. Expressions of these proteins are higher in later stages of NHLs, and expressions of p53(+) and Bcl-2(+) are higher in more aggressive NHLs.

  4. Oridonin induces apoptosis and senescence in colorectal cancer cells by increasing histone hyperacetylation and regulation of p16, p21, p27 and c-myc

    Directory of Open Access Journals (Sweden)

    Zhao Ying-Zheng

    2010-11-01

    Full Text Available Abstract Background Oridonin, a tetracycline diterpenoid compound, has the potential antitumor activities. Here, we evaluate the antitumor activity and action mechanisms of oridonin in colorectal cancer. Methods Effects of oridonin on cell proliferation were determined by using a CCK-8 Kit. Cell cycle distribution was determined by flow cytometry. Apoptosis was examined by analyzing subdiploid population and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assay. Senescent cells were determined by senescence-associated β-galactosidase activity analysis. Semi-quantitative RT-PCR was used to examine the changes of mRNA of p16, p21, p27 and c-myc. The concomitant changes of protein expression were analyzed with Western blot. Expression of AcH3 and AcH4 were examined by immunofluorescence staining and Western blots. Effects of oridonin on colony formation of SW1116 were examined by Soft Agar assay. The in vivo efficacy of oridonin was detected using a xenograft colorectal cancer model in nude mice. Results Oridonin induced potent growth inhibition, cell cycle arrest, apoptosis, senescence and colony-forming inhibition in three colorectal cancer cell lines in a dose-dependent manner in vitro. Daily i.p. injection of oridonin (6.25, 12.5 or 25 mg/kg for 28 days significantly inhibited the growth of SW1116 s.c. xenografts in BABL/C nude mice. With western blot and reverse transcription-PCR, we further showed that the antitumor activities of oridonin correlated with induction of histone (H3 and H4 hyperacetylation, activation of p21, p27 and p16, and suppression of c-myc expression. Conclusion Oridonin possesses potent in vitro and in vivo anti-colorectal cancer activities that correlated with induction of histone hyperacetylation and regulation of pathways critical for maintaining growth inhibition and cell cycle arrest. Therefore, oridonin may represent a novel therapeutic option in colorectal cancer treatment.

  5. USP10 Antagonizes c-Myc Transcriptional Activation through SIRT6 Stabilization to Suppress Tumor Formation

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    Zhenghong Lin

    2013-12-01

    Full Text Available The reduced protein expression of SIRT6 tumor suppressor is involved in tumorigenesis. The molecular mechanisms underlying SIRT6 protein downregulation in human cancers remain unknown. Using a proteomic approach, we have identified the ubiquitin-specific peptidase USP10, another tumor suppressor, as one of the SIRT6-interacting proteins. USP10 suppresses SIRT6 ubiquitination to protect SIRT6 from proteasomal degradation. USP10 antagonizes the transcriptional activity of the c-Myc oncogene through SIRT6, as well as p53, to inhibit cell-cycle progression, cancer cell growth, and tumor formation. To support this conclusion, we detected significant reductions in both USP10 and SIRT6 protein expression in human colon cancers. Our study discovered crosstalk between two tumor-suppressive genes in regulating cell-cycle progression and proliferation and showed that dysregulated USP10 function promotes tumorigenesis through SIRT6 degradation.

  6. Localisation of lung cancer by a radiolabelled monoclonal antibody against the c-myc oncogene product.

    Science.gov (United States)

    Chan, S. Y.; Evan, G. I.; Ritson, A.; Watson, J.; Wraight, P.; Sikora, K.

    1986-01-01

    A set of mouse nonoclonal antibodies against the c-myc oncogene product, a 62,000 dalton nuclear binding protein involved in cell cycle control, has been constructed by immunisation with synthetic peptide fragments. One such antibody, CT14, was radiolabelled with 131I and administered to 20 patients with different malignant diseases. Good tumour localisation was observed in 12 out of 14 patients with primary bronchial carcinoma but not in patients with pulmonary metastases from primary tumours elsewhere. Successfully localised tumours were all 3 cm or more in diameter. Monoclonal antibodies against oncogene products may provide novel selective tools for the diagnosis and therapy of cancer. Images Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:3801273

  7. Transplantation of induced pluripotent stem cells without C-Myc attenuates retinal ischemia and reperfusion injury in rats.

    Science.gov (United States)

    Fang, I-Mo; Yang, Chung-May; Yang, Chang-Hao; Chiou, Shih-Hwa; Chen, Muh-Shy

    2013-08-01

    Induced pluripotent stem cells (iPSC) are novel stem cell populations, but the role of iPSC in retinal ischemia and reperfusion (I/R) injury remains unknown. Since oncogene c-Myc is substantially contributed to tumor formation, in this study, we investigated the effects, mechanisms and safety of subretinal transplantation of iPSC without c-Myc (non-c-Myc iPSC) in a rat model of retinal I/R injury. Retinal I/R injury was induced by raising the intraocular pressure of Sprague-Dawley rats to 110 mmHg for 60 min. A subretinal injection of non-c-Myc iPSC or murine epidermal fibroblast was given 2 h after I/R injury. Electroretinograms (ERG) were performed to determine the functionality of the retinas. The surviving retinal ganglion cells (RGCs) and retinal apoptosis following I/R injury were determined by counting NeuN-positive cells in whole-mounted retinas and TUNEL staining, respectively. The generation of reactive oxygen species (ROS) and the activities of superoxide dismutase (SOD) and catalase (CAT) in the retinal tissues were determined by lucigenin- and luminol-enhanced chemiluminescence and enzyme-linked immunosorbent assay (ELISA). The degree of retinal oxidative damage was assessed by nitrotyrosine, acrolein, and 8-hydroxy-2'-deoxyguanosine (8-OHdG) staining. The expression of brain-derived neurotrophic factor (BDNF), ciliary neurotrophic factor (CNTF) and basic fibroblast growth factor (bFGF) in retinas was measured by immunohistochemistry and ELISA. We found that subretinal transplantation of non-c-Myc iPSC significantly suppressed the I/R-induced reduction in the ERG a- and b-wave ratio, attenuated I/R-induced loss of RGCs and remarkably ameliorated retinal morphological changes. Non-c-Myc iPSC potentially increased the activities of SOD and CAT, decreased the levels of ROS, which may account for preventing retinal cells from apoptotic cell death. In addition, the levels of BDNF and CNTF in retina were significantly elevated in non-c-Myc iPSC-treated eyes

  8. c-Myc Enhances Sonic Hedgehog-Induced Medulloblastoma Formation from Nestin-Expressing Neural Progenitors in Mice

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    Ganesh Rao

    2003-05-01

    Full Text Available Medulloblastomas are malignant brain tumors that arise in the cerebella of children. The presumed cellsof-origin are undifferentiated precursors of granule neurons that occupy the external granule layer (EGL of the developing cerebellum. The overexpression of proteins that normally stimulate proliferation of neural progenitor cells may initiate medulloblastoma formation. Two known mitogens for neural progenitors are the c-Myc oncoprotein and Sonic hedgehog (Shh, a crucial determinant of embryonic pattern formation in the central nervous system. We modeled the ability of c-Myc and Shh to induce medulloblastoma in mice using the RCAS/tv-a system, which allows postnatal gene transfer and expression in a cell type-specific manner. We targeted the expression of Shh and c-Myc to nestin-expressing neural progenitor cells by injecting replication-competent ALV splice acceptor (RCAS vectors into the cerebella of newborn mice. Following injection with RCAS-Shh alone, 3/32 (9% mice developed medulloblastomas and 5/32 showed multifocal hyperproliferation of the EGL, possibly a precursor stage of medulloblastoma. Following injection with RCAS-Shh plus RCAS-Myc, 9/39 (23% mice developed medulloblastomas. We conclude that nestin-expressing neural progenitors, present in the cerebellum at birth, can act as the cells-of-origin for medulloblastoma, and that c-Myc cooperates with Shh to enhance tumorigenicity.

  9. Expression of merlin, NDRG2, ERBB2, and c-MYC in meningiomas: relationship with tumor grade and recurrence.

    Science.gov (United States)

    Ongaratti, B R; Silva, C B O; Trott, G; Haag, T; Leães, C G S; Ferreira, N P; Oliveira, M C; Pereira-Lima, J F S

    2016-01-01

    Meningiomas are common, usually benign tumors of the central nervous system that have a high rate of post-surgical recurrence or regrowth. We determined expression of the proteins merlin, NDRG2, ERBB2, and c-MYC in meningiomas using immunohistochemistry and assessed relationships between protein expression and gender, age, tumor grade, and recurrence or regrowth. The study sample comprised 60 patients, (44 women and 16 men) with a mean age of 53.2 ± 12.7 years. Tumors were classified as grade I (n=48) or grades II and III (n=12). Expression of merlin, NDRG2, ERBB2, and c-MYC was not significantly different statistically with relation to gender, age, or meningioma recurrence or regrowth. Merlin was expressed in 100% of the cases. No statistically significant difference between tumor grade and recurrence or regrowth was identified. Statistically significant differences were identified between the mean age of patients with grade I (54.83 ± 11.60) and grades II and III (46.58 ± 15.08) meningiomas (P=0.043), between strong c-MYC expression and grades II and III (Pmerlin on tumorigenesis, the association of c-MYC with aggressive meningiomas, and that partial surgical resection is associated with tumor recurrence or regrowth.

  10. Amino-terminal domains of c-myc and N-myc proteins mediate binding to the retinoblastoma gene product

    Science.gov (United States)

    Rustgi, Anil K.; Dyson, Nicholas; Bernards, Rene

    1991-08-01

    THE proteins encoded by the myc gene family are involved in the control of cell proliferation and differentiation, and aberrant expression of myc proteins has been implicated in the genesis of a variety of neoplasms1. In the carboxyl terminus, myc proteins have two domains that encode a basic domain/helix-loop-helix and a leucine zipper motif, respectively. These motifs are involved both in DNA binding and in protein dimerization2-5. In addition, myc protein family members share several regions of highly conserved amino acids in their amino termini that are essential for transformation6,7. We report here that an N-terminal domain present in both the c-myc and N-myc proteins mediates binding to the retinoblastoma gene product, pRb. We show that the human papilloma virus E7 protein competes with c-myc for binding to pRb, indicating that these proteins share overlapping binding sites on pRb. Furthermore, a mutant Rb protein from a human tumour cell line that carried a 35-amino-acid deletion in its C terminus failed to bind to c-myc. Our results suggest that c-myc and pRb cooperate through direct binding to control cell proliferation.

  11. Alterations of C-MYC, NKX3.1, and E-cadherin expression in canine prostate carcinogenesis

    DEFF Research Database (Denmark)

    Fonseca-Alves, Carlos E; Rodrigues, Marcela M P; de Moura, Veridiana M B D

    2013-01-01

    using the peroxidase and DAB methods. The C-MYC protein expression was elevated in the cytoplasm and nuclei of the canine PCa and PIA compared with the normal prostate (P = 0.004. The NKX3.1 protein expression was reduced in 94.75% of the PCa and 100% of the PIA compared with the normal prostate (P = 0...

  12. Expression of the c-myc oncogene and the presence of HPV 18: possible surrogate markers for cervical cancer?

    Science.gov (United States)

    Rughooputh, S; Manraj, S; Eddoo, R; Greenwell, P

    2009-01-01

    The study aims to evaluate the cause of cervical cancer in a cohort of patients and to establish whether or not human papillomavirus (HPV) is the leading risk factor and to determine whether or not c-myc oncogene over-expression is a predicative marker for the disease. Cone biopsy samples are examined from 53 patients diagnosed with either adenocarcinoma or squamous cell carcinoma of the cervix. Results showed that 19% of the patients studied were positive for high-grade HPV 18 DNA by polymerase chain reaction (PCR). For the c-myc gene expression, only three (23%) of the 13 control slides were positive. Of 49 known cervical cancer patients examined, 41% were positive, 51% were negative and 8% were doubtful. Of those who were positive for HPV, only two were positive for a mutation in the c-myc gene and one slide gave a doubtful result. P value for hysterectomy patients was 0.23 and for cancer patients was 0.48. In the cervical cancer patients studied, the HPV 18 prevalence rate was very low compared to that found in other studies. Therefore, the presence of HPV and expression of the c-myc oncogene cannot be used as surrogate markers for cervical cancer.

  13. Occurrence and expression of p53 suppressor gene and c-Myc oncogene in dog eyelid tumors.

    Science.gov (United States)

    Lopes, Rodrigo Antonio; Cardoso, Tereza Cristina; Luvizotto, Maria Cecília Rui; de Andrade, Alexandre Lima

    2010-03-01

    To detect the occurrence and expression of the suppressor gene p53 and of the oncogene c-Myc in eyelid tumors of dogs using the PCR, RT-PCR, PCR-ELISA and RT-PCR-ELISA techniques. These genes have not been described in dog eyelid tumors before. Nine samples of eyelid or third eyelid epithelial tumors were obtained from the archives of the Department of Veterinary Pathology. Tumor diagnosis was confirmed by evaluation of hematoxylin-eosin stained sections, and immunohistochemistry for cytokeratin AE1/AE3 and vimentin V9. A canine mammary tumor was used for positive control. Agarose gel electrophoresis, PCR-ELISA and RT-PCR-ELISA were used to detect p53 and c-Myc genes. The occurrence of p53 was detected in most of the eyelid tumors and third eyelid tumors studied (88.8%, n = 8) and was expressed in 75% of the positive samples, as indicated by ELISA. The c-Myc gene was found in 77.7% (n = 7) of the samples and was expressed in eight samples. Eyelid and third eyelid tumors of dogs express both the p53 and the c-Myc genes as shown by PCR and RT-PCR. However, PCR ELISA and RT-PCR ELISA were more efficient in assessing occurrence and expression of these genes because they identified amplified products that were not detected by agarose gel electrophoresis.

  14. Effects of c-myc oncogene modulation on differentiation of human small cell lung carcinoma cell lines

    NARCIS (Netherlands)

    Van Waardenburg, RCAM; Meijer, C; Pinto-Sietsma, SJ; De Vries, EGE; Timens, W; Mulder, NM

    1998-01-01

    Amplification and over-expression of oncogenes of the myc family are related to the prognosis of certain solid tumors such as small cell lung cancer (SCLC). For SCLC, c-myc is the oncogene most consistently found to correlate with the end stage behaviour of the tumour, in particular with survival

  15. Rapid loss of intestinal crypts upon conditional deletion of the Wnt/Tcf-4 target gene c-Myc.

    NARCIS (Netherlands)

    Muncan, V.; Sansom, O.J.; Tertoolen, L.; Phesse, T.J.; Begthel, H.; Sancho, E.; Cole, A.M.; Gregorieff, A.; Alboran, I.M. de; Clevers, J.C.; Clarke, A.R.

    2006-01-01

    Inhibition of the mutationally activated Wnt cascade in colorectal cancer cell lines induces a rapid G1 arrest and subsequent differentiation. This arrest can be overcome by maintaining expression of a single Tcf4 target gene, the proto-oncogene c-Myc. Since colorectal cancer cells share many

  16. The 5T mouse multiple myeloma model: absence of c-myc oncogene rearrangement in early transplant generations.

    Science.gov (United States)

    Radl, J.; Punt, Y. A.; van den Enden-Vieveen, M. H.; Bentvelzen, P. A.; Bakkus, M. H.; van den Akker, T. W.; Benner, R.

    1990-01-01

    Consistent chromosomal translocations involving the c-myc cellular oncogene and one of the three immunoglobin loci are typical for human Burkitt's lymphoma, induced mouse plasmacytoma (MPC) and spontaneously arising rat immunocytoma (RIC). Another plasma cell malignancy, multiple myeloma (MM), arising spontaneously in the ageing C57BL/KaLwRij mice, was investigated in order to see whether the MM cells contain c-myc abnormalities of the MPC or RIC type. Rearrangement of the c-myc oncogene was found in the bone marrow cells only in 5T2 MM transplantation line in a mouse of the 24th generation and in none of the seven other MM of the 5T series which were of earlier generations. Since the mouse 5T MM resembles the human MM very closely, including the absence of consistent structural c-myc oncogene abnormalities, it can serve as a useful experimental model for studies on the aetiopathogenesis of this disease. Images Figure 2 Figure 3 PMID:2310679

  17. TATA-binding protein and the retinoblastoma gene product bind to overlapping epitopes on c-Myc and adenovirus E1A protein

    NARCIS (Netherlands)

    Hateboer, G.; Timmers, H.T.M.; Rustgi, A.K.; Billaud, Marc; Veer, L.J. Van 't; Bernards, R.A.

    1993-01-01

    Using a protein binding assay, we show that the amino-teminal 204 amino acids of the c-Myc protein interact di y with a key component of the basal p tdon factor TFID, the TATA box-binding protein (TBP). Essentialy the same region of the c-Myc protein alo binds the product of the retinoblatoma

  18. Isolation and characterization of c-myc, a cellular homolog of the oncogene (v-myc) of avian myelocytomatosis virus strain 29.

    Science.gov (United States)

    Vennstrom, B; Sheiness, D; Zabielski, J; Bishop, J M

    1982-01-01

    The chicken genome contains nucleotide sequences homologous to transforming genes (oncogenes) of a number of avian retroviruses. We have isolated chicken DNA (c-myc) that is homologous to the oncogene (v-myc) of the avian myelocytomatosis virus MC29 and have compared the structures of the cellular and viral genes. Results from restriction endonuclease mapping of c-myc and from analysis of heteroduplexes between the DNAs of the cellular and viral genes show that c-myc is homologous to 1,500 nucleotides in v-myc DNA. This homologous region is interrupted in c-myc by an intron-like sequence of 1,100 nucleotides which is absent from v-myc. Nuclear RNA from normal chicken cells contains at least five species of transcripts from c-myc ranging from 2.5 to 6.5 kilobases in length. By contrast, cytoplasm contains only the 2.5-kilobase c-myc RNA. These features of the c-myc gene and its nuclear transcripts are characteristic of normal cellular genes and suggest that the myc gene is of cellular rather than viral origin. The exons in c-myc may define two functional domains in the gene and may therefore facilitate the dissection of the different oncogenic potentials of the MC29 virus. Images PMID:6284994

  19. Non-Hodgkin's lymphomas with Burkitt-like cells are associated with c-Myc amplification and poor prognosis.

    Science.gov (United States)

    Mossafa, H; Damotte, D; Jenabian, A; Delarue, R; Vincenneau, A; Amouroux, I; Jeandel, R; Khoury, E; Martelli, J M; Samson, T; Tapia, S; Flandrin, G; Troussard, X

    2006-09-01

    Out of 344 patients with newly diagnosed non-Hodgkin's lymphoma (NHL), this study identified 16 patients presenting Burkitt-like cells (BLCs) after cytological and/or histological review. Conventional cytogenetic analysis showed at diagnosis complex chromosomal abnormalities in 13 cases and a normal karyotype in three cases. However, neither t(8;14)(q24;q32) nor the variants t(2;8)(p12;q24) or t(8;22)(q24;q11) was detected. FISH studies showed c-MYC amplification in all cases with four to more than seven copies in 10 - 77% metaphase or inter-phase cells. This study did not observe any gene fusion signal for c-MYC/IgH excluding a t(8;14) translocation and partial tri or polysomy of chromosome 8. It also excluded in that cases a break apart for the c-MYC locus. This study also never detected IgL/c-MYC, IgK/c-MYC or X-c-MYC. The BLCs were present whatever the lymphoma sub-type: follicular lymphoma (FL) was diagnosed in six out of 16 patients, mantle cell lymphoma (MCL) in four out of 16 patients, marginal zone lymphoma (MZL) in two out of 16 patients and diffuse large B-cell lymphomas (DLBCL) in three out of 16 patients. One additional patient presented a T-cell lymphoma. The clinical course was aggressive with a poor prognosis, as death occurred in nine patients, within 6 months after diagnosis for eight of them. These data could suggest a sub-group of NHL patients (15 B-NHL, 1 T-NHL) have been identified with a poor prognosis characterized by the association of Burkitt-like cells and c-MYC amplification without t(8;14)(q24;q32) or its variants. The possibility that this profile may represent a distinct morphologic NHL sub-set remains to be determined on a large cohort of patients.

  20. K-RAS point mutation, and amplification of C-MYC and C-ERBB2 in colon adenocarcinoma.

    Directory of Open Access Journals (Sweden)

    Tadeusz Pawełczyk

    2004-10-01

    Full Text Available The routine multidisciplinary management of colon cancer is based mainly on tumor staging, histology, grading and vascular invasion. In this approach, important individual information derived from molecular characteristics of the tumor may be missed, especially since significant heterogeneity of molecular aberrations in cancer cells has been observed, and recognition of every of relationships between them may be of value. K-RAS, C-MYC and C-ERBB2 are protooncogenes taking part in carcinogenesis and tumor progression in the colon. They influence cell proliferation, differentiation and survival. K-RAS point mutation, as well as amplification of C-MYC and C-ERBB2 were searched in 84 primary colon adenocarcinomas resected with curative intent. Multiplex polymerase-chain reaction and restriction fragment length polymorphism were performed to assess codon 12 K-RAS point mutation. Amplification of C-MYC and C-ERBB2 genes was evaluated by densitometry after agarose gel separation of the respective multiplex PCR products. No relation was found among mutated and/or amplified genes, and between searched molecular aberrations and pathoclinical features. In multivariate analysis, nodal status appeared to be the only independent prognostic indicator. In colon adenocarcinoma, codon 12 K-RAS point mutation and amplification of C-MYC and C-ERBB2 seem to occur independently in the process of tumor progression. Amplification of C-ERBB2 tends to associate with more advanced stage of disease. Concomitant occurrence of codon 12 K-RAS mutation, C-MYC and C-ERBB2 amplification was of no prognostic value in respect to survival.

  1. Spontaneous DNA lesions modulate DNA structural transitions occurring at nuclease hypersensitive element III(1) of the human c-myc proto-oncogene.

    Science.gov (United States)

    Beckett, Joshua; Burns, Jacob; Broxson, Christopher; Tornaletti, Silvia

    2012-07-03

    G quadruplex (G4) DNA is a noncanonical four-stranded DNA structure that can form in G repeats by stacking of planar arrays of four hydrogen-bonded guanines called G quartets, in the presence of potassium ions. In addition to a presumed function in the regulation of gene expression, G4 DNA also localizes to regions often characterized by genomic instability. This suggests that formation of this structure may interfere with DNA transactions, including processing of DNA damage at these sites. Here we have studied the effect of two spontaneous DNA lesions, the abasic site and 8-oxoguanine, on the transition from duplex to quadruplex DNA structure occurring at nuclease hypersensitive element III(1) (NHEIII(1)) of the human c-myc promoter. We show by dimethyl sulfate footprinting and RNA polymerase arrest assays that at physiological concentrations of potassium ions NHEIII(1) folds into two coexisting G4 DNA structures, myc-1245 and myc-2345, depending on which G runs are utilized for G quartet formation. We found that a single substitution of G12 of NHEIII(1) with a single abasic site or a single 8-oxoguanine prevented formation of G4 structure myc-2345 in favor of structure myc-1245, where the lesion was accommodated in a DNA loop formed by G11-AP12/(or 8-oxoG12)-G13-G14. Surprisingly, when an additional G to A base substitution was introduced at position 3 of NHEIII(1), we observed formation of myc-2345. The extent of this structural transition was modulated by the location and type of lesion within the G11-G14 repeat. Our data indicate that spontaneous lesions formed in the G4-forming sequence of c-myc NHEIII(1) affect the structural transitions occurring at this regulatory site, potentially altering transcription factor binding and DNA repair of lesions formed in this highly regulated sequence.

  2. Suppression of c-Myc induces apoptosis via an AMPK/mTOR-dependent pathway by 4-O-methyl-ascochlorin in leukemia cells.

    Science.gov (United States)

    Shin, Jae-Moon; Jeong, Yun-Jeong; Cho, Hyun-Ji; Magae, Junji; Bae, Young-Seuk; Chang, Young-Chae

    2016-05-01

    4-O-Methyl-ascochlorin (MAC) is a methylated derivative of the prenyl-phenol antibiotic ascochlorin, which was isolated from an incomplete fungus, Ascochyta viciae. Although the effects of MAC on apoptosis have been reported, the underlying mechanisms remain unknown. Here, we show that MAC promoted apoptotic cell death and downregulated c-Myc expression in K562 human leukemia cells. The effect of MAC on apoptosis was similar to that of 10058-F4 (a c-Myc inhibitor) or c-Myc siRNA, suggesting that the downregulation of c-Myc expression plays a role in the apoptotic effect of MAC. Further investigation showed that MAC downregulated c-Myc by inhibiting protein synthesis. MAC promoted the phosphorylation of AMP-activated protein kinase (AMPK) and inhibited the phosphorylation of mammalian target of rapamycin (mTOR) and its target proteins, including p70S6 K and 4E-BP-1. Treatment of cells with AICAR (an AMPK activator), rapamycin (an mTOR inhibitor), or mTOR siRNA downregulated c-Myc expression and induced apoptosis to a similar extent to that of MAC. These results suggest that the effect of MAC on apoptosis induction in human leukemia cells is mediated by the suppression of c-Myc protein synthesis via an AMPK/mTOR-dependent mechanism.

  3. Patients with high c-MYC-expressing squamous cell carcinomas of the tongue show better survival than those with low- and medium-expressing tumours.

    Science.gov (United States)

    Strindlund, Klas; Troiano, Giuseppe; Sgaramella, Nicola; Coates, Philip J; Gu, Xiaolian; Boldrup, Linda; Califano, Luigi; Fahraeus, Robin; Muzio, Lorenzo Lo; Ardito, Fatima; Colella, Giuseppe; Tartaro, Gianpaolo; Franco, Renato; Norberg-Spaak, Lena; Saadat, Mohammad; Nylander, Karin

    2017-11-01

    c-MYC is a potent oncoprotein with roles in a wide range of cellular processes such as differentiation, apoptosis and growth control. Deregulation of the MYC gene is commonly seen in human tumours resulting in overexpression of the protein. Here we studied expression of c-MYC in correlation to clinical outcome in patients with primary squamous cell carcinoma of the mobile tongue. Immunohistochemistry was used to identify c-MYC in a group of 104 tongue squamous cell carcinomas with an antibody directed against the N-terminal part of the protein. Staining was evaluated by multiplying the percentage of c-MYC-expressing cells with staining intensity, giving a quick score for each tumour. All 104 tumours expressed c-MYC at varying levels. Quantitation according to per cent of positive cells and staining intensity revealed that most (15/21; 71%) high-expressing tumours were seen in males. Within the group of high c-MYC-expressing tumours, the majority were alive 2 and 5 years after treatment. The present findings show that expression of c-MYC has prognostic value in squamous cell carcinoma of the tongue, and could be useful in choice of therapy. © 2017 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  4. [Effect of isorhmnetin on circadian rhythms of DNA synthesis and expression of c-myc gene in Eca-109 cells of human oesophageal cancer].

    Science.gov (United States)

    Yang, Chunlei; Peng, Tao; Qu, Yi; Tao, Dachang; Wang, Zhengrong; Zhu, Bin

    2005-12-01

    This study was focused on the circadian rhythms of DNA synthesis and the expression of c-myc gene in untreated and treated Eca-109 cells in human oesophageal cancer with isorhmnetin. The circadian rhythms of 3H-TdR incorporation and expression of c-myc gene in untreated and treated Eca-109 cells were measured by 3H-thymidine uptake assay and flow cytometry. The data collected were analyzed by ANOVA and Cosinor method. DNA synthesis and expression of c-myc gene in untreated group varied according to circadian time with statistical significance, the distribution curves of both DNA synthesis and the expression level of c-myc were fit for cosinor changes. The circadian rhythms of DNA synthesis and circadian parameters of c-myc expression in treated Eca-109 cells changed. The circadian parameters of DNA synthesis and expression level of c-myc varied after treatment by isorhmnetin. The effects of isorhmnetin on cell proliferation and c-myc expression reached the highest level from 20: 00 to 0: 00. The results provide a guidance for instituting the chemotherapy and chronotherapy of human tumors, when isorhmnetin is for use as anti-cancer agent.

  5. The FUSE binding proteins FBP1 and FBP3 are potential c-myc regulators in renal, but not in prostate and bladder cancer

    Directory of Open Access Journals (Sweden)

    Meyer Hellmuth-Alexander

    2008-12-01

    Full Text Available Abstract Background The three far-upstream element (FUSE binding proteins (FBP1, FBP2, and FBP3 belong to an ancient family of single-stranded DNA binding proteins which are required for proper regulation of the c-myc proto-oncogene. Whereas it is known that c-myc alterations play a completely different role in various carcinomas of the urogenital tract, the relevance of FBPs is unclear. Methods FBP1, FBP3 and c-myc expression was studied in 105 renal cell, 95 prostate and 112 urinary bladder carcinomas by immunohistochemistry using tissue microarrays. Results High rates of FBP1 and FBP3 expression were observed in all cancer types. There was a concomitant up-regulation of FBP1 and FBP3 in renal cell and prostate carcinomas (p C-myc expression was detectable in 21% of prostate, 30% of renal and 34% of urothelial carcinomas. Interestingly, strong FBP1 and FBP3 expression was associated with c-myc up-regulation in clear cell renal cell carcinomas (p Conclusion The correlation between FBP1/FBP3, c-myc and high proliferation rate in renal cell carcinoma provides strong in vivo support for the suggested role of FBP1 and FBP3 as activators of c-myc. The frequent up-regulation of FBP1 and FBP3 in urothelial and prostate carcinoma suggests that FBPs also have an important function in gene regulation of these tumors.

  6. Activating and sustaining c-Myc by depletion of miR-144/451 gene locus contributes to B-lymphomagenesis.

    Science.gov (United States)

    Ding, Lan; Zhang, Yanqing; Han, Lingling; Fu, Lei; Mei, Xia; Wang, Jijun; Itkow, Jacobi; Elabid, Afaf Elabid Ibrahim; Pang, Lei; Yu, Duonan

    2017-12-29

    Hyper activity of protooncogene c-Myc is one of the hallmarks of highly aggressive lymphomas. However, the mechanism of how c-Myc is subjected to activation and amplification is still not well defined. In this study, we use gene knockout strategy to show that targeted depletion of a well-conserved microRNA gene locus miR-144/451 initiates tumorigenesis including B-lymphoma development in aged mice. This is due, at least in part, to the direct activation of the c-Myc gene by loss of miR-144/451 expression in hematopoietic cells. Moreover, oncoprotein c-Myc inversely regulates miR-144/451 expression by directly binding to the miR-144/451 promoter region, forming a miRNA-Myc positive feedback loop to safeguard the high level of c-Myc in B-lymphocytes. We also demonstrate that this miRNA-Myc crosstalk is disrupted in human diffuse large B-cell lymphomas with aberrant c-Myc expression. Therefore, our findings provide strong evidence, for the first time, that deficiency of miR-144/451 expression may play a bona fide role in derepression of silenced c-Myc, which contributes to tumor development including B-lymphomagenesis.

  7. Detection of C-MYC oncogene translocation and copy number change in the normal-dysplasia-carcinoma sequence of the larynx by fluorescence in situ hybridization.

    Science.gov (United States)

    Liu, Yu; Gong, Li-Ping; Dong, Xiao-Li; Liu, Hong-Gang

    2013-06-01

    The aim of this study was to determine the translocation and copy number change of the C-MYC gene in patients with laryngeal dysplasia and laryngeal squamous cell carcinoma (LSCC), and to evaluate the prevalence of such expression in relation to the normal-dysplasia-carcinoma sequence. Fluorescent in situ hybridization (FISH) was applied on formalin-fixed paraffin-embedded blocks of 93 laryngeal lesion specimens (14 normal epithelium, 15 mild dysplasia, 18 moderate dysplasia, 16 severe dysplasia, 9 carcinoma in situ, and 21 invasive carcinoma). C-MYC translocation was not observed in all laryngeal tissue. The high frequency for C-MYC copy-number increased (100%) in invasive carcinoma: 57.14% amplifications and 42.86% gains, and the positive rate of C-MYC amplification and copy-number change increased with the increasing severity of laryngeal lesions (P < 0.0001). The results suggest that C-MYC may be activated by gain/amplification in LSCC and precancerous lesions. Thus, C-MYC may play an important role in promoting LSCC progression, and early FISH detection of C-MYC may be exploited to set a screening test for laryngeal dysplasia. Copyright © 2012 Wiley Periodicals, Inc.

  8. Driving gradual endogenous c-myc overexpression by flow-sorting: intracellular signaling and tumor cell phenotype correlate with oncogene expression.

    Science.gov (United States)

    Knudsen, Kasper Jermiin; Nelander Holm, Gitte-Mai; Krabbe, Jonas S; Listov-Saabye, Nicolai; Kiehr, Benedicte; Dufva, Martin; Svendsen, Jette E; Oleksiewicz, Martin B

    2009-12-01

    Insulin-exposed rat mammary cancer cells were flow sorted based on a c-myc reporter plasmid encoding a destabilized green fluorescent protein. Sorted cells exhibited gradual increases in c-myc levels. Cells overexpressing c-myc by only 10% exhibited phenotypic changes attributable to c-myc overexpression, such as cell cycle disturbances, increased cell size, and overexpression of the S6 ribosomal protein. Cells overexpressing c-myc by 70% exhibited additional phenotypic changes typical of c-myc overexpression, such as increased histone H3 phosphorylation, and reduced adherence. Sorted cells also exhibited overexpression of the IGF-1R, and slightly elevated expression of the IR. Increased susceptibility to the mitogenic effect of insulin was seen in a small proportion of the sorted cells, and insulin was more effective in activating the p44/42 MAPK pathway, but not the PI3K pathway, in the sorted cells than in the nonsorted cell population. To our knowledge, this is the first in vitro system allowing functional coupling between mitogenic signaling by a well-defined growth factor and gradual overexpression of the normal, endogenous c-myc gene. Thus, our flow-sorting approach provides an alternative modeling of the receptor-mediated carcinogenic process, compared to the currently used approaches of recombinant constitutive or conditional overexpression of oncogenic transmembrane receptor tyrosine kinases or oncogenic transcription factors.

  9. Tumor suppressor DYRK1A effects on proliferation and chemoresistance of AML cells by downregulating c-Myc.

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    Qiang Liu

    Full Text Available Acute myeloid leukemia (AML, caused by abnormal proliferation and accumulation of hematopoietic progenitor cells, is one of the most common malignancies in adults. We reported here DYRK1A expression level was reduced in the bone marrow of adult AML patients, comparing to normal controls. Overexpression of DYRK1A inhibited the proliferation of AML cell lines by increasing the proportion of cells undergoing G0/G1 phase. We reasoned that the proliferative inhibition was due to downregulation of c-Myc by DYRK1A, through mediating its degradation. Moreover, overexpression of c-Myc markedly reversed AML cell growth inhibition induced by DYRK1A. DYRK1A also had significantly lower expression in relapsed/refractory AML patients, comparing to newly-diagnosed AML patients, which indicated the role of DYRK1A in chemoresistance of AML. Our study provided functional evidences for DYRK1A as a potential tumor suppressor in AML.

  10. Focal Adhesion Kinase Is Required for Intestinal Regeneration and Tumorigenesis Downstream of Wnt/c-Myc Signaling

    Science.gov (United States)

    Ashton, Gabrielle H.; Morton, Jennifer P.; Myant, Kevin; Phesse, Toby J.; Ridgway, Rachel A.; Marsh, Victoria; Wilkins, Julie A.; Athineos, Dimitris; Muncan, Vanesa; Kemp, Richard; Neufeld, Kristi; Clevers, Hans; Brunton, Valerie; Winton, Douglas J.; Wang, Xiaoyan; Sears, Rosalie C.; Clarke, Alan R.; Frame, Margaret C.; Sansom, Owen J.

    2012-01-01

    SUMMARY The intestinal epithelium has a remarkable capacity to regenerate after injury and DNA damage. Here, we show that the integrin effector protein Focal Adhesion Kinase (FAK) is dispensable for normal intestinal homeostasis and DNA damage signaling, but is essential for intestinal regeneration following DNA damage. Given Wnt/c-Myc signaling is activated following intestinal regeneration, we investigated the functional importance of FAK following deletion of the Apc tumor suppressor protein within the intestinal epithelium. Following Apc loss, FAK expression increased in a c-Myc-dependent manner. Codeletion of Apc and Fak strongly reduced proliferation normally induced following Apc loss, and this was associated with reduced levels of phospho-Akt and suppression of intestinal tumorigenesis in Apc heterozygous mice. Thus, FAK is required downstream of Wnt Signaling, for Akt/mTOR activation, intestinal regeneration, and tumorigenesis. Importantly, this work suggests that FAK inhibitors may suppress tumorigenesis in patients at high risk of developing colorectal cancer. PMID:20708588

  11. Using a Novel Transgenic Mouse Model to Study c-Myc Oncogenic Pathway in Castration Resistance and Chemoresistance of Prostate Cancer

    Science.gov (United States)

    2015-10-01

    AWARD NUMBER: W81XWH-13-1-0162 TITLE: Using a Novel Transgenic Mouse Model to Study c-Myc Oncogenic Pathway in Castration Resistance and...TELEPHONE NUMBER (include area code) Standard Form 298 (Rev. 8-98) Prescribed by ANSI Std. Z39.18 Using a Novel Transgenic Mouse Model to Study c-Myc... Transgenic Mouse Model to Study c-Myc Oncogenic Pathway in Castration Resistance and Chemoresistance of Prostate Cancer Feng Yang, Ph.D. Department of

  12. Targeting C-myc G-Quadruplex: Dual Recognition by Aminosugar-Bisbenzimidazoles with Varying Linker Lengths

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    Nihar Ranjan

    2013-11-01

    Full Text Available G-quadruplexes are therapeutically important biological targets. In this report, we present biophysical studies of neomycin-Hoechst 33258 conjugates binding to a G-quadruplex derived from the C-myc promoter sequence. Our studies indicate that conjugation of neomycin to a G-quadruplex binder, Hoechst 33258, enhances its binding. The enhancement in G-quadruplex binding of these conjugates varies with the length and composition of the linkers joining the neomycin and Hoechst 33258 units.

  13. Acinar-to-ductal metaplasia accompanies c-myc-induced exocrine pancreatic cancer progression in transgenic rodents.

    Science.gov (United States)

    Grippo, Paul J; Sandgren, Eric P

    2012-09-01

    Several important characteristics of exocrine pancreatic tumor pathogenesis remain incompletely defined, including identification of the cell of origin. Most human pancreatic neoplasms are ductal adenocarcinomas. However, acinar cells have been proposed as the source of some ductal neoplasms through a process of acinar-to-ductal metaplasia. The oncogenic transcription factor c-myc is associated with human pancreatic neoplasms. Transgenic mice overexpressing c-myc under control of acinar cell-specific elastase (Ela) gene regulatory elements not only develop acinar cell carcinomas but also mixed neoplasms that display both acinar-like neoplastic cells and duct-like neoplastic cells. In this report, we demonstrate that, first, c-myc is sufficient to induce acinar hyperplasia, though neoplastic lesions develop focally. Second, cell proliferation remains elevated in the neoplastic duct cell compartment of mixed neoplasms. Third, the proliferation/apoptosis ratio in cells from all lesion types remains constant, suggesting that differential regulation of these processes is not a feature of cancer progression in this model. Fourth, before the development of mixed neoplasms, there is transcriptional activation of the duct cell-specific cytokeratin-19 gene promoter in multicellular foci of amylase-positive acinar neoplasms. This observation provides direct evidence for metaplasia as the mechanism underlying development of ductal neoplastic cells within the context of an acinar neoplasm and suggests that the stimulus for this transformation acts over a multicellular domain or field within a neoplasm. Finally, focal ductal elements develop in some acinar cell carcinomas in Ela-c-myc transgenic rats, indicating that myc-associated acinar-to-ductal metaplasia is not restricted to the mouse. Copyright © 2011 UICC.

  14. Activation of nuclear β-catenin/c-Myc axis promotes oxidative stress injury in streptozotocin-induced diabetic cardiomyopathy.

    Science.gov (United States)

    Liu, Peng; Su, Jianfang; Song, Xixi; Wang, Shixiao

    2017-12-02

    Myocardial oxidative stress injury plays a crucial role in the pathogenesis of diabetic cardiomyopathy (DCM). Wnt/β-catenin signaling has been reported to involve in various heart diseases. However, the underlying mechanism associated with β-catenin in DCM remains elusive. This study intended to explore the effect of β-catenin on oxidative damage of DCM by establishing streptozotocin (STZ)-induced diabetic mouse model and hydrogen peroxide (H 2 O 2 )-treated myocardial cell model. Cardiac oxidative stress in DCM was detected by measurements of lipid peroxidation and anti-oxidative enzyme activities as well as DHE staining. Nuclear β-catenin activity and oxidative damage degree were measured by western blotting, qPCR, MTT assay and TUNEL staining. Cardiac function and morphology were evaluated by echocardiography and histopathology. Under diabetic oxidative stress or H 2 O 2 stimulation, nuclear β-catenin accumulation upregulated downstream c-Myc and further facilitated DNA damage and p53-mediated apoptosis as well as cell viability reduction, followed by phenotypic changes of cardiac dysfunction, interstitial fibrosis deposition and myocardial atrophy. Conversely, through directly inhibiting nuclear β-catenin/c-Myc axis, not only did siRNA knockdown of β-catenin or c-Myc attenuate cell injury in H 2 O 2 -stimulated cardiomyocytes, but also diabetic cardiac-specific β-catenin-knockout mice displayed the same prevention of heart injury as insulin-treated diabetic mice. The present study demonstrated that activated nuclear β-catenin/c-Myc axis was responsible for oxidative cardiac impairment of DCM. Therefore, repressing functional nuclear β-catenin may provide a hopeful therapeutic strategy for DCM. Copyright © 2017 Elsevier Inc. All rights reserved.

  15. Conserved features of cancer cells define their sensitivity to HAMLET-induced death; c-Myc and glycolysis.

    Science.gov (United States)

    Storm, P; Aits, S; Puthia, M K; Urbano, A; Northen, T; Powers, S; Bowen, B; Chao, Y; Reindl, W; Lee, D Y; Sullivan, N L; Zhang, J; Trulsson, M; Yang, H; Watson, J D; Svanborg, C

    2011-12-01

    HAMLET is the first member of a new family of tumoricidal protein-lipid complexes that kill cancer cells broadly, while sparing healthy, differentiated cells. Many and diverse tumor cell types are sensitive to the lethal effect, suggesting that HAMLET identifies and activates conserved death pathways in cancer cells. Here, we investigated the molecular basis for the difference in sensitivity between cancer cells and healthy cells. Using a combination of small-hairpin RNA (shRNA) inhibition, proteomic and metabolomic technology, we identified the c-Myc oncogene as one essential determinant of HAMLET sensitivity. Increased c-Myc expression levels promoted sensitivity to HAMLET and shRNA knockdown of c-Myc suppressed the lethal response, suggesting that oncogenic transformation with c-Myc creates a HAMLET-sensitive phenotype. Furthermore, HAMLET sensitivity was modified by the glycolytic state of tumor cells. Glucose deprivation sensitized tumor cells to HAMLET-induced cell death and in the shRNA screen, hexokinase 1 (HK1), 6-phosphofructo-2-kinase/fructose-2,6-biphosphatase 1 and hypoxia-inducible factor 1α modified HAMLET sensitivity. HK1 was shown to bind HAMLET in a protein array containing ∼8000 targets, and HK activity decreased within 15 min of HAMLET treatment, before morphological signs of tumor cell death. In parallel, HAMLET triggered rapid metabolic paralysis in carcinoma cells. Tumor cells were also shown to contain large amounts of oleic acid and its derivatives already after 15 min. The results identify HAMLET as a novel anti-cancer agent that kills tumor cells by exploiting unifying features of cancer cells such as oncogene addiction or the Warburg effect.

  16. Conserved features of cancer cells define their sensitivity of HAMLET-induced death; c-Myc and glycolysis

    Science.gov (United States)

    Storm, Petter; Puthia, Manoj Kumar; Aits, Sonja; Urbano, Alexander; Northen, Trent; Powers, Scott; Bowen, Ben; Chao, Yinxia; Reindl, Wolfgang; Lee, Do Yup; Sullivan, Nancy Liu; Zhang, Jianping; Trulsson, Maria; Yang, Henry; Watson, James; Svanborg, Catharina

    2014-01-01

    HAMLET is the first member of a new family of tumoricidal protein-lipid complexes that kill cancer cells broadly, while sparing healthy, differentiated cells. Many and diverse tumor cell types are sensitive to the lethal effect, suggesting that HAMLET identifies and activates conserved death pathways in cancer cells. Here we investigated the molecular basis for the difference in sensitivity between cancer cells and healthy cells. Using a combination of small hairpin RNA inhibition, proteomic and metabolomic technology we identified the c-Myc oncogene as one essential determinant of HAMLET sensitivity. Increased c-Myc expression levels promoted the sensitivity to HAMLET and shRNA knockdown of c-Myc suppressed the lethal response, suggesting that oncogenic transformation with c-Myc creates a HAMLET-sensitive phenotype. Furthermore, the HAMLET sensitivity was modified by the glycolytic state of the tumor cells. Glucose deprivation sensitized tumor cells to HAMLET-induced cell death and in the shRNA screen Hexokinase 1, PFKFB1 and HIF1α modified HAMLET sensitivity. Hexokinase 1 was shown to bind HAMLET in a protein array containing approximately 8000 targets and Hexokinase activity decreased within 15 minutes of HAMLET treatment, prior to morphological signs of tumor cell death. In parallel, HAMLET triggered rapid metabolic paralysis in carcinoma cells. The glycolytic machinery was modified and glycolysis was shifted towards the pentose phosphate pathway. Tumor cells were also shown to contain large amounts of oleic acid and its derivatives already after 15 minutes. The results identify HAMLET as a novel anti-cancer agent that kills tumor cells by exploiting unifying features of cancer cells such as oncogene-addiction or the Warburg effect. PMID:21643007

  17. Shikonin regulates C-MYC and GLUT1 expression through the MST1-YAP1-TEAD1 axis

    Czech Academy of Sciences Publication Activity Database

    Vališ, Karel; Talacko, Pavel; Grobárová, Valeria; Černý, J.; Novák, Petr

    2016-01-01

    Roč. 349, č. 2 (2016), s. 273-281 ISSN 0014-4827 R&D Projects: GA ČR(CZ) GP14-21095P; GA ČR GA13-16565S; GA MŠk(CZ) LQ1604; GA MŠk(CZ) ED1.1.00/02.0109 Institutional support: RVO:61388971 Keywords : Hippo * Glycolysis * C-MYC Subject RIV: EE - Microbiology, Virology Impact factor: 3.546, year: 2016

  18. MYCT1-TV, A Novel MYCT1 Transcript, Is Regulated by c-Myc and May Participate in Laryngeal Carcinogenesis

    Science.gov (United States)

    Fu, Shuang; Guo, Yan; Chen, Hong; Xu, Zhen-Ming; Qiu, Guang-Bin; Zhong, Ming; Sun, Kai-Lai; Fu, Wei-Neng

    2011-01-01

    Background MYCT1, a putative target of c-Myc, is a novel candidate tumor suppressor gene cloned from laryngeal squamous cell carcinoma (LSCC). Its transcriptional regulation and biological effects on LSCC have not been clarified. Methodology/Principal Findings Using RACE assay, we cloned a 1106 bp transcript named Myc target 1 transcript variant 1 (MYCT1-TV) and confirmed its transcriptional start site was located at 140 bp upstream of the ATG start codon of MYCT1-TV. Luciferase, electrophoretic mobility shift and chromatin immunoprecipitation assays confirmed c-Myc could regulate the promoter activity of MYCT1-TV by specifically binding to the E-box elements within −886 to −655 bp region. These results were further verified by site-directed mutagenesis and RNA interference (RNAi) assays. MYCT1-TV and MYCT1 expressed lower in LSCC than those in paired adjacent normal laryngeal tissues, and overexpression of MYCT1-TV and MYCT1 could inhibit cell proliferation and invasion and promote apoptosis in LSCC cells. Conclusions/Significance Our data indicate that MYCT1-TV, a novel MYCT1 transcript, is regulated by c-Myc and down-regulation of MYCT1-TV/MYCT1 could contribute to LSCC development and function. PMID:21998677

  19. Repression of PLA2R1 by c-MYC and HIF-2alpha promotes cancer growth.

    Science.gov (United States)

    Vindrieux, David; Devailly, Guillaume; Augert, Arnaud; Le Calvé, Benjamin; Ferrand, Mylène; Pigny, Pascal; Payen, Léa; Lambeau, Gérard; Perrais, Michael; Aubert, Sébastien; Simonnet, Hélène; Dante, Robert; Bernard, David

    2014-02-28

    Loss of secreted phospholipase A2 receptor (PLA2R1) has recently been found to render human primary cells more resistant to senescence whereas increased PLA2R1 expression is able to induce cell cycle arrest, cancer cell death or blockage of cancer cell transformation in vitro, suggesting that PLA2R1 displays tumor suppressive activities. Here we report that PLA2R1 expression strongly decreases in samples of human renal cell carcinoma (RCC). Knockdown of PLA2R1 increases renal cancer cell tumorigenicity supporting a role of PLA2R1 loss to promote in vivo RCC growth. Most RCC result from Von Hippel-Lindau (VHL) tumor suppressor loss-of-function and subsequent gain-of-function of the oncogenic HIF-2alpha/c-MYC pathway. Here, by genetically manipulating VHL, HIF-2alpha and c-MYC, we demonstrate that loss of VHL, stabilization of HIF-2alpha and subsequent increased c-MYC activity, binding and transcriptional repression, through induction of PLA2R1 DNA methylation closed to PLA2R1 transcriptional start site, results in decreased PLA2R1 transcription. Our results describe for the first time an oncogenic pathway leading to PLA2R1 transcriptional repression and the importance of this repression for tumor growth.

  20. Immunodetection of rasP21 and c-myc oncogenes in oral mucosal swab preparation from clove cigarette smokers

    Directory of Open Access Journals (Sweden)

    Silvi Kintawati

    2008-12-01

    Full Text Available Background: Smoking is the biggest factor for oral cavity malignancy. Some carcinogens found in cigar will stimulate epithel cell in oral cavity and cause mechanism disturbance on tissue resistance and produce abnormal genes (oncogenes. Oncogenes ras and myc are found on malignant tumor in oral cavity which are associated with smoking. Purpose: This research is to find the expression of oncogenes rasP21 and c-myc in oral mucosa epithelial of smoker with immunocytochemistry reaction. Methods: An oral mucosal swab was performed to 30 smokers categorized as light, moderate, and chain, and 10 non smokers which was followed by immunocytochemistry reaction using antibody towards oncogene rasP21 and c-myc is reacted to identify the influence of smoking towards malignant tumor in oral cavity. The result is statistically analyzed using Kruskal-Wallis test. Result: Based on the observation result of oncogene rasP21reaction, it shows that there is significant difference between non smoker group and light smoker, compared to moderate and chain smoker group (p < 0.01. On the other side, the observation result of oncogene c-myc indicates that there is no significant difference between the group of non smokers and the group of light, moderate, and chain smokers (p > 0.05. Conclusion: The higher the possibility of oral cavity malignancy and that the antibody for rasP21 oncogene can be used as a marker for early detection of oral cavity malignancy caused by smoking.

  1. Reevesioside A, a cardenolide glycoside, induces anticancer activity against human hormone-refractory prostate cancers through suppression of c-myc expression and induction of G1 arrest of the cell cycle.

    Directory of Open Access Journals (Sweden)

    Wohn-Jenn Leu

    Full Text Available In the past decade, there has been a profound increase in the number of studies revealing that cardenolide glycosides display inhibitory activity on the growth of human cancer cells. The use of potential cardenolide glycosides may be a worthwhile approach in anticancer research. Reevesioside A, a cardenolide glycoside isolated from the root of Reevesia formosana, displayed potent anti-proliferative activity against human hormone-refractory prostate cancers. A good correlation (r² = 0.98 between the expression of Na⁺/K⁺-ATPase α₃ subunit and anti-proliferative activity suggested the critical role of the α₃ subunit. Reevesioside A induced G1 arrest of the cell cycle and subsequent apoptosis in a thymidine block-mediated synchronization model. The data were supported by the down-regulation of several related cell cycle regulators, including cyclin D1, cyclin E and CDC25A. Reevesioside A also caused a profound decrease of RB phosphorylation, leading to an increased association between RB and E2F1 and the subsequent suppression of E2F1 activity. The protein and mRNA levels of c-myc, which can activate expression of many downstream cell cycle regulators, were dramatically inhibited by reevesioside A. Transient transfection of c-myc inhibited the down-regulation of both cyclin D1 and cyclin E protein expression to reevesioside A action, suggesting that c-myc functioned as an upstream regulator. Flow cytometric analysis of JC-1 staining demonstrated that reevesioside A also induced the significant loss of mitochondrial membrane potential. In summary, the data suggest that reevesioside A inhibits c-myc expression and down-regulates the expression of CDC25A, cyclin D1 and cyclin E, leading to a profound decrease of RB phosphorylation. G1 arrest is, therefore, induced through E2F1 suppression. Consequently, reevesioside A causes mitochondrial damage and an ultimate apoptosis in human hormone-refractory prostate cancer cells.

  2. C-Myc regulates substrate oxidation patterns during early pressure-overload hypertrophy

    Energy Technology Data Exchange (ETDEWEB)

    Ledee, Dolena R. [Seattle Children' s Research Inst., Seattle, WA (United States); Smith, Lincoln [Seattle Children' s Hospital, Seattle, WA (United States); Kajimoto, Masaki [Seattle Children' s Research Inst., Seattle, WA (United States); Bruce, Margaret [Seattle Children' s Research Inst., Seattle, WA (United States); Isern, Nancy G. [Pacific Northwest National Lab. (PNNL), Richland, WA (United States). Environmental Molecular Sciences Lab. (EMSL); Xu, Chun [Seattle Children' s Research Inst., Seattle, WA (United States); Portman, Michael A. [Seattle Children' s Research Inst., Seattle, WA (United States); Olson, Aaron [Seattle Children' s Research Inst., Seattle, WA (United States)

    2013-11-26

    Pressure overload cardiac hypertrophy alters substrate metabolism. Prior work showed that myocardial inactivation of c-Myc (Myc) attenuated hypertrophy and decreased expression of glycolytic genes after aortic constriction. Accordingly, we hypothesize that Myc regulates substrate preferences for the citric acid cycle during pressure overload hypertrophy from transverse aortic constriction (TAC) and that these metabolic changes impact cardiac function and growth. To test this hypothesis, we subjected FVB mice with cardiac specific, inducible Myc inactivation (MycKO-TAC) and non-transgenic littermates (Cont-TAC) to transverse aortic constriction (n=7/group). A separate group underwent sham surgery (Sham, n=5). After two weeks, function was measured in isolated working hearts along with substrate fractional contributions to the citric acid cycle by using perfusate with 13C labeled mixed fatty acids, lactate, ketones and unlabeled glucose and insulin. Western blots were used to evaluate metabolic enzymes. Cardiac function was similar between groups after TAC although +dP/dT and -dP/dT trended towards improvement in MycKO-TAC versus Cont-TAC. Compared to Sham, Cont-TAC had increased free fatty acid fractional contribution with a concurrent decrease in unlabeled (presumably glucose) contribution. Myc inactivation (MycKO-TAC) inhibited these metabolic changes. Hypertrophy in general increased protein levels of PKM2; however this change was not linked to Myc status. Protein post-translation modification by O-GlcNAc was significantly greater in Cont-TAC versus both Sham and MycKO-TAC. In conclusion, Myc regulates substrate utilization during early pressure overload hypertrophy. Our results show that the metabolic switch during hypertrophy is not necessary to maintain cardiac function, but it may be important mechanism to promote cardiomyocyte growth. Myc also regulates protein O-GlcNAcylation during hypertrophy.

  3. c-Myc and AMPK Control Cellular Energy Levels by Cooperatively Regulating Mitochondrial Structure and Function.

    Directory of Open Access Journals (Sweden)

    Lia R Edmunds

    Full Text Available The c-Myc (Myc oncoprotein and AMP-activated protein kinase (AMPK regulate glycolysis and oxidative phosphorylation (Oxphos although often for different purposes. Because Myc over-expression depletes ATP with the resultant activation of AMPK, we explored the potential co-dependency of and cross-talk between these proteins by comparing the consequences of acute Myc induction in ampk+/+ (WT and ampk-/- (KO murine embryo fibroblasts (MEFs. KO MEFs showed a higher basal rate of glycolysis than WT MEFs and an appropriate increase in response to activation of a Myc-estrogen receptor (MycER fusion protein. However, KO MEFs had a diminished ability to increase Oxphos, mitochondrial mass and reactive oxygen species in response to MycER activation. Other differences between WT and KO MEFs, either in the basal state or following MycER induction, included abnormalities in electron transport chain function, levels of TCA cycle-related oxidoreductases and cytoplasmic and mitochondrial redox states. Transcriptional profiling of pathways pertinent to glycolysis, Oxphos and mitochondrial structure and function also uncovered significant differences between WT and KO MEFs and their response to MycER activation. Finally, an unbiased mass-spectrometry (MS-based survey capable of quantifying ~40% of all mitochondrial proteins, showed about 15% of them to be AMPK- and/or Myc-dependent in their steady state. Significant differences in the activities of the rate-limiting enzymes pyruvate kinase and pyruvate dehydrogenase, which dictate pyruvate and acetyl coenzyme A abundance, were also differentially responsive to Myc and AMPK and could account for some of the differences in basal metabolite levels that were also detected by MS. Thus, Myc and AMPK are highly co-dependent and appear to engage in significant cross-talk across numerous pathways which support metabolic and ATP-generating functions.

  4. Inducement of G-quadruplex DNA forming and down-regulation of oncogene c-myc by bile acid-amino acid conjugate-BAA.

    Science.gov (United States)

    Tian, Mingyue; Zhang, Xiufeng; Li, Yan; Ju, Yong; Xiang, Junfeng; Zhao, Changqi; Tang, Yalin

    2010-03-01

    Human c-myc gene is a central regulator of cellular proliferation and cell growth, and G-quadruplexes have been proven to be the transcriptional controller of this gene. In this study, the interaction of bile acid-amino acid conjugate (BAA) with G-quadruplexes in c-myc was investigated by circular dichroism spectroscopy, nuclear magnetic resonance (NMR) measurement, and quantitative real-time polymerase chain reaction (PCR) assay. The experimental results indicated that BAA has the ability to selectively induce the formation of parallel G-quadruplexes in c-myc, which leads to down-regulation of c-myc transcription in the human breast cancer cell MCF-7.

  5. 9-N-Substituted berberine derivatives: stabilization of G-quadruplex DNA and down-regulation of oncogene c-myc.

    Science.gov (United States)

    Ma, Yan; Ou, Tian-Miao; Hou, Jin-Qiang; Lu, Yu-Jing; Tan, Jia-Heng; Gu, Lian-Quan; Huang, Zhi-Shu

    2008-08-15

    A series of 9-N-substituted berberine derivatives (2a-j) were synthesized and evaluated as a new class of G-quadruplex binding ligands. G-quadruplex of DNA had been proven to be the transcription controller of human c-myc gene. The interaction of 9-N-substituted berberine derivatives with G-quadruplex DNA in c-myc was examined via EMSA, CD spectroscopy, FRET-melting method, PCR-stop assay, competitive dialysis, cell proliferation assay, and RT-PCR assay. The experiment results indicated that these derivatives could selectively induce and stabilize the formation of intramolecular parallel G-quadruplex in c-myc, which led to down-regulation of transcription of the c-myc in the HL60 lymphomas cell line. The related structure-activity relationships were also discussed.

  6. A re-emerging marker for prognosis in hepatocellular carcinoma: the add-value of fishing c-myc gene for early relapse.

    Science.gov (United States)

    Pedica, Federica; Ruzzenente, Andrea; Bagante, Fabio; Capelli, Paola; Cataldo, Ivana; Pedron, Serena; Iacono, Calogero; Chilosi, Marco; Scarpa, Aldo; Brunelli, Matteo; Tomezzoli, Anna; Martignoni, Guido; Guglielmi, Alfredo

    2013-01-01

    Hepatocellular carcinoma is one leading cause of cancer-related death and surgical resection is still one of the major curative therapies. Recently, there has been a major effort to find mechanisms involved in carcinogenesis and early relapse. c-myc gene abnormality is found in hepatocarcinogenesis. Our aim was to analyze the role of c-myc as prognostic factor in terms of overall survival and disease-free survival and to investigate if c-myc may be an important target for therapy. We studied sixty-five hepatocellular carcinomas submitted to surgical resection with curative intent. Size, macro-microvascular invasion, necrosis, number of nodules, grading and serum alfa-fetoprotein level were registered for all cases. We evaluated the c-myc aberrations by using break-apart FISH probes. Probes specific for the centromeric part of chromosome 8 and for the locus specific c-myc gene (8q24) were used to assess disomy, gains of chromosomes (polysomy due to polyploidy) and amplification. c-myc gene amplification was scored as 8q24/CEP8 > 2. Statistical analysis for disease-free survival and overall survival were performed. At molecular level, c-myc was amplified in 19% of hepatocellular carcinoma, whereas showed gains in 55% and set wild in 26% of cases. The 1- and 3-year disease-free survival and overall survival for disomic, polysomic and amplified groups were significantly different (p=0.020 and p=.018 respectively). Multivariate analysis verified that the AFP and c-myc status (amplified vs. not amplified) were significant prognostic factors for overall patients survival. c-myc gene amplification is significantly correlated with disease-free survival and overall survival in patients with hepatocellular carcinoma after surgical resection and this model identifies patients with risk of early relapse (≤12 months). We suggest that c-myc assessment may be introduced in the clinical practice for improving prognostication (high and low risk of relapse) routinely and may have

  7. A re-emerging marker for prognosis in hepatocellular carcinoma: the add-value of fishing c-myc gene for early relapse.

    Directory of Open Access Journals (Sweden)

    Federica Pedica

    Full Text Available Hepatocellular carcinoma is one leading cause of cancer-related death and surgical resection is still one of the major curative therapies. Recently, there has been a major effort to find mechanisms involved in carcinogenesis and early relapse. c-myc gene abnormality is found in hepatocarcinogenesis. Our aim was to analyze the role of c-myc as prognostic factor in terms of overall survival and disease-free survival and to investigate if c-myc may be an important target for therapy. We studied sixty-five hepatocellular carcinomas submitted to surgical resection with curative intent. Size, macro-microvascular invasion, necrosis, number of nodules, grading and serum alfa-fetoprotein level were registered for all cases. We evaluated the c-myc aberrations by using break-apart FISH probes. Probes specific for the centromeric part of chromosome 8 and for the locus specific c-myc gene (8q24 were used to assess disomy, gains of chromosomes (polysomy due to polyploidy and amplification. c-myc gene amplification was scored as 8q24/CEP8 > 2. Statistical analysis for disease-free survival and overall survival were performed. At molecular level, c-myc was amplified in 19% of hepatocellular carcinoma, whereas showed gains in 55% and set wild in 26% of cases. The 1- and 3-year disease-free survival and overall survival for disomic, polysomic and amplified groups were significantly different (p=0.020 and p=.018 respectively. Multivariate analysis verified that the AFP and c-myc status (amplified vs. not amplified were significant prognostic factors for overall patients survival. c-myc gene amplification is significantly correlated with disease-free survival and overall survival in patients with hepatocellular carcinoma after surgical resection and this model identifies patients with risk of early relapse (≤12 months. We suggest that c-myc assessment may be introduced in the clinical practice for improving prognostication (high and low risk of relapse routinely

  8. C-Myc Induced Compensated Cardiac Hypertrophy Increases Free Fatty Acid Utilization for the Citric Acid Cycle

    Energy Technology Data Exchange (ETDEWEB)

    Olson, Aaron; Ledee, Dolena; Iwamoto, Kate; Kajimoto, Masaki; O' Kelly-Priddy, Colleen M.; Isern, Nancy G.; Portman, Michael A.

    2013-02-01

    The protooncogene C-Myc (Myc) regulates cardiac hypertrophy. Myc promotes compensated cardiac function, suggesting that the operative mechanisms differ from those leading to heart failure. Myc regulation of substrate metabolism is a reasonable target, as Myc alters metabolism in other tissues. We hypothesize that Myc-induced shifts in substrate utilization signal and promote compensated hypertrophy. We used cardiac specific Myc-inducible C57/BL6 male mice between 4-6 months old that develop hypertrophy with tamoxifen (tam). Isolated working hearts and 13Carbon (13C )-NMR were used to measure function and fractional contributions (Fc) to the citric acid cycle by using perfusate containing 13C-labeled free fatty acids, acetoacetate, lactate, unlabeled glucose and insulin. Studies were performed at pre-hypertrophy (3-days tam, 3dMyc), established hypertrophy (7-days tam, 7dMyc) or vehicle control (cont). Non-transgenic siblings (NTG) received 7-days tam or vehicle to assess drug effect. Hypertrophy was confirmed by echocardiograms and heart weights. Western blots were performed on key metabolic enzymes. Hypertrophy occurred in 7dMyc only. Cardiac function did not differ between groups. Tam alone did not affect substrate contribution in NTG. Substrate utilization was not significantly altered in 3dMyc versus cont. The free fatty acid FC was significantly greater in 7dMyc vs cont with decreased unlabeled Fc, which is predominately exogenous glucose. Free fatty acid flux to the citric acid cycle increased while lactate flux was diminished in 7dMyc compared to cont. Total protein levels of a panel of key metabolic enzymes were unchanged; however total protein O-GlcNAcylation was increased in 7dMyc. Substrate utilization changes did not precede hypertrophy; therefore they are not the primary signal for cardiac growth in this model. Free fatty acid utilization and oxidation increase at established hypertrophy. Understanding the mechanisms whereby this change maintained

  9. Reversible lysine-specific demethylase 1 antagonist HCI-2509 inhibits growth and decreases c-MYC in castration- and docetaxel-resistant prostate cancer cells.

    Science.gov (United States)

    Gupta, S; Weston, A; Bearrs, J; Thode, T; Neiss, A; Soldi, R; Sharma, S

    2016-12-01

    Lysine-specific demethylase 1 (LSD1 or KDM1A) overexpression correlates with poor survival and castration resistance in prostate cancer. LSD1 is a coregulator of ligand-independent androgen receptor signaling promoting c-MYC expression. We examined the antitumor efficacy of LSD1 inhibition with HCI-2509 in advanced stages of prostate cancer. Cell survival, colony formation, histone methylation, c-MYC level, c-MYC expression, cell cycle changes and in vivo efficacy were studied in castration-resistant prostate cancer cells upon treatment with HCI-2509. In vitro combination studies, using HCI-2509 and docetaxel, were performed to assess the synergy. Cell survival, colony formation, histone methylation and c-myc levels were studied in docetaxel-resistant prostate cancer cells treated with HCI-2509. HCI-2509 is cytotoxic and inhibits colony formation in castration-resistant prostate cancer cells. HCI-2509 treatment causes a dose-dependent increase in H3K9me2 (histone H3lysine 9) levels, a decrease in c-MYC protein, inhibition of c-MYC expression and accumulation in the G0/G1 phase of the cell cycle in these cells. PC3 xenografts in mice have a significant reduction in tumor burden upon treatment with HCI-2509 with no associated myelotoxicity or weight loss. More synergy is noted at sub-IC50 (half-maximal inhibitory concentration) doses of docetaxel and HCI-2509 in PC3 cells than in DU145 cells. HCI-2509 has growth-inhibitory efficacy and decreases the c-myc level in docetaxel-resistant prostate cancer cells. LSD1 inhibition with HCI-2509 decreases the c-MYC level in poorly differentiated prostate cancer cell lines and has a therapeutic potential in castration- and docetaxel-resistant prostate cancer.

  10. Decoding c-Myc networks of cell cycle and apoptosis regulated genes in a transgenic mouse model of papillary lung adenocarcinomas

    Science.gov (United States)

    Ciribilli, Yari; Singh, Prashant; Spanel, Reinhard; Inga, Alberto; Borlak, Jürgen

    2015-01-01

    The c-Myc gene codes for a basic-helix-loop-helix-leucine zipper transcription factor protein and is reported to be frequently over-expressed in human cancers. Given that c-Myc plays an essential role in neoplastic transformation we wished to define its activity in lung cancer and therefore studied its targeted expression to respiratory epithelium in a transgenic mouse disease model. Using histological well-defined tumors, transcriptome analysis identified novel c-Myc responsive cell cycle and apoptosis genes that were validated as direct c-Myc targets using EMSA, Western blotting, gene reporter and ChIP assays. Through computational analyses c-Myc cooperating transcription factors emerged for repressed and up-regulated genes in cancer samples, namely Klf7, Gata3, Sox18, p53 and Elf5 and Cebpα, respectively. Conversely, at promoters of genes regulated in transgenic but non-carcinomatous lung tissue enriched binding sites for c-Myc, Hbp1, Hif1 were observed. Bioinformatic analysis of tumor transcriptomic data revealed regulatory gene networks and highlighted mortalin and moesin as master regulators while gene reporter and ChIP assays in the H1299 lung cancer cell line as well as cross-examination of published ChIP-sequence data of 7 human and 2 mouse cell lines provided strong evidence for the identified genes to be c-Myc targets. The clinical significance of findings was established by evaluating expression of orthologous proteins in human lung cancer. Taken collectively, a molecular circuit for c-Myc-dependent cellular transformation was identified and the network analysis broadened the perspective for molecularly targeted therapies. PMID:26427040

  11. Cytoplasmic calcium increase via fusion with inactivated Sendai virus induces apoptosis in human multiple myeloma cells by downregulation of c-Myc oncogene.

    Science.gov (United States)

    Jiang, Yingzhe; Saga, Kotaro; Miyamoto, Yasuhide; Kaneda, Yasufumi

    2016-06-14

    Because the emergence of drug resistance is a major limitation of current treatments for multiple myeloma (MM), it is necessary to continuously develop novel anticancer strategies. Here, using an inactivated Sendai virus (Hemagglutinating Virus of Japan; HVJ) envelope (HVJ-E), we discovered that increase of cytoplasmic Ca2+ by virus-cell fusion significantly induced apoptosis against human MM cells but not peripheral blood mononuclear cells from healthy donors. Interaction of F protein of HVJ-E with MM cells increased intracellular Ca2+ level of MMs by the induction of Ca2+ efflux from endoplasmic reticulum but not influx from extracellular region. The elevation of the Ca2+ cytoplasmic level induced SMAD1/5/8 phosphorylation and translocation into the nucleus, and SMAD1/5/8 and SMAD4 complex suppressed c-Myc transcription. Meanwhile, HVJ-E decreases S62 phosphorylation of c-Myc and promotes c-Myc protein degradation. Thus, HVJ-E-induced cell death of MM resulted from suppression of c-Myc by both destabilization of c-Myc protein and downregulation of c-Myc transcription. This study indicates that HVJ-E will be a promising tool for MM therapy.

  12. Enforced expression of the c-myc oncogene inhibits cell differentiation by precluding entry into a distinct predifferentiation state in G/sub 0//G/sub 1/

    Energy Technology Data Exchange (ETDEWEB)

    Freytag, S.O.

    1988-04-01

    A broad base of data has implicated a role for the c-myc proto-oncogene in the control of the cell cycle and cell differentiation. To further define the role of myc in these processes, the authors examined the effect of enforced myc expression on several events that are thought to be important steps leading to the terminally differentiated state: (i) the ability to arrest growth in G/sub 0//G/sub 1/, (ii) the ability to replicate the genome upon initiation of the differentiation program, and (iii) the ability to loose responsiveness to mitogens and withdraw from the cell cycle. 3T3-L1 preadipocyte cell lines expressing various levels of myc mRNA were established by transfection with a recombinant myc gene under the transcriptional control of the Rous sarcoma virus (RSV) promoter. Cells that expressed high constitutive levels of pRSV myc mRNA arrested in G/sub 0//G/sub 1/ at densities similar to those of normal cells at confluence. Upon initiation of the differentiation program, such cells traversed the cell cycle with kinetics similar to those of normal cells and subsequently arrested in G/sub 0//G/sub 1/. Thus, enforced expression of myc had no effect on the ability of cells to arrest growth in G/sub 0//G/sub 1/ or to replicate the genome upon initiation of the differentiation program. Cells were then tested for their ability to reenter the cell cycle upon exposure to high concentrations of serum and for their capacity to differentiate. In contrast to normal cells, cells expressing high constitutive levels of myc RNA reentered the cell cycle when challenged with 30% serum and failed to terminally differentiate.

  13. Guttiferone K impedes cell cycle re-entry of quiescent prostate cancer cells via stabilization of FBXW7 and subsequent c-MYC degradation.

    Science.gov (United States)

    Xi, Z; Yao, M; Li, Y; Xie, C; Holst, J; Liu, T; Cai, S; Lao, Y; Tan, H; Xu, H-X; Dong, Q

    2016-06-02

    Cell cycle re-entry by quiescent cancer cells is an important mechanism for cancer progression. While high levels of c-MYC expression are sufficient for cell cycle re-entry, the modality to block c-MYC expression, and subsequent cell cycle re-entry, is limited. Using reversible quiescence rendered by serum withdrawal or contact inhibition in PTEN(null)/p53(WT) (LNCaP) or PTEN(null)/p53(mut) (PC-3) prostate cancer cells, we have identified a compound that is able to impede cell cycle re-entry through c-MYC. Guttiferone K (GUTK) blocked resumption of DNA synthesis and preserved the cell cycle phase characteristics of quiescent cells after release from the quiescence. In vehicle-treated cells, there was a rapid increase in c-MYC protein levels upon release from the quiescence. However, this increase was inhibited in the presence of GUTK with an associated acceleration in c-MYC protein degradation. The inhibitory effect of GUTK on cell cycle re-entry was significantly reduced in cells overexpressing c-MYC. The protein level of FBXW7, a subunit of E3 ubiquitin ligase responsible for degradation of c-MYC, was reduced upon the release from the quiescence. In contrast, GUTK stabilized FBXW7 protein levels during release from the quiescence. The critical role of FBXW7 was confirmed using siRNA knockdown, which impaired the inhibitory effect of GUTK on c-MYC protein levels and cell cycle re-entry. Administration of GUTK, either in vitro prior to transplantation or in vivo, suppressed the growth of quiescent prostate cancer cell xenografts. Furthermore, elevation of FBXW7 protein levels and reduction of c-MYC protein levels were found in the xenografts of GUTK-treated compared with vehicle-treated mice. Hence, we have identified a compound that is capable of impeding cell cycle re-entry by quiescent PTEN(null)/p53(WT) and PTEN(null)/p53(mut) prostate cancer cells likely by promoting c-MYC protein degradation through stabilization of FBXW7. Its usage as a clinical modality to

  14. SAP155-mediated splicing of FUSE-binding protein-interacting repressor serves as a molecular switch for c-myc gene expression.

    Science.gov (United States)

    Matsushita, Kazuyuki; Kajiwara, Toshiko; Tamura, Mai; Satoh, Mamoru; Tanaka, Nobuko; Tomonaga, Takeshi; Matsubara, Hisahiro; Shimada, Hideaki; Yoshimoto, Rei; Ito, Akihiro; Kubo, Shuji; Natsume, Tohru; Levens, David; Yoshida, Minoru; Nomura, Fumio

    2012-06-01

    The Far UpStream Element (FUSE)-binding protein-interacting repressor (FIR), a c-myc transcriptional suppressor, is alternatively spliced removing the transcriptional repression domain within exon 2 (FIRΔexon2) in colorectal cancers. SAP155 is a subunit of the essential splicing factor 3b (SF3b) subcomplex in the spliceosome. This study aims to study the significance of the FIR-SAP155 interaction for the coordination of c-myc transcription, pre-mRNA splicing, and c-Myc protein modification, as well as to interrogate FIRΔexon2 for other functions relating to altered FIR pre-mRNA splicing. Knockdown of SAP155 or FIR was used to investigate their reciprocal influence on each other and on c-myc transcription, pre-mRNA splicing, and protein expression. Pull down from HeLa cell nuclear extracts revealed the association of FIR, FIRΔexon2, and SF3b subunits. FIR and FIRΔexon2 were coimmunoprecipitated with SAP155. FIR and FIRΔexon2 adenovirus vector (Ad-FIR and Ad-FIRΔexon2, respectively) were prepared to test for their influence on c-myc expression. FIR, SAP155, SAP130, and c-myc were coordinately upregulated in human colorectal cancer. These results reveal that SAP155 and FIR/FIRΔexon2 form a complex and are mutually upregulating. Ad-FIRΔexon2 antagonized Ad-FIR transcriptional repression of c-myc in HeLa cells. Because FIRΔexon2 still carries RRM1 and RRM2 and binding activity to FUSE, it is able to displace repression competent FIR from FUSE in electrophoretic mobility shift assays, thus thwarting FIR-mediated transcriptional repression by FUSE. Thus aberrant FIRΔexon2 production in turn sustained c-Myc expression. In conclusion, altered FIR and c-myc pre-mRNA splicing, in addition to c-Myc expression by augmented FIR/FIRΔexon2-SAP155 complex, potentially contribute to colorectal cancer development. 2012 AACR

  15. C-MYC overexpression is required for continuous suppression of oncogene-induced senescence in melanoma cells.

    Science.gov (United States)

    Zhuang, D; Mannava, S; Grachtchouk, V; Tang, W-H; Patil, S; Wawrzyniak, J A; Berman, A E; Giordano, T J; Prochownik, E V; Soengas, M S; Nikiforov, M A

    2008-11-06

    Malignant melanomas often harbor activating mutations in BRAF (V600E) or, less frequently, in NRAS (Q61R). Intriguingly, the same mutations have been detected at higher incidences in benign nevi, which are largely composed of senescent melanocytes. Overexpression of BRAF(V600E) or NRAS(Q61R) in human melanocytes in vitro has been shown to induce senescence, although via different mechanisms. How oncogene-induced senescence is overcome during melanoma progression remains unclear. Here, we report that in the majority of analysed BRAF(V600E)- or NRAS(Q61R)-expressing melanoma cells, C-MYC depletion induced different yet overlapping sets of senescence phenotypes that are characteristic of normal melanocytes undergoing senescence due to overexpression of BRAF(V600E) or NRAS(Q61R), respectively. These senescence phenotypes were p16(INK4A)- or p53-independent, however, several of them were suppressed by genetic or pharmacological inhibition of BRAF(V600E) or phosphoinositide 3-kinase pathways, including rapamycin-mediated inhibition of mTOR-raptor in NRAS(Q61R)-expressing melanoma cells. Reciprocally, overexpression of C-MYC in normal melanocytes suppressed BRAF(V600E)-induced senescence more efficiently than NRAS(Q61R)-induced senescence, which agrees with the generally higher rates of activating mutations in BRAF than NRAS gene in human cutaneous melanomas. Our data suggest that one of the major functions of C-MYC overexpression in melanoma progression is to continuous suppress BRAF(V600E)- or NRAS(Q61R)-dependent senescence programs.

  16. Gene expression profiles in primary pancreatic tumors and metastatic lesions of Ela-c-myc transgenic mice

    Directory of Open Access Journals (Sweden)

    Liao Dezhong J

    2008-01-01

    Full Text Available Abstract Background Pancreatic carcinoma usually is a fatal disease with no cure, mainly due to its invasion and metastasis prior to diagnosis. We analyzed the gene expression profiles of paired primary pancreatic tumors and metastatic lesions from Ela-c-myc transgenic mice in order to identify genes that may be involved in the pancreatic cancer progression. Differentially expressed selected genes were verified by semi-quantitative and quantitative RT-PCR. To further evaluate the relevance of some of the selected differentially expressed genes, we investigated their expression pattern in human pancreatic cancer cell lines with high and low metastatic potentials. Results Data indicate that genes involved in posttranscriptional regulation were a major functional category of upregulated genes in both primary pancreatic tumors (PT and liver metastatic lesions (LM compared to normal pancreas (NP. In particular, differential expression for splicing factors, RNA binding/pre-mRNA processing factors and spliceosome related genes were observed, indicating that RNA processing and editing related events may play critical roles in pancreatic tumor development and progression. High expression of insulin growth factor binding protein-1 (Igfbp1 and Serine proteinase inhibitor A1 (Serpina1, and low levels or absence of Wt1 gene expression were exclusive to liver metastatic lesion samples. Conclusion We identified Igfbp1, Serpina1 and Wt1 genes that are likely to be clinically useful biomarkers for prognostic or therapeutic purposes in metastatic pancreatic cancer, particularly in pancreatic cancer where c-Myc is overexpressed.

  17. Gene expression profiles in primary pancreatic tumors and metastatic lesions of Ela-c-myc transgenic mice.

    Science.gov (United States)

    Thakur, Archana; Bollig, Aliccia; Wu, Jiusheng; Liao, Dezhong J

    2008-01-24

    Pancreatic carcinoma usually is a fatal disease with no cure, mainly due to its invasion and metastasis prior to diagnosis. We analyzed the gene expression profiles of paired primary pancreatic tumors and metastatic lesions from Ela-c-myc transgenic mice in order to identify genes that may be involved in the pancreatic cancer progression. Differentially expressed selected genes were verified by semi-quantitative and quantitative RT-PCR. To further evaluate the relevance of some of the selected differentially expressed genes, we investigated their expression pattern in human pancreatic cancer cell lines with high and low metastatic potentials. Data indicate that genes involved in posttranscriptional regulation were a major functional category of upregulated genes in both primary pancreatic tumors (PT) and liver metastatic lesions (LM) compared to normal pancreas (NP). In particular, differential expression for splicing factors, RNA binding/pre-mRNA processing factors and spliceosome related genes were observed, indicating that RNA processing and editing related events may play critical roles in pancreatic tumor development and progression. High expression of insulin growth factor binding protein-1 (Igfbp1) and Serine proteinase inhibitor A1 (Serpina1), and low levels or absence of Wt1 gene expression were exclusive to liver metastatic lesion samples. We identified Igfbp1, Serpina1 and Wt1 genes that are likely to be clinically useful biomarkers for prognostic or therapeutic purposes in metastatic pancreatic cancer, particularly in pancreatic cancer where c-Myc is overexpressed.

  18. Target guided synthesis using DNA nano-templates for selectively assembling a G-quadruplex binding c-MYC inhibitor

    Science.gov (United States)

    Panda, Deepanjan; Saha, Puja; Das, Tania; Dash, Jyotirmayee

    2017-07-01

    The development of small molecules is essential to modulate the cellular functions of biological targets in living system. Target Guided Synthesis (TGS) approaches have been used for the identification of potent small molecules for biological targets. We herein demonstrate an innovative example of TGS using DNA nano-templates that promote Huisgen cycloaddition from an array of azide and alkyne fragments. A G-quadruplex and a control duplex DNA nano-template have been prepared by assembling the DNA structures on gold-coated magnetic nanoparticles. The DNA nano-templates facilitate the regioselective formation of 1,4-substituted triazole products, which are easily isolated by magnetic decantation. The G-quadruplex nano-template can be easily recovered and reused for five reaction cycles. The major triazole product, generated by the G-quadruplex inhibits c-MYC expression by directly targeting the c-MYC promoter G-quadruplex. This work highlights that the nano-TGS approach may serve as a valuable strategy to generate target-selective ligands for drug discovery.

  19. Quadruplex forming promoter region of c-myc oncogene as a potential target for a telomerase inhibitory plant alkaloid, chelerythrine.

    Science.gov (United States)

    Ghosh, Saptaparni; Dasgupta, Dipak

    2015-03-27

    Guanine rich sequences present in the promoter region of oncogenes could fold into G-quadruplexes and modulate transcription. Equilibrium between folding and unfolding of the quadruplexes in these regions play important role in disease processes. We have studied the effect of a putative anticancer agent chelerythrine on G-rich NHE III1 present in the promoter region of c-myc oncogene. We have demonstrated the ability of chelerythrine, a telomerase inhibitor, to block the hybridization of Pu27 with its complementary strand via folding it into a quadruplex structure. Calorimetry shows that the association of Pu27 with chelerythrine is primarily enthalpy driven with high binding affinity (∼10(5) M(-1)). The association does not lead to any major structural perturbation of Pu27. The resulting 2:1 complex has enhanced stability as compared to free Pu27. Another notable feature is that the presence of molecular crowding agent like ficoll 70 does not change the mode of recognition though the binding affinity decreases. We suggest that the anticancer activity of chelerythrine could be ascribed to its ability to stabilize the quadruplex structure in the c-myc promoter region thereby downregulating its transcription. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Analysis of xanthine dehydrogenase mRNA levels in mutants affecting the expression of the rosy locus.

    OpenAIRE

    Covington, M; Fleenor, D; Devlin, R B

    1984-01-01

    Xanthine dehydrogenase (XDH) mRNA levels were measured in a number of mutants and natural variants affecting XDH gene expression. Two variants, ry+4 and ry+10, contain cis-acting elements which map to a region flanking the 5' end of the XDH gene. Ry+4, which has 2-3 times more XDH protein than a wild type strain, has 3.2 times more XDH mRNA. Ry+10 has 50% of the wild type XDH level and 54% of the wild type XDH mRNA level. Three rosy mutants which map within the structural gene were also exami...

  1. Combined use of nuclear phosphoprotein c-Myc and cellular phosphoprotein p53 for hepatocellular carcinoma detection in high-risk chronic hepatitis C patients.

    Science.gov (United States)

    Attallah, A M; El-Far, M; Abdelrazek, M A; Omran, M M; Attallah, A A; Elkhouly, A A; Elkenawy, H M; Farid, K

    2017-10-01

    Hepatocellular carcinoma (HCC) is a multistage process resulting from various genetic changes. We aimed to determine nuclear phosphoprotein c-Myc and cellular phosphoprotein p53 expression and to evaluate their importance in HCC diagnosis. One hundred and twenty chronic hepatitis C (CHC) patients (60 non-HCC CHC patients and 60 HCC patients who had a single small (p53 were identified in liver tissues and serum samples using immunostaining, western blot and ELISA. Immunohistochemical detection of c-Myc and p53 with monospecific antibodies revealed intense and diffuse cytoplasmic staining patterns. Accumulated mutant proteins, released from tumour cells into the extracellular serum, were detected at 62 KDa, for c-Myc, and 53 KDa, for p53, using western blotting. In contrast to alpha feto-protein, there was a significant increase (p p53 (78.3% vs. 8.3%) in the malignant vs. non-malignant patients. The parallel combination of c-Myc and p53 reach the absolute sensitivity (100%), for more accurate and reliable HCC detection (specificity was 87%). c-Myc and p53 are potential HCC diagnostic biomarkers, and convenient combinations of them could improve diagnostic accuracy of HCC.

  2. I-motif structures formed in the human c-MYC promoter are highly dynamic--insights into sequence redundancy and I-motif stability.

    Directory of Open Access Journals (Sweden)

    Jixun Dai

    Full Text Available The GC-rich nuclease hypersensitivity element III1 (NHE III1 of the c-MYC promoter largely controls the transcriptional activity of the c-MYC oncogene. The C-rich strand in this region can form I-motif DNA secondary structures. We determined the folding pattern of the major I-motif formed in the NHE III1, which can be formed at near-neutral pH. While we find that the I-motif formed in the four 3' consecutive runs of cytosines appears to be the most favored, our results demonstrate that the C-rich strand of the c-MYC NHE III1 exhibits a high degree of dynamic equilibration. Using a trisubstituted oligomer of this region, we determined the formation of two equilibrating loop isomers, one of which contains a flipped-out cytosine. Our results indicate that the intercalative cytosine+-cytosine base pairs are not always necessary for an intramolecular I-motif. The dynamic character of the c-MYC I-motif is intrinsic to the NHE III1 sequence and appears to provide stability to the c-MYC I-motif.

  3. Human RNA polymerase II associated factor 1 complex promotes tumorigenesis by activating c-MYC transcription in non-small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Zhi, Xiuyi [Department of Thoracic Surgery, Xuanwu Hospital, Capital Medical University, Beijing, 100053 (China); Giroux-Leprieur, Etienne [ER2 GRC UPMC04 Theranoscan, Pierre et Marie Curie University, Tenon Hospital, 4 Rue de La Chine, 75020, Paris (France); Respiratory Diseases and Thoracic Oncology Department, Ambroise Pare Hospital – APHP, Versailles Saint Quentin en Yvelines University, 9 Avenue Charles de Gaulle, 92100, Boulogne-Billancourt (France); Wislez, Marie [ER2 GRC UPMC04 Theranoscan, Pierre et Marie Curie University, Tenon Hospital, 4 Rue de La Chine, 75020, Paris (France); Hu, Mu; Zhang, Yi [Department of Thoracic Surgery, Xuanwu Hospital, Capital Medical University, Beijing, 100053 (China); Shi, Huaiyin [Department of Pathology, Chinese PLA General Hospital, Fu-xing Road #28, Beijing, 100853 (China); Du, Kaiqi, E-mail: kaiqidu_zhejiang@163.com [Department of Cardiothoracic Surgery, Chinese People' s Armed Police Force, Zhejiang Corps Hospital, Jiaxing, Zhejiang Province (China); Wang, Lei, E-mail: leiwang_hebei@163.com [Department of Human Anatomy, Hebei Medical University, Hebei Province (China)

    2015-10-02

    Human RNA polymerase II (RNAPII)-associated factor 1 complex (hPAF1C) plays a crucial role in protein-coding gene transcription. Overexpression of hPAF1C has been implicated in the initiation and progression of various human cancers. However, the molecular pathways involved in tumorigenesis through hPAF1C remain to be elucidated. The current study suggested hPAF1C expression as a prognostic biomarker for early stage non-small cell lung cancer (NSCLC) and patients with low hPAF1C expression levels had significantly better overall survival. Furthermore, the expression of hPAF1C was found to be positively correlated with c-MYC expression in patient tumor samples and in cancer cell lines. Mechanistic studies indicated that hPAF1C could promote lung cancer cell proliferation through regulating c-MYC transcription. These results demonstrated the prognostic value of hPAF1C in early-stage NSCLC and the role of hPAF1C in the transcriptional regulation of c-MYC oncogene during NSCLC tumorigenesis. - Highlights: • hPAF1C expression is a prognostic biomarker for early stage non-small cell lung cancer. • The expression of hPAF1C was positively correlated with c-MYC in tumor samples of patients and in several NSCLC cell lines. • hPAF1C could promote lung cancer cell proliferation through regulating c-MYC transcription.

  4. Transcriptional regulation of microsomal prostaglandin E synthase 1 by the proto-oncogene, c-myc, in the pathogenesis of inflammation and cancer.

    Science.gov (United States)

    Ramanan, M; Pilli, V S; Aradhyam, G K; Doble, M

    2017-01-22

    Pro-inflammatory molecules play a key role in the progression of various types of cancers highlighting the importance of studying the pathways that regulate the inflammatory cytokine production. To this end, prostaglandins have been reported to correlate with exacerbated cancer phenotypes that may be prevented by using anti-inflammatory drugs in humans. To understand how the prostaglandin E synthase 1 (mPGES1) may be regulated we analyzed its promoter sequence and identified myc-binding sites. Functional validation was performed by mutating the sites that led to attenuated promoter activation of mPGES1. The known c-myc inhibitor (10058-F4) also blocked PGE2 activity, indicating the importance of c-Myc in PGE2 synthesis. Isocoumarin analogs were able to reduce the expressions of both c-myc as well as mPGES1 and also inhibit the production of PGE2. Based on these data and the well-established role of c-myc in oncogenesis, we have demonstrated an additional role of c-myc in exacerbating cancers via PGE2 production, which may provide a therapeutic opportunity to treat these diseases. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. miR-196b targets c-myc and Bcl-2 expression, inhibits proliferation and induces apoptosis in endometriotic stromal cells.

    Science.gov (United States)

    Abe, Wakana; Nasu, Kaei; Nakada, Chisato; Kawano, Yukie; Moriyama, Masatsugu; Narahara, Hisashi

    2013-03-01

    What is the global expression pattern of microRNAs (miRNAs) in endometriotic stromal cells and is miR-196b involved in the pathogenesis of endometriosis? Several miRNAs are aberrantly expressed in endometriotic cyst stromal cells (ECSCs), including miR-196b whose expression is repressed in endometriotic stromal cells. Although, histologically, endometriotic tissues and normal proliferative endometrium are similar, a number of distinct molecular differences have been reported to date. The anti-apoptotic and excessive proliferative properties of endometriotic cells are considered to be involved in the development and progression of endometriosis. ECSCs and normal endometrial stromal cells (NESCs) were isolated from ovarian endometriotic tissues and eutopic endometrial tissues, respectively and compared. Aberrantly expressed miRNAs in ECSCs were identified by a global miRNA microarray technique. The roles of miR-196b in ECSC proliferation, apoptosis, and c-myc and B-cell lymphoma/leukemia (Bcl)-2 mRNA expression were investigated with precursor hsa-miR-196b transfection. The methylation status of the miR-196b gene in ECSCs and the effect of a DNA demethylating agent on miR-196b expression were also examined. miRNA microarray analysis identified eight down-regulated miRNAs (including miR-196b) and four up-regulated miRNAs in ECSCs. Compulsory expression of miR-196b directed the inhibition of proliferation and the induction of apoptosis in ECSCs. miR-196b was found to suppress c-myc and Bcl-2 mRNA expression in ECSCs, and there was a significant correlation between miR-196b and HOXA10 expression in ECSCs and NESCs. The miR-196b gene was hypermethylated in ECSCs when compared with NESCs, and the treatment with a DNA demethylating agent restored the expression of miR-196b in ECSCs. miRNA expression profiles were investigated only in the stromal component of ectopic and eutopic endometrium samples. In addition to miR-196b, the roles of other miRNAs aberrantly expressed in

  6. Targeted deletion of multiple CTCF-binding elements in the human C-MYC gene reveals a requirement for CTCF in C-MYC expression.

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    Wendy M Gombert

    Full Text Available BACKGROUND: Insulators and domain boundaries both shield genes from adjacent enhancers and inhibit intrusion of heterochromatin into transgenes. Previous studies examined the functional mechanism of the MYC insulator element MINE and its CTCF binding sites in the context of transgenes that were randomly inserted into the genome by transfection. However, the contribution of CTCF binding sites to both gene regulation and maintenance of chromatin has not been tested at the endogenous MYC gene. METHODOLOGY/PRINCIPAL FINDINGS: To determine the impact of CTCF binding on MYC expression, a series of mutant human chromosomal alleles was prepared in homologous recombination-efficient DT40 cells and individually transferred by microcell fusion into murine cells. Functional tests reported here reveal that deletion of CTCF binding elements within the MINE does not impact the capacity of this locus to correctly organize an 'accessible' open chromatin domain, suggesting that these sites are not essential for the formation of a competent, transcriptionally active locus. Moreover, deletion of the CTCF site at the MYC P2 promoter reduces transcription but does not affect promoter acetylation or serum-inducible transcription. Importantly, removal of either CTCF site leads to DNA methylation of flanking sequences, thereby contributing to progressive loss of transcriptional activity. CONCLUSIONS: These findings collectively demonstrate that CTCF-binding at the human MYC locus does not repress transcriptional activity but is required for protection from DNA methylation.

  7. Identification of a c-Jun N-terminal kinase-2-dependent signal amplification cascade that regulates c-Myc levels in ras transformation

    DEFF Research Database (Denmark)

    Mathiasen, D.P.; Egebjerg, C.; Andersen, S.H.

    2012-01-01

    that is critical for ras transformation in murine embryonic fibroblasts. This cascade is coordinated by ERK and JNK2 MAPKs, whose Ras-mediated activation leads to the enhanced levels of three oncogenic transcription factors, namely, c-Myc, activating transcription factor 2 (ATF2) and ATF3, all of which...... are essential for ras transformation. Previous studies show that ERK-mediated serine 62 phosphorylation protects c-Myc from proteasomal degradation. ERK is, however, not alone sufficient to stabilize c-Myc but requires the cooperation of cancerous inhibitor of protein phosphatase 2A (CIP2A), an oncogene......Ras is one of the most frequently activated oncogenes in cancer. Two mitogen-activated protein kinases (MAPKs) are important for ras transformation: extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase 2 (JNK2). Here we present a downstream signal amplification cascade...

  8. cMyc/miR-125b-5p signalling determines sensitivity to bortezomib in preclinical model of cutaneous T-cell lymphomas

    DEFF Research Database (Denmark)

    Manfè, Valentina; Biskup, Edyta; Willumsgaard, Ayalah

    2013-01-01

    improve their clinical efficacy. Using cutaneous T-cell lymphoma (CTCL) as a model of the chemotherapy-resistant peripheral lymphoid malignancy, we demonstrated that resistance to proteasome inhibition involved a signaling between the oncogene cMyc and miR-125b-5p. Bortezomib repressed c......Myc and simultaneously induced miR-125b-5p that exerted a cytoprotective effect through the downmodulation of MAD4. Overexpression of cMyc repressed miR-125b-5p transcription and sensitized lymphoma cells to bortezomib. The central role of miR-125b-5p was further confirmed in a mouse model of T-cell lymphoma, where...

  9. Regulation of the equilibrium between G-quadruplex and duplex DNA in promoter of human c-myc oncogene by a pyrene derivative.

    Science.gov (United States)

    Zhang, Zhenjiang; He, Xiangwei; Yuan, Gu

    2011-12-01

    It has been established that the equilibrium between duplex and G-quadruplex of the nuclease hypersensitivity element III1 (NHE III1) in human c-myc promoter is linked with this gene's transcription. Using NMR and ESI-MS, we have found a pyrene derivative, DMAPP, is able to modulate this equilibrium and, thus, might have the potential to regulate this oncogene's transcription. DMAPP has shown as a G-quadruplex binding agent and could induce c-myc G-quadruplex formation out of duplex. These results provide new clue for rational drug design to target transcription control of c-myc. Copyright © 2011 Elsevier B.V. All rights reserved.

  10. Silencing c-Myc translation as a therapeutic strategy through targeting PI3Kδ and CK1ε in hematological malignancies.

    Science.gov (United States)

    Deng, Changchun; Lipstein, Mark R; Scotto, Luigi; Jirau Serrano, Xavier O; Mangone, Michael A; Li, Shirong; Vendome, Jeremie; Hao, Yun; Xu, Xiaoming; Deng, Shi-Xian; Realubit, Ronald B; Tatonetti, Nicholas P; Karan, Charles; Lentzsch, Suzanne; Fruman, David A; Honig, Barry; Landry, Donald W; O'Connor, Owen A

    2017-01-05

    Phosphoinositide 3-kinase (PI3K) and the proteasome pathway are both involved in activating the mechanistic target of rapamycin (mTOR). Because mTOR signaling is required for initiation of messenger RNA translation, we hypothesized that cotargeting the PI3K and proteasome pathways might synergistically inhibit translation of c-Myc. We found that a novel PI3K δ isoform inhibitor TGR-1202, but not the approved PI3Kδ inhibitor idelalisib, was highly synergistic with the proteasome inhibitor carfilzomib in lymphoma, leukemia, and myeloma cell lines and primary lymphoma and leukemia cells. TGR-1202 and carfilzomib (TC) synergistically inhibited phosphorylation of the eukaryotic translation initiation factor 4E (eIF4E)-binding protein 1 (4E-BP1), leading to suppression of c-Myc translation and silencing of c-Myc-dependent transcription. The synergistic cytotoxicity of TC was rescued by overexpression of eIF4E or c-Myc. TGR-1202, but not other PI3Kδ inhibitors, inhibited casein kinase-1 ε (CK1ε). Targeting CK1ε using a selective chemical inhibitor or short hairpin RNA complements the effects of idelalisib, as a single agent or in combination with carfilzomib, in repressing phosphorylation of 4E-BP1 and the protein level of c-Myc. These results suggest that TGR-1202 is a dual PI3Kδ/CK1ε inhibitor, which may in part explain the clinical activity of TGR-1202 in aggressive lymphoma not found with idelalisib. Targeting CK1ε should become an integral part of therapeutic strategies targeting translation of oncogenes such as c-Myc. © 2017 by The American Society of Hematology.

  11. DNA repair in the c-myc proto-oncogene locus: Possible involvement in susceptibility or resistance to plasmacytoma induction in BALB/c mice

    Energy Technology Data Exchange (ETDEWEB)

    Beecham, E.J.; Mushinski, J.F.; Shacter, E.; Potter, M.; Bohr, V.A. (Laboratory of Molecular Pharmacology, National Cancer Institute, Bethesda, MD (USA))

    1991-06-01

    This report describes an unexpected difference in the efficiency of removal of UV-induced DNA damage in the c-myc locus in splenic B lymphoblasts from two inbred strains of mice. In cells from plasmacytoma-resistant DBA/2N mice, 35% of UV-induced damage in the regulatory and 5{prime} flank of c-myc is removed by 12 h. However, in cells from plasmacytoma-susceptible BALB/cAn mice, damage is not removed from this region. In the protein-encoding region and 3{prime} flank of c-myc as well as in two dihydrofolate reductase gene fragments, UV damage is repaired with similar efficiency in B lymphoblasts from both strains of mice. Furthermore, in the protein-encoding portion and 3{prime} flank of c-myc, damage is selectively removed from only the transcribed strand. No repair is detected in the nontranscribed strand. In contrast, DNA repair in the 5{prime} flank of c-myc is not strand specific; in DNA from DBA/2N cells, UV damage is rapidly removed from both the transcribed and nontranscribed strands. In BALB/cAn cells no repair was detected in either strand in the 5'flank, consistent with the results with double-stranded, nick-translated probes to this region of c-myc. In addition to the repair studies, we have detected post-UV-damage formation: in most of the genes studied, we find that additional T4 endonuclease-sensitive sites are formed in the DNA 2 h after irradiation. Our findings provide new insights into the details of gene-specific and strand-specific DNA repair and suggest that there may be close links between DNA repair and B-cell neoplastic development.

  12. MicroRNA-184 inhibits proliferation and promotes apoptosis of human colon cancer SW480 and HCT116 cells by downregulating C-MYC and BCL-2.

    Science.gov (United States)

    Wang, Yong-Bing; Zhao, Xiao-Hui; Li, Gang; Zheng, Jun-Hua; Qiu, Wei

    2018-02-01

    This study aimed to investigate the effects of microRNA-184 (miR-184) on the proliferation and apoptosis of human colon cancer cells through the regulation of C-MYC and BCL-2. Human colon cancer tissues were selected as case group, and adjacent normal tissues were as control group. Human colon cancer SW480 and HCT116 cells were allocated into blank, miR-184 mimic negative control (mimic-NC), miR-184 inhibitor NC (inhibitor-NC), miR-184 mimic, and miR-184 inhibitor groups. Flow cytometry, Annexin V/PI and MTT assay were used to examine the cell cycle, apoptosis and viability. The expressions of C-MYC, BCL-2 and miR-184 were detected via immunohistochemistry, Western blotting and reverse transcription quantitative polymerase chain reaction (RT-qPCR). C-MYC and BCL-2 were direct targets to miR-184. The growth of colon cancer cells in the miR-184 mimic group was inhibited and exhibited an increase in apoptosis. Cell growth in the miR-184 mimic group was increased in addition to the inhibition of apoptosis. Compared with miR-184 mimic group, the expressions of C-MYC and BCL-2 in miR-184 inhibitor group were increased. The expressions of C-MYC and BCL-2 in colon cancer tissues exhibited high levels of expression, while miR-184 displayed relatively low levels in comparison to the adjacent normal tissues. An association was detected regarding the expressions of miR-184, C-MYC and BCL-2 with the differentiation, invasion depth and lymph node metastasis. MiR-184 expression was negatively related to C-MYC and BCL-2 expressions. Our study suggested that miR-184 could inhibit proliferation and promote apoptosis of colon cancer cells by down-regulating expressions of C-MYC and BCL-2. © 2017 Wiley Periodicals, Inc.

  13. Identification of functional networks of estrogen- and c-Myc-responsive genes and their relationship to response to tamoxifen therapy in breast cancer.

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    Elizabeth A Musgrove

    Full Text Available BACKGROUND: Estrogen is a pivotal regulator of cell proliferation in the normal breast and breast cancer. Endocrine therapies targeting the estrogen receptor are effective in breast cancer, but their success is limited by intrinsic and acquired resistance. METHODOLOGY/PRINCIPAL FINDINGS: With the goal of gaining mechanistic insights into estrogen action and endocrine resistance, we classified estrogen-regulated genes by function, and determined the relationship between functionally-related genesets and the response to tamoxifen in breast cancer patients. Estrogen-responsive genes were identified by transcript profiling of MCF-7 breast cancer cells. Pathway analysis based on functional annotation of these estrogen-regulated genes identified gene signatures with known or predicted roles in cell cycle control, cell growth (i.e. ribosome biogenesis and protein synthesis, cell death/survival signaling and transcriptional regulation. Since inducible expression of c-Myc in antiestrogen-arrested cells can recapitulate many of the effects of estrogen on molecular endpoints related to cell cycle progression, the estrogen-regulated genes that were also targets of c-Myc were identified using cells inducibly expressing c-Myc. Selected genes classified as estrogen and c-Myc targets displayed similar levels of regulation by estrogen and c-Myc and were not estrogen-regulated in the presence of siMyc. Genes regulated by c-Myc accounted for 50% of all acutely estrogen-regulated genes but comprised 85% (110/129 genes in the cell growth signature. siRNA-mediated inhibition of c-Myc induction impaired estrogen regulation of ribosome biogenesis and protein synthesis, consistent with the prediction that estrogen regulates cell growth principally via c-Myc. The 'cell cycle', 'cell growth' and 'cell death' gene signatures each identified patients with an attenuated response in a cohort of 246 tamoxifen-treated patients. In multivariate analysis the cell death signature

  14. Changes in the gene expression of C-myc and CD38 in HL-60 cells during differentiation induced by nicotinic acid-related compounds.

    Science.gov (United States)

    Ida, Chieri; Ogata, Shin; Okumura, Katsuzumi; Taguchi, Hiroshi

    2008-03-01

    Changes in gene expression levels of c-myc and CD38 were examined during the differentiation of HL-60 cells to granulocytes due to three nicotinic acid-related compounds. CD38 expression was increased by isonicotinic acid and all-trans-retinoic acid (ATRA). Nicotinamide and nicotinamide N-oxide drastically decreased c-myc expression, but isonicotinic acid had no effect, suggesting that these compounds differentiate HL-60 to granulocytes through different pathways. These results should provide useful information as to the mechanisms of cell differentiation.

  15. Long-term cultivation of in vitro Apis mellifera cells by gene transfer of human c-myc proto-oncogene.

    Science.gov (United States)

    Kitagishi, Yasuko; Okumura, Naoko; Yoshida, Hitomi; Nishimura, Yuri; Takahashi, Jun-ichi; Matsuda, Satoru

    2011-08-01

    Establishment of cell lines representative of honeybee character would greatly assist in their analysis. Here, we show that immortalized cell line, designated as MYN9, has been generated from honeybee embryo by the gene transfer of human c-myc proto-oncogene. The morphology of the cell is characteristic of embryonic stem cell, although the cell is stable and does not spontaneously differentiate. Polymerase chain reaction analyses show that the cell is originated from authentic honeybee cell. It is proposed that the integration of human c-myc gene into honeybee precursor populations results in the establishment of stable cell line suitable for cellular and molecular studies.

  16. The reducing agent Dithiothreitol (DTT) increases expression of c-myc and c- fos protooncogenes in human cells

    DEFF Research Database (Denmark)

    Skouv, J.; Sørensen, Ilona Kryspin; Frandsen, H.

    1995-01-01

    The objective of the present study was to assess the possible tumour promoting activity of the food mutagen 2-hydroxyamino-1-methyl-6-phenylimidazo[4,5-b]pyridine (N-OH-PhIP), by studying its influence on the expression of three genes considered to be of relevance in the tumour promotion step....... However, when cells were treated with DTT alone, the expression of c-fos and c-myc was also transiently induced. We therefore conclude that DTT, and not N-OH-PhIP, induced oncogene expression. Induction of both c-fos and c-mye expression by a reducing agent, DTT, which is frequently used in in vitro...

  17. Nucleolus disassembly in mitosis and apoptosis: dynamic redistribution of phosphorylated-c-Myc, fibrillarin and Ki-67

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    C Soldani

    2009-06-01

    Full Text Available The nucleolus may undergo disassembly either reversibly during mitosis, or irreversibly in apoptosis, thus allowing the redistribution of the nucleolar proteins.We investigated here by immunocytochemistry the fate of three representative proteins, namely phosphorylated c-Myc, fibrillarin and Ki-67, and found that they behave independently in both processes: they relocate in distinct compartments during mitosis, whereas during apoptosis they may either be cleaved (Ki-67 or be extruded into the cytoplasm with a different kinetics and following an ordered, non chaotic program. The separation of these nucleolar proteins which occurs in early apoptotic nuclei continues also in the cytoplasm, and culminates in the final formation of apoptotic blebs containing different nucleolar proteins: this evidence confirms that the apoptotic bodies may be variable in size, content and surface reactivity, and include heterogeneous aggregates of nuclear proteins and/or nucleic acids.

  18. c-MYC-Induced Sebaceous Gland Differentiation Is Controlled by an Androgen Receptor/p53 Axis

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    Denny L. Cottle

    2013-02-01

    Full Text Available Although the sebaceous gland (SG plays an important role in skin function, the mechanisms regulating SG differentiation and carcinoma formation are poorly understood. We previously reported that c-MYC overexpression stimulates SG differentiation. We now demonstrate roles for the androgen receptor (AR and p53. MYC-induced SG differentiation was reduced in mice lacking a functional AR. High levels of MYC triggered a p53-dependent DNA damage response, leading to accumulation of proliferative SG progenitors and inhibition of AR signaling. Conversely, testosterone treatment or p53 deletion activated AR signaling and restored MYC-induced differentiation. Poorly differentiated human sebaceous carcinomas exhibited high p53 and low AR expression. Thus, the consequences of overactivating MYC in the SG depend on whether AR or p53 is activated, as they form a regulatory axis controlling proliferation and differentiation.

  19. The Proto-Oncogene c-myc Is a Direct Target Gene of Epstein-Barr Virus Nuclear Antigen 2

    Science.gov (United States)

    Kaiser, Carmen; Laux, Gerhard; Eick, Dirk; Jochner, Nicola; Bornkamm, Georg W.; Kempkes, Bettina

    1999-01-01

    Epstein-Barr virus (EBV) infects and transforms primary B lymphocytes in vitro. Viral infection initiates the cell cycle entry of the resting B lymphocytes. The maintenance of proliferation in the infected cells is strictly dependent on functional EBNA2. We have recently developed a conditional immortalization system for EBV by rendering the function of EBNA2, and thus proliferation of the immortalized cells, dependent on estrogen. This cellular system was used to identify early events preceding induction of proliferation. We show that LMP1 and c-myc are directly activated by EBNA2, indicating that all cellular factors essential for induction of these genes by EBNA2 are present in the resting cells. In contrast, induction of the cell cycle regulators cyclin D2 and cdk4 are secondary events, which require de novo protein synthesis. PMID:10196351

  20. Silencing c-Myc translation as a therapeutic strategy through targeting PI3Kδ and CK1ε in hematological malignancies

    OpenAIRE

    Deng, Changchun; Lipstein, Mark R.; Scotto, Luigi; Jirau Serrano, Xavier O.; Mangone, Michael A.; Li, Shirong; Vendome, Jeremie; Hao, Yun; Xu, Xiaoming; Deng, Shi-Xian; Realubit, Ronald B.; Tatonetti, Nicholas P.; Karan, Charles; Lentzsch, Suzanne; Fruman, David A.

    2017-01-01

    A novel PI3Kδ inhibitor TGR-1202 synergizes with proteasome inhibitor carfilzomib by silencing c-Myc in preclinical models of lymphoma.The unique activity of TGR-1202 as a single agent and in combination with carfilzomib is driven by an unexpected activity targeting CK1ε.

  1. Gene therapy of c-myc suppressor FUSE-binding protein-interacting repressor by Sendai virus delivery prevents tracheal stenosis.

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    Daisuke Mizokami

    Full Text Available Acquired tracheal stenosis remains a challenging problem for otolaryngologists. The objective of this study was to determine whether the Sendai virus (SeV-mediated c-myc suppressor, a far upstream element (FUSE-binding protein (FBP-interacting repressor (FIR, modulates wound healing of the airway mucosa, and whether it prevents tracheal stenosis in an animal model of induced mucosal injury. A fusion gene-deleted, non-transmissible SeV vector encoding FIR (FIR-SeV/ΔF was prepared. Rats with scraped airway mucosae were administered FIR-SeV/ΔF through the tracheostoma. The pathological changes in the airway mucosa and in the tracheal lumen were assessed five days after scraping. Untreated animals showed hyperplasia of the airway epithelium and a thickened submucosal layer with extensive fibrosis, angiogenesis, and collagen deposition causing lumen stenosis. By contrast, the administration of FIR-SeV/ΔF decreased the degree of tracheal stenosis (P < 0.05 and improved the survival rate (P < 0.05. Immunohistochemical staining showed that c-Myc expression was downregulated in the tracheal basal cells of the FIR-SeV/ΔF-treated animals, suggesting that c-myc was suppressed by FIR-SeV/ΔF in the regenerating airway epithelium of the injured tracheal mucosa. The airway-targeted gene therapy of the c-myc suppressor FIR, using a recombinant SeV vector, prevented tracheal stenosis in a rat model of airway mucosal injury.

  2. Simultaneous detection of human papillomavirus integration and c-MYC gene amplification in cervical lesions: an emerging marker for the risk to progression.

    Science.gov (United States)

    Gimenes, Fabrícia; Souza, Raquel Pantarotto; de Abreu, André Luelsdorf Pimenta; Pereira, Monalisa Wolski; Consolaro, Marcia Edilaine Lopes; da Silva, Vânia Ramos Sela

    2016-04-01

    The persistence of high-risk oncogenic human papillomavirus (HR-HPV) infection and its integration into the host genome are key steps in the induction of malignant alterations. c-MYC chromosome region is a frequent localization for HPV insertion that has been observed in chromosome band 8q24 by fluorescence in situ hybridization (FISH). We report the HPV viral integration and amplification patterns of the c-MYC gene in cytological smears with FISH as a potential biomarker for the progression of squamous intraepithelial lesions (SIL). HPV detection and genotyping by polymerase chain reaction (PCR) and FISH analysis by "Vysis Cervical FISH Probe" kit (ABBOTT Molecular Inc.) were performed in 37 cervical samples including 8 NILM, 7 ASC-US, 7 LSIL, 3 ASC-H, 7 HSIL and 5 SCC. The results show concordance between FISH and PCR techniques for HPV detection. The majority of the samples contained HR-HPV, the majority being -16 and -18 genotypes. HPV integration as determined by FISH was most frequent in high-risk lesions. The c-MYC gene amplification was found only in HPV-positive samples and was detected primarily in high-risk lesions and in cells with an integrated form of HPV. HPV integration and c-MYC gene amplification detected by FISH could be an important biomarker for use in clinical practice to determine SIL with a risk of progression.

  3. Analyzing the Effect of c-myc Oncogene and Matrix Mettalloproteinase-2 Enzyme Ekspression on Metastasis and Prognosis of Malignant Melanoma

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    Abdulcebbar Siyer

    2016-09-01

    Full Text Available Objectives: The aim of this study is to measure the effect of c-myc oncogene and matrix metalloproteinase-2 enzyme expression on metastasis and prognosis of malignant melanoma. At the end of this study, we hope to get information about the prognosis of melanoma, to be helpful for choosing the treatment strategy. Methods: Sixty-three patients treated in our hospital during 2006-2015 were included in this study. All clinical and histological parameters were collected from each patient’s records and survival rate is assessed. Forty-seven suitable tumor specimens were assessed by immunohistochemical stains and analyzed for the c-myc and MMP-2 positivity. All results were evaluated statistically for the effect on melanoma metastasis and survival rate. Results: C-myc positivity and MMP-2 positivity decreases the survival rate of melanoma. MMP-2 positivity increases the risk of death four more times. Conclusions: So this study confirms that we can evaluate c-myc and MMP-2 as a prognostic factor.

  4. Effect of Neem Leaf Extract (Azadirachta indica) on c-Myc Oncogene Expression in 4T1 Breast Cancer Cells of BALB/c Mice.

    Science.gov (United States)

    Othman, Fauziah; Motalleb, Gholamreza; Lam Tsuey Peng, Sally; Rahmat, Asmah; Basri, Rusliza; Pei Pei, Chong

    2012-01-01

    Breast cancer is the most common cause of cancer-related deaths in women both worldwide and in Malaysia. Azadirachta indica (A. Juss), commonly known as neem, is one of the most versatile medicinal plants that has gained worldwide prominence due to its medicinal properties. However, the anticancer effect of ethanolic neem leaf extract against breast cancer has not been documented. The purpose of the present study is to investigate the effect of neem leaf extract on c-Myc oncogene expression in 4T1 breast cancer BALB/c mice. In this experimental study, A total of 48 female BALB/c mice were divided randomly into four groups of 12 mice per group: i.cancer control (CC) treated with 0.5% Tween 20 in PBS, ii. 0.5 µg/mL tamoxifen citrate (CT), iii. 250 mg/kg neem leaf extract (C250), and iv. 500 mg/kg neem leaf extract (C500). in situ reverse transcription polymerase chain reaction (in situ RT-PCR) was applied to evaluate suppression of c-Myc oncogene expression in breast cancer tissue. The C500 group showed significant (p<0.05) suppression of c-Myc oncogene expression compared to the CC group. c-Myc was found to be down regulated under the effect of 500 mg/kg ethanolic neem leaf extract.

  5. Microwave-assisted synthesis of ruthenium(II) complexes with alkynes as potential inhibitor by selectively recognizing c-myc G-quadruplex DNA.

    Science.gov (United States)

    Zhang, Shuangyan; Wu, Qiong; Zhang, Hao; Wang, Qi; Wang, Xicheng; Mei, Wenjie; Wu, Xiaohui; Zheng, Wenjie

    2017-11-01

    Herein, two polypyridyl ruthenium(II) complexes with alkynes, [Ru(bpy)2L](ClO4)2 (L=p-TEPIP (1) and p-BEPIP (2); bpy=2,2'-bipyridine; p-TEPIP=2-(4-trimethylsilylpropargyl)-1H-imidazo[4,5f][1,10]phenanthroline; p-BEPIP=2-(4-phenyacetylenephenyl)-1H-imidazo[4,5f][1,10]phenanthroline) have been successfully achieved in yields of 32%-89% by a Sonogashira coupling reaction under microwave irradiation. We studied these complexes as potential stabilizers of c-myc G-quadruplex DNA. Observations revealed that both complexes could selectively bind to and stabilize c-myc G-quadruplex DNA with a constant of approximately 1.61±0.78 and 9.47±4.20×10(3)M(-1), respectively, as determined from ITC (isothermal ttitration calorimetry) experiments, FRET (fluorescence resonance energy ttransfer) assay and competitive FRET assay. Moreover, the melting point (Tm) of the c-myc G-quadruplex DNA increased in the presence of 1 and 2 ([Ru]=0.2μM) by approximately 9 and 19.9°C, respectively. It is noteworthy that the conformation of the c-myc G-quadruplex DNA appeared to change when titrated with 1 and 2, which was accompanied by a negative-induced CD (circular dichroism) signal that appeared at a wavelength of 295nm. Furthermore, the conformational change in c-myc G-quadruplex DNA induced by 1 and 2have also been confirmed by TEM (transmission electron microscopy) and AFM (atomic force microscopy). Consequently, the replication of c-myc DNA was blocked by 1 and 2, and especially by 2, as verified by PCR (polymerase chain reaction) -stop assay and Western-blot assay. Thus, these ruthenium(II) complexes can be developed as potential inhibitors in chemotherapy through their binding and stabilization of c-myc G-quadruplex DNA. Copyright © 2017 Elsevier Inc. All rights reserved.

  6. Structure-based design of platinum(II) complexes as c-myc oncogene down-regulators and luminescent probes for G-quadruplex DNA.

    Science.gov (United States)

    Wang, Ping; Leung, Chung-Hang; Ma, Dik-Lung; Yan, Siu-Cheong; Che, Chi-Ming

    2010-06-18

    A series of platinum(II) complexes with tridentate ligands was synthesized and their interactions with G-quadruplex DNA within the c-myc gene promoter were evaluated. Complex 1, which has a flat planar 2,6-bis(benzimidazol-2-yl)pyridine (bzimpy) scaffold, was found to stabilize the c-myc G-quadruplex structure in a cell-free system. An in silico G-quadruplex DNA model has been constructed for structure-based virtual screening to develop new Pt(II)-based complexes with superior inhibitory activities. By using complex 1 as the initial structure for hit-to-lead optimization, bzimpy and related 2,6-bis(pyrazol-3-yl)pyridine (dPzPy) scaffolds containing amine side-chains emerge as the top candidates. Six of the top-scoring complexes were synthesized and their interactions with c-myc G-quadruplex DNA have been investigated. The results revealed that all of the complexes have the ability to stabilize the c-myc G-quadruplex. Complex 3 a ([Pt(II)L2R](+); L2=2,6-bis[1-(3-piperidinepropyl)-1H-enzo[d]imidazol-2-yl]pyridine, R=Cl) displayed the strongest inhibition in a cell-free system (IC(50)=2.2 microM) and was 3.3-fold more potent than that of 1. Complexes 3 a and 4 a ([Pt(II)L3R](+); L3=2,6-bis[1-(3-morpholinopropyl)-1H-pyrazol-3-yl]pyridine, R=Cl) were found to effectively inhibit c-myc gene expression in human hepatocarcinoma cells with IC(50) values of approximately 17 microM, whereas initial hit 1 displayed no significant effect on gene expression at concentrations up to 50 microM. Complexes 3 a and 4 a have a strong preference for G-quadruplex DNA over duplex DNA, as revealed by competition dialysis experiments and absorption titration; 3 a and 4 a bind G-quadruplex DNA with binding constants (K) of approximately 10(6)-10(7) dm(3) mol(-1), which are at least an order of magnitude higher than the K values for duplex DNA. NMR spectroscopic titration experiments and molecular modeling showed that 4 a binds c-myc G-quadruplex DNA through an external end-stacking mode at

  7. Stabilization of G-quadruplex DNA with platinum(II) Schiff base complexes: luminescent probe and down-regulation of c-myc oncogene expression.

    Science.gov (United States)

    Wu, Peng; Ma, Dik-Lung; Leung, Chung-Hang; Yan, Siu-Cheong; Zhu, Nianyong; Abagyan, R; Che, Chi-Ming

    2009-12-07

    The interactions of a series of platinum(II) Schiff base complexes with c-myc G-quadruplex DNA were studied. Complex [PtL(1a)] (1 a; H(2)L(1a)=N,N'-bis(salicylidene)-4,5-methoxy-1,2-phenylenediamine) can moderately inhibit c-myc gene promoter activity in a cell-free system through stabilizing the G-quadruplex structure and can inhibit c-myc oncogene expression in cultured cells. The interaction between 1 a and G-quadruplex DNA has been examined by (1)H NMR spectroscopy. By using computer-aided structure-based drug design for hit-to-lead optimization, an in silico G-quadruplex DNA model has been constructed for docking-based virtual screening to develop new platinum(II) Schiff base complexes with improved inhibitory activities. Complex [PtL(3)] (3; H(2)L(3)=N,N'-bis{4-[1-(2-propylpiperidine)oxy]salicylidene}-4,5-methoxy-1,2-phenylenediamine) has been identified with a top score in the virtual screening. This complex was subsequently prepared and experimentally tested in vitro for its ability to stabilize or induce the formation of the c-myc G-quadruplex. The inhibitory activity of 3 (IC(50)=4.4 muM) is tenfold more than that of 1 a. The interaction between 1 a or 3 with c-myc G-quadruplex DNA has been examined by absorption titration, emission titration, molecular modeling, and NMR titration experiments, thus revealing that both 1 a and 3 bind c-myc G-quadruplex DNA through an external end-stacking mode at the 3' terminal face of the G-quadruplex. Such binding of G-quadruplex DNA with 3 is accompanied by up to an eightfold increase in the intensity of photoluminescence at lambda(max)=652 nm. Complex 3 also effectively down-regulated the expression of c-myc in human hepatocarcinoma cells.

  8. The Effect of Molecular Crowding on the Stability of Human c-MYC Promoter Sequence I-Motif at Neutral pH

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    Edwin A. Lewis

    2013-10-01

    Full Text Available We have previously shown that c-MYC promoter sequences can form stable i-motifs in acidic solution (pH 4.5–5.5. In terms of drug targeting, the question is whether c-MYC promoter sequence i-motifs will exist in the nucleus at neutral pH. In this work, we have investigated the stability of a mutant c-MYC i-motif in solutions containing a molecular crowding agent. The crowded nuclear environment was modeled by the addition of up to 40% w/w polyethylene glycols having molecular weights up to 12,000 g/mol. CD and DSC were used to establish the presence and stability of c-MYC i-motifs in buffer solutions over the pH range 4 to 7. We have shown that the c-MYC i-motif can exist as a stable structure at pH values as high as 6.7 in crowded solutions. Generic dielectric constant effects, e.g., a shift in the pKa of cytosine by more than 2 units (e.g., 4.8 to 7.0, or the formation of non-specific PEG/DNA complexes appear to contribute insignificantly to i-motif stabilization. Molecular crowding, largely an excluded volume effect of added PEG, having a molecular weight in excess of 1,000 g/mol, appears to be responsible for stabilizing the more compact i-motif over the random coil at higher pH values.

  9. MicroRNA-451 induces epithelial-mesenchymal transition in docetaxel-resistant lung adenocarcinoma cells by targeting proto-oncogene c-Myc.

    Science.gov (United States)

    Chen, Dongqin; Huang, Jiayuan; Zhang, Kai; Pan, Banzhou; Chen, Jing; De, Wei; Wang, Rui; Chen, Longbang

    2014-11-01

    Epithelial-mesenchymal transition (EMT) has been reported to play a significant role in tumour metastasis as well as chemoresistance. However, the molecular mechanisms involved in chemotherapy-induced EMT are still unclear. MicroRNA (miRNA) expression and functions have been reported to contribute to phenotypic features of tumour cells. To investigate the roles of miRNAs in chemotherapy-induced EMT, we established two docetaxel-resistant lung adenocarcinoma (LAD) cell models (SPC-A1/DTX and H1299/DTX), which display EMT-like properties and gain increased invasion or migration activity. MiR-451 was found to be significantly downregulated in docetaxel-resistant LAD cells, and re-expression of miR-451 could reverse EMT to mesenchymal-epithelial transition (MET) and inhibit invasion and metastasis of docetaxel-resistant LAD cells both in vitro and in vivo. The proto-oncogene c-Myc was identified as a direct and functional target of miR-451, and further researches confirmed that overexpression of c-Myc which induced extracellular-signal-regulated kinase (ERK)-dependent glycogen synthase kinase-3 beta (GSK-3β) inactivation and subsequent snail activation is essential for acquisition of EMT phenotype induced by loss of miR-451. Furthermore, c-Myc was significantly upregulated in docetaxel-non-responding LAD tissues in comparison with docetaxel-responding tissues, and its expression was inversely correlated with miR-451 expression. This study first reported the involvement of miR-451/c-Myc/ERK/GSK-3β signalling axis in the acquisition of EMT phenotype in docetaxel-resistant LAD cells, suggesting that re-expression of miR-451 or targeting c-Myc will be a potential strategy for the treatment of chemoresistant LAD patients. Copyright © 2014 Elsevier Ltd. All rights reserved.

  10. Cooperation between the polyomavirus Middle-T-antigen gene and the human c-myc oncogene in a rat thyroid epithelial differentiated cell line: Model of in vitro progression

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    Berlingieri, M.T.; Portella, G.; Grieco, M.; Santoro, M.; Fusco, A.

    1988-05-01

    Two rat thyroid epithelial differentiated cell lines, PC CI 3 and PC myc, were infected with the polyoma murine leukemia virus (PyMLV) carrying the Middle-T-antigen gene of polyomavirus. After infection, both cell lines acquired the typical markers of neoplastic transformation; however, the PC myc cells showed a greater malignant phenotype. Furthermore, the thyroid differentiated functions were completely suppressed in PC myc cells transformed by PyMLV, whereas they were, at least partially, retained in PC CI 3 cells transformed by PyMLV, and in particular, thyroglobulin synthesis and secretion were not affected at all. Since no differences in the expression of the middle-T-antigen gene were observed in the two PyMLV-transformed cell lines, the different properties shown by these two infected cell lines must be ascribed to the expression of the c-myc oncogene.

  11. c-myc and N-myc promote active stem cell metabolism and cycling as architects of the developing brain.

    Science.gov (United States)

    Wey, Alice; Knoepfler, Paul S

    2010-06-01

    myc genes are associated with a wide variety of human cancers including most types of nervous system tumors. While the mechanisms by which myc overexpression causes tumorigenesis are multifaceted and have yet to be clearly elucidated, they are at least in part related to endogenous myc function in normal cells. Knockout (KO) of either c-myc or N-myc genes in neural stem and precursor cells (NSC) driven by nestin-cre impairs mouse brain growth and mutation of N-myc also causes microcephaly in humans in Feingold Syndrome. To further define myc function in NSC and nervous system development, we created a double KO (DKO) for c- and N-myc using nestin-cre. The DKO mice display profoundly impaired overall brain growth associated with decreased cell cycling and migration of NSC, which are strikingly decreased in number. The DKO brain also exhibits specific changes in gene expression including downregulation of genes involved in protein and nucleotide metabolism, mitosis, and chromatin structure as well as upregulation of genes associated with differentiation. Together these data support a model of nervous system tumorigenesis in which excess myc aberrantly locks in a developmentally active chromatin state characterized by overactive cell cycling, and metabolism as well as blocked differentiation.

  12. PIK3CD promoted proliferation in diffuse large B cell lymphoma through upregulation of c-myc.

    Science.gov (United States)

    Cui, Wenli; Zheng, Shutao; Li, Xinxia; Ma, Yuqing; Sang, Wei; Liu, Ming; Zhang, Wei; Zhou, Xiaoyan

    2016-09-01

    Despite PIK3CD has been extensively reported in cancers, however, little evidence has been available regarding its role in the setting of diffuse large B cell lymphoma (DLBCL). In the present study, to investigate the role of PIK3CD in DLBCL, relevant experiments were carried out on both in vivo clinical tissue level and in vitro cell line level. Prognostic and clinicopathological significance were analyzed after immunohistochemical assay of PIK3CD expression on DLBCL tissue microarray. MTT assay and flow cytometry were employed to evaluate the proliferative variation, cell cycle, and apoptosis. Athymic nude mice xenografted with DLBCL cell line were employed to confirm the role of PIK3CD. It was found that there was a significant difference between expression of PIK3CD and international prognosis index (IPI), performance state (PS), and inferior overall prognosis. Furthermore, PIK3CD can promote proliferation and prevent apoptosis in DLBCL cells in vitro through upregulation of c-myc and p-AKT and in contrast downregulation of p21 and p27. In nude mice model, knock-down of PIK3CD was shown to be able to suppress the proliferation of DLBCL but not significantly compared with control group. Taken together, our study showed that PIK3CD can promote proliferation of DLBCL cells both in vitro and in vivo, suggesting that PIK3CD could be druggable in the therapy of DLBCL.

  13. HER2, TOP2A, CCND1, EGFR and C-MYC oncogene amplification in colorectal cancer.

    Science.gov (United States)

    Al-Kuraya, Khawla; Novotny, Hedvika; Bavi, Prashant; Siraj, Abdul K; Uddin, Shahab; Ezzat, Adnan; Sanea, Nasser Al; Al-Dayel, Fouad; Al-Mana, Hadeel; Sheikh, Salwa S; Mirlacher, Martina; Tapia, Coya; Simon, Ronald; Sauter, Guido; Terracciano, Luigi; Tornillo, Luigi

    2007-07-01

    Recent studies had suggested substantial molecular differences between tumours from different ethnic groups. In this study, the molecular differences between the incidences of colorectal carcinoma in Saudi and Swiss populations are investigated. 518 cases of colon cancer tumours (114 from Saudi Arabia and 404 from Switzerland) were analysed in a tissue microarray format. Fluorescence in situ hybridisation (FISH) was used to estimate frequencies of copy number changes of known oncogenes, including HER2, TOPO2A, CCND1, EGFR and C-MYC. Using FISH, amplifications were mostly low level (gene-to-centromere ratio 2 to 4), which is in contrast with other tumour types with more frequent gene amplifications. The amplifications were particularly frequent for MYC (Saudi 9% and Swiss 14.2%) but unrelated to clinical outcome and pathological information. Remarkably, there were four tumours exhibiting classic high-level gene amplification for HER2 (Swiss 1.3%), a pattern often accompanied by response to trastuzumab (Herceptin) in breast cancer. Occasional high-level amplifications were also observed for CCND1 (Saudi 1/106, 0.9%; Swiss 2/373, 0.5%) and EGFR (Swiss 2/355; 0.6%). Rare high-level amplifications of therapeutic target genes were found in patients with colon cancer. Although no molecular differences were found between incidences of colon cancer cases in Swiss and Saudi populations, these observations emphasise the urgent need for clinical studies investigating the effect of targeted therapies.

  14. mRNA pseudouridylation affects RNA metabolism in the parasite Toxoplasma gondii.

    Science.gov (United States)

    Nakamoto, Margaret A; Lovejoy, Alexander F; Cygan, Alicja M; Boothroyd, John C

    2017-12-01

    RNA contains over 100 modified nucleotides that are created post-transcriptionally, among which pseudouridine (Ψ) is one of the most abundant. Although it was one of the first modifications discovered, the biological role of this modification is still not fully understood. Recently, we reported that a pseudouridine synthase (TgPUS1) is necessary for differentiation of the single-celled eukaryotic parasite Toxoplasma gondii from active to chronic infection. To better understand the biological role of pseudouridylation, we report here gel-based and deep-sequencing methods to identify TgPUS1-dependent Ψ's in Toxoplasma RNA, and the use of TgPUS1 mutants to examine the effect of this modification on mRNAs. In addition to identifying conserved sites of pseudouridylation in Toxoplasma rRNA, tRNA, and snRNA, we also report extensive pseudouridylation of Toxoplasma mRNAs, with the Ψ's being relatively depleted in the 3'-UTR but enriched at position 1 of codons. We show that many Ψ's in tRNA and mRNA are dependent on the action of TgPUS1 and that TgPUS1-dependent mRNA Ψ's are enriched in developmentally regulated transcripts. RNA-seq data obtained from wild-type and TgPUS1-mutant parasites shows that genes containing a TgPUS1-dependent Ψ are relatively more abundant in mutant parasites, while pulse/chase labeling of RNA with 4-thiouracil shows that mRNAs containing TgPUS1-dependent Ψ have a modest but statistically significant increase in half-life in the mutant parasites. These data are some of the first evidence suggesting that mRNA Ψ's play an important biological role. © 2017 Nakamoto et al.; Published by Cold Spring Harbor Laboratory Press for the RNA Society.

  15. Quinolino-benzo-[5, 6]-dihydroisoquindolium compounds derived from berberine: a new class of highly selective ligands for G-quadruplex DNA in c-myc oncogene.

    Science.gov (United States)

    Ma, Yan; Ou, Tian-Miao; Tan, Jia-Heng; Hou, Jin-Qiang; Huang, Shi-Liang; Gu, Lian-Quan; Huang, Zhi-Shu

    2011-05-01

    A series of quinolino-benzo-[5, 6]-dihydroisoquindolium compounds (3a, 3f, 3g, and 3j) derived from alkaloid berberine were designed and synthesized as novel G-quadruplex ligands. Subsequent biophysical and biochemical evaluation demonstrated that the addition of pyridine ring and amino group into berberine improved the binding ability and selectivity towards G-quadruplex DNA in comparison with the previously reported 9-N-substituted berberine derivatives. Furthermore, qRT-PCR assay showed compound 3j led the down-regulation of c-myc gene transcription in leukemia cell line HL60, while little effect on normal cell line ECV-304, which was consistent with the behavior of an effective G-quadruplex ligand targeting c-myc oncogene. Copyright © 2011 Elsevier Masson SAS. All rights reserved.

  16. Cytotoxic effect of γ-sitosterol from Kejibeling (Strobilanthes crispus and its mechanism of action towards c-myc gene expression and apoptotic pathway

    Directory of Open Access Journals (Sweden)

    Susi Endrini

    2015-01-01

    Full Text Available Background: This study aimed to analyze the cytotoxicity effect of γ-sitosterol isolated from “Kejibeling” (Strobilanthes crispus, a medicinal plant, on several cancer cell lines. The mechanisms of the effects were studied through the expression of cancer-caused gene, c-myc and apoptotic pathways.Methods: This in vitro study was done using human colon cancer cell lines (Caco-2, liver cancer cell lines (HepG2, hormone-dependent breast cancer cell lines (MCF-7 and the normal liver cell lines (Chang Liver. The cytotoxic effect was measured through MTT assay and the potential cytotoxic value was calculated by determining the toxic concentration which may kill up to 50% of the total cell used (IC50. Meanwhile, the cytotoxic mechanism was studied by determining the effect of adding γ-sitosterol to the c-myc gene expression by reverse transciptase-polymerase chain reaction (RT-PCR. The effect of γ-sitosterol through apoptotic pathway was studied by using terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL assay.Results: γ-sitosterol was cytotoxic against Caco-2, HepG2, and MCF-7 with IC50-values of 8.3, 21.8, and 28.8 μg/mL, respectively. There were no IC50-values obtained from this compound against Chang Liver cell line. This compound induced apotosis on Caco-2 and HepG2 cell lines and suppressed the c-myc genes expression in both cells.Conclusion: γ-sitosterol was cytotoxic against colon and liver cancer cell lines and the effect was mediated by down-regulation of c-myc expression and induction of the apoptotic pathways.

  17. Diminished WNT → β-catenin → c-MYC signaling is a barrier for malignant progression of BRAFV600E-induced lung tumors

    Science.gov (United States)

    Juan, Joseph; Muraguchi, Teruyuki; Iezza, Gioia; Sears, Rosalie C.; McMahon, Martin

    2014-01-01

    Oncogene-induced senescence (OIS) is proposed as a cellular defense mechanism that restrains malignant progression of oncogene-expressing, initiated tumor cells. Consistent with this, expression of BRAFV600E in the mouse lung epithelium elicits benign tumors that fail to progress to cancer due to an apparent senescence-like proliferative arrest. Here we demonstrate that nuclear β-catenin → c-MYC signaling is essential for early stage proliferation of BRAFV600E-induced lung tumors and is inactivated in the subsequent senescence-like state. Furthermore, either β-catenin silencing or pharmacological blockade of Porcupine, an acyl-transferase essential for WNT ligand secretion and activity, significantly inhibited BRAFV600E-initiated lung tumorigenesis. Conversely, sustained activity of β-catenin or c-MYC significantly enhanced BRAFV600E-induced lung tumorigenesis and rescued the anti-tumor effects of Porcupine blockade. These data indicate that early stage BRAFV600E-induced lung tumors are WNT-dependent and suggest that inactivation of WNT → β-catenin → c-MYC signaling is a trigger for the senescence-like proliferative arrest that constrains the expansion and malignant progression of BRAFV600E-initiated lung tumors. Moreover, these data further suggest that the trigger for OIS in initiated BRAFV600E-expressing lung tumor cells is not simply a surfeit of signals from oncogenic BRAF but an insufficiency of WNT → β-catenin → c-MYC signaling. These data have implications for understanding how genetic abnormalities cooperate to initiate and promote lung carcinogenesis. PMID:24589553

  18. SirT1 knockdown potentiates radiation-induced bystander effect through promoting c-Myc activity and thus facilitating ROS accumulation

    Energy Technology Data Exchange (ETDEWEB)

    Xie, Yuexia [Institute of Radiation Medicine, Fudan University, Shanghai (China); Central Laboratory, Renji Hospital, School of Medicine, Shanghai Jiaotong University, Shanghai (China); Tu, Wenzhi; Zhang, Jianghong; He, Mingyuan; Ye, Shuang; Dong, Chen [Institute of Radiation Medicine, Fudan University, Shanghai (China); Shao, Chunlin, E-mail: clshao@shmu.edu.cn [Institute of Radiation Medicine, Fudan University, Shanghai (China)

    2015-02-15

    Highlights: • γ-Irradiation induced bystander effects between hepatoma cells and hepatocyte cells. • SirT1 played a protective role in regulating this bystander effect. • SirT1 contributed to the protective effects via elimination the accumulation of ROS. • The activity of c-Myc is critical for maintaining the protective role of SirT1. - Abstract: Radiation-induced bystander effect (RIBE) has important implications for secondary cancer risk assessment during cancer radiotherapy, but the bystander signaling processes, especially under hypoxic condition, are still largely unclear. The present study found that micronuclei (MN) formation could be induced in the non-irradiated HL-7702 hepatocyte cells after being treated with the conditioned medium from irradiated hepatoma HepG2 and SK-Hep-1 cells under either normoxia or hypoxia. This bystander response was dramatically diminished or enhanced when the SirT1 gene of irradiated hepatoma cells was overexpressed or knocked down, respectively, especially under hypoxia. Meanwhile, SirT1 knockdown promoted transcriptional activity for c-Myc and facilitated ROS accumulation. But both of the increased bystander responses and ROS generation due to SirT1-knockdown were almost completely suppressed by c-Myc interference. Moreover, ROS scavenger effectively abolished the RIBE triggered by irradiated hepatoma cells even with SirT1 depletion. These findings provide new insights that SirT1 has a profound role in regulating RIBE where a c-Myc-dependent release of ROS may be involved.

  19. Effect of teicoplanin on the expression of c-myc and c-fos proto-oncogenes in MCF-7 breast cancer cell line

    Directory of Open Access Journals (Sweden)

    Saeideh Ashouri

    2016-01-01

    Conclusion: it could be concluded that although teicoplanin is considered as an enhancing cell growth and proliferation, but probably its effect is not through MAP kinase signaling pathway or perhaps even has inhibitory effect on the expression of some genes such as c-myc and c-fos in this pathway. Hence, the mechanism of action of teicoplanin for increasing cell propagation, through cell signaling pathways or chromosomal abnormalities, remains unclear, and further studies should be conducted.

  20. Mechanisms for c-myc Induced Mouse Mammary Gland Carcinogenesis and for the Synergistic Role of TGF(alpha) in the Process

    Science.gov (United States)

    2001-07-01

    al. 1999, Di Cristofano & Pandolfi 2000). In a cells, and that co-transfection with another oncogene or myc-CAT ( chloramphenicol acetyl transferase...sexes of as well, with a 50% incidence at about 7 months, slightly animals suggest that the strong synergism between the two earlier than that in MMTV...tgfci nor c-myc been reported to be associated with a reduced survival in alone is a sufficient carcinogenic factor, their synergism is some studies

  1. Mutant p53 Gains Its Function via c-Myc Activation upon CDK4 Phosphorylation at Serine 249 and Consequent PIN1 Binding.

    Science.gov (United States)

    Liao, Peng; Zeng, Shelya X; Zhou, Xiang; Chen, Tianjian; Zhou, Fen; Cao, Bo; Jung, Ji Hoon; Del Sal, Giannino; Luo, Shiwen; Lu, Hua

    2017-11-23

    TP53 missense mutations significantly influence the development and progression of various human cancers via their gain of new functions (GOF) through different mechanisms. Here we report a unique mechanism underlying the GOF of p53-R249S (p53-RS), a p53 mutant frequently detected in human hepatocellular carcinoma (HCC) that is highly related to hepatitis B infection and aflatoxin B1. A CDK inhibitor blocks p53-RS's nuclear translocation in HCC, whereas CDK4 interacts with p53-RS in the G1/S phase of the cells, phosphorylates it, and enhances its nuclear localization. This is coupled with binding of a peptidyl-prolyl cis-trans isomerase NIMA-interacting 1 (PIN1) to p53-RS, but not the p53 form with mutations of four serines/threonines previously shown to be crucial for PIN1 binding. As a result, p53-RS interacts with c-Myc and enhances c-Myc-dependent rDNA transcription key for ribosomal biogenesis. These results unveil a CDK4-PIN1-p53-RS-c-Myc pathway as a novel mechanism for the GOF of p53-RS in HCC. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Ketogenic HMGCS2 Is a c-Myc target gene expressed in differentiated cells of human colonic epithelium and down-regulated in colon cancer.

    Science.gov (United States)

    Camarero, Nuria; Mascaró, Cristina; Mayordomo, Cristina; Vilardell, Felip; Haro, Diego; Marrero, Pedro F

    2006-09-01

    HMGCS2, the gene that regulates ketone body production, is expressed in liver and several extrahepatic tissues, such as the colon. In CaCo-2 colonic epithelial cells, the expression of this gene increases with cell differentiation. Accordingly, immunohistochemistry with specific antibodies shows that HMGCS2 is expressed mainly in differentiated cells of human colonic epithelium. Here, we used a chromatin immunoprecipitation assay to study the molecular mechanism responsible for this expression pattern. The assay revealed that HMGCS2 is a direct target of c-Myc, which represses HMGCS2 transcriptional activity. c-Myc transrepression is mediated by blockade of the transactivating activity of Miz-1, which occurs mainly through a Sp1-binding site in the proximal promoter of the gene. Accordingly, the expression of human HMGCS2 is down-regulated in 90% of Myc-dependent colon and rectum tumors. HMGCS2 protein expression is down-regulated preferentially in moderately and poorly differentiated carcinomas. In addition, it is also down-regulated in 80% of small intestine Myc-independent tumors. Based on these findings, we propose that ketogenesis is an undesirable metabolic characteristic of the proliferating cell, which is down-regulated through c-Myc-mediated repression of the key metabolic gene HMGCS2.

  3. A Novel PTEN/Mutant p53/c-Myc/Bcl-XL Axis Mediates Context-Dependent Oncogenic Effects of PTEN with Implications for Cancer Prognosis and Therapy

    Directory of Open Access Journals (Sweden)

    Xiaoping Huang

    2013-08-01

    Full Text Available Phosphatase and tensin homolog located on chromosome 10 (PTEN is one of the most frequently mutated tumor suppressors in human cancer including in glioblastoma. Here, we show that PTEN exerts unconventional oncogenic effects in glioblastoma through a novel PTEN/mutant p53/c-Myc/Bcl-XL molecular and functional axis. Using a wide array of molecular, genetic, and functional approaches, we demonstrate that PTEN enhances a transcriptional complex containing gain-of-function mutant p53, CBP, and NFY in human glioblastoma cells and tumor tissues. The mutant p53/CBP/NFY complex transcriptionally activates the oncogenes c-Myc and Bcl-XL, leading to increased cell proliferation, survival, invasion, and clonogenicity. Disruption of the mutant p53/c-Myc/Bcl-XL axis or mutant p53/CBP/NFY complex reverses the transcriptional and oncogenic effects of PTEN and unmasks its tumor-suppressive function. Consistent with these data, we find that PTEN expression is associated with worse patient survival than PTEN loss in tumors harboring mutant p53 and that a small molecule modulator of p53 exerts greater antitumor effects in PTEN-expressing cancer cells. Altogether, our study describes a new signaling pathway that mediates context-dependent oncogenic/tumor-suppressive role of PTEN. The data also indicate that the combined mutational status of PTEN and p53 influences cancer prognosis and anticancer therapies that target PTEN and p53.

  4. K-Ras and B-Raf oncogenes inhibit colon epithelial polarity establishment through up-regulation of c-myc.

    Science.gov (United States)

    Magudia, Kirti; Lahoz, Aurelia; Hall, Alan

    2012-07-23

    KRAS, BRAF, and PI3KCA are the most frequently mutated oncogenes in human colon cancer. To explore their effects on morphogenesis, we used the colon cancer-derived cell line Caco-2. When seeded in extracellular matrix, individual cells proliferate and generate hollow, polarized cysts. The expression of oncogenic phosphatidylinositol 3-kinase (PI3KCA H1047R) in Caco-2 has no effect, but K-Ras V12 or B-Raf V600E disrupts polarity and tight junctions and promotes hyperproliferation, resulting in large, filled structures. Inhibition of mitogen-activated protein/extracellular signal-regulated kinase (ERK) kinase blocks the disruption of morphology, as well as the increased levels of c-myc protein induced by K-Ras V12 and B-Raf V600E. Apical polarity is already established after the first cell division (two-cell stage) in Caco-2 three-dimensional cultures. This is disrupted by expression of K-Ras V12 or B-Raf V600E but can be rescued by ribonucleic acid interference-mediated depletion of c-myc. We conclude that ERK-mediated up-regulation of c-myc by K-Ras or B-Raf oncogenes disrupts the establishment of apical/basolateral polarity in colon epithelial cells independently of its effect on proliferation.

  5. Keratinocyte transcriptional regulation of the human c-Myc promoter occurs via a novel Lef/Tcf binding element distinct from neoplastic cells.

    Science.gov (United States)

    Kolly, Carine; Zakher, Antony; Strauss, Christian; Suter, Maja M; Müller, Eliane J

    2007-05-15

    The proto-oncogene c-Myc is involved in early neoplastic transformations. Two consensus Lef/Tcf binding elements (TBE) were found to be prerequisite for transcriptional transactivation by the armadillo proteins beta-catenin and plakoglobin (PG) together with Tcf4 in human neoplastic cells. In epidermal keratinocytes, c-Myc was reported to be repressed by Lef-1 and PG. Using reporter gene assays, here we demonstrate that deletion of the two consensus TBE fails to abrogate transcriptional regulation by Lef-1/PG in wildtype and beta-catenin-/- keratinocytes, while it reduces transcription in pre-neoplastic PG-/- keratinocytes. We identified a TBE sequence variant downstream of the major transcriptional initiation site that binds Lef-1 in vitro and in vivo, and its mutation compromised transcriptional regulation by Lef-1/PG. Collectively, this study demonstrates that the two consensus TBE's reported in neoplastic cells are dispensable for c-Myc regulation in normal keratinocytes, which instead use a novel TBE sequence variant. This unprecedented finding may have important implications for armadillo target genes involved in carcinogenesis.

  6. DNA Methylation Mediated Downregulation of miR-449c Controls Osteosarcoma Cell Cycle Progression by Directly Targeting Oncogene c-Myc.

    Science.gov (United States)

    Li, Qing; Li, Hua; Zhao, Xueling; Wang, Bing; Zhang, Lin; Zhang, Caiguo; Zhang, Fan

    2017-01-01

    MicroRNAs (miRNAs) are critical regulators of gene expression, and they have broad roles in the pathogenesis of different diseases including cancer. Limited studies and expression profiles of miRNAs are available in human osteosarcoma cells. By applying a miRNA microarray analysis, we observed a number of miRNAs with abnormal expression in cancerous tissues from osteosarcoma patients. Of particular interest in this study was miR-449c, which was significantly downregulated in osteosarcoma cells and patients, and its expression was negatively correlated with tumor size and tumor MSTS stages. Ectopic expression of miR-449c significantly inhibited osteosarcoma cell proliferation and colony formation ability, and caused cell cycle arrest at the G1 phase. Further analysis identified that miR-449c was able to directly target the oncogene c-Myc and negatively regulated its expression. Overexpression of c-Myc partially reversed miR-449c-mimic-inhibited cell proliferation and colony formation. Moreover, DNA hypermethylation was observed in two CpG islands adjacent to the genomic locus of miR-449c in osteosarcoma cells. Conversely, treatment with the DNA methylation inhibitor AZA caused induction of miR-449c. In conclusion, our results support a model that DNA methylation mediates downregulation of miR-449c, diminishing miR-449c mediated inhibition of c-Myc and thus leading to the activation of downstream targets, eventually contributing to osteosarcoma tumorigenesis.

  7. SirT1 knockdown potentiates radiation-induced bystander effect through promoting c-Myc activity and thus facilitating ROS accumulation.

    Science.gov (United States)

    Xie, Yuexia; Tu, Wenzhi; Zhang, Jianghong; He, Mingyuan; Ye, Shuang; Dong, Chen; Shao, Chunlin

    2015-02-01

    Radiation-induced bystander effect (RIBE) has important implications for secondary cancer risk assessment during cancer radiotherapy, but the bystander signaling processes, especially under hypoxic condition, are still largely unclear. The present study found that micronuclei (MN) formation could be induced in the non-irradiated HL-7702 hepatocyte cells after being treated with the conditioned medium from irradiated hepatoma HepG2 and SK-Hep-1 cells under either normoxia or hypoxia. This bystander response was dramatically diminished or enhanced when the SirT1 gene of irradiated hepatoma cells was overexpressed or knocked down, respectively, especially under hypoxia. Meanwhile, SirT1 knockdown promoted transcriptional activity for c-Myc and facilitated ROS accumulation. But both of the increased bystander responses and ROS generation due to SirT1-knockdown were almost completely suppressed by c-Myc interference. Moreover, ROS scavenger effectively abolished the RIBE triggered by irradiated hepatoma cells even with SirT1 depletion. These findings provide new insights that SirT1 has a profound role in regulating RIBE where a c-Myc-dependent release of ROS may be involved. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. The Dysregulation of Polyamine Metabolism in Colorectal Cancer Is Associated with Overexpression of c-Myc and C/EBPβ rather than Enterotoxigenic Bacteroides fragilis Infection

    Directory of Open Access Journals (Sweden)

    Anastasiya V. Snezhkina

    2016-01-01

    Full Text Available Colorectal cancer is one of the most common cancers in the world. It is well known that the chronic inflammation can promote the progression of colorectal cancer (CRC. Recently, a number of studies revealed a potential association between colorectal inflammation, cancer progression, and infection caused by enterotoxigenic Bacteroides fragilis (ETBF. Bacterial enterotoxin activates spermine oxidase (SMO, which produces spermidine and H2O2 as byproducts of polyamine catabolism, which, in turn, enhances inflammation and tissue injury. Using qPCR analysis, we estimated the expression of SMOX gene and ETBF colonization in CRC patients. We found no statistically significant associations between them. Then we selected genes involved in polyamine metabolism, metabolic reprogramming, and inflammation regulation and estimated their expression in CRC. We observed overexpression of SMOX, ODC1, SRM, SMS, MTAP, c-Myc, C/EBPβ (CREBP, and other genes. We found that two mediators of metabolic reprogramming, inflammation, and cell proliferation c-Myc and C/EBPβ may serve as regulators of polyamine metabolism genes (SMOX, AZIN1, MTAP, SRM, ODC1, AMD1, and AGMAT as they are overexpressed in tumors, have binding site according to ENCODE ChIP-Seq data, and demonstrate strong coexpression with their targets. Thus, increased polyamine metabolism in CRC could be driven by c-Myc and C/EBPβ rather than ETBF infection.

  9. Establishment of c-myc-immortalized Kupffer cell line from a C57BL/6 mouse strain

    Directory of Open Access Journals (Sweden)

    Hiroshi Kitani

    2014-01-01

    Full Text Available We recently demonstrated in several mammalian species, a novel procedure to obtain liver-macrophages (Kupffer cells in sufficient numbers and purity using a mixed primary culture of hepatocytes. In this study, we applied this method to the C57BL/6 mouse liver and established an immortalized Kupffer cell line from this mouse strain. The hepatocytes from the C57BL/6 adult mouse liver were isolated by a two-step collagenase perfusion method and cultured in T25 culture flasks. Similar to our previous studies, the mouse hepatocytes progressively changed their morphology into a fibroblastic appearance after a few days of culture. After 7–10 days of culture, Kupffer-like cells, which were contaminants in the hepatocyte fraction at the start of the culture, actively proliferated on the mixed fibroblastic cell sheet. At this stage, a retroviral vector containing the human c-myc oncogene and neomycin resistance gene was introduced into the mixed culture. Gentle shaking of the culture flask, followed by the transfer and brief incubation of the culture supernatant, resulted in a quick and selective adhesion of Kupffer cells to a plastic dish surface. After selection with G418 and cloning by limiting dilutions, a clonal cell line (KUP5 was established. KUP5 cells displayed typical macrophage morphology and were stably passaged at 4–5 days intervals for more than 5 months, with a population doubling time of 19 h. KUP5 cells are immunocytochemically positive for mouse macrophage markers, such as Mac-1, F4/80. KUP5 cells exhibited substantial phagocytosis of polystyrene microbeads and the release of inflammatory cytokines upon lipopolysaccharide stimulation. Taken together, KUP5 cells provide a useful means to study the function of Kupffer cells in vitro.

  10. Transgenic rabbits with lymphocytic leukemia induced by the c-myc oncogene fused with the immunoglobulin heavy chain enhancer.

    Science.gov (United States)

    Knight, K L; Spieker-Polet, H; Kazdin, D S; Oi, V T

    1988-01-01

    Transgenic rabbits with the rabbit c-myc oncogene fused with the rabbit immunoglobulin heavy chain enhancer region (E mu) DNA were developed by microinjecting pronuclei of single cell zygotes with the gene construct and implanting the microinjected eggs into pseudopregnant females. At age 17-20 days, 3 of 21 offspring born to seven females were found to have peripheral blood leukocyte counts of greater than 100,000 per mm3. Histology analyses showed extensive lymphocytic infiltration in the liver and kidney, indicating that these rabbits had a malignant lymphocytic leukemia. Genomic DNA analyses of thymus and peripheral blood lymphocytes revealed that the leukemic rabbits were transgenic and that both immunoglobulin heavy and kappa light chain genes were rearranged in the leukemic cells; thus, the leukemic cells are of B-cell lineage. One to four heavy and light chain gene rearrangements were observed, suggesting that the B-cell tumors were oligoclonal. Stable tissue culture cell lines from the bone marrow and peripheral blood of one of the transgenic rabbits have been developed. The development of B-cell leukemias in rabbits with the E mu-myc transgene contrasts with the development of B-cell lymphomas in mice carrying a similar transgene. The lymphomas in mice develop in adults and are monoclonal in origin. The leukemias in rabbits develop in juveniles, less than 3 weeks after birth, and appear oligoclonal in origin. The leukemias seem to develop in rabbit at a specific stage of development, and we suggest that a normal developmental signal may be involved in the oncogenesis. Images PMID:2834733

  11. Marek's disease virus-encoded analog of microRNA-155 activates the oncogene c-Myc by targeting LTBP1 and suppressing the TGF-β signaling pathway.

    Science.gov (United States)

    Chi, Jia-Qi; Teng, Man; Yu, Zu-Hua; Xu, Hui; Su, Jing-Wei; Zhao, Pu; Xing, Guang-Xu; Liang, Hong-De; Deng, Rui-Guang; Qu, Liang-Hu; Zhang, Gai-Ping; Luo, Jun

    2015-02-01

    Marek's disease virus (MDV) is a representative alpha herpes virus able to induce rapid-onset T-cell lymphoma in its natural host and regarded as an ideal model for the study of virus-induced tumorigenesis. Recent studies have shown that the mdv1-miR-M4-5p, a viral analog of cellular miR-155, is critical for MDV׳s oncogenicity. However, the precise mechanism whereby it was involved in MD lymphomagenesis remained unknown. We have presently identified the host mRNA targets of mdv1-miR-M4-5 and identified the latent TGF-β binding protein 1 (LTBP1) as a critical target for it. We found that during MDV infection, down-regulation of LTBP1 expression by mdv1-miR-M4-5p led to a significant decrease of the secretion and activation of TGF-β1, with suppression of TGF-β signaling and a significant activation of expression of c-Myc, a well-known oncogene which is critical for virus-induced tumorigenesis. Our findings reveal a novel and important mechanism of how mdv1-miR-M4-5p potentially contributes to MDV-induced tumorigenesis. Copyright © 2014 The Authors. Published by Elsevier Inc. All rights reserved.

  12. Feedback circuitry via let-7c between lncRNA CCAT1 and c-Myc is involved in cigarette smoke extract-induced malignant transformation of HBE cells.

    Science.gov (United States)

    Lu, Lu; Qi, Hong; Luo, Fei; Xu, Hui; Ling, Min; Qin, Yu; Yang, Ping; Liu, Xinlu; Yang, Qianlei; Xue, Junchao; Chen, Chao; Lu, Jiachun; Xiang, Quanyong; Liu, Qizhan; Bian, Qian

    2017-03-21

    Cigarette smoking is a primary risk factor for the development of lung cancer, which is regarded as the leading cause of cancer-related deaths. The process of malignant transformation of cells, however, is complex and elusive. The present study investigated the roles of an lncRNA, CCAT1, and a transcriptional factor, c-Myc, in human bronchial epithelial (HBE) cell transformation induced by cigarette smoke extract. With acute and chronic treatment of HBE cells, cigarette smoke extract induced increases of CCAT1 and c-Myc levels and decreases of levels of let-7c, a microRNA. Down-regulation of c-Myc reduced the degree of malignancy and the invasion/migration capacity of HBE cells transformed by cigarette smoke extract. ChIP assays established that c-Myc, increased by cigarette smoke extract, binds to the promoter of CCAT1, activating its transcription. Further, let-7c suppressed the expression of c-Myc through binding to its 3'-UTR. In turn, CCAT1 promoted the accumulation of c-Myc through binding to let-7c and decreasing free let-7c, which influenced the neoplastic capacity of HBE cells transformed by cigarette smoke extract. These results indicate that a positive feedback loop ensures expression of cigarette smoke extract-induced CCAT1 and c-Myc via let-7c, which is involved in cigarette smoke extract-induced malignant transformation of HBE cells. Thus, the present research establishes a new mechanism for the reciprocal regulation between CCAT1 and c-Myc and provides an understanding of cigarette smoke extract-induced lung carcinogenesis.

  13. Increases in iPS Transcription Factor (Oct4, Sox2, c-Myc, and Klf4) Gene Expression after Modified Electroconvulsive Therapy.

    Science.gov (United States)

    Nishiguchi, Masaki; Kikuyama, Hiroki; Kanazawa, Tetsufumi; Tsutsumi, Atsushi; Kaneko, Takao; Uenishi, Hiroyuki; Kawabata, Yasuo; Kawashige, Seiya; Koh, Jun; Yoneda, Hiroshi

    2015-10-01

    Electroconvulsive therapy (ECT) is a reasonable option for intractable depression or schizophrenia, but a mechanism of action has not been established. One credible hypothesis is related to neural plasticity. Three genes (Oct4, Sox2, c-Myc) involved in the induction of induced pluripotent stem (iPS) cells are Wnt-target genes, which constitute a key gene group involved in neural plasticity through the TCF family. Klf4 is the other gene among Yamanaka's four transcription factors, and increases in its expression are induced by stimulation of the canonical Wnt pathway. We compared the peripheral blood gene expression of the four iPS genes (Oct4, Sox2, c-Myc, and Klf4) before and after modified ECT (specifically ECT with general anesthesia) of patients with intractable depression (n=6) or schizophrenia (n=6). Using Thymatron ten times the total bilateral electrical stimulation was evoked. Both assessments of the symptoms demonstrated significant improvement after mECT stimulation. Expression of all four genes was confirmed to increase after initial stimulation. The gene expression levels after treatment were significantly different from the initial gene expression in all twelve cases at the following treatment stages: at the 3rd mECT for Oct4; at the 6th and 10th mECT for Sox2; and at the 3rd, 6th and 10th mECT for c-Myc. These significant differences were not present after correction for multiple testing; however, our data have the potential to explain the molecular mechanisms of mECT from a unique perspective. Further studie should be conducted to clarify the pathophysiological involvement of iPS-inducing genes in ECT.

  14. The BET Bromodomain Inhibitor JQ1 Suppresses Chondrosarcoma Cell Growth via Regulation of YAP/p21/c-Myc Signaling.

    Science.gov (United States)

    Zhang, Huan-Tian; Gui, Tao; Sang, Yuan; Yang, Jie; Li, Yu-Hang; Liang, Gui-Hong; Li, Thomas; He, Qing-Yu; Zha, Zhen-Gang

    2017-08-01

    Chondrosarcoma, the second-most frequent primary bone malignancy, is generally more resistant to conventional chemotherapy and radiotherapy. Therefore, the development of an effective adjuvant therapy is necessary. Recently, targeting the epigenetic regulator such as bromodomain and extraterminal domain (BET) proteins has achieved great success. For instance, the bromodomain inhibitor JQ1 has been shown to inhibit the growth of several cancer cells both in vitro and in vivo. Herein, we demonstrated that JQ1 significantly inhibited chondrosarcoma cell growth and colony formation. JQ1 also induced marked G1-phase cell cycle arrest coincided with the up-regulation of p21WAF1/CIP1 , p27Kip1 , and Cyclin D1 expression, and the down-regulation of Cyclin E2 expression. Moreover, JQ1 induced the premature senescence of SW 1353 cells, and that prolong treatment of JQ1 caused cell apoptosis. Mechanistically, the JQ1-induced cell growth inhibition was correlated with the suppression of c-Myc and Bcl-xL, which are the prime genes for cell cycle control and anti-apoptosis. Furthermore, we demonstrated that p21 negatively regulated the expression of c-Myc and Bcl-xL upon JQ1 treatment, and that the growth inhibition of SW 1353 and Hs 819.T cells and induction of p21 were predominantly regulated by the LATS1/YAP signaling but not through a p53-dependent manner. In conclusion, we disclosed a novel mechanism that JQ1 inhibits cell proliferation, induces cell senescence and apoptosis of chondrosarcoma cells through the regulation of the YAP/p21/c-Myc/Bcl-xL signaling axis. J. Cell. Biochem. 118: 2182-2192, 2017. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  15. Receptor for hyaluronic acid- mediated motility (RHAMM) regulates HT1080 fibrosarcoma cell proliferation via a β-catenin/c-myc signaling axis.

    Science.gov (United States)

    Kouvidi, Katerina; Berdiaki, Aikaterini; Tzardi, Maria; Karousou, Evgenia; Passi, Alberto; Nikitovic, Dragana; Tzanakakis, George N

    2016-04-01

    High levels of hyaluronan (HA) synthesis in various cancer tissues, including sarcomas, are correlated with tumorigenesis and malignant transformation. RHAMM (receptor for hyaluronic acid-mediated motility) is overexpressed during tumor development in different malignancies. β-Catenin is a crucial downstream mediator of the Wnt signaling cascade which facilitates carcinogenic events characterized by deregulated cell proliferation. Real-time PCR, in vitro cell proliferation assay, siRNA transfection, flow cytometry, immunoprecipitation, western blotting and immunofluorescence were utilized. The reduction of RHAMM expression was strongly correlated with an inhibition of HT1080 fibrosarcoma cell growth (p≤0.01). LMWHA, in a RHAMM-dependent manner increases cell growth of HT1080 cells (p≤0.01). Both basal and LMWHA dependent growth of HT1080 cells was attenuated by β-catenin deficiency (p≤0.01). β-Catenin cytoplasmatic deposition is positively regulated by RHAMM (p≤0.01). Immunoflourescence and immunoprecipitation suggest that RHAMM/β-catenin form an intracellular complex. Transfection experiments identified c-myc as candidate downstream mediator of RHAMM/β-catenin effects on HT1080 fibrosarcoma cell proliferation. LMWHA/RHAMM downstream signaling regulates fibrosarcoma cell growth in a β-catenin/c-myc dependent manner. The present study suggests that RHAMM is a novel β-catenin intracellular binding partner, protecting β-catenin from degradation and supporting the nuclear translocation of this key cellular mediator, which results in c-myc activation and enhanced fibrosarcoma cell growth. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. The effect of non-coding DNA variations on P53 and cMYC competitive inhibition at cis-overlapping motifs.

    Science.gov (United States)

    Kin, Katherine; Chen, Xi; Gonzalez-Garay, Manuel; Fakhouri, Walid D

    2016-04-15

    Non-coding DNA variations play a critical role in increasing the risk for development of common complex diseases, and account for the majority of SNPs highly associated with cancer. However, it remains a challenge to identify etiologic variants and to predict their pathological effects on target gene expression for clinical purposes. Cis-overlapping motifs (COMs) are elements of enhancer regions that impact gene expression by enabling competitive binding and switching between transcription factors. Mutations within COMs are especially important when the involved transcription factors have opposing effects on gene regulation, like P53 tumor suppressor and cMYC proto-oncogene. In this study, genome-wide analysis of ChIP-seq data from human cancer and mouse embryonic cells identified a significant number of putative regulatory elements with signals for both P53 and cMYC. Each co-occupied element contains, on average, two COMs, and one common SNP every two COMs. Gene ontology of predicted target genes for COMs showed that the majority are involved in DNA damage, apoptosis, cell cycle regulation, and RNA processing. EMSA results showed that both cMYC and P53 bind to cis-overlapping motifs within a ChIP-seq co-occupied region in Chr12. In vitro functional analysis of selected co-occupied elements verified enhancer activity, and also showed that the occurrence of SNPs within three COMs significantly altered enhancer activity. We identified a list of COM-associated functional SNPs that are in close proximity to SNPs associated with common diseases in large population studies. These results suggest a potential molecular mechanism to identify etiologic regulatory mutations associated with common diseases. © The Author 2016. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  17. Liver tumor formation by a mutant retinoblastoma protein in the transgenic mice is caused by an upregulation of c-Myc target genes

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Bo; Hikosaka, Keisuke; Sultana, Nishat; Sharkar, Mohammad Tofael Kabir [Department of Biochemistry, Hamamatsu University School of Medicine, 1-20-1 Handa-yama, Higashi-ku, Hamamatsu 431-3192 (Japan); Noritake, Hidenao [Department of Biochemistry, Hamamatsu University School of Medicine, 1-20-1 Handa-yama, Higashi-ku, Hamamatsu 431-3192 (Japan); Department of Internal Medicine, Hamamatsu University School of Medicine, 1-20-1 Handa-yama, Higashi-ku, Hamamatsu 431-3192 (Japan); Kimura, Wataru; Wu, Yi-Xin [Department of Biochemistry, Hamamatsu University School of Medicine, 1-20-1 Handa-yama, Higashi-ku, Hamamatsu 431-3192 (Japan); Kobayashi, Yoshimasa [Department of Internal Medicine, Hamamatsu University School of Medicine, 1-20-1 Handa-yama, Higashi-ku, Hamamatsu 431-3192 (Japan); Uezato, Tadayoshi [Department of Biochemistry, Hamamatsu University School of Medicine, 1-20-1 Handa-yama, Higashi-ku, Hamamatsu 431-3192 (Japan); Miura, Naoyuki, E-mail: nmiura@hama-med.ac.jp [Department of Biochemistry, Hamamatsu University School of Medicine, 1-20-1 Handa-yama, Higashi-ku, Hamamatsu 431-3192 (Japan)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Fifty percent of the mutant Rb transgenic mice produced liver tumors. Black-Right-Pointing-Pointer In the tumor, Foxm1, Skp2, Bmi1 and AP-1 mRNAs were up-regulated. Black-Right-Pointing-Pointer No increase in expression of the Myc-target genes was observed in the non-tumorous liver. Black-Right-Pointing-Pointer Tumor formation depends on up-regulation of the Myc-target genes. -- Abstract: The retinoblastoma (Rb) tumor suppressor encodes a nuclear phosphoprotein that regulates cellular proliferation, apoptosis and differentiation. In order to adapt itself to these biological functions, Rb is subjected to modification cycle, phosphorylation and dephosphorylation. To directly determine the effect of phosphorylation-resistant Rb on liver development and function, we generated transgenic mice expressing phosphorylation-resistant human mutant Rb (mt-Rb) under the control of the rat hepatocyte nuclear factor-1 gene promoter/enhancer. Expression of mt-Rb in the liver resulted in macroscopic neoplastic nodules (adenomas) with {approx}50% incidence within 15 months old. Interestingly, quantitative reverse transcriptase-PCR analysis showed that c-Myc was up-regulated in the liver of mt-Rb transgenic mice irrespective of having tumor tissues or no tumor. In tumor tissues, several c-Myc target genes, Foxm1, c-Jun, c-Fos, Bmi1 and Skp2, were also up-regulated dramatically. We determined whether mt-Rb activated the Myc promoter in the HTP9 cells and demonstrated that mt-Rb acted as an inhibitor of wild-type Rb-induced repression on the Myc promoter. Our results suggest that continued upregulation of c-Myc target genes promotes the liver tumor formation after about 1 year of age.

  18. Parallel folding topology-selective label-free detection and monitoring of conformational and topological changes of different G-quadruplex DNAs by emission spectral changes via FRET of mPPE-Ala-Pt(ii) complex ensembleElectronic supplementary information (ESI) available: Experimental details, electronic absorption spectra and resonance light scattering spectra of mPPE-Ala at different concentrations of 1; emission spectra and time-resolved emission decay profiles of mPPE-Ala at different concentrations of 1; parameters obtained from the emission spectra and time-resolved emission decay profiles of mPPE-Ala at different concentrations of 1; electronic absorption spectra of 1 at different concentrations of c-myc; circular dichroism (CD) spectrum of c-myc; electronic absorption spectra and UV melting profile of c-myc at different temperatures in the absence and in the presence of 1; table listing the UV melting temperatures of c-myc, bcl-2, c-kit1 and human telomeric DNA in the absence and in the presence of 1; emission spectra of mPPE-Ala-1 ensemble at different concentrations of c-myc-c, pre-formed duplex of c-myc and c-myc-c, pre-formed duplex of human telomeric DNA and complementary sequence of human telomeric DNA, bcl-2, c-kit1 and human telomeric DNA; CD spectra of pre-formed duplex of c-myc and c-myc-c at different concentrations of 1; emission spectra of 1 at different concentrations of c-myc; emission spectra of mPPE-Ala-1 ensemble at different concentrations of c-myc in the presence of 36% (v/v) PEG-200; changes in relative emission intensities of mPPE-Ala-1 ensemble at different concentrations of c-myc at different volume percentages of PEG-200; emission spectra of mPPE-Ala, 1 and mPPE-Ala-1 ensemble at different volume percentages of PEG-200; CD spectra of c-myc at different volume percentages of PEG-200; changes in relative emission intensities of mPPE-Ala-1 ensemble at different concentrations of human telomeric DNA at different volume percentages of PEG

    National Research Council Canada - National Science Library

    Chan, Kevin; Yik-Sham Chung, Clive; Wing-Wah Yam, Vivian

    2016-01-01

    ..., such as c-myc , in aqueous buffer solution. By the modulation of the aggregation/deaggregation of the polymer-metal complex aggregates and hence the FRET from the m PPE-Ala donor to the aggregated 1 as acceptor, the ensemble has been...

  19. Parallel folding topology-selective label-free detection and monitoring of conformational and topological changes of different G-quadruplex DNAs by emission spectral changes via FRET of mPPE-Ala-Pt(ii) complex ensembleElectronic supplementary information (ESI) available: Experimental details, electronic absorption spectra and resonance light scattering spectra of mPPE-Ala at different concentrations of 1; emission spectra and time-resolved emission decay profiles of mPPE-Ala at different concentrations of 1; parameters obtained from the emission spectra and time-resolved emission decay profiles of mPPE-Ala at different concentrations of 1; electronic absorption spectra of 1 at different concentrations of c-myc; circular dichroism (CD) spectrum of c-myc; electronic absorption spectra and UV melting profile of c-myc at different temperatures in the absence and in the presence of 1; table listing the UV melting temperatures of c-myc, bcl-2, c-kit1 and human telomeric DNA in the absence and in the presence of 1; emission spectra of mPPE-Ala-1 ensemble at different concentrations of c-myc-c, pre-formed duplex of c-myc and c-myc-c, pre-formed duplex of human telomeric DNA and complementary sequence of human telomeric DNA, bcl-2, c-kit1 and human telomeric DNA; CD spectra of pre-formed duplex of c-myc and c-myc-c at different concentrations of 1; emission spectra of 1 at different concentrations of c-myc; emission spectra of mPPE-Ala-1 ensemble at different concentrations of c-myc in the presence of 36% (v/v) PEG-200; changes in relative emission intensities of mPPE-Ala-1 ensemble at different concentrations of c-myc at different volume percentages of PEG-200; emission spectra of mPPE-Ala, 1 and mPPE-Ala-1 ensemble at different volume percentages of PEG-200; CD spectra of c-myc at different volume percentages of PEG-200; changes in relative emission intensities of mPPE-Ala-1 ensemble at different concentrations of human telomeric DNA at different volume percentages of PEG

    National Research Council Canada - National Science Library

    Chan, Kevin; Yik-Sham Chung, Clive; Wing-Wah Yam, Vivian

    2016-01-01

    ..., such as c-myc , in aqueous buffer solution. By the modulation of the aggregation/deaggregation of the polymer-metal complex aggregates and hence the FRET from the m PPE-Ala donor to the aggregated 1 as acceptor, the ensemble...

  20. Phospholipase D1 Couples CD4+ T Cell Activation to c-Myc-Dependent Deoxyribonucleotide Pool Expansion and HIV-1 Replication.

    Directory of Open Access Journals (Sweden)

    Harry E Taylor

    2015-05-01

    Full Text Available Quiescent CD4+ T cells restrict human immunodeficiency virus type 1 (HIV-1 infection at early steps of virus replication. Low levels of both deoxyribonucleotide triphosphates (dNTPs and the biosynthetic enzymes required for their de novo synthesis provide one barrier to infection. CD4+ T cell activation induces metabolic reprogramming that reverses this block and facilitates HIV-1 replication. Here, we show that phospholipase D1 (PLD1 links T cell activation signals to increased HIV-1 permissivity by triggering a c-Myc-dependent transcriptional program that coordinates glucose uptake and nucleotide biosynthesis. Decreasing PLD1 activity pharmacologically or by RNA interference diminished c-Myc-dependent expression during T cell activation at the RNA and protein levels. PLD1 inhibition of HIV-1 infection was partially rescued by adding exogenous deoxyribonucleosides that bypass the need for de novo dNTP synthesis. Moreover, the data indicate that low dNTP levels that impact HIV-1 restriction involve decreased synthesis, and not only increased catabolism of these nucleotides. These findings uncover a unique mechanism of action for PLD1 inhibitors and support their further development as part of a therapeutic combination for HIV-1 and other viral infections dependent on host nucleotide biosynthesis.

  1. SIRT1 activation by a c-MYC oncogenic network promotes the maintenance and drug resistance of human FLT3-ITD acute myeloid leukemia stem cells.

    Science.gov (United States)

    Li, Ling; Osdal, Tereza; Ho, Yinwei; Chun, Sookhee; McDonald, Tinisha; Agarwal, Puneet; Lin, Allen; Chu, Su; Qi, Jing; Li, Liang; Hsieh, Yao-Te; Dos Santos, Cedric; Yuan, Hongfeng; Ha, Trung-Quang; Popa, Mihaela; Hovland, Randi; Bruserud, Øystein; Gjertsen, Bjørn Tore; Kuo, Ya-Huei; Chen, Wenyong; Lain, Sonia; McCormack, Emmet; Bhatia, Ravi

    2014-10-02

    The FLT3-ITD mutation is frequently observed in acute myeloid leukemia (AML) and is associated with poor prognosis. In such patients, FLT3 tyrosine kinase inhibitors (TKIs) are only partially effective and do not eliminate the leukemia stem cells (LSCs) that are assumed to be the source of treatment failure. Here, we show that the NAD-dependent SIRT1 deacetylase is selectively overexpressed in primary human FLT3-ITD AML LSCs. This SIRT1 overexpression is related to enhanced expression of the USP22 deubiquitinase induced by c-MYC, leading to reduced SIRT1 ubiquitination and enhanced stability. Inhibition of SIRT1 expression or activity reduced the growth of FLT3-ITD AML LSCs and significantly enhanced TKI-mediated killing of the cells. Therefore, these results identify a c-MYC-related network that enhances SIRT1 protein expression in human FLT3-ITD AML LSCs and contributes to their maintenance. Inhibition of this oncogenic network could be an attractive approach for targeting FLT3-ITD AML LSCs to improve treatment outcomes. Copyright © 2014 Elsevier Inc. All rights reserved.

  2. miR-23b/SP1/c-myc forms a feed-forward loop supporting multiple myeloma cell growth.

    Science.gov (United States)

    Fulciniti, M; Amodio, N; Bandi, R L; Cagnetta, A; Samur, M K; Acharya, C; Prabhala, R; D'Aquila, P; Bellizzi, D; Passarino, G; Adamia, S; Neri, A; Hunter, Z R; Treon, S P; Anderson, K C; Tassone, P; Munshi, N C

    2016-01-15

    Deregulated microRNA (miR)/transcription factor (TF)-based networks represent a hallmark of cancer. We report here a novel c-Myc/miR-23b/Sp1 feed-forward loop with a critical role in multiple myeloma (MM) and Waldenstrom's macroglobulinemia (WM) cell growth and survival. We have found miR-23b to be downregulated in MM and WM cells especially in the presence of components of the tumor bone marrow milieu. Promoter methylation is one mechanism of miR-23b suppression in myeloma. In gain-of-function studies using miR-23b mimics-transfected or in miR-23b-stably expressing MM and WM cell lines, we observed a significant decrease in cell proliferation and survival, along with induction of caspase-3/7 activity over time, thus supporting a tumor suppressor role for miR-23b. At the molecular level, miR-23b targeted Sp1 3'UTR and significantly reduced Sp1-driven nuclear factor-κB activity. Finally, c-Myc, an important oncogenic transcription factor known to stimulate MM cell proliferation, transcriptionally repressed miR-23b. Thus MYC-dependent miR-23b repression in myeloma cells may promote activation of oncogenic Sp1-mediated signaling, representing the first feed-forward loop with critical growth and survival role in myeloma.

  3. Expression of c-erbB-2, p53 and c-myc proteins in male breast carcinoma: Comparison with traditional prognostic factors and survival

    Directory of Open Access Journals (Sweden)

    Mourão Netto M.

    2001-01-01

    Full Text Available There are few data evaluating biological markers for men with breast cancer. The purpose of the present study was to analyze the expression of the oncogenes c-erbB-2 and c-myc and of the suppressor gene p53 by immunohistochemical techniques in archival paraffin-embedded tissue blocks of 48 male breast cancer patients, treated at the A.C. Camargo Cancer Hospital, São Paulo, SP, Brazil. The results were compared with clinicopathological prognostic features. Immunopositivity of c-erbB-2, p53 and c-myc was detected in 62.5, 16.7 and 20.8% of the cases analyzed, respectively. Estrogen and progesterone receptors were positive in 75 and 69% of the cases, respectively. Increasing staging was statistically associated with c-erbB-2 (P = 0.04 and weakly related to p53 positivity (P = 0.06. No significant correlation between specific survival rate (determined by the log rank test and the molecular markers analyzed was found, whereas the number of compromised lymph nodes and advanced TNM (tumor, node, metastasis staging were associated with diminished survival.

  4. Expression Analysis of p16, c-Myc, and mSin3A in Non-small Cell Lung Cancer by Computer Aided Scoring and Analysis (CASA).

    Science.gov (United States)

    Salmaninejad, Arash; Estiar, Mehrdad Asghari; Gill, Rajbir K; Shih, Joanna H; Hewitt, Stephen; Jeon, Hyo-Sung; Fukuoka, Junya; Shilo, Konstantin; Shakoori, Abbas; Jen, Jin

    2015-01-01

    Immunohistochemical analysis (IHC) of tissue microarray (TMA) slides enables large sets of tissue samples to be analyzed simultaneously on a single slide. However, manual evaluation of small cores on a TMA slide is time consuming and error prone. We describe a computer aided scoring and analysis (CASA) method to allow facile and reliable scoring of IHC staining using TMA containing 300 non-small cell lung cancer (NSCLC) cases. In the two previous published papers utilizing our TMA slides of lung cancer we examined 18 proteins involved in the chromatin machinery. We developed our study using more proteins of the chromatin complex and several transcription factors that facilitate the chromatin machinery. Then, a total of 78 antibodies were evaluated by CASA to derive a normalized intensity value that correlated with the overall staining status of the targeting protein. The intensity values for TMA cores were then examined for association to clinical variables and predictive significance individually and with other factors. RESULTs: Using our TMA, the intensity of several protein pairs were significantly correlated with an increased risk of death in NSCLC. These included c-Myc with p16, mSin3A with p16 and c-Myc with mSinA. Predictive values of these pairs remained significant when evaluated based on standard IHC scores. Our results demonstrate the usefulness of CASA as a valuable tool for systematic assessment of TMA slides to identify potential predictive biomarkers using a large set of primary human tissues.

  5. Real-time monitoring of PCR amplification of proto-oncogene c-MYC using a Ta₂O₅ electrolyte-insulator-semiconductor sensor.

    Science.gov (United States)

    Branquinho, Rita; Veigas, Bruno; Pinto, Joana V; Martins, Rodrigo; Fortunato, Elvira; Baptista, Pedro V

    2011-10-15

    We present a new approach for real-time monitoring of PCR amplification of a specific sequence from the human c-MYC proto-oncogene using a Ta(2)O(5) electrolyte-insulator-semiconductor (EIS) sensor. The response of the fabricated EIS sensor to cycle DNA amplification was evaluated and compared to standard SYBR-green fluorescence incorporation, showing it was possible to detect DNA concentration variations with 30 mV/μM sensitivity. The sensor's response was then optimized to follow in real-time the PCR amplification of c-MYC sequence from a genomic DNA sample attaining an amplification profile comparable to that of a standard real-time PCR. Owing to the small size, ease of fabrication and low-cost, the developed Ta(2)O(5) sensor may be incorporated onto a microfluidic device and then used for real-time PCR. Our approach may circumvent the practical and economical obstacles posed by current platforms that require an external fluorescence detector difficult to miniaturize and incorporate into a lab-on-chip system. Copyright © 2011 Elsevier B.V. All rights reserved.

  6. Elevated proto-oncogene and collagen mRNA expression in PVR retinas.

    Science.gov (United States)

    Hollborn, Margrit; Faude, Frank; Wiedemann, Peter; Kohen, Leon

    2003-05-01

    Retinal detachment is often accompanied by proliferation and migration of retinal cells and by increased synthesis of structural proteins, known as proliferative vitreoretinopathy (PVR). Herein we investigate the messenger RNA (mRNA) expression of proto-oncogenes responsible for cell proliferation and of structural proteins that have a role in membrane formation. Retinal samples were obtained from patients undergoing vitreoretinal surgery for the treatment of retinal detachment complicated by PVR. Normal human control retinas were obtained from cornea donors. The mRNA expression of the proto-oncogenes c- myc, c- fos and the proliferation marker Ki67, as well as of collagen type III and type IV, were investigated using the ribonuclease protection assay. Ki67 mRNA expression was not detectable in either sample type, but c- fos and c- myc mRNA expression was found in normal and PVR retinas. Whereas the expression of c- myc showed a marginal increase, the up-regulation in c- fos expression was strongly significant (5.07-fold). The mRNA of collagen type III was detectable at widely varying levels in all the PVR retinas but was found in only 2 of the 16 analysed normal samples. Collagen type IV mRNA was expressed in both PVR and control samples but was higher (2.21-fold) in the PVR retinas. These results indicate that an up-regulation of the proto-oncogene c- fos occurs in human PVR retinas. An increase in mRNA expression of collagen types III and IV takes place simultaneously. These changes in mRNA expression appear to be mainly connected to the initiation of cell proliferation, dedifferentiation and formation of tractional membranes.

  7. Histone deacetylase inhibitor romidepsin induces efficient tumor cell lysis via selective down-regulation of LMP1 and c-myc expression in EBV-positive diffuse large B-cell lymphoma.

    Science.gov (United States)

    Shin, Dong-Yeop; Kim, Areumnuri; Kang, Hye Jin; Park, Sunhoo; Kim, Dong Wan; Lee, Seung-Sook

    2015-08-10

    We investigated the role of the histone deacetylase inhibitor, romidepsin, in Epstein-Barr virus (EBV)-positive diffuse large B-cell lymphoma (DLBCL), an aggressive non-Hodgkin lymphoma with poor clinical outcomes. We used EBV-positive and EBV-negative DLBCL cell lines and generated two EBV-transfected cell lines, LY7/EBV and U2932/EBV. Romidepsin was cytotoxic to cultured EBV-positive cells via the activation of the caspase cascade. Moreover, in vivo mice xenograft models demonstrated the cytotoxicity of romidepsin to EBV-positive DLBCL cells. Romidepsin induced cytotoxicity via the reduction of LMP1 and c-myc expression in EBV-positive cells. Inhibiting either LMP1 or c-myc using small inhibitory RNAs caused partial cytotoxicity in EBV-positive Farage and U2932/EBV lines. The dual inhibition of LMP1 and c-myc showed a synergistic cytotoxic effect in EBV-positive cells similar in magnitude to that of romidepsin alone. In addition, either double blockade of LMP1 and c-myc activity or romidepsin single treatment activated EBV lytic cycle in EBV-positive cells. In conclusion, romidepsin exerts strong anti-tumor activity in EBV-positive DLBCL via the inhibition of both LMP1 and c-myc. Our findings indicate that romidepsin might be a promising treatment for EBV-positive DLBCL. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  8. PI3Kδ activates E2F1 synthesis in response to mRNA translation stress.

    Science.gov (United States)

    Gnanasundram, Sivakumar Vadivel; Pyndiah, Slovénie; Daskalogianni, Chrysoula; Armfield, Kate; Nylander, Karin; Wilson, Joanna B; Fåhraeus, Robin

    2017-12-13

    The c-myc oncogene stimulates ribosomal biogenesis and protein synthesis to promote cellular growth. However, the pathway by which cells sense and restore dysfunctional mRNA translation and how this is linked to cell proliferation and growth is not known. We here show that mRNA translation stress in cis triggered by the gly-ala repeat sequence of Epstein-Barr virus (EBV)-encoded EBNA1, results in PI3Kδ-dependent induction of E2F1 mRNA translation with the consequent activation of c-Myc and cell proliferation. Treatment with a specific PI3Kδ inhibitor Idelalisib (CAL-101) suppresses E2F1 and c-Myc levels and causes cell death in EBNA1-induced B cell lymphomas. Suppression of PI3Kδ prevents E2F1 activation also in non-EBV-infected cells. These data illustrate an mRNA translation stress-response pathway for E2F1 activation that is exploited by EBV to promote cell growth and proliferation, offering new strategies to treat EBV-carrying cancers.

  9. Down-regulation of 5S rRNA by miR-150 and miR-383 enhances c-Myc-rpL11 interaction and inhibits proliferation of esophageal squamous carcinoma cells.

    Science.gov (United States)

    Wang, Xinyu; Ren, Yanli; Wang, Zhiqiong; Xiong, Xiangyu; Han, Sichong; Pan, Wenting; Chen, Hongwei; Zhou, Liqing; Zhou, Changchun; Yuan, Qipeng; Yang, Ming

    2015-12-21

    5S rRNA plays an important part in ribosome biology and is over-expression in multiple cancers. In this study, we found that 5S rRNA is a direct target of miR-150 and miR-383 in esophageal squamous cell carcinoma (ESCC). Overexpression of miR-150 and miR-383 inhibited ESCC cell proliferation in vitro and in vivo. Moreover, 5S rRNA silencing by miR-150 and miR-383 might intensify rpL11-c-Myc interaction, which attenuated role of c-Myc as an oncogenic transcriptional factor and dysregulation of multiple c-Myc target genes. Taken together, our results highlight the involvement of miRNAs in ribosomal regulation during tumorigenesis. Copyright © 2015 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  10. Concordant genetic distinctness of the phylogroup of the Siberian chipmunk from the Korean peninsula (Tamias sibiricus barberi), reexamined with nuclear DNA c-myc gene exon 2 and mtDNA control region sequences.

    Science.gov (United States)

    Koh, Hung Sun; Zhang, Minghai; Bayarlkhagva, Damdingiin; Ham, Eui Jeong; Kim, Jin Seong; Jang, Kyung Hee; Park, Nam Jeong

    2010-08-01

    We reexamined Tamias sibiricus barberi from Korea by sequencing c-myc exon 2 and the mtDNA control region. In the c-myc exon, the monogenic T. s. barberi differed from the monogenic T. s. orientalis (nucleotide distance 0.48%; 3 variable sites at 168, 306, and 552), whereas T. s. orientalis was identical to T. s. sibiricus. In the control region, T. s. barberi differed from T. s. orientalis (distance 6.84%) and T. s. sibiricus (9.35%). We considered the concordant, extensive gaps between the phylogroup of T. s. barberi and other subspecies of T. sibiricus in the c-myc gene, control region, and cytochrome b gene to be evidence of a lack of intergradation through North Korea from T. s. barberi to T. s. orientalis. Our results, showing the genetic and morphological distinctness of T. s. barberi, support that this phylogroup is a distinct species.

  11. Orientation and position of avian leukosis virus DNA relative to the cellular oncogene c-myc in B-lymphoma tumors of highly susceptible 15I5 X 7(2) chickens.

    Science.gov (United States)

    Fung, Y K; Crittenden, L B; Kung, H J

    1982-01-01

    We previously reported our characterizations of the B-lymphoma tumors induced in a highly susceptible line of chickens (15I5 X 7(2)) by the avian leukosis virus RAV-1 (Proc. Natl. Acad. Sci. 78:3418-3422, 1981). We demonstrated that in greater than 90% of the tumors, the RAV-1 provirus is integrated near a cellular oncogene, c-myc. In the present study, we devised a simple approach, relying on SalI digestion, for further defining the locations and orientations of the proviruses with respect to the c-myc gene. We report here that in the great majority of cases the provirus is situated upstream from and in the same transcriptional direction as the c-myc gene--a configuration compatible with the promoter-insertion model proposed by Hayward et al. (Nature [London] 290:475-480, 1981). Images PMID:6292531

  12. Incidence of HPV Infection in Oral Squamous Cell Carcinoma and Its Association with the Presence of p53 & c-myc Mutation : A Case Control Study in Muwardi Hospital Surakarta

    Directory of Open Access Journals (Sweden)

    Adi Prayitno

    2012-12-01

    Full Text Available Introduction: Annual incidence rates for oral and pharyngeal cancer are estimated at 25 cases per 100,000 in developing countries. Human papilloma virus (HPV was implicated in pathogenesis of Cancer. The mutations of p53 and c-myc are found 50% in cancer. Objective: Aims of this research were to know the incidence of OSSC patient which realized HPV infection without p53 and c-myc gene mutation. Materials and Methods: Tissue biopsy frozen sections were taken from BOSC (Benign Oral Squamous Cell and OSCC (Oral Squamous Cell Carcinoma patients collected from Oral and Dental Departement of dr Muwardi Distric Hospital in Surakarta from January 2007 to January 2008. To amplify L1-HPV gene for fixed the HPV stressor. To amplify p53 and c-myc genes, continued with SSCP (Single Strand Conformational Polymorphisme analysis and followed with measurement using densitometer, to see mutation existence. The collected data were analyzed with Chi Square. Results: BOSC patient identified 23% with HPV infections and OSCC patient identified 73% with HPV infections. Hundred percent BOSC patient with HPV infection without mutation in p53 gene and c-myc gene, 81% OSCC patient with HPV infection without mutation in p53 gene and 91% OSCC patient with HPV infection without mutation in c-myc gene. Chi  square analysis showed significant difference between BOSC and OSCC patients with HPV infection without mutation in p53 and c-myc gene. Conclusion: HPV is a factor for pathogenesis of OSCC.DOI: 10.14693/jdi.v17i2.44

  13. Tumor-suppressive microRNA-135a inhibits cancer cell proliferation by targeting the c-MYC oncogene in renal cell carcinoma.

    Science.gov (United States)

    Yamada, Yasutoshi; Hidaka, Hideo; Seki, Naohiko; Yoshino, Hirofumi; Yamasaki, Takeshi; Itesako, Toshihiko; Nakagawa, Masayuki; Enokida, Hideki

    2013-03-01

    Recently, many studies have suggested that microRNAs (miRNAs) are involved in cancer cell development, invasion, and metastasis of various types of human cancers. In a previous study, miRNA expression signatures from renal cell carcinoma (RCC) revealed that expression of microRNA-135a (miR-135a) was significantly reduced in cancerous tissues. The aim of this study was to investigate the functional significance of miR-135a and to identify miR-135a-mediated molecular pathways in RCC cells. Restoration of mature miR-135a significantly inhibited cancer cell proliferation and induced G0 /G1 arrest in the RCC cell lines caki2 and A498, suggesting that miR-135a functioned as a potential tumor suppressor. We then examined miR-135a-mediated molecular pathways using genome-wide gene expression analysis and in silico analysis. A total of 570 downregulated genes were identified in miR-135a transfected RCC cell lines. To investigate the biological significance of potential miR-135a-mediated pathways, we classified putative miR-135a-regulated genes according to the Kyoto Encyclopedia of Genes and Genomics pathway database. From our in silico analysis, 25 pathways, including the cell cycle, pathways in cancer, DNA replication, and focal adhesion, were significantly regulated by miR-135a in RCC cells. Moreover, based on the results of this analysis, we investigated whether miR-135a targeted the c-MYC gene in RCC. Gain-of-function and luciferase reporter assays showed that c-MYC was directly regulated by miR-135a in RCC cells. Furthermore, c-MYC expression was significantly upregulated in RCC clinical specimens. Our data suggest that elucidation of tumor-suppressive miR-135a-mediated molecular pathways could reveal potential therapeutic targets in RCC. © 2012 Japanese Cancer Association.

  14. Redox regulation of cardiomyocyte cell cycling via an ERK1/2 and c-Myc-dependent activation of cyclin D2 transcription

    Science.gov (United States)

    Murray, Thomas V.A.; Smyrnias, Ioannis; Schnelle, Moritz; Mistry, Rajesh K.; Zhang, Min; Beretta, Matteo; Martin, Daniel; Anilkumar, Narayana; de Silva, Shana M.; Shah, Ajay M.; Brewer, Alison C.

    2015-01-01

    Adult mammalian cardiomyocytes have a very limited capacity to proliferate, and consequently the loss of cells after cardiac stress promotes heart failure. Recent evidence suggests that administration of hydrogen peroxide (H2O2), can regulate redox-dependent signalling pathway(s) to promote cardiomyocyte proliferation in vitro, but the potential relevance of such a pathway in vivo has not been tested. We have generated a transgenic (Tg) mouse model in which the H2O2-generating enzyme, NADPH oxidase 4 (Nox4), is overexpressed within the postnatal cardiomyocytes, and observed that the hearts of 1–3 week old Tg mice pups are larger in comparison to wild type (Wt) littermate controls. We demonstrate that the cardiomyocytes of Tg mouse pups have increased cell cycling capacity in vivo as determined by incorporation of 5-bromo-2′-deoxyuridine. Further, microarray analyses of the transcriptome of these Tg mouse hearts suggested that the expression of cyclin D2 is significantly increased. We investigated the molecular mechanisms which underlie this more proliferative phenotype in isolated neonatal rat cardiomyocytes (NRCs) in vitro, and demonstrate that Nox4 overexpression mediates an H2O2-dependent activation of the ERK1/2 signalling pathway, which in turn phosphorylates and activates the transcription factor c-myc. This results in a significant increase in cyclin D2 expression, which we show to be mediated, at least in part, by cis-acting c-myc binding sites within the proximal cyclin D2 promoter. Overexpression of Nox4 in NRCs results in an increase in their proliferative capacity that is ablated by the silencing of cyclin D2. We further demonstrate activation of the ERK1/2 signalling pathway, increased phosphorylation of c-myc and significantly increased expression of cyclin D2 protein in the Nox4 Tg hearts. We suggest that this pathway acts to maintain the proliferative capacity of cardiomyocytes in Nox4 Tg pups in vivo and so delays their exit from the cell

  15. DJ-1, an oncogene and causative gene for familial Parkinson's disease, is essential for SV40 transformation in mouse fibroblasts through up-regulation of c-Myc.

    Science.gov (United States)

    Kim, Yun Chul; Kitaura, Hirotake; Iguchi-Ariga, Sanae M M; Ariga, Hiroyoshi

    2010-09-24

    Simian virus 40 (SV40) is a tumor virus and its early gene product large T-antigen (LT) is responsible for the transforming activity of SV40. Parkinson's disease causative gene DJ-1 is also a ras-dependent oncogene, but the mechanism of its oncogene function is still not known. In this study, we found that there were no transformed foci when fibroblasts from DJ-1-knockout mice were transfected with LT. We also found that DJ-1 directly bound to LT and that the expression level of c-Myc in transformed cells was parallel to that of DJ-1. These findings indicate that DJ-1 is essential for SV40 transformation. Copyright © 2010 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  16. Protein kinase A antagonist inhibits β-catenin nuclear translocation, c-Myc and COX-2 expression and tumor promotion in ApcMin/+ mice

    Directory of Open Access Journals (Sweden)

    Brudvik Kristoffer W

    2011-12-01

    Full Text Available Abstract Background The adenomatous polyposis coli (APC protein is part of the destruction complex controlling proteosomal degradation of β-catenin and limiting its nuclear translocation, which is thought to play a gate-keeping role in colorectal cancer. The destruction complex is inhibited by Wnt-Frz and prostaglandin E2 (PGE2 - PI-3 kinase pathways. Recent reports show that PGE2-induced phosphorylation of β-catenin by protein kinase A (PKA increases nuclear translocation indicating two mechanisms of action of PGE2 on β-catenin homeostasis. Findings Treatment of ApcMin/+ mice that spontaneously develop intestinal adenomas with a PKA antagonist (Rp-8-Br-cAMPS selectively targeting only the latter pathway reduced tumor load, but not the number of adenomas. Immunohistochemical characterization of intestines from treated and control animals revealed that expression of β-catenin, β-catenin nuclear translocation and expression of the β-catenin target genes c-Myc and COX-2 were significantly down-regulated upon Rp-8-Br-cAMPS treatment. Parallel experiments in a human colon cancer cell line (HCT116 revealed that Rp-8-Br-cAMPS blocked PGE2-induced β-catenin phosphorylation and c-Myc upregulation. Conclusion Based on our findings we suggest that PGE2 act through PKA to promote β-catenin nuclear translocation and tumor development in ApcMin/+ mice in vivo, indicating that the direct regulatory effect of PKA on β-catenin nuclear translocation is operative in intestinal cancer.

  17. Genetic dissimilarity between primary colorectal carcinomas and their lymph node metastases: ploidy, p53, bcl-2, and c-myc expression--a pilot study.

    Science.gov (United States)

    Zalata, Khaled Refaat; Elshal, Mohamed Farouk; Foda, Abd AlRahman Mohammad; Shoma, Ashraf

    2015-08-01

    The current paradigm of metastasis proposes that rare cells within primary tumors acquire metastatic capability via sequential mutations, suggesting that metastases are genetically dissimilar from their primary tumors. This study investigated the changes in the level of expression of a well-defined panel of cell proliferation, differentiation, and apoptosis markers between the primary colorectal cancer (CRC) and the corresponding synchronous lymph node (LN) metastasis from the same patients. DNA flow cytometry and immunostaining of p53, bcl-2, and c-myc were carried out on 36 cases of CRC radical resection specimens with their corresponding LN metastases. There was very low probability that the histological patterns of primary tumors and LN metastases are independent (p < 0.001). Metastatic tumors were significantly more diffusely positive for p53 than the primary tumors (p < 0.001). Conversely, primary tumors were significantly more diffusely positive for c-myc than metastatic tumors (p = 0.011). No significant difference was found between the LNs and the primary tumors in bcl-2 positivity (p = 0.538) and DNA aneuploidy (p = 0.35), with a tendency towards negative bcl-2 and less aneuploidy in LN metastases than primary tumors. In conclusion, LN metastatic colorectal carcinomas have a tendency of being less differentiated, with a higher incidence of diffuse p53 staining, lower incidence of bcl-2 staining, and less aneuploidy in comparison to their primary counterparts suggesting a more aggressive biological behavior, which could indicate the necessity for more aggressive adjuvant therapy.

  18. Predictive value of BRCA1, ERCC1, ATP7B, PKM2, TOPOI, TOPΟ-IIA, TOPOIIB and C-MYC genes in patients with small cell lung cancer (SCLC who received first line therapy with cisplatin and etoposide.

    Directory of Open Access Journals (Sweden)

    Niki Karachaliou

    Full Text Available BACKGROUND: The aim of the study was to evaluate the predictive value of genes involved in the action of cisplatin-etoposide in Small Cell Lung Cancer (SCLC. METHODS: 184 SCLC patients' primary tumour samples were analyzed for ERCCI, BRCA1, ATP7B, PKM2 TOPOI, TOPOIIA, TOPOIIB and C-MYC mRNA expression. All patients were treated with cisplatin-etoposide. RESULTS: The patients' median age was 63 years and 120 (65% had extended stage, 75 (41% had increased LDH serum levels and 131 (71% an ECOG performance status was 0-1. Patients with limited stage, whose tumours expressed high ERCC1 (p=0.028, PKM2 (p=0.046, TOPOI (p=0.008, TOPOIIA (p=0.002 and TOPOIIB (p<0.001 mRNA had a shorter Progression Free Survival (PFS. In limited stage patients, high expression of ERCC1 (p=0.014, PKM2 (p=0.026, TOPOIIA (p=0.021 and TOPOIIB (p=0.019 was correlated with decreased median overall survival (mOS while in patients with extended stage, only high TOPOIIB expression had a negative impact on Os (p=0.035. The favorable expression signature expression signature (low expression of ERCC1, PKM2, TOPOIIA and TOPOIIB was correlated with significantly better PFS and Os in both LS-SCLC (p<0.001 and p=0.007, respectively and ES-SCLC (p=0.007 and (p=0.011, respectively group. The unfavorable expression signature was an independent predictor for poor PFS (HR: 3.18; p=0.002 and HR: 3.14; p=0.021 and Os (HR: 4.35; p=0.001and HR: 3.32; p=0.019 in both limited and extended stage, respectively. CONCLUSIONS: Single gene's expression analysis as well as the integrated analysis of ERCC1, PKM2, TOPOIIA and TOPOIIB may predict treatment outcome in patients with SCLC. These findings should be further validated in a prospective study.

  19. The 5' untranslated mRNA region base content can greatly affect translation initiation in the absence of secondary structures in Prevotella bryantii TC1-1.

    Science.gov (United States)

    Seničar, Lenart; Accetto, Tomaž

    2015-01-01

    It has become clear lately that many bacteria and even whole bacterial phyla do not use the classical Shine-Dalgarno sequence mediated pathway of protein translation initiation. The prominent phylum Bacteroidetes is one of them, and this was shown not only using bioinformatic but also functional reporter gene studies in its representative Prevotella bryantii. The latter studies revealed much higher sensitivity toward secondary structures in 5(') untranslated mRNA regions (5(') UTRs) during translation initiation compared to Escherichia coli. It was proposed that in the absence of Shine-Dalgarno sequence interaction the key elements enabling translation initiation are local absence of secondary structures in 5(') UTRs, and the ribosomal protein S1 which binds to mRNA. Here, we evaluate the 5(') UTRs devoid of secondary structures but containing divergent nucleotide compositions in P. bryantii reporter assay. We show that base composition profoundly affects the amount of the reporter synthesized, and further that these amounts were in agreement with S1 protein binding affinity for adenine/uracil bases in mRNA. This is the first, though indirect, clue that S1 is actually involved in translation initiation in Bacteroidetes and adds the second layer of control beside mRNA secondary structure affecting translation initiation in this phylum. © FEMS 2014. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  20. Snipper, an Eri1 homologue, affects histone mRNA abundance and is crucial for normal Drosophila melanogaster development.

    Science.gov (United States)

    Alexiadis, Anastasios; Delidakis, Christos; Kalantidis, Kriton

    2017-07-01

    The conserved 3'-5' RNA exonuclease ERI1 is implicated in RNA interference inhibition, 5.8S rRNA maturation and histone mRNA maturation and turnover. The single ERI1 homologue in Drosophila melanogaster Snipper (Snp) is a 3'-5' exonuclease, but its in vivo function remains elusive. Here, we report Snp requirement for normal Drosophila development, since its perturbation leads to larval arrest and tissue-specific downregulation results in abnormal tissue development. Additionally, Snp directly interacts with histone mRNA, and its depletion results in drastic reduction in histone transcript levels. We propose that Snp protects the 3'-ends of histone mRNAs and upon its absence, histone transcripts are readily degraded. This in turn may lead to cell cycle delay or arrest, causing growth arrest and developmental perturbations. © 2017 Federation of European Biochemical Societies.

  1. High intensity interval training favourably affects antioxidant and inflammation mRNA expression in early-stage chronic kidney disease.

    Science.gov (United States)

    Tucker, Patrick S; Briskey, David R; Scanlan, Aaron T; Coombes, Jeff S; Dalbo, Vincent J

    2015-12-01

    Increased levels of oxidative stress and inflammation have been linked to the progression of chronic kidney disease. To reduce oxidative stress and inflammation related to chronic kidney disease, chronic aerobic exercise is often recommended. Data suggests high intensity interval training may be more beneficial than traditional aerobic exercise. However, appraisals of differing modes of exercise, along with explanations of mechanisms responsible for observed effects, are lacking. This study assessed effects of eight weeks of high intensity interval training (85% VO2max), versus low intensity exercise (45-50% VO2max) and sedentary behaviour, in an animal model of early-stage chronic kidney disease. We examined kidney-specific mRNA expression of genes related to endogenous antioxidant enzyme activity (glutathione peroxidase 1; Gpx1, superoxide dismutase 1; Sod1, and catalase; Cat) and inflammation (kidney injury molecule 1; Kim1 and tumour necrosis factor receptor super family 1b; Tnfrsf1b), as well as plasma F2-isoprostanes, a marker of lipid peroxidation. Compared to sedentary behaviour, high intensity interval training resulted in increased mRNA expression of Sod1 (p=0.01) and Cat (pintensity exercise, high intensity interval training resulted in increased mRNA expression of Cat (phigh intensity interval training was superior to sedentary behaviour and low intensity exercise as high intensity interval training beneficially influenced expression of genes related to endogenous antioxidant enzyme activity and inflammation. Copyright © 2015 Elsevier Inc. All rights reserved.

  2. Controlled and localized delivery of c-myc AS-ODN to cells by 3-aminopropyl-trimethoxylsilane modified SBA-15 mesoporous silica

    Science.gov (United States)

    Zhang, Juan; Chen, Minmin; Zhao, Xiqiu; Zhang, Min; Mao, Jinxiang; Cao, Xichuan; Zhang, Zhuoqi

    2018-01-01

    SBA-15 mesoporous silicate was synthesized and functionalized with 3-aminopropyl organic groups through a post-synthesis method. The materials were characterized consecutively by powder X-ray diffraction (XRD), N2 adsorption/desorption analysis and solid-state magic-angle spinning 29Si nuclear magnetic resonance (MAS NMR). Human c-myc anti-sense oligodeoxyneucleotide (AS-ODN) was selected as a model molecule to be loaded onto the surface of bare and functionalized SBA-15 via different loading conditions. It has been found that the amount of AS-ODN incorporated into the porous matrix is strongly dependent on the surface properties, pH of the loading solvent and AS-ODN concentration. The release behaviour of AS-ODN from modified SBA-15 materials was also investigated and depended on conditions chosen. Cellular uptake of the eluted AS-ODN into Hela cells was observed by fluorescent microscopy. The materials showed excellent cytocompatibility. The AS-ODN keeps full transfection and expression activities indicating its structural integrity. The functionalized SBA-15 is an excellent prospect as a biomedical material candidate for the future.

  3. Melatonin promotes circadian rhythm-induced proliferation through Clock/histone deacetylase 3/c-Myc interaction in mouse adipose tissue.

    Science.gov (United States)

    Liu, Zhenjiang; Gan, Lu; Luo, Dan; Sun, Chao

    2017-05-01

    Melatonin is synthesized in the pineal gland and controls circadian rhythm of peripheral adipose tissue, resulting in changes in body weight. Although core regulatory components of clock rhythmicity have been defined, insight into the mechanisms of circadian rhythm-mediated proliferation in adipose tissue is still limited. Here, we showed that melatonin (20 mg/kg/d) promoted circadian and proliferation processes in white adipose tissue. The circadian amplitudes of brain and muscle aryl hydrocarbon receptor nuclear translocator-like 1 (Bmal1, Pmelatonin in adipose tissue. Melatonin also promoted cell cycle and increased cell numbers (PMelatonin also attenuated circadian disruption and promoted adipocyte proliferation in chronic jet-lagged mice and obese mice. Thus, our study found that melatonin promoted adipocyte proliferation by forming a Clock/HDAC3/c-Myc complex and subsequently driving the circadian amplitudes of proliferation genes. Our data reveal a novel mechanism that links circadian rhythm to cell proliferation in adipose tissue. These findings also identify a new potential means for melatonin to prevent and treat sleep deprivation-caused obesity. © 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  4. Candidate tumour suppressor CCDC19 regulates miR-184 direct targeting of C-Myc thereby suppressing cell growth in non-small cell lung cancers.

    Science.gov (United States)

    Liu, Zhen; Mai, Chunping; Yang, Huiling; Zhen, Yan; Yu, Xiaoli; Hua, Shengni; Wu, Qiangyun; Jiang, Qinping; Zhang, Yajie; Song, Xin; Fang, Weiyi

    2014-08-01

    We previously reported and revised the nasopharyngeal epithelium specific protein CCDC19 and identified it as a potential tumour suppressor in nasopharyngeal carcinoma. The purpose of this study was to investigate the involvement of CCDC19 in the pathogenesis of human non-small cell lung cancers (NSCLC). Down-regulated CCDC19 expression was observed in NSCLC tissues and cells compared to normal tissues. However, reduced protein expression did not correlate with the status of NSCLC progression. Instead, we observed that patients with lower CCDC19 expression had a shorter overall survival than did patients with higher CCDC19 expression. Lentiviral-mediated CCDC19 overexpression significantly suppressed cell proliferation and cell cycle transition from G1 to S and G2 phases in NSCLC cells. Knocking down CCDC19 expression significantly restored the ability of cell growth in CCDC19 overexpressing NSCLC cells. Mechanistically CCDC19 functions as a potential tumour suppressor by stimulating miR-184 suppression of C-Myc thus blocking cell growth mediated by the PI3K/AKT/C-Jun pathway. Our studies are the first to demonstrate that reduced expression of CCDC19 is an unfavourable factor in NSCLC. © 2014 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  5. [Effects of lithium chloride and harringtonine on the differentiation, proliferation and c-myc proto-oncogene expression of HL-60 cells].

    Science.gov (United States)

    Li, W; Jiang, D; Tan, M

    1997-05-01

    This research was to observe the effects of lithium chloride (LiCl) and Harringtonine (HT) on the proliferation and differentiation of HL-60 leukemia cells. The results obtained by liquid suspension culture, semi-solid colony culture and 3H-TdR incorporation into HL-60 cells indicated that different concentrations of LiCl (5-20 mmol/L) and HT (10(-8)-10(-5)mol/L) exerted the inhibitory effects in a dose-dependent manner on HL-60 cell proliferation respectively. When LiCl (10 mmol/L) and HT (10(-7) mol/L) were added together in the liquid culture or semi-solid culture of HL-60 cells, they showed much greater inhibitory effect than that by each agent separately. It was discovered that there was induction of the differentiation of HL-60 cells by lithium and HT and the induction of HL-60 cells differentiation by HT was markedly enhanced by the addition of low concentration of lithium. This work also showed that by treating HL-60 cells with lithium and HT, the expression of the c-myc proto-oncogene was markedly decreased as measured by RT/PCR-mRNA (P lithium and HT in the treatment of leukemia and in vitro purging of leukemic cells for autologous bone marrow transplantation.

  6. Synthetic artificial microRNAs targeting UCA1-MALAT1 or c-Myc inhibit malignant phenotypes of bladder cancer cells T24 and 5637.

    Science.gov (United States)

    Fu, Xing; Liu, Yuchen; Zhuang, Chengle; Liu, Li; Cai, Zhiming; Huang, Weiren

    2015-05-01

    The biggest concern of using natural microRNAs for treating cancer is that they usually cause few phenotypic changes due to the divergent functions of their target genes. Based on the engineering principles of synthetic biology, we provided a standard platform for constructing artificial microRNAs that can target one or few specific genes and silence both protein-coding genes and long non-coding genes. To prove the utility of this platform, we chose MALAT1, UCA1, and c-Myc as the functional targets and used the bladder cancer cell lines T24 and 5637 as the test models. The relative expression level of the target genes was measured by qRT-PCR. Cell proliferation and migration were determined by MTT assay and wound-healing assay, respectively. Cell apoptosis was revealed by both Hoechst 33258 staining assay and ELISA assay. We found that the artificial microRNAs can effectively silence their target genes and induce anti-cancer effects in T24 and 5637 cells. These devices can inhibit proliferation, induce apoptosis, and suppress migration of the two bladder cancer cell lines. The synthetic artificial microRNAs may represent a kind of novel genetic devices for treating human bladder cancer.

  7. [Amplification of the erbb-2 (Her-2/NEU), erbb-1 (HER-1) and c-myc oncogenes is often combined with the deletion of the short arm of chromosome 17 in human carcinoma].

    Science.gov (United States)

    Imianitov, E N; Chernitsa, O I; Nikiforova, I F; Serova, O M; Sokolov, S I; Laur, O Iu; Togo, A V; Kniazev, P G

    1993-01-01

    Amplification of oncogenes erbb-2, erbb-1, c-myc and losses of heterozygosity (LOH) at chromosomes 11p (probe hras-1), 17p (probe ynz-22) and 17q (probe thh-59) were studied in 165 human tumours (60 breast, 22 ovary, 40 colorectal, 23 lung, and 20 thyroid carcinomas). The correlation (P < 0.01) between the increased copy number of mentioned oncogenes and LOH at 17p was demonstrated for tumours tested: extra copies of these oncogenes were revealed in 11 of 46 DNA specimens with LOH on ynz-22, but only in 3 of 61 without LOH. This correlation was mostly due to frequent combinations between erbb-2 amplification and 17p deletions; the incidence of increased copy number of erbb-1 and c-myc oncogenes was not high enough for final conclusions about the association of their alterations with LOH at chromosome 17p.

  8. Primary central nervous system diffuse large B-cell lymphoma shows an activated B-cell-like phenotype with co-expression of C-MYC, BCL-2, and BCL-6.

    Science.gov (United States)

    Li, Xiaomei; Huang, Ying; Bi, Chengfeng; Yuan, Ji; He, Hong; Zhang, Hong; Yu, QiuBo; Fu, Kai; Li, Dan

    2017-06-01

    Diffuse large B-cell lymphoma (DLBCL) is the most common non-Hodgkin lymphoma, whose main prognostic factor is closely related to germinal center B-cell-like subtype (GCB- DLBCL) or activated B-cell-like type (non-GCB-DLBCL). The most common type of primary central nervous system lymphoma is diffuse large B-cell type with poor prognosis and the reason is unclear. This study aims to stratify primary central nervous system diffuse large B-cell lymphoma (PCNS-DLBCL) according to the cell-of-origin (COO) and to investigate the multiple proteins expression of C-MYC, BCL-6, BCL-2, TP53, further to elucidate the reason why primary central nervous system diffuse large B-cell lymphoma possesses a poor clinical outcome as well. Nineteen cases of primary central nervous system DLBCL were stratified according to immunostaining algorithms of Hans, Choi and Meyer (Tally) and we investigated the multiple proteins expression of C-MYC, BCL-6, BCL-2, TP53. The Epstein-Barr virus and Borna disease virus infection were also detected. Among nineteen cases, most (15-17 cases) were assigned to the activated B-cell-like subtype, highly expression of C-MYC (15 cases, 78.9%), BCL-2 (10 cases, 52.6%), BCL-6 (15 cases, 78.9%). Unfortunately, two cases were positive for PD-L1 while PD-L2 was not expressed in any case. Two cases infected with BDV but no one infected with EBV. In conclusion, most primary central nervous system DLBCLs show an activated B-cell-like subtype characteristic and have multiple expressions of C-MYC, BCL-2, BCL-6 protein, these features might be significant factor to predict the outcome and guide treatment of PCNS-DLBCLs. Copyright © 2017 Elsevier GmbH. All rights reserved.

  9. Forced expression of miR-143 represses ERK5/c-Myc and p68/p72 signaling in concert with miR-145 in gut tumors of Apc(Min) mice.

    Science.gov (United States)

    Takaoka, Yuji; Shimizu, Yuko; Hasegawa, Hitoki; Ouchi, Yasuo; Qiao, Shanlou; Nagahara, Miki; Ichihara, Masatoshi; Lee, Jiing-Dwan; Adachi, Koichi; Hamaguchi, Michinari; Iwamoto, Takashi

    2012-01-01

    Recently, miR-143 and miR-145 have been shown to belong to a subset of microRNAs whose expression is controlled by a complex of a tumor suppressor p53 and DEAD-box RNA helicase subunits p68/p72. While accumulating studies have acknowledged that both miRNAs function as tumor suppressors and are similarly regulated, evidence of their coordinated action against tumorigenesis has been poorly presented. Herein, we establish transgenic mice that express miR-143 under the control of the CAG regulatory unit. When crossbred with Apc(Min/+) mice, the development of tumors in the small intestines is significantly attenuated. In the transgenic small intestine tumors, the endogenous miR-145 is also enhanced and the expression of c-Myc and p68/p72, both of which have been reported to be pivotal for gut tumor development, is suppressed, corresponding to the downregulation of ERK5. We demonstrate that the combination of miR-143 and miR-145 inhibits the expression of c-Myc in human colon cancer cells, whereas miR-145 retards that of p72. Moreover, we show the possibilities that miR-145 modulates p72 expression through its 3' untranslated region and that c-Myc downregulation is involved in both p68 suppression and miR-145 induction. These findings suggest that forced expression of miR-143, probably interacting with endogenous miR-145, inhibits ERK5/c-Myc and p68/p72/β-catenin signaling and hampers small intestine tumor development in Apc(Min/+) mice. This unique cascade, in turn, may prevent overproduction of a subset of tumor suppressive miRNAs by repressing their own modulators, p68/p72.

  10. Forced expression of miR-143 represses ERK5/c-Myc and p68/p72 signaling in concert with miR-145 in gut tumors of Apc(Min mice.

    Directory of Open Access Journals (Sweden)

    Yuji Takaoka

    Full Text Available Recently, miR-143 and miR-145 have been shown to belong to a subset of microRNAs whose expression is controlled by a complex of a tumor suppressor p53 and DEAD-box RNA helicase subunits p68/p72. While accumulating studies have acknowledged that both miRNAs function as tumor suppressors and are similarly regulated, evidence of their coordinated action against tumorigenesis has been poorly presented. Herein, we establish transgenic mice that express miR-143 under the control of the CAG regulatory unit. When crossbred with Apc(Min/+ mice, the development of tumors in the small intestines is significantly attenuated. In the transgenic small intestine tumors, the endogenous miR-145 is also enhanced and the expression of c-Myc and p68/p72, both of which have been reported to be pivotal for gut tumor development, is suppressed, corresponding to the downregulation of ERK5. We demonstrate that the combination of miR-143 and miR-145 inhibits the expression of c-Myc in human colon cancer cells, whereas miR-145 retards that of p72. Moreover, we show the possibilities that miR-145 modulates p72 expression through its 3' untranslated region and that c-Myc downregulation is involved in both p68 suppression and miR-145 induction. These findings suggest that forced expression of miR-143, probably interacting with endogenous miR-145, inhibits ERK5/c-Myc and p68/p72/β-catenin signaling and hampers small intestine tumor development in Apc(Min/+ mice. This unique cascade, in turn, may prevent overproduction of a subset of tumor suppressive miRNAs by repressing their own modulators, p68/p72.

  11. Targeted overexpression of an activated N-ras gene results in B-cell and plasma cell lymphoproliferation and cooperates with c-myc to induce fatal B-cell neoplasia.

    Science.gov (United States)

    Linden, Michael A; Kirchhof, Nicole; Carlson, Cathy S; Van Ness, Brian G

    2012-03-01

    Multiple myeloma is an incurable malignant expansion of plasma cells in the bone marrow. Although there is no pathognomonic genetic lesion among multiple myeloma patients, activation of the ras gene has been identified as a common mutation. We have previously described the use of the 3' κ immunoglobulin light chain enhancer (3'KE) to target transgenic expression in murine B and plasma cells, resulting in bcl-X(L) and c-myc-driven murine models of multiple myeloma. In this report, we characterize the role of activated mutant N-ras in B and plasma cells in transgenic mice. We constructed transgenic mice that use 3'KE to direct expression of a mutant activated N-ras. We also crossed the N-ras mice with mice bearing a c-myc transgene to study the cooperative effects of the transgenic constructs. Mice were sacrificed when moribund or at specific time intervals and characterized by serology, light microscopy, and flow cytometry. The transgenic N-ras animals develop B- and plasma cell lymphoproliferation, and aged mice develop immunoglobulinemia, renal hyaline tubular casts, and microscopic foci of abnormal plasma cells in extramedullary sites, including the liver and kidney. Bitransgenic 3'KE/N-Ras V12 × Eμ-c-Myc mice develop fatal B-cell neoplasia, with a median survival of 10 weeks. These data indicate that activated N-ras can play a role in B- and plasma cell homeostasis and that activated N-Ras and c-Myc can cooperate to induce B-cell neoplasia. Copyright © 2012 ISEH - Society for Hematology and Stem Cells. Published by Elsevier Inc. All rights reserved.

  12. Suppression of c-Myc is involved in multi-walled carbon nanotubes' down-regulation of ATP-binding cassette transporters in human colon adenocarcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Zhaojing [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 430030 Wuhan (China); Xu, Yonghong [Institute of Ophthalmological Research, Department of Ophthalmology, Renmin Hospital of Wuhan University, 430060 Wuhan (China); Meng, Xiangning [School of Materials and Metallurgy, Northeastern University, Shenyang 110819 (China); Watari, Fumio [Department of Biomedical, Dental Materials and Engineering, Graduate School of Dental Medicine, Hokkaido University, Sapporo 060-8586 (Japan); Liu, Hudan, E-mail: hudanliu@hust.edu.cn [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 430030 Wuhan (China); Chen, Xiao, E-mail: mornsmile@yahoo.com [Department of Pharmacology, School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, 430030 Wuhan (China)

    2015-01-01

    Over-expression of ATP-binding cassette (ABC) transporters, a large family of integral membrane proteins that decrease cellular drug uptake and accumulation by active extrusion, is one of the major causes of cancer multi-drug resistance (MDR) that frequently leads to failure of chemotherapy. Carbon nanotubes (CNTs)-based drug delivery devices hold great promise in enhancing the efficacy of cancer chemotherapy. However, CNTs' effects on the ABC transporters remain under-investigated. In this study, we found that multiwalled carbon nanotubes (MWCNTs) reduced transport activity and expression of ABC transporters including ABCB1/Pgp and ABCC4/MRP4 in human colon adenocarcinoma Caco-2 cells. Proto-oncogene c-Myc, which directly regulates ABC gene expression, was concurrently decreased in MWCNT-treated cells and forced over-expression of c-Myc reversed MWCNTs' inhibitory effects on ABCB1 and ABCC4 expression. MWCNT-cell membrane interaction and cell membrane oxidative damage were observed. However, antioxidants such as vitamin C, β-mecaptoethanol and dimethylthiourea failed to antagonize MWCNTs' down-regulation of ABC transporters. These data suggest that MWCNTs may act on c-Myc, but not through oxidative stress, to down-regulate ABC transporter expression. Our findings thus shed light on CNTs' novel cellular effects that may be utilized to develop CNTs-based drug delivery devices to overcome ABC transporter-mediated cancer chemoresistance.

  13. Ku70 acetylation and modulation of c-Myc/ATF4/CHOP signaling axis by SIRT1 inhibition lead to sensitization of HepG2 cells to TRAIL through induction of DR5 and down-regulation of c-FLIP

    DEFF Research Database (Denmark)

    Kim, Mi-Ju; Hong, Kyung-Soo; Kim, Hak-Bong

    2013-01-01

    In this study, we investigated the role of c-Myc/ATF4/CHOP signaling pathway in sensitization of human hepatoma HepG2 cells to TRAIL. Knockdown of SIRT1 or treatment with SIRT1 inhibitor caused the up-regulation of DR5 and down-regulation of c-FLIP through modulation of c-Myc/ATF4/CHOP pathway, a...

  14. Anguillicola crassus infection significantly affects the silvering related modifications in steady state mRNA levels in gas gland tissue of the European eel

    Directory of Open Access Journals (Sweden)

    Bernd ePelster

    2016-05-01

    Full Text Available Using Illumina sequencing, transcriptional changes occurring during silvering in swimbladder tissue of the European eel have been analyzed by comparison of yellow eel and silver eel tissue samples. Functional annotation analysis based on GO terms revealed significant expression changes in a number of genes related to the extracellular matrix, important for the control of gas permeability of the swimbladder, and to ROS (reactive oxygen species defense, important to cope with reactive oxygen species generated under hyperbaric oxygen partial pressures. Focusing on swimbladder tissue metabolism, levels of several mRNA species encoding glucose transport proteins were several-fold higher in silver eels, while enzymes of the glycolytic pathway were not affected. The significantly higher steady state level of a transcript encoding for membrane bound carbonic anhydrase, however, suggested that CO2 production in the pentose phosphate shunt and diffusion of CO2 was of particular importance in silver eel swimbladder. In addition, the mRNA level of a large number of genes related to immune response and to sexual maturation was significantly modified in the silver eel swimbladder. The modification of several processes related to protein metabolism and transport, cell cycle, and apoptosis suggested that these changes in swimbladder metabolism and permeability were achieved by increasing cell turn-over. The impact of an infection of the swimbladder with the nematode Anguillicola crassus has been assessed by comparing these expression changes with expression changes observed between uninfected yellow eel swimbladder tissue and infected silver eel swimbladder tissue. In contrast to uninfected silver eel swimbladder tissue, in infected tissue the mRNA level of several glycolytic enzymes was significantly elevated, and with respect to extracellular matrix, several mucin genes were many-fold higher in their mRNA level. Modification of many immune related genes and

  15. Mutations affecting mRNA splicing define distinct clinical phenotypes and correlate with patient outcome in myelodysplastic syndromes.

    Science.gov (United States)

    Damm, Frederik; Kosmider, Olivier; Gelsi-Boyer, Véronique; Renneville, Aline; Carbuccia, Nadine; Hidalgo-Curtis, Claire; Della Valle, Véronique; Couronné, Lucile; Scourzic, Laurianne; Chesnais, Virginie; Guerci-Bresler, Agnes; Slama, Bohrane; Beyne-Rauzy, Odile; Schmidt-Tanguy, Aline; Stamatoullas-Bastard, Aspasia; Dreyfus, François; Prébet, Thomas; de Botton, Stéphane; Vey, Norbert; Morgan, Michael A; Cross, Nicholas C P; Preudhomme, Claude; Birnbaum, Daniel; Bernard, Olivier A; Fontenay, Michaela

    2012-04-05

    A cohort of MDS patients was examined for mutations affecting 4 splice genes (SF3B1, SRSF2, ZRSR2, and U2AF35) and evaluated in the context of clinical and molecular markers. Splice gene mutations were detected in 95 of 221 patients. These mutations were mutually exclusive and less likely to occur in patients with complex cytogenetics or TP53 mutations. SF3B1(mut) patients presented with lower hemoglobin levels, increased WBC and platelet counts, and were more likely to have DNMT3A mutations. SRSF2(mut) patients clustered in RAEB-1 and RAEB-2 subtypes and exhibited pronounced thrombocytopenias. ZRSR2(mut) patients clustered in International Prognostic Scoring System intermediate-1 and intermediate-2 risk groups, had higher percentages of bone marrow blasts, and more often displayed isolated neutropenias. SRSF2 and ZRSR2 mutations were more common in TET2(mut) patients. U2AF35(mut) patients had an increased prevalence of chromosome 20 deletions and ASXL1 mutations. Multivariate analysis revealed an inferior overall survival and a higher AML transformation rate for the genotype ZRSR2(mut)/TET2(wt) (overall survival: hazard ratio = 3.3; 95% CI, 1.4-7.7; P = .006; AML transformation: hazard ratio = 3.6; 95% CI, 2-4.2; P = .026). Our results demonstrate that splice gene mutations are among the most frequent molecular aberrations in myelodysplastic syndrome, define distinct clinical phenotypes, and show preferential associations with mutations targeting transcriptional regulation.

  16. BIN1 reverses PD-L1-mediated immune escape by inactivating the c-MYC and EGFR/MAPK signaling pathways in non-small cell lung cancer.

    Science.gov (United States)

    Wang, J; Jia, Y; Zhao, S; Zhang, X; Wang, X; Han, X; Wang, Y; Ma, M; Shi, J; Liu, L

    2017-11-09

    Non-small cell lung cancer (NSCLC) is one of the most common and malignant carcinoma worldwide, and the incidence and mortality are increasing rapidly. Immunotherapy targeting programmed death 1/programmed death ligand 1 (PD-L1) signaling has shown prominent clinical effects in treating NSCLC; however, a poor understanding of the associated regulating molecular mechanisms of PD-L1 has become one of the biggest obstacles for further improving efficacy. Bridging integrator-1 (BIN1) can regulate numerous cancer-related molecules to exert multiple tumor-suppressing effects by either interacting or not interacting with c-MYC. In the present study, we observed that there exists a negative correlation between the expression of PD-L1 and BIN1 in NSCLC tissues. The expression levels of BIN1 and PD-L1 were significantly related to the tumor, lymph node and metastasis grade (TNM) stage, invasion range and lymph node metastasis. Simultaneously, for NSCLC patients, the expression statuses of BIN1 and PD-L1 might be independent prognostic factors. Furthermore, the expression of tumor-infiltrating lymphocytes was positively associated with BIN1 expression and negatively related to PD-L1 expression in NSCLC tissues. Importantly, we showed that PD-L1 was under the control of BIN1. In addition, the overexpression of BIN1 could inhibit the c-MYC and epithelial growth factor receptor (EGFR)-dependent PD-L1 expression and reverse the suppressive immuno-microenvironment in vivo. Taken together, our findings indicated that BIN1 restoration in NSCLC could reverse PD-L1-mediated immune escape by inactivating the c-MYC and EGFR/mitogen-activated protein kinase pathways.

  17. A mouse strain defective in both T cells and NK cells has enhanced sensitivity to tumor induction by plasmid DNA expressing both activated H-Ras and c-Myc.

    Directory of Open Access Journals (Sweden)

    Li Sheng-Fowler

    Full Text Available As part of safety studies to evaluate the risk of residual cellular DNA in vaccines manufactured in tumorigenic cells, we have been developing in vivo assays to detect and quantify the oncogenic activity of DNA. We generated a plasmid expressing both an activated human H-ras gene and murine c-myc gene and showed that 1 µg of this plasmid, pMSV-T24-H-ras/MSV-c-myc, was capable of inducing tumors in newborn NIH Swiss mice. However, to be able to detect the oncogenicity of dominant activated oncogenes in cellular DNA, a more sensitive system was needed. In this paper, we demonstrate that the newborn CD3 epsilon transgenic mouse, which is defective in both T-cell and NK-cell functions, can detect the oncogenic activity of 25 ng of the circular form of pMSV-T24-H-ras/MSV-c-myc. When this plasmid was inoculated as linear DNA, amounts of DNA as low as 800 pg were capable of inducing tumors. Animals were found that had multiple tumors, and these tumors were independent and likely clonal. These results demonstrate that the newborn CD3 epsilon mouse is highly sensitive for the detection of oncogenic activity of DNA. To determine whether it can detect the oncogenic activity of cellular DNA derived from four human tumor-cell lines (HeLa, A549, HT-1080, and CEM, DNA (100 µg was inoculated into newborn CD3 epsilon mice both in the presence of 1 µg of linear pMSV-T24-H-ras/MSV-c-myc as positive control and in its absence. While tumors were induced in 100% of mice with the positive-control plasmid, no tumors were induced in mice receiving any of the tumor DNAs alone. These results demonstrate that detection of oncogenes in cellular DNA derived from four human tumor-derived cell lines in this mouse system was not possible; the results also show the importance of including a positive-control plasmid to detect inhibitory effects of the cellular DNA.

  18. IS THE AMPLIFICATION OF c-MYC, MLL AND RUNX1 GENES IN AML AND MDS PATIENTS WITH TRISOMY 8, 11 AND 21 A FACTOR FOR A CLONAL EVOLUTION IN THEIR KARYOTYPE?

    Science.gov (United States)

    Angelova, S; Spassov, B; Nikolova, V; Christov, I; Tzvetkov, N; Simeonova, M

    2015-01-01

    The aim of our study was 1) to define if the amplification of c-MYC, MLL and RUNX1 genes is related to the progressive changes of the karyotype in patients with AML and MDS with trisomy 8, 11 and 21 (+8, +11 and +21) in bone marrow and 2) can that amplification be accepted as part of the clonal evolution (CE). Karyotype analysis was performed in 179 patients with AML or MDS with the different chromosomal aberrations (CA) aged 16-81. The findings were distributed as follow: initiating balanced CA (n = 60), aneuploidia (n = 55), unbalanced CA (n = 64). Amplification of c-MYC, MLL and RUNX1 genes by means of fluorescence in situ hybridization (FISH) was found in 35% (7 out of 20) of AML and MDS patients with +8, +11 u +21 as single CA in their karyotype; in 63.6% of pts (7 out of 11)--with additional numerical or structural CA and in 75% (9 out of 12)--with complex karyotype. We assume that the amplification of the respective chromosomal regions in patients with +8, +11 and +21 is related to CE. Considering the amplification as a factor of CE, we established 3 patterns of karyotype development depending on the type of the initiating CA in it. Significant statistical differences were found between the three patterns regarding the karyotype distribution in the different stages of progression (p < 0.001).

  19. Sodium arsenite induces ROS generation, DNA oxidative damage, HO-1 and c-Myc proteins, NF-kappaB activation and cell proliferation in human breast cancer MCF-7 cells.

    Science.gov (United States)

    Ruiz-Ramos, Ruben; Lopez-Carrillo, Lizbeth; Rios-Perez, Alfonso D; De Vizcaya-Ruíz, Andrea; Cebrian, Mariano E

    2009-03-31

    Epidemiological evidence has associated exposure to arsenic (As) in drinking water with an increased incidence of human cancers in the skin, bladder, liver, kidney and lung. Sodium arsenite mimics the effects of estradiol and induces cell proliferation in the estrogen responsive breast cancer cell line MCF-7. Therefore, our aim was to further explore the ability of sodium arsenite to induce MCF-7 epithelial breast cell proliferation and some of its underlying mechanisms by studying ROS production, c-Myc and HO-1 protein levels, 8-OHdG formation and NF-kappaB activation. Low arsenite concentrations (0.5-5 microM) induced ROS production and ROS-related depolarization of the mitochondrial membrane suggesting that mitochondria played an important role in the oxidative effects of As. ROS-mediated DNA damage as measured by the presence of 8-OHdG DNA-adducts in their nuclei, IkappaB phosphorylation, NF-kappaB activation and increases in c-Myc and HO-1 protein levels were also observed, suggesting that these factors play a relevant role in the arsenite induced MCF-7 cell recruitment into the S-phase of the cell cycle and cell proliferation observed. In conclusion, arsenite activates several pathways involved in MCF-7 cell proliferation suggesting that arsenite exposure may pose a risk for breast cancer in human exposed populations notwithstanding that most studies to date have not yet implicated this metalloid as a cofactor in the etiology of this disease.

  20. Proliferation, cell cycle exit, and onset of terminal differentiation in cultured keratinocytes: pre-programmed pathways in control of C-Myc and Notch1 prevail over extracellular calcium signals.

    Science.gov (United States)

    Kolly, Carine; Suter, Maja M; Müller, Eliane J

    2005-05-01

    So far it was reported that a switch from low to high extracellular calcium induces growth arrest and terminal differentiation in cultured human and mouse keratinocytes. We had observed that both canine and mouse keratinocytes proliferate in high (1.8 mM, respectively, 1.2 mM) or low (0.09 and 0.06 mM) calcium-containing medium. In-depth analysis of this phenomenon revealed, as reported here, that the switch between proliferation and terminal differentiation occurred irrespective of calcium conditions when the canine and murine keratinocytes reach confluency. The "confluency switch" coincided with transcriptional upregulation of cell cycle inhibitors p21(WAF1) and p27(KIP1) as well as proteins marking onset of terminal differentiation. It was further accompanied by downregulation and nuclear clearance of c-Myc, and conversely activation of Notch1, which are shown to be critical determinants of this process. Together, this study demonstrates that even in the absence of and similar to their in vivo environment, cultured canine and mouse keratinocytes follow a pre-defined differentiation program. This program is in control of c-Myc and Notch1 and does not require complementary signals for onset of terminal differentiation except those given by cell-cell contact. Once triggered, completion of the terminal differentiation process depends on elevated extracellular calcium to stabilize intercellular junctions and components of the cornified envelope.

  1. Detection of three nonsense mutations and one missense mutation in the interleukin-2 receptor [gamma] chain gene in SCIDX1 that differently affect the mRNA processing

    Energy Technology Data Exchange (ETDEWEB)

    Markiewicz, S.; Fischer, A.; Saint Basile, G. de (INSERM, Paris (France)); Subtil, A.; Dautry-Varsat, A. (Institut Pasteur, Paris (France))

    1994-05-01

    The interleukin-2 receptor [gamma] (IL-2R[gamma]) chain gene encodes a 64-kDa protein that not only composes the high-affinity form of the IL-2 binding receptor in association with the 2R [alpha] and [beta] chains, but also participates in at least the IL-4 and IL-7 receptor complexes. Mutations in this gene have recently been shown to cause X-linked severe combined immunodeficiency (SCIDX1). This disease of the immune system results from an early block of T lymphocyte and natural killer (NK) cell differentiation, which leads to a severe cellular and humoral immune defect that is lethal unless treated by bone marrow transplantation. Analysis of the IL-2R[gamma] gene in SCIDX1 patients has revealed the presence of heterogeneous mutations principally located in the extracellular domain of the molecule. We report here three intraexonic mutations and one deletion in the IL-2R[gamma] gene in four SCIDX1 patients. These mutations appear to differentially affect RNA processing, either by decreasing IL-2R[gamma] mRNA level or by the skipping of a constitutive exon. 16 refs., 1 fig.

  2. Direct short-term cytotoxic effects of BIBR 1532 on acute promyelocytic leukemia cells through induction of p21 coupled with downregulation of c-Myc and hTERT transcription.

    Science.gov (United States)

    Bashash, D; Ghaffari, S H; Zaker, F; Hezave, K; Kazerani, M; Ghavamzadeh, A; Alimoghaddam, K; Mosavi, S A; Gharehbaghian, A; Vossough, P

    2012-01-01

    Acute promyelocytic leukemia (APL) is characterized by specific t(15;17), distinct morphologic picture, and clinical coagulopathy that contribute to the morbidity and mortality of the disease. This study aims to investigate the effects of antitelomerase compound BIBR1532 on APL cells (NB4). BIBR 1532 exerts a direct short-term growth suppressive effect in a concentration-dependent manner probably through downregulation of c-Myc and hTERT expression. Our results also suggest that induction of p21 and subsequent disturbance of Bax/Bcl-2 balanced ratio as well as decreased telomerase activity may be rational mechanisms for the potent/direct short-term cytotoxicity of high doses of BIBR1532 against NB4 cells.

  3. The Phosphoinositide 3-Kinase p110α Isoform Regulates Leukemia Inhibitory Factor Receptor Expression via c-Myc and miR-125b to Promote Cell Proliferation in Medulloblastoma

    Science.gov (United States)

    von Bueren, André O.; Ćwiek, Paulina; Rehrauer, Hubert; Djonov, Valentin; Anderle, Pascale; Arcaro, Alexandre

    2015-01-01

    Medulloblastoma (MB) is the most common malignant brain tumor in childhood and represents the main cause of cancer-related death in this age group. The phosphoinositide 3-kinase (PI3K) pathway has been shown to play an important role in the regulation of medulloblastoma cell survival and proliferation, but the molecular mechanisms and downstream effectors underlying PI3K signaling still remain elusive. The impact of RNA interference (RNAi)-mediated silencing of PI3K isoforms p110α and p110δ on global gene expression was investigated by DNA microarray analysis in medulloblastoma cell lines. A subset of genes with selectively altered expression upon p110α silencing in comparison to silencing of the closely related p110δ isoform was revealed. Among these genes, the leukemia inhibitory factor receptor α (LIFR α) was validated as a novel p110α target in medulloblastoma. A network involving c-Myc and miR-125b was shown to be involved in the control of LIFRα expression downstream of p110α. Targeting the LIFRα by RNAi, or by using neutralizing reagents impaired medulloblastoma cell proliferation in vitro and induced a tumor volume reduction in vivo. An analysis of primary tumors revealed that LIFRα and p110α expression were elevated in the sonic hedgehog (SHH) subgroup of medulloblastoma, indicating its clinical relevance. Together, these data reveal a novel molecular signaling network, in which PI3K isoform p110α controls the expression of LIFRα via c-Myc and miR-125b to promote MB cell proliferation. PMID:25915540

  4. The Phosphoinositide 3-Kinase p110α Isoform Regulates Leukemia Inhibitory Factor Receptor Expression via c-Myc and miR-125b to Promote Cell Proliferation in Medulloblastoma.

    Directory of Open Access Journals (Sweden)

    Fabiana Salm

    Full Text Available Medulloblastoma (MB is the most common malignant brain tumor in childhood and represents the main cause of cancer-related death in this age group. The phosphoinositide 3-kinase (PI3K pathway has been shown to play an important role in the regulation of medulloblastoma cell survival and proliferation, but the molecular mechanisms and downstream effectors underlying PI3K signaling still remain elusive. The impact of RNA interference (RNAi-mediated silencing of PI3K isoforms p110α and p110δ on global gene expression was investigated by DNA microarray analysis in medulloblastoma cell lines. A subset of genes with selectively altered expression upon p110α silencing in comparison to silencing of the closely related p110δ isoform was revealed. Among these genes, the leukemia inhibitory factor receptor α (LIFR α was validated as a novel p110α target in medulloblastoma. A network involving c-Myc and miR-125b was shown to be involved in the control of LIFRα expression downstream of p110α. Targeting the LIFRα by RNAi, or by using neutralizing reagents impaired medulloblastoma cell proliferation in vitro and induced a tumor volume reduction in vivo. An analysis of primary tumors revealed that LIFRα and p110α expression were elevated in the sonic hedgehog (SHH subgroup of medulloblastoma, indicating its clinical relevance. Together, these data reveal a novel molecular signaling network, in which PI3K isoform p110α controls the expression of LIFRα via c-Myc and miR-125b to promote MB cell proliferation.

  5. Two rare deletions upstream of the NRXN1 gene (2p16.3) affecting the non-coding mRNA AK127244 segregate with diverse psychopathological phenotypes in a family

    DEFF Research Database (Denmark)

    Duong, L. T. T.; Hoeffding, L. K.; Petersen, K. B.

    2015-01-01

    susceptibility. In this study, we describe a family affected by a wide range of psychiatric disorders including early onset schizophrenia, schizophreniform disorder, and affective disorders. Microarray analysis identified two rare deletions immediately upstream of the NRXN1 gene affecting the non-coding mRNA AK......127244 in addition to the pathogenic 15q11.2 deletion in distinct family members. The two deletions upstream of the NRXN1 gene were found to segregate with psychiatric disorders in the family and further similar deletions have been observed in patients diagnosed with autism spectrum disorder. Thus, we...

  6. Differential expression of leptin and leptin's receptor isoform (Ob-Rb) mRNA between advanced and minimally affected osteoarthritic cartilage; effect on cartilage metabolism.

    Science.gov (United States)

    Simopoulou, T; Malizos, K N; Iliopoulos, D; Stefanou, N; Papatheodorou, L; Ioannou, M; Tsezou, A

    2007-08-01

    To investigate leptin's effect on cartilage metabolism and the pathophysiology of osteoarthritis (OA). Messenger RNA (mRNA) expression and protein levels of leptin and leptin's receptor isoforms were measured by real-time reverse transcription-PCR and Western blot in osteoarthritic and normal cartilage. Osteoarthritic cartilage samples were obtained from two locations of the knee (n=11) and hip (n=6); from the main defective area (advanced OA) and from adjacent macroscopically and histological intact regions (minimal OA). Paired serum and synovial fluid (SF) leptin levels were measured. The effect of leptin was evaluated on chondrocyte proliferation, IL-1beta (interleukin-1beta), NO and metalloproteinases 9 and 13 (MMP-9, MMP-13) protein expression. Leptin's and leptin's receptor (Ob-Rb) expression levels were significantly increased in advanced OA cartilage compared to minimal. Leptin was significantly increased in SF than serum samples. Also, leptin had a detrimental effect on chondrocyte proliferation and induced IL-1beta production and MMP-9 and MMP-13 protein expression. Furthermore, leptin's mRNA expression in advanced OA cartilage was significantly correlated with BMI of the patients. The increased leptin levels in SF point toward a local effect of leptin in articular cartilage, while the observed intrajoint differences of leptin and Ob-Rb mRNA expression may be related to the grade of cartilage destruction. The observed production of IL-1beta, MMP-9 and MMP-13 by chondrocytes after leptin treatment indicates a pro-inflammatory and catabolic role of leptin on cartilage metabolism. Furthermore, the observed correlation of leptin's mRNA expression with BMI suggests that leptin may be a metabolic link between obesity and OA.

  7. Polymorphisms in the 3' UTR in the neurocalcin delta gene affect mRNA stability, and confer susceptibility to diabetic nephropathy

    DEFF Research Database (Denmark)

    Kamiyama, Masumi; Kobayashi, Masaaki; Araki, Shin-ichi

    2007-01-01

    of NCALD (rs1131863: exon 4 +1340 A vs. G, P = 0.00004, odds ratio = 1.59, 95% CI 1.27-1.98). We also discovered two other SNPs in exon 4 of this gene (+999 T/A, +1307 A/G) that showed absolute linkage disequilibrium to the landmark SNP. Subsequent in vitro functional analysis revealed that synthetic mRNA...... corresponding to the disease susceptible haplotype (exon 4 +1340 G, +1307 G, +999 A) was degraded faster than mRNA corresponding to the major haplotype (exon 4 +1340 A, +1307 A, +999 T), and allelic mRNA expression of the disease susceptibility allele was significantly lower than that of the major allele...... in normal kidney tissues. In an experiment using a short interfering RNA targeting NCALD, we found that silencing of the NCALD led to a considerable enhancement of cell migration, accompanied by a significant reduction in E-cadherin expression, and by an elevation of alpha smooth muscle actin expression...

  8. Postovulatory aging affects dynamics of mRNA, expression and localization of maternal effect proteins, spindle integrity and pericentromeric proteins in mouse oocytes.

    Science.gov (United States)

    Trapphoff, T; Heiligentag, M; Dankert, D; Demond, H; Deutsch, D; Fröhlich, T; Arnold, G J; Grümmer, R; Horsthemke, B; Eichenlaub-Ritter, U

    2016-01-01

    Is the postovulatory aging-dependent differential decrease of mRNAs and polyadenylation of mRNAs coded by maternal effect genes associated with altered abundance and distribution of maternal effect and RNA-binding proteins (MSY2)? Postovulatory aging results in differential reduction in abundance of maternal effect proteins, loss of RNA-binding proteins from specific cytoplasmic domains and critical alterations of pericentromeric proteins without globally affecting protein abundance. Oocyte postovulatory aging is associated with differential alteration in polyadenylation and reduction in abundance of mRNAs coded by selected maternal effect genes. RNA-binding and -processing proteins are involved in storage, polyadenylation and degradation of mRNAs thus regulating stage-specific recruitment of maternal mRNAs, while chromosomal proteins that are stage-specifically expressed at pericentromeres, contribute to control of chromosome segregation and regulation of gene expression in the zygote. Germinal vesicle (GV) and metaphase II (MII) oocytes from sexually mature C57B1/6J female mice were investigated. Denuded in vivo or in vitro matured MII oocytes were postovulatory aged and analyzed by semiquantitative confocal microscopy for abundance and localization of polyadenylated RNAs, proteins of maternal effect genes (transcription activator BRG1 also known as ATP-dependent helicase SWI/SNF related, matrix associated, actin dependent regulator of chromatin, subfamily a, member 4 (SMARCA4) and NOD-like receptor family pyrin domain containing 5 (NLRP5) also known as MATER), RNA-binding proteins (MSY2 also known as germ cell-specific Y-box-binding protein, YBX2), and post-transcriptionally modified histones (trimethylated histone H3K9 and acetylated histone H4K12), as well as pericentromeric ATRX (alpha thalassemia/mental retardation syndrome X-linked, also termed ATP-dependent helicase ATRX or X-linked nuclear protein (XNP)). For proteome analysis five replicates of 30 mouse

  9. Comparative analysis of the EGFR, HER2, c-MYC, and MET variations in colorectal cancer determined by three different measures: gene copy number gain, amplification status and the 2013 ASCO/CAP guideline criterion for HER2 testing of breast cancer.

    Science.gov (United States)

    Kwak, Yoonjin; Yun, Sumi; Nam, Soo Kyung; Seo, An Na; Lee, Kyu Sang; Shin, Eun; Oh, Heung-Kwon; Kim, Duck Woo; Kang, Sung Bum; Kim, Woo Ho; Lee, Hye Seung

    2017-08-01

    The purpose of this study was to explore gene copy number (GCN) variation of EGFR, HER2, c-MYC, and MET in patients with primary colorectal cancer (CRC). Dual-colour silver-enhanced in situ hybridization was performed in tissue samples of 334 primary CRC patients. The amplification status (GCN ratio ≥2) and GCN gain (average GCN ≥4) data for the EGFR, HER2, c-MYC and MET genes were obtained. GCN variation was also assessed by the criterion of the 2013 ASCO/CAP guidelines for HER2 testing. Amplification of EGFR, HER2, c-MYC and MET was detected in 8 (2.4%), 20 (6.0%), 29 (8.7%), and 14 (4.2%) patients, respectively. Of 66 patients with at least one amplified gene, five exhibited co-amplification of genes studied (HER2-MET co-amplification: two patients; HER2-c-MYC co-amplification: two patients; EGFR-c-MYC co-amplification: one patient). There were 109 patients with GCN gains of one or more genes (EGFR: 11/334, HER2: 29/334, c-MYC; 60/334, MET: 48/334) and 32.1% (35/109) had multiple GCN gains. When each GCN was assessed by the criterion of the ASCO/CAP 2013 guideline for HER2 testing, 116 people showed positive or equivocal results for one or more genes. The cumulative amplification status had no association with patients' outcome. However, the cumulative results of the GCN gain and GCN status determined according to the ASCO/CAP guideline had a significant prognostic correlation in the univariate analysis (P values of 0.006 and 0.022, respectively). In the multivariate analysis, GCN gain and GCN status were independent prognostic factors (P values of 0.010 and 0.017, respectively). In this study, we evaluated GCN variation of four genes in a large sample of Korean CRC patients. The amplification status was not related to patient outcome. However, the GCN gain and GCN status according to the ASCO/CAP 2013 guideline were independent prognostic factors.

  10. Complex Biological Systems Analysis of Cell Cycling Models in Carcinogenesis: I. The essential roles of modifications in the c-Myc, TP53/p53, p27 and hTERT modules in Cancer Initiation and Progression

    CERN Document Server

    Prisecaru, V I

    2004-01-01

    A new approach to the integration of results from a modular, complex biological systems analysis of nonlinear dynamics in cell cycling network transformations that are leading to carcinogenesis is proposed. Carcinogenesis is a complex process that involves dynamically inter-connected biomolecules in the intercellular, membrane, cytosolic, nuclear and nucleolar compartments that form numerous inter-related pathways referred to as networks. One such network module contains the cell cyclins whose functions are essential to cell cycling and division. Cyclins are proteins that also link to several critical pro-apoptotic and other cell cycling/division components, such as: c-Myc, p27, the tumor suppressor gene TP53 and its product-- the p53 protein with key roles in controlling DNA repair, inducing apoptosis and activating p21 (which can depress cell cyclins if activated), mdm2(with its biosynthesis activated by p53 and also, in its turn, inhibiting p53), p21, the Thomsen-Friedenreich antigen(T- antigen),Rb,Bax, Ba...

  11. PRIMA-1 induces caspase-mediated apoptosis in acute promyelocytic leukemia NB4 cells by inhibition of nuclear factor-κB and downregulation of Bcl-2, XIAP, and c-Myc.

    Science.gov (United States)

    Farhadi, Elham; Safa, Majid; Sharifi, Ali M; Bashash, Davood

    2017-01-01

    Restoration of p53 function triggers cell death and eliminates tumors in vivo. Identification of p53-reactivating small molecules such as PRIMA-1 holds promise for effective new anticancer therapies. Here, we investigated the effects of small molecule PRIMA-1 on cell viability and expression of p53-regulated genes and proteins in the acute promyelocytic leukemia-derived NB4 cell line. Our results showed that PRIMA-1 had antileukemic properties in acute promyelocytic leukemia-derived NB4 cells. PRIMA-1-triggered apoptosis in a dose-dependent and time-dependent manner as indicated by the MTT assay and annexin-V staining. Apoptosis induction by PRIMA-1 was associated with caspase-9, caspase-7 activation and PARP cleavage. p21 protein expression was increased after PRIMA-1 treatment and real-time PCR analysis of proapoptotic p53 target genes indicated upregulation of Bax and Noxa. Western blot analysis showed that IκBα phosphorylation and its degradation were inhibited by PRIMA-1. Moreover, protein expression of nuclear factor-κB-regulated antiapoptotic (Bcl-2 and XIAP) and proliferative (c-Myc) gene products was decreased. Importantly, PRIMA-1 did not show any significant apoptotic effect in normal human peripheral blood mononuclear cells. These in-vitro studies imply that p53 reactivation by small compounds may become a novel anticancer therapy in acute promyelocytic leukemia.

  12. CAPER is vital for energy and redox homeostasis by integrating glucose-induced mitochondrial functions via ERR-α-Gabpa and stress-induced adaptive responses via NF-κB-cMYC.

    Directory of Open Access Journals (Sweden)

    Yun Kyoung Kang

    2015-04-01

    Full Text Available Ever since we developed mitochondria to generate ATP, eukaryotes required intimate mito-nuclear communication. In addition, since reactive oxygen species are a cost of mitochondrial oxidative phosphorylation, this demands safeguards as protection from these harmful byproducts. Here we identified a critical transcriptional integrator which eukaryotes share to orchestrate both nutrient-induced mitochondrial energy metabolism and stress-induced nuclear responses, thereby maintaining carbon-nitrogen balance, and preserving life span and reproductive capacity. Inhibition of nutrient-induced expression of CAPER arrests nutrient-dependent cell proliferation and ATP generation and induces autophagy-mediated vacuolization. Nutrient signaling to CAPER induces mitochondrial transcription and glucose-dependent mitochondrial respiration via coactivation of nuclear receptor ERR-α-mediated Gabpa transcription. CAPER is also a coactivator for NF-κB that directly regulates c-Myc to coordinate nuclear transcriptome responses to mitochondrial stress. Finally, CAPER is responsible for anaplerotic carbon flux into TCA cycles from glycolysis, amino acids and fatty acids in order to maintain cellular energy metabolism to counter mitochondrial stress. Collectively, our studies reveal CAPER as an evolutionarily conserved 'master' regulatory mechanism by which eukaryotic cells control vital homeostasis for both ATP and antioxidants via CAPER-dependent coordinated control of nuclear and mitochondrial transcriptomic programs and their metabolisms. These CAPER dependent bioenergetic programs are highly conserved, as we demonstrated that they are essential to preserving life span and reproductive capacity in human cells-and even in C. elegans.

  13. [Complex characteristics of the alterations of oncogenes HER-2/ERBB-2, HER-1/ERBB-1, HRAS-1, C-MYC and antioncogenes p53, RB1, as well as deletions of loci of chromosome 17 in colon carcinoma].

    Science.gov (United States)

    Kniazev, P G; Imianitov, E N; Chernitsa, O I; Nikiforova, I F; Babenko, V I; Bruderliaĭn, S; Plutalo, O V; Kuz'min, A I; Kaboev, O K; Berlin, Iu A

    1992-01-01

    Abnormalities of some oncogenes, antioncogenes and losses of heterozygosity (LOH) of chromosome 11p, 17p, and 17q in colorectal carcinomas (CC) was studied. Amplification of ERBB-1/HER-1 oncogene was detected in 2 of 56 cases; ERBB-2/HER-2- in 4 of 62. There was a lack of evidence for C-MYC oncogene amplification (67 cases). LOH of chromosome 11p (HRAS-1 probe) was found in 2 of 37 informative (heterozygous) cases; such events were not accompanied by point mutations in "hot" codons (12th or 61st) in the remaining allele. Prevalence of A3 and A4 alleles of HRAS-1 oncogene (68 cases) as compared to healthy donors was noted. RB-1 (41 cases) and p53 (62 cases) suppressor genes did not show any alterations in Southern-blot analysis. LOH of chromosome 17p (YNZ-22 probe) was found in 15 of 26 heterozygous CC; 17q (THH-59 probe)--in 4 of 16. Analysis of 175th codon of p53 gene revealed only one case of mutation in 35 CC studied. Finally, we were able to detect genetic alterations in 23 of 40 (58%) CC, that were studied on each parameter using Southern-blot. We failed to find any correlation between various molecular abnormalities or clinical characteristics. The data obtained are in disagreement with the view concerning frequent involvement of p53 antioncogene in chromosome 17p deletions.

  14. High Intensity Interval Training Favourably Affects Angiotensinogen mRNA Expression and Markers of Cardiorenal Health in a Rat Model of Early-Stage Chronic Kidney Disease

    Directory of Open Access Journals (Sweden)

    Patrick S. Tucker

    2015-01-01

    Full Text Available The majority of CKD-related complications stem from cardiovascular pathologies such as hypertension. To help reduce cardiovascular complications, aerobic exercise is often prescribed. Emerging evidence suggests high intensity interval training (HIIT may be more beneficial than traditional aerobic exercise. However, appraisals of varying forms of aerobic exercise, along with descriptions of mechanisms responsible for health-related improvements, are lacking. This study examined the effects of 8 weeks of HIIT (85% VO2max, versus low intensity aerobic exercise (LIT; 45–50% VO2max and sedentary behaviour (SED, in an animal model of early-stage CKD. Tissue-specific mRNA expression of RAAS-related genes and CKD-related clinical markers were examined. Compared to SED, HIIT resulted in increased plasma albumin (p=0.001, reduced remnant kidney weight (p=0.028, and reduced kidney weight-body weight ratios (p=0.045. Compared to LIT, HIIT resulted in reduced Agt mRNA expression (p=0.035, reduced plasma LDL (p=0.001, triglycerides (p=0.029, and total cholesterol (p=0.002, increased plasma albumin (p=0.047, reduced remnant kidney weight (p=0.005, and reduced kidney weight-body weight ratios (p=0.048. These results suggest HIIT is a more potent regulator of several markers that describe and influence health in CKD.

  15. The Influence of Spirulina platensis Filtrates on Caco-2 Proliferative Activity and Expression of Apoptosis-Related microRNAs and mRNA.

    Science.gov (United States)

    Śmieszek, Agnieszka; Giezek, Ewa; Chrapiec, Martyna; Murat, Martyna; Mucha, Aleksandra; Michalak, Izabela; Marycz, Krzysztof

    2017-03-07

    Spirulina platensis (SP) is a blue-green microalga that has recently raised attention not only as a nutritional component, but also as a source of bioactivities that have therapeutic effects and may find application in medicine, including cancer treatment. In the present study we determined the cytotoxic effect of S. platensis filtrates (SPF) on human colon cancer cell line Caco-2. Three concentrations of SPF were tested-1.25%, 2.5%, and 5% (v/v). We have found that the highest concentration of SPF exerts the strongest anti-proliferative and pro-apoptotic effect on Caco-2 cultures. The SPF negatively affected the morphology of Caco-2 causing colony shrinking and significant inhibition of metabolic and proliferative activity of cells. The wound-healing assay showed that the SPF impaired migratory capabilities of Caco-2. This observation was consistent with lowered mRNA levels for metalloproteinases. Furthermore, SPF decreased the transcript level of pro-survival genes (cyclin D1, surviving, and c-Myc) and reduced the autocrine secretion of Wnt-10b. The cytotoxic effect of SPF involved the modulation of the Bax and Bcl-2 ratio and a decrease of mitochondrial activity, and was related with increased levels of intracellular reactive oxygen species (ROS) and nitric oxide (NO). Moreover, the SPF also caused an increased number of cells in the apoptotic sub-G0 phase and up-regulated expression of mir-145, simultaneously decreasing expression of mir-17 and 146. Obtained results indicate that SPF can be considered as an agent with anti-cancer properties that may be used for colon cancer prevention and treatment.

  16. The Influence of Spirulina platensis Filtrates on Caco-2 Proliferative Activity and Expression of Apoptosis-Related microRNAs and mRNA

    Science.gov (United States)

    Śmieszek, Agnieszka; Giezek, Ewa; Chrapiec, Martyna; Murat, Martyna; Mucha, Aleksandra; Michalak, Izabela; Marycz, Krzysztof

    2017-01-01

    Spirulina platensis (SP) is a blue-green microalga that has recently raised attention not only as a nutritional component, but also as a source of bioactivities that have therapeutic effects and may find application in medicine, including cancer treatment. In the present study we determined the cytotoxic effect of S. platensis filtrates (SPF) on human colon cancer cell line Caco-2. Three concentrations of SPF were tested—1.25%, 2.5%, and 5% (v/v). We have found that the highest concentration of SPF exerts the strongest anti-proliferative and pro-apoptotic effect on Caco-2 cultures. The SPF negatively affected the morphology of Caco-2 causing colony shrinking and significant inhibition of metabolic and proliferative activity of cells. The wound-healing assay showed that the SPF impaired migratory capabilities of Caco-2. This observation was consistent with lowered mRNA levels for metalloproteinases. Furthermore, SPF decreased the transcript level of pro-survival genes (cyclin D1, surviving, and c-Myc) and reduced the autocrine secretion of Wnt-10b. The cytotoxic effect of SPF involved the modulation of the Bax and Bcl-2 ratio and a decrease of mitochondrial activity, and was related with increased levels of intracellular reactive oxygen species (ROS) and nitric oxide (NO). Moreover, the SPF also caused an increased number of cells in the apoptotic sub-G0 phase and up-regulated expression of mir-145, simultaneously decreasing expression of mir-17 and 146. Obtained results indicate that SPF can be considered as an agent with anti-cancer properties that may be used for colon cancer prevention and treatment. PMID:28272349

  17. Non-secreted clusterin isoforms are translated in rare amounts from distinct human mRNA variants and do not affect Bax-mediated apoptosis or the NF-κB signaling pathway.

    Directory of Open Access Journals (Sweden)

    Hans Prochnow

    Full Text Available Clusterin, also known as apolipoprotein J, is expressed from a variety of tissues and implicated in pathological disorders such as neurodegenerative diseases, ischemia and cancer. In contrast to secretory clusterin (sCLU, which acts as an extracellular chaperone, the synthesis, subcellular localization and function(s of intracellular CLU isoforms is currently a matter of intense discussion. By investigating human CLU mRNAs we here unravel mechanisms leading to the synthesis of distinct CLU protein isoforms and analyze their subcellular localization and their impact on apoptosis and on NF-κB-activity. Quantitative PCR-analyses revealed the expression of four different stress-inducible CLU mRNA variants in non-cancer and cancer cell lines. In all cell lines variant 1 represents the most abundant mRNA, whereas all other variants collectively account for no more than 0.34% of total CLU mRNA, even under stressed conditions. Overexpression of CLU cDNAs combined with in vitro mutagenesis revealed distinct translational start sites including a so far uncharacterized non-canonical CUG start codon. We show that all exon 2-containing mRNAs encode sCLU and at least three non-glycosylated intracellular isoforms, CLU1‑449, CLU21‑449 and CLU34‑449, which all reside in the cytosol of unstressed and stressed HEK‑293 cells. The latter is the only form expressed from an alternatively spliced mRNA variant lacking exon 2. Functional analysis revealed that none of these cytosolic CLU forms modulate caspase-mediated intrinsic apoptosis or significantly affects TNF-α-induced NF-κB-activity. Therefore our data challenge some of the current ideas regarding the physiological functions of CLU isoforms in pathologies.

  18. Severe fluoropyrimidine toxicity due to novel and rare DPYD missense mutations, deletion and genomic amplification affecting DPD activity and mRNA splicing

    DEFF Research Database (Denmark)

    van Kuilenburg, André B P; Meijer, Judith; Maurer, Dirk

    2017-01-01

    in 9 cancer patients presenting with severe fluoropyrimidine-associated toxicity. All patients possessed a strongly reduced DPD activity, ranging from 9 to 53% of controls. Analysis of the DPD gene (DPYD) showed the presence of 21 variable sites including 4 novel and 4 very rare aberrations: 3 missense.......G284V mutation showed residual DPD activities of 30% and 0.5%, respectively. Analysis of a DPD homology model indicated that the p.I948T and p.G284V mutations may affect electron transfer and the binding of FAD, respectively. cDNA analysis showed that the c.321+1G>A mutation in DPYD leads to skipping...

  19. siRNA down-regulation of FGA mRNA in HepG2 cells demonstrated that heterozygous abnormality of the Aα-chain gene does not affect the plasma fibrinogen level.

    Science.gov (United States)

    Takezawa, Yuka; Matsuda, Kazuyuki; Terasawa, Fumiko; Sugano, Mitsutoshi; Honda, Takayuki; Okumura, Nobuo

    2013-04-01

    We encountered two afibrinogenemia patients with homozygous and compound heterozygous FGA mutation. Of interest, the patients' parents, who are heterozygous, had normal levels of plasma fibrinogen; thus, we hypothesized that liver FGA mRNA levels were higher than those of FGB and/or FGG mRNA. To test the hypothesis, we quantitated mRNA levels of a normal liver and a human hepatocyte cell line, HepG2 cells, and performed siRNA-mediated down-regulation of the fibrinogen gene in HepG2 cells. mRNA levels were determined using real-time quantitative RT- PCR for three normal livers and HepG2 cells. Down-regulation of FGA, FGB, or FGG in HepG2 cells was performed by the addition of siRNA corresponding to each of the three genes, and the mRNA levels determined in the cells and the secreted fibrinogen concentration in media. The mRNA level of normal human liver was FGA=FGB>FGG and the FGG mRNA level was about 2-fold lower than the others, that of HepG2 cells was FGA>FGG>FGB and FGA mRNA was approximately 2- or 4-fold higher than FGG mRNA and FGB mRNA. When FGA, FGB, or FGG mRNA expression levels were down-regulated by nearby 50%, fibrinogen concentrations in media were 78%, 49%, or 57% of the control, respectively. Our results suggest that FGG mRNA levels limit fibrinogen expression in normal liver and HepG2 cells and that 50% reduction of FGA mRNA levels would not limit fibrinogen expression in normal liver and HepG2 cells. Copyright © 2013 Elsevier Ltd. All rights reserved.

  20. Amount and source of dietary copper affects small intestine morphology, duodenal lipid peroxidation, hepatic oxidative stress,and mRNA expression of hepatic copper regulatory proteins in weanling pigs.

    Science.gov (United States)

    Fry, R S; Ashwell, M S; Lloyd, K E; O'Nan, A T; Flowers, W L; Stewart, K R; Spears, J W

    2012-09-01

    Thirty weanling, crossbred barrows (SUS SCROFA) were used to determine the effects of amount and source of dietary Cu on small intestinal morphology and lipid peroxidation, Cu metabolism, and mRNA expression of proteins involved in hepatic Cu homeostasis. At 21 d of age, pigs were stratified by BW (6.33 ± 0.23 kg) and allocated to 1 of the following dietary treatments: i) control (no supplemental Cu; 6.7 mg Cu/kg), ii) 225 mg supplemental Cu/kg diet from Cu sulfate (CuSO(4)), or iii) 225 mg supplemental Cu/kg diet from tribasic Cu chloride (TBCC). Pigs were housed 2 pigs per pen and were fed a 3-phase diet regimen until d 35 or 36 of the study. During harvest, bile and liver were obtained for mineral analysis, and liver samples were also obtained for analysis of liver glutathione (GSH) and mRNA expression of Cu regulatory proteins. Segments of duodenum, proximal jejunum, and ileum were obtained for mucosal morphology, and duodenal mucosal scrapings were collected from all pigs for analysis of malondialdehyde (MDA). Duodenal villus height was reduced in CuSO(4) pigs compared with control (P = 0.001) and TBCC (P = 0.03) pigs. Villus height in the proximal jejunum of CuSO(4) pigs was reduced (P = 0.03) compared with control pigs, but ileal villus height was not affected (P = 0.82) by treatment. Duodenal MDA concentrations were greater (P = 0.03) in CuSO(4) pigs and tended to be greater (P = 0.10) in pigs supplemented with TBCC compared with control pigs. Liver Cu was greater (P = 0.01) in CuSO(4) vs. control pigs, and tended (P = 0.07) to be greater in TBCC pigs than control pigs. Bile Cu concentrations were greater (P stress in the duodenum than CuSO(4). Feeding weanling pigs increased Cu resulted in modulation of certain Cu transporters and chaperones at the transcription level.

  1. Does MW Radiation Affect Gene Expression, Apoptotic Level, and Cell Cycle Progression of Human SH-SY5Y Neuroblastoma Cells?

    Science.gov (United States)

    Kayhan, Handan; Esmekaya, Meric Arda; Saglam, Atiye Seda Yar; Tuysuz, Mehmed Zahid; Canseven, Ayşe Gulnihal; Yagci, Abdullah Munci; Seyhan, Nesrin

    2016-06-01

    Neuroblastoma (NB) is a cancer that occurs in sympathetic nervous system arising from neuroblasts and nerve tissue of the adrenal gland, neck, chest, or spinal cord. It is an embryonal malignancy and affects infants and children. In this study, we investigated the effects of microwave (MW) radiation on apoptotic activity, cell viability, and cell cycle progression in human SH-SY5Y NB cells which can give information about MW radiation effects on neural cells covering the period from the embryonic stages to infants. SH-SY5Y NB cells were exposed to 2.1 GHz W-CDMA modulated MW radiation for 24 h at a specific absorption rate of 0.491 W/kg. Control samples were in the same conditions with MW-exposed samples but they were not exposed to MW radiation. The apoptotic activity of cells was measured by Annexin-V-FITC and propidium iodide staining. Moreover, mRNA levels of proliferative and cell cycle proteins were determined by real-time RT-PCR. The change in cell cycle progression was observed by using CycleTest-Plus DNA reagent. No significant change was observed in apoptotic activity of MW-exposed cells compared to control cells. The mRNA levels of c-myc and cyclin D1 were significantly reduced in MW group (p cells in G1 phase was significantly higher than the percentage of control cells in G1 phase. MW radiation caused cell cycle arrest in G1 phase. These results showed that 2.1 GHz W-CDMA modulated MW radiation did not cause apoptotic cell death but changed cell cycle progression.

  2. Inhibition of bromodomain and extra-terminal (BET proteins increases NKG2D ligand MICA expression and sensitivity to NK cell-mediated cytotoxicity in multiple myeloma cells: role of cMYC-IRF4-miR-125b interplay

    Directory of Open Access Journals (Sweden)

    Maria Pia Abruzzese

    2016-12-01

    Full Text Available Abstract Background Anti-cancer immune responses may contribute to the control of tumors after conventional chemotherapy, and different observations have indicated that chemotherapeutic agents can induce immune responses resulting in cancer cell death and immune-stimulatory side effects. Increasing experimental and clinical evidence highlight the importance of natural killer (NK cells in immune responses toward multiple myeloma (MM, and combination therapies able to enhance the activity of NK cells against MM are showing promise in treating this hematologic cancer. The epigenetic readers of acetylated histones bromodomain and extra-terminal (BET proteins are critical regulators of gene expression. In cancer, they can upregulate transcription of key oncogenes such as cMYC, IRF4, and BCL-2. In addition, the activity of these proteins can regulate the expression of osteoclastogenic cytokines during cancer progression. Here, we investigated the effect of BET bromodomain protein inhibition, on the expression of NK cell-activating ligands in MM cells. Methods Five MM cell lines [SKO-007(J3, U266, RPMI-8226, ARP-1, JJN3] and CD138+ MM cells isolated from MM patients were used to investigate the activity of BET bromodomain inhibitors (BETi (JQ1 and I-BET151 and of the selective BRD4-degrader proteolysis targeting chimera (PROTAC (ARV-825, on the expression and function of several NK cell-activating ligands (NKG2DLs and DNAM-1Ls, using flow cytometry, real-time PCR, transient transfections, and degranulation assays. Results Our results indicate that inhibition of BET proteins via small molecule inhibitors or their degradation via a hetero-bifunctional PROTAC probe can enhance the expression of MICA, a ligand of the NKG2D receptor, in human MM cell lines and primary malignant plasma cells, rendering myeloma cells more efficient to activate NK cell degranulation. Noteworthy, similar results were obtained using selective CBP/EP300 bromodomain inhibition

  3. The siRNA-mediated knockdown of GluN3A in 46C-derived neural stem cells affects mRNA expression levels of neural genes, including known iGluR interactors

    Science.gov (United States)

    Eilebrecht, Elke; Schöneborn, Hendrik; Neumann, Sebastian; Benecke, Arndt G.; Hollmann, Michael

    2018-01-01

    For years, GluN3A was solely considered to be a dominant-negative modulator of NMDARs, since its incorporation into receptors alters hallmark features of conventional NMDARs composed of GluN1/GluN2 subunits. Only recently, increasing evidence has accumulated that GluN3A plays a more diversified role. It is considered to be critically involved in the maturation of glutamatergic synapses, and it might act as a molecular brake to prevent premature synaptic strengthening. Its expression pattern supports a putative role during neural development, since GluN3A is predominantly expressed in early pre- and postnatal stages. In this study, we used RNA interference to efficiently knock down GluN3A in 46C-derived neural stem cells (NSCs) both at the mRNA and at the protein level. Global gene expression profiling upon GluN3A knockdown revealed significantly altered expression of a multitude of neural genes, including genes encoding small GTPases, retinal proteins, and cytoskeletal proteins, some of which have been previously shown to interact with GluN3A or other iGluR subunits. Canonical pathway enrichment studies point at important roles of GluN3A affecting key cellular pathways involved in cell growth, proliferation, motility, and survival, such as the mTOR pathway. This study for the first time provides insights into transcriptome changes upon the specific knockdown of an NMDAR subunit in NSCs, which may help to identify additional functions and downstream pathways of GluN3A and GluN3A-containing NMDARs. PMID:29438442

  4. AKT (v-akt murine thymoma viral oncogene homolog 1) and N-Ras (neuroblastoma ras viral oncogene homolog) coactivation in the mouse liver promotes rapid carcinogenesis by way of mTOR (mammalian target of rapamycin complex 1), FOXM1 (forkhead box M1)/SKP2, and c-Myc pathways.

    Science.gov (United States)

    Ho, Coral; Wang, Chunmei; Mattu, Sandra; Destefanis, Giulia; Ladu, Sara; Delogu, Salvatore; Armbruster, Julia; Fan, Lingling; Lee, Susie A; Jiang, Lijie; Dombrowski, Frank; Evert, Matthias; Chen, Xin; Calvisi, Diego F

    2012-03-01

    Activation of v-akt murine thymoma viral oncogene homolog (AKT) and Ras pathways is often implicated in carcinogenesis. However, the oncogenic cooperation between these two cascades in relationship to hepatocellular carcinoma (HCC) development remains undetermined. To investigate this issue, we generated a mouse model characterized by combined overexpression of activated forms of AKT and neuroblastoma Ras viral oncogene homolog (N-Ras) protooncogenes in the liver by way of hydrodynamic gene transfer. The molecular mechanisms underlying crosstalk between AKT and N-Ras were assessed in the mouse model and further evaluated in human and murine HCC cell lines. We found that coexpression of AKT and N-Ras resulted in a dramatic acceleration of liver tumor development when compared with mice overexpressing AKT alone, whereas N-Ras alone did not lead to tumor formation. At the cellular level, concomitant up-regulation of AKT and N-Ras resulted in increased proliferation and microvascularization when compared with AKT-injected mice. Mechanistic studies suggested that accelerated hepatocarcinogenesis driven by AKT and N-Ras resulted from a strong activation of mammalian target of rapamycin complex 1 (mTORC1). Furthermore, elevated expression of FOXM1/SKP2 and c-Myc also contributed to rapid tumor growth in AKT/Ras mice, yet by way of mTORC1-independent mechanisms. The biological effects of coactivation of AKT and N-Ras were then recapitulated in vitro using HCC cell lines, which supports the functional significance of mTORC1, FOXM1/SKP2, and c-Myc signaling cascades in mediating AKT and N-Ras-induced liver tumor development. Our data demonstrate the in vivo crosstalk between the AKT and Ras pathways in promoting liver tumor development, and the pivotal role of mTORC1-dependent and independent pathways in mediating AKT and Ras induced hepatocarcinogenesis. Copyright © 2011 American Association for the Study of Liver Diseases.

  5. High frequency electromagnetic fields (GSM signals) affect gene expression levels in tumor suppressor p53-deficient embryonic stem cells.

    Science.gov (United States)

    Czyz, Jaroslaw; Guan, Kaomei; Zeng, Qinghua; Nikolova, Teodora; Meister, Armin; Schönborn, Frank; Schuderer, Jürgen; Kuster, Niels; Wobus, Anna M

    2004-05-01

    Effects of electromagnetic fields (EMF) simulating exposure to the Global System for Mobile Communications (GSM) signals were studied using pluripotent embryonic stem (ES) cells in vitro. Wild-type ES cells and ES cells deficient for the tumor suppressor p53 were exposed to pulse modulated EMF at 1.71 GHz, lower end of the uplink band of GSM 1800, under standardized and controlled conditions, and transcripts of regulatory genes were analyzed during in vitro differentiation. Two dominant GSM modulation schemes (GSM-217 and GSM-Talk), which generate temporal changes between GSM-Basic (active during talking phases) and GSM-DTX (active during listening phases thus simulating a typical conversation), were applied to the cells at and below the basic safety limits for local exposures as defined for the general public by the International Commission on Nonionizing Radiation Protection (ICNIRP). GSM-217 EMF induced a significant upregulation of mRNA levels of the heat shock protein, hsp70 of p53-deficient ES cells differentiating in vitro, paralleled by a low and transient increase of c-jun, c-myc, and p21 levels in p53-deficient, but not in wild-type cells. No responses were observed in either cell type after EMF exposure to GSM-Talk applied at similar slot-averaged specific absorption rates (SAR), but at lower time-averaged SAR values. Cardiac differentiation and cell cycle characteristics were not affected in embryonic stem and embryonic carcinoma cells after exposure to GSM-217 EMF signals. Our data indicate that the genetic background determines cellular responses to GSM modulated EMF. Bioelectromagnetics 25:296-307, 2004. Copyright 2004 Wiley-Liss, Inc.

  6. The VSV polymerase can initiate at mRNA start sites located either up or downstream of a transcription termination signal but size of the intervening intergenic region affects efficiency of initiation.

    Science.gov (United States)

    Barr, J N; Tang, Xiaoling; Hinzman, Edward; Shen, Ruizhong; Wertz, Gail W

    2008-05-10

    Transcription by the vesicular stomatitis virus (VSV) polymerase has been characterized as obligatorily sequential with transcription of each downstream gene dependent on termination of the gene immediately upstream. In studies described here we investigated the ability of the VSV RNA-dependent RNA polymerase (RdRp) to access mRNA initiation sites located at increasing distances either downstream or upstream of a transcription termination signal. Bi-cistronic subgenomic replicons were constructed containing progressively extended intergenic regions preceding the initiation site of a downstream gene. The ability of the RdRp to access the downstream sites was progressively reduced as the length of the intergenic region increased. Alternatively, bi-cistronic replicons were constructed containing an mRNA start signal located at increasing distances upstream of a termination site. Analysis of transcription of these "overlapped" genes showed that for an upstream mRNA start site to be recognized it had to contain not only the canonical 3'-UUGUCnnUAG-5' gene start signal, but that signal needed also to be preceded by a U7 tract. Access of these upstream mRNA initiation sites by the VSV RdRp was proportionately reduced with increasing distance between the termination site and the overlapped initiation signal. Possible mechanisms for how the RdRp accesses these upstream start sites are discussed.

  7. Wnt signaling and c-Myc in intestinal epithelium

    NARCIS (Netherlands)

    Muncan, V.

    2007-01-01

    constantly produce cells from a stem cell reservoir that give rise to proliferating transit amplifying cells, which subsequently differentiate and are positioned in their proper compartments. This process has to be under stringent control to ensure life-long tissue homeostasis. It has now become

  8. Metformin targets c-MYC oncogene to prevent prostate cancer

    OpenAIRE

    Akinyeke, Tunde; Matsumura, Satoko; Wang, Xinying; Wu, Yingjie; Schalfer, Eric D.; Saxena, Anjana; Yan, Wenbo; Logan, Susan K; Li, Xin

    2013-01-01

    Prostate cancer (PCa) is the second leading cause of cancer-related death in American men and many PCa patients develop skeletal metastasis. Current treatment modalities for metastatic PCa are mostly palliative with poor prognosis. Epidemiological studies indicated that patients receiving the diabetic drug metformin have lower PCa risk and better prognosis, suggesting that metformin may have antineoplastic effects. The mechanism by which metformin acts as chemopreventive agent to impede PCa i...

  9. mRNA Quantification of NIPBL Isoforms A and B in Adult and Fetal Human Tissues, and a Potentially Pathological Variant Affecting Only Isoform A in Two Patients with Cornelia de Lange Syndrome.

    Science.gov (United States)

    Puisac, Beatriz; Teresa-Rodrigo, María-Esperanza; Hernández-Marcos, María; Baquero-Montoya, Carolina; Gil-Rodríguez, María-Concepción; Visnes, Torkild; Bot, Christopher; Gómez-Puertas, Paulino; Kaiser, Frank J; Ramos, Feliciano J; Ström, Lena; Pié, Juan

    2017-02-23

    Cornelia de Lange syndrome (CdLS) is a congenital developmental disorder characterized by craniofacial dysmorphia, growth retardation, limb malformations, and intellectual disability. Approximately 60% of patients with CdLS carry a recognizable pathological variant in the NIPBL gene, of which two isoforms, A and B, have been identified, and which only differ in the C-terminal segment. In this work, we describe the distribution pattern of the isoforms A and B mRNAs in tissues of adult and fetal origin, by qPCR (quantitative polymerase chain reaction). Our results show a higher gene expression of the isoform A, even though both seem to have the same tissue distribution. Interestingly, the expression in fetal tissues is higher than that of adults, especially in brain and skeletal muscle. Curiously, the study of fibroblasts of two siblings with a mild CdLS phenotype and a pathological variant specific of the isoform A of NIPBL (c.8387A > G; P.Tyr2796Cys), showed a similar reduction in both isoforms, and a normal sensitivity to DNA damage. Overall, these results suggest that the position of the pathological variant at the 3´ end of the NIPBL gene affecting only isoform A, is likely to be the cause of the atypical mild phenotype of the two brothers.

  10. mRNA Quantification of NIPBL Isoforms A and B in Adult and Fetal Human Tissues, and a Potentially Pathological Variant Affecting Only Isoform A in Two Patients with Cornelia de Lange Syndrome

    Directory of Open Access Journals (Sweden)

    Beatriz Puisac

    2017-02-01

    Full Text Available Cornelia de Lange syndrome (CdLS is a congenital developmental disorder characterized by craniofacial dysmorphia, growth retardation, limb malformations, and intellectual disability. Approximately 60% of patients with CdLS carry a recognizable pathological variant in the NIPBL gene, of which two isoforms, A and B, have been identified, and which only differ in the C-terminal segment. In this work, we describe the distribution pattern of the isoforms A and B mRNAs in tissues of adult and fetal origin, by qPCR (quantitative polymerase chain reaction. Our results show a higher gene expression of the isoform A, even though both seem to have the same tissue distribution. Interestingly, the expression in fetal tissues is higher than that of adults, especially in brain and skeletal muscle. Curiously, the study of fibroblasts of two siblings with a mild CdLS phenotype and a pathological variant specific of the isoform A of NIPBL (c.8387A > G; P.Tyr2796Cys, showed a similar reduction in both isoforms, and a normal sensitivity to DNA damage. Overall, these results suggest that the position of the pathological variant at the 3´ end of the NIPBL gene affecting only isoform A, is likely to be the cause of the atypical mild phenotype of the two brothers.

  11. Nebulisation of IVT mRNA Complexes for Intrapulmonary Administration.

    Directory of Open Access Journals (Sweden)

    Sarah M Johler

    Full Text Available During the last years the potential role of in vitro transcribed (IVT mRNA as a vehicle to deliver genetic information has come into focus. IVT mRNA could be used for anti-cancer therapies, vaccination purposes, generation of pluripotent stem cells and also for genome engineering or protein replacement. However, the administration of IVT mRNA into the target organ is still challenging. The lung with its large surface area is not only of interest for delivery of genetic information for treatment of e.g. for cystic fibrosis or alpha-1-antitrypsin deficiency, but also for vaccination purposes. Administration of IVT mRNA to the lung can be performed by direct intratracheal instillation or by aerosol inhalation/nebulisation. The latter approach shows a non-invasive tool, although it is not known, if IVT mRNA is resistant during the process of nebulisation. Therefore, we investigated the transfection efficiency of non-nebulised and nebulised IVT mRNA polyplexes and lipoplexes in human bronchial epithelial cells (16HBE. A slight reduction in transfection efficiency was observed for lipoplexes (Lipofectamine 2000 in the nebulised part compared to the non-nebulised which can be overcome by increasing the amount of Lipofectamine. However, Lipofectamine was more than three times more efficient in transfecting 16HBE than DMRIE and linear PEI performed almost 10 times better than its branched derivative. By contrast, the nebulisation process did not affect the cationic polymer complexes. Furthermore, aerosolisation of IVT mRNA complexes did neither affect the protein duration nor the toxicity of the cationic complexes. Taken together, these data show that aerosolisation of cationic IVT mRNA complexes constitute a potentially powerful means to transfect cells in the lung with the purpose of protein replacement for genetic diseases such as cystic fibrosis or alpha-1-antitrypsin deficiency or for infectious disease vaccines, while bringing along the advantages

  12. Collagen mRNA levels changes during colorectal cancer carcinogenesis

    DEFF Research Database (Denmark)

    Skovbjerg, Hanne; Anthonsen, Dorit; Lothe, Inger M B

    2009-01-01

    . In addition, corresponding tissue was examined from healthy volunteers (n = 20). mRNA levels were normalized to beta-actin. Immunohistochemical analysis of the distributions of type IV and type VII collagens were performed on normal and affected tissues from colorectal cancer patients. RESULTS: The alpha1(IV......). The level of alpha 6(IV) was 5-fold lower in colorectal cancer tissue as compared to healthy individuals (p alpha 6(IV) mRNA coincides...... zone of stratified epithelia. Immunohistochemical studies have previously reported changes in steady-state levels of different alpha(IV) chains in several epithelial cancer types. In the present study we aimed to quantitatively determine the mRNA levels of type IV collagen (alpha1/alpha 4/alpha 6...

  13. CIP2A influences survival in colon cancer and is critical for maintaining Myc expression.

    Directory of Open Access Journals (Sweden)

    Armin Wiegering

    Full Text Available The cancerous inhibitor of protein phosphatase 2A (CIP2A is an oncogenic factor that stabilises the c-Myc protein. CIP2A is overexpressed in several tumours, and expression levels are an independent marker for long-term outcome. To determine whether CIP2A expression is elevated in colon cancer and whether it might serve as a prognostic marker for survival, we analysed CIP2A mRNA expression by real-time PCR in 104 colon cancer samples. CIP2A mRNA was overexpressed in colon cancer samples and CIP2A expression levels correlated significantly with tumour stage. We found that CIP2A serves as an independent prognostic marker for disease-free and overall survival. Further, we investigated CIP2A-dependent effects on levels of c-Myc, Akt and on cell proliferation in three colon cancer cell lines by silencing CIP2A using small interfering (si and short hairpin (sh RNAs. Depletion of CIP2A substantially inhibited growth of colon cell lines and reduced c-Myc levels without affecting expression or function of the upstream regulatory kinase, Akt. Expression of CIP2A was found to be dependent on MAPK activity, linking elevated c-Myc expression to deregulated signal transduction in colon cancer.

  14. Occupational health hazards of trichloroethylene among workers in relation to altered mRNA expression of cell cycle regulating genes (p53, p21, bax and bcl-2) and PPARA.

    Science.gov (United States)

    Varshney, Meenu; Chandra, Abhijit; Jain, Rajeev; Ahmad, Riaz; Bihari, Vipin; Chandran, C Keshava; Mudiam, Mohana K R; Patnaik, Satykam; Goel, S K

    2015-01-01

    Trichloroethylene (TCE) is widely used as a metal degreaser in industrial processes. The present study reports on the effects of TCE exposure on workers employed in the lock industries. To ensure exposure of the workers to TCE, its toxic metabolites, trichloroacetic acid (TCA), dichloroacetic acid (DCA) and trichloroethanol (TCEOH) were detected in the plasma of the subjects through solid phase microextraction-gas chromatography-electron capture detection. TCA, DCA and TCEOH were detected in the range of 0.004-2.494 μg/mL, 0.01-3.612 μg/mL and 0.002-0.617 μg/mL, respectively. Quantitative reverse transcription polymerase chain reaction analysis revealed up-regulated expression of p53 (2.4-fold; p  < 0.05), p21 (2-fold; p  < 0.01), bax (2.9-fold; p  < 0.01) mRNAs and down-regulated expression of bcl-2 (67%; p  < 0.05) mRNAs, indicating DNA damaging potential of these metabolites. No effects were observed on the levels of p16 and c-myc mRNAs. Further, as TCA and DCA, the ligand of peroxisome proliferator activated receptor alpha (PPARA), are involved in the process of hepatocarcinogenesis in rodents, we examined expression of PPARA mRNA and let-7c miRNA in the workers. No statistically significant differences in expression of PPARA mRNA and let-7c miRNA in patients were observed as compared to values in controls. Dehydroepiandosterone sulfate (DHEAS) is a reported endogenous ligand of PPARA so its competitive role was also studied. We observed decreased levels of DHEAS hormone in the subjects. Hence, its involvement in mediation of the observed changes in the levels of various mRNAs analyzed in this study appears unlikely.

  15. Self-amplifying mRNA vaccines.

    Science.gov (United States)

    Brito, Luis A; Kommareddy, Sushma; Maione, Domenico; Uematsu, Yasushi; Giovani, Cinzia; Berlanda Scorza, Francesco; Otten, Gillis R; Yu, Dong; Mandl, Christian W; Mason, Peter W; Dormitzer, Philip R; Ulmer, Jeffrey B; Geall, Andrew J

    2015-01-01

    This chapter provides a brief introduction to nucleic acid-based vaccines and recent research in developing self-amplifying mRNA vaccines. These vaccines promise the flexibility of plasmid DNA vaccines with enhanced immunogenicity and safety. The key to realizing the full potential of these vaccines is efficient delivery of nucleic acid to the cytoplasm of a cell, where it can amplify and express the encoded antigenic protein. The hydrophilicity and strong net negative charge of RNA impedes cellular uptake. To overcome this limitation, electrostatic complexation with cationic lipids or polymers and physical delivery using electroporation or ballistic particles to improve cellular uptake has been evaluated. This chapter highlights the rapid progress made in using nonviral delivery systems for RNA-based vaccines. Initial preclinical testing of self-amplifying mRNA vaccines has shown nonviral delivery to be capable of producing potent and robust innate and adaptive immune responses in small animals and nonhuman primates. Historically, the prospect of developing mRNA vaccines was uncertain due to concerns of mRNA instability and the feasibility of large-scale manufacturing. Today, these issues are no longer perceived as barriers in the widespread implementation of the technology. Currently, nonamplifying mRNA vaccines are under investigation in human clinical trials and can be produced at a sufficient quantity and quality to meet regulatory requirements. If the encouraging preclinical data with self-amplifying mRNA vaccines are matched by equivalently positive immunogenicity, potency, and tolerability in human trials, this platform could establish nucleic acid vaccines as a versatile new tool for human immunization. Copyright © 2015 Elsevier Inc. All rights reserved.

  16. Heat shock response in yeast involves changes in both transcription rates and mRNA stabilities.

    Directory of Open Access Journals (Sweden)

    Laia Castells-Roca

    Full Text Available We have analyzed the heat stress response in the yeast Saccharomyces cerevisiae by determining mRNA levels and transcription rates for the whole transcriptome after a shift from 25 °C to 37 °C. Using an established mathematical algorithm, theoretical mRNA decay rates have also been calculated from the experimental data. We have verified the mathematical predictions for selected genes by determining their mRNA decay rates at different times during heat stress response using the regulatable tetO promoter. This study indicates that the yeast response to heat shock is not only due to changes in transcription rates, but also to changes in the mRNA stabilities. mRNA stability is affected in 62% of the yeast genes and it is particularly important in shaping the mRNA profile of the genes belonging to the environmental stress response. In most cases, changes in transcription rates and mRNA stabilities are homodirectional for both parameters, although some interesting cases of antagonist behavior are found. The statistical analysis of gene targets and sequence motifs within the clusters of genes with similar behaviors shows that both transcriptional and post-transcriptional regulons apparently contribute to the general heat stress response by means of transcriptional factors and RNA binding proteins.

  17. A selective splicing variant of hepcidin mRNA in hepatocellular carcinoma cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Toki, Yasumichi [Division of Gastroenterology and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, Hokkaido 078-8510 (Japan); Sasaki, Katsunori, E-mail: k-sasaki@asahikawa-med.ac.jp [Department of Gastrointestinal Immunology and Regenerative Medicine, Asahikawa Medical University, Hokkaido 078-8510 (Japan); Tanaka, Hiroki [Department of Legal Medicine, Asahikawa Medical University, Hokkaido 078-8510 (Japan); Yamamoto, Masayo; Hatayama, Mayumi; Ito, Satoshi; Ikuta, Katsuya; Shindo, Motohiro; Hasebe, Takumu; Nakajima, Shunsuke; Sawada, Koji; Fujiya, Mikihiro [Division of Gastroenterology and Hematology/Oncology, Department of Medicine, Asahikawa Medical University, Hokkaido 078-8510 (Japan); Torimoto, Yoshihiro [Oncology Center, Asahikawa Medical University Hospital, Hokkaido 078-8510 (Japan); Ohtake, Takaaki; Kohgo, Yutaka [Department of Gastroenterology, International University of Health and Welfare Hospital, Tochigi 329-2763 (Japan)

    2016-08-05

    Hepcidin is a main regulator of iron metabolism, of which abnormal expression affects intestinal absorption and reticuloendothelial sequestration of iron by interacting with ferroportin. It is also noted that abnormal iron accumulation is one of the key factors to facilitate promotion and progression of cancer including hepatoma. By RT-PCR/agarose gel electrophoresis of hepcidin mRNA in a hepatocellular carcinoma cell line HLF, a smaller mRNA band was shown in addition to the wild-type hepcidin mRNA. From sequencing analysis, this additional band was a selective splicing variant of hepcidin mRNA lacking exon 2 of HAMP gene, producing the transcript that encodes truncated peptide lacking 20 amino acids at the middle of preprohepcidin. In the present study, we used the digital PCR, because such a small amount of variant mRNA was difficult to quantitate by the conventional RT-PCR amplification. Among seven hepatoma-derived cell lines, six cell lines have significant copy numbers of this variant mRNA, but not in one cell line. In the transient transfection analysis of variant-type hepcidin cDNA, truncated preprohepcidin has a different character comparing with native preprohepcidin: its product is insensitive to digestion, and secreted into the medium as a whole preprohepcidin form without maturation. Loss or reduction of function of HAMP gene by aberrantly splicing may be a suitable phenomenon to obtain the proliferating advantage of hepatoma cells. - Highlights: • An aberrant splicing variant of hepcidin mRNA lacking exon 2 of HAMP gene. • Absolute quantification of hepcidin mRNA by digital PCR amplification. • Hepatoma-derived cell lines have significant copies of variant-type hepcidin mRNA. • Truncated preprohepcidin is secreted from cells without posttranslational cleavage.

  18. Impairment of FOS mRNA stabilization following translation arrest in granulocytes from myelodysplastic syndrome patients.

    Science.gov (United States)

    Feng, Xiaomin; Shikama, Yayoi; Shichishima, Tsutomu; Noji, Hideyoshi; Ikeda, Kazuhiko; Ogawa, Kazuei; Kimura, Hideo; Takeishi, Yasuchika; Kimura, Junko

    2013-01-01

    Although quantitative and qualitative granulocyte defects have been described in myelodysplastic syndromes (MDS), the underlying molecular basis of granulocyte dysfunction in MDS is largely unknown. We recently found that FOS mRNA elevation under translation-inhibiting stimuli was significantly smaller in granulocytes from MDS patients than in healthy individuals. The aim of this study is to clarify the cause of the impaired FOS induction in MDS. We first examined the mechanisms of FOS mRNA elevation using granulocytes from healthy donors cultured with the translation inhibitor emetine. Emetine increased both transcription and mRNA stability of FOS. p38 MAPK inhibition abolished the emetine-induced increase of FOS transcription but did not affect FOS mRNA stabilization. The binding of an AU-rich element (ARE)-binding protein HuR to FOS mRNA containing an ARE in 3'UTR was increased by emetine, and the knockdown of HuR reduced the FOS mRNA stabilizing effect of emetine. We next compared the emetine-induced transcription and mRNA stabilization of FOS between MDS patients and healthy controls. Increased rates of FOS transcription by emetine were similar in MDS and controls. In the absence of emetine, FOS mRNA decayed to nearly 17% of initial levels in 45 min in both groups. In the presence of emetine, however, 76.7±19.8% of FOS mRNA remained after 45 min in healthy controls, versus 37.9±25.5% in MDS (Pgranulocytes.

  19. Viscum album-mediated COX-2 inhibition implicates destabilization of COX-2 mRNA.

    Science.gov (United States)

    Saha, Chaitrali; Hegde, Pushpa; Friboulet, Alain; Bayry, Jagadeesh; Kaveri, Srinivas V

    2015-01-01

    Extensive use of Viscum album (VA) preparations in the complementary therapy of cancer and in several other human pathologies has led to an increasing number of cellular and molecular approaches to explore the mechanisms of action of VA. We have recently demonstrated that, VA preparations exert a potent anti-inflammatory effect by selectively down-regulating the COX-2-mediated cytokine-induced secretion of prostaglandin E2 (PGE2), one of the important molecular signatures of inflammatory reactions. In this study, we observed a significant down-regulation of COX-2 protein expression in VA-treated A549 cells however COX-2 mRNA levels were unaltered. Therefore, we hypothesized that VA induces destabilisation of COX-2 mRNA, thereby depleting the available functional COX-2 mRNA for the protein synthesis and for the subsequent secretion of PGE2. To address this question, we analyzed the molecular degradation of COX-2 protein and its corresponding mRNA in A549 cell line. Using cyclohexamide pulse chase experiment, we demonstrate that, COX-2 protein degradation is not affected by the treatment with VA whereas experiments on transcriptional blockade with actinomycin D, revealed a marked reduction in the half life of COX-2 mRNA due to its rapid degradation in the cells treated with VA compared to that in IL-1β-stimulated cells. These results thus demonstrate that VA-mediated inhibition of PGE2 implicates destabilization of COX-2 mRNA.

  20. Viscum album-mediated COX-2 inhibition implicates destabilization of COX-2 mRNA.

    Directory of Open Access Journals (Sweden)

    Chaitrali Saha

    Full Text Available Extensive use of Viscum album (VA preparations in the complementary therapy of cancer and in several other human pathologies has led to an increasing number of cellular and molecular approaches to explore the mechanisms of action of VA. We have recently demonstrated that, VA preparations exert a potent anti-inflammatory effect by selectively down-regulating the COX-2-mediated cytokine-induced secretion of prostaglandin E2 (PGE2, one of the important molecular signatures of inflammatory reactions. In this study, we observed a significant down-regulation of COX-2 protein expression in VA-treated A549 cells however COX-2 mRNA levels were unaltered. Therefore, we hypothesized that VA induces destabilisation of COX-2 mRNA, thereby depleting the available functional COX-2 mRNA for the protein synthesis and for the subsequent secretion of PGE2. To address this question, we analyzed the molecular degradation of COX-2 protein and its corresponding mRNA in A549 cell line. Using cyclohexamide pulse chase experiment, we demonstrate that, COX-2 protein degradation is not affected by the treatment with VA whereas experiments on transcriptional blockade with actinomycin D, revealed a marked reduction in the half life of COX-2 mRNA due to its rapid degradation in the cells treated with VA compared to that in IL-1β-stimulated cells. These results thus demonstrate that VA-mediated inhibition of PGE2 implicates destabilization of COX-2 mRNA.

  1. mRNA transcripts as molecular biomarkers in medicine and nutrition

    Science.gov (United States)

    Sunde, Roger A.

    2010-01-01

    In medicine, mRNA transcripts are being developed as molecular biomarkers for the diagnosis and treatment of a number of diseases. These biomarkers offer early and more accurate prediction and diagnosis of disease and disease progression, and ability to identify individuals at risk. Use of microarrays also offers opportunity to identify orthogonal (uncorrelated) biomarkers not known to be linked with conventional biomarkers. Investigators are increasingly using blood as a surrogate tissue for biopsy and analysis; total RNA isolated from whole blood is predominantly from erythroid cells, and whole blood mRNA share more than 80% of the transcriptome with major tissues. Thus blood mRNA biomarkers for individualized disease prediction and diagnosis are an exciting area in medicine; mRNA biomarkers in nutrition have potential application that parallel these opportunities. Assessment of selenium (Se) status and requirements is one area where tissue mRNA levels have been used successfully. Selenoprotein-H and selenoprotein-W as well as glutathione peroxidase-1 (Gpx1) mRNAs are highly down-regulated in Se deficiency in rat liver, and the minimum dietary Se requirement is 0.06–0.07 μg Se/g based on these biomarkers, similar to requirements determined using conventional biomarkers. Blood Gpx1 mRNA can also be used to determine Se requirements in rats, showing that blood mRNA has potential for assessment of nutrient status. Future research is needed to develop mRNA biomarker panels for all nutrients that will discriminate between deficient, marginal, adequate, and supernutritional individuals and populations, and differentiate between individuals that will benefit versus be adversely affected by nutrient supplementation. PMID:20303730

  2. Responses of mRNA expression of PepT1 in small intestine to ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-05-04

    May 4, 2009 ... tides by duodenal fistula, and changed the concentration of circulation peptides, and then monitored how mRNA expression of Pep T1 in duodenum, ileum and jejunum tissue was affected. MATERIALS AND METHODS. Experimental animals and diets. Sixteen Chinese Saanen dairy goats, with an average ...

  3. Increased IL-10 mRNA and IL-23 mRNA expression in multiple sclerosis

    DEFF Research Database (Denmark)

    Krakauer, Martin; Sorensen, P; Khademi, M

    2008-01-01

    volunteers served to confirm initial findings. mRNA was analyzed by real-time reverse transcriptase polymerase chain reaction (PCR). RESULTS: We found elevated expression of interleukin (IL)-23 and IL-10 in untreated MS patients. IFN-beta therapy increased IL-10 and decreased IL-23 expression independently...... of the regulatory cytokine IL-10. The elevated IL-23 mRNA levels in MS patients are noteworthy in view of the newly discovered IL-23-driven Th17 T-cell subset, which is crucial in animal models of MS. Since IFN-beta therapy resulted in decreased IL-23 mRNA levels, the Th17 axis could be another target of IFN...

  4. Stress granules: sites of mRNA triage that regulate mRNA stability and translatability.

    Science.gov (United States)

    Kedersha, N; Anderson, P

    2002-11-01

    Mammalian stress granules (SGs) are cytoplasmic domains into which mRNAs are sorted dynamically in response to phosphorylation of eukaryotic initiation factor (eIF) 2alpha, a key regulatory step in translational initiation. The activation of one or more of the eIF2alpha kinases leads to SG assembly by decreasing the levels of eIF2-GTP-tRNA(Met), the ternary complex that is normally required for loading the initiator methionine onto the 48 S preinitiation complex to begin translation. This stress-induced scarcity of eIF2-GTP-tRNA(Met) allows the RNA-binding proteins TIA-1 (T-cell internal antigen-1) and TIAR (TIA-1-related protein) to bind the 48 S complex in lieu of the ternary complex, thereby promoting polysome disassembly and the concurrent routing of the mRNA into a SG. The actual formation of SGs occurs upon auto-aggregation of the prion-like C-termini of TIA-1 proteins; this aggregation is reversed in vivo by overexpression of the heat-shock protein (HSP) chaperone HSP70. Remarkably, HSP70 mRNA is excluded from SGs and is preferentially translated during stress, indicating that the RNA composition of the SG is selective. Moreover, the effects of HSP70 on TIA aggregation suggest a feedback loop whereby HSP70 synthesis is auto-regulated. Proteins that promote mRNA stability [e.g. HuR (Hu protein R)] and destabilize mRNA [i.e. tristetraprolin (TTP)] are also recruited to SGs, suggesting that SGs effect a process of mRNA triage, by promoting polysome disassembly and routing mRNAs to cytoplasmic domains enriched for HuR and TTP. This model reveals connections between the eIF2alpha kinase system, mRNA stability and cellular chaperone levels.

  5. mRNA quality control pathways in Saccharomyces cerevisiae

    Indian Academy of Sciences (India)

    2013-07-10

    Jul 10, 2013 ... mRNPs in the nucleus very often leads to the formation of aberrant and faulty messages along with their functional .... Figure 1. mRNA life-cycle in eukaryotic cell: Schematic view of the nuclear and cytoplasmic phases of mRNA life cycle, namely ..... structure is characteristic and critical feature of an mRNA.

  6. Messenger RNA (mRNA) nanoparticle tumour vaccination

    Science.gov (United States)

    Phua, Kyle K. L.; Nair, Smita K.; Leong, Kam W.

    2014-06-01

    Use of mRNA-based vaccines for tumour immunotherapy has gained increasing attention in recent years. A growing number of studies applying nanomedicine concepts to mRNA tumour vaccination show that the mRNA delivered in nanoparticle format can generate a more robust immune response. Advances in the past decade have deepened our understanding of gene delivery barriers, mRNA's biological stability and immunological properties, and support the notion for engineering innovations tailored towards a more efficient mRNA nanoparticle vaccine delivery system. In this review we will first examine the suitability of mRNA for engineering manipulations, followed by discussion of a model framework that highlights the barriers to a robust anti-tumour immunity mediated by mRNA encapsulated in nanoparticles. Finally, by consolidating existing literature on mRNA nanoparticle tumour vaccination within the context of this framework, we aim to identify bottlenecks that can be addressed by future nanoengineering research.

  7. Mammalian tissues defective in nonsense-mediated mRNA decay display highly aberrant splicing patterns

    DEFF Research Database (Denmark)

    Weischenfeldt, Joachim Lütken; Waage, Johannes Eichler; Tian, Geng

    2012-01-01

    a bioinformatic pipeline that maps RNA-seq data to a combinatorial exon database, predicts NMD-susceptibility for mRNA isoforms and calculates the distribution of major splice isoform classes. We present a catalog of NMD-regulated alternative splicing events, showing that isoforms of 30% of all expressed genes......ABSTRACT: BACKGROUND: Nonsense-mediated mRNA decay (NMD) affects the outcome of alternative splicing by degrading mRNA isoforms with premature termination codons. Splicing regulators constitute important NMD targets; however, the extent to which loss of NMD causes extensive deregulation...... of alternative splicing has not previously been assayed in a global, unbiased manner. Here, we combine mouse genetics and RNA-seq to provide the first in vivo analysis of the global impact of NMD on splicing patterns in two primary mouse tissues ablated for the NMD factor UPF2. RESULTS: We developed...

  8. EXPRESSION OF AID-SPECIFIC mRNA IN PERIPHERAL BLOOD LYMPHOCYTES IN BRONCHIAL ASTHMA

    Directory of Open Access Journals (Sweden)

    V. N. Mineev

    2015-01-01

    Full Text Available The aim of this study was to evaluate a possible role of AID (AICDA – activation-induced (cytidine deaminase in the bronchial asthma (BA pathogenesis. Materials and methods. We have examined twelve healthy control persons, forty-two patients with allergic bronchial asthma (ABA and twenty-seven patients with non-allergic bronchial asthma (NABA. The AID mRNA expression was evaluated by means of RT-PCR. Results: AID mRNA was more significantly expressed in BA, than in healthy controls. Meanwhile, no significant differences were revealed between ABA and NABA groups. Correlation analysis of AID mRNA expression levels in peripheral blood lymphocytes has shown that CHε mRNA and total serum IgE revealed significant negative correlation, in the NABA group only. The levels of AID mRNA expression in peripheral blood lymphocytes exhibited positive and significant correlations with clinical characteristics, reflecting severity and stage of the disease, in BA patients. Moreover, a significant positive correlation was revealed with eosinophil contents in sputum. Conclusion. It was concluded that normal regulation of IgE class switching normally based on feedback regulation, was impaired in ABA but not affected in NABA. 

  9. Perylene and coronene derivatives binding to G-rich promoter oncogene sequences efficiently reduce their expression in cancer cells.

    Science.gov (United States)

    Micheli, Emanuela; Altieri, Alessandro; Cianni, Lorenzo; Cingolani, Chiara; Iachettini, Sara; Bianco, Armandodoriano; Leonetti, Carlo; Cacchione, Stefano; Biroccio, Annamaria; Franceschin, Marco; Rizzo, Angela

    2016-06-01

    A novel approach to cancer therapeutics is emerging in the field of G-quadruplex (G4) ligands, small molecules designed to stabilize four-stranded structures that can form at telomeres as well as in other genomic sequences, including oncogene promoter sequences, 5'-UTR regions and introns. In this study, we investigated the binding activity of perylene and coronene derivatives PPL3C, CORON and EMICORON to G4 structures formed within the promoter regions of two important cancer-related genes, c-MYC and BCL-2, and their biochemical effects on gene and protein expression. In order to fully characterize the ability of the selected ligands to bind and stabilize the G4 structures originated by the c-MYC and BCL-2 promoter sequences, we performed electrospray ionization mass spectrometry (ESI-MS), Fluorescence Resonance Energy Transfer (FRET) measurements, Circular Dichroism (CD) spectra and polymerase stop assay. Altogether our results showed that the ligands had a high capacity in binding and stabilizing the G4 structures within the c-MYC and BCL-2 promoter sequences in vitro. Notably, when we evaluated by quantitative real-time PCR and western blotting analysis, the effects of treatment with the different G4 ligands on c-MYC and BCL2 expression in a human melanoma cell line, EMICORON appeared the most effective compound in reducing the mRNA and protein levels of both genes. These results encourage to consider EMICORON as a promising example of multimodal class of an antineoplastic drug, affecting different tumor crucial pathways simultaneously: telomere maintenance (as previously described), cell proliferation and apoptosis via down-regulation of both c-MYC and BCL-2 (this paper). Copyright © 2016 Elsevier B.V. and Société française de biochimie et biologie Moléculaire (SFBBM). All rights reserved.

  10. Ammonium Chloride Ingestion Attenuates Exercise-Induced mRNA Levels in Human Muscle.

    Directory of Open Access Journals (Sweden)

    Johann Edge

    Full Text Available Minimizing the decrease in intracellular pH during high-intensity exercise training promotes greater improvements in mitochondrial respiration. This raises the intriguing hypothesis that pH may affect the exercise-induced transcription of genes that regulate mitochondrial biogenesis. Eight males performed 10x2-min cycle intervals at 80% VO2speak intensity on two occasions separated by ~2 weeks. Participants ingested either ammonium chloride (ACID or calcium carbonate (PLA the day before and on the day of the exercise trial in a randomized, counterbalanced order, using a crossover design. Biopsies were taken from the vastus lateralis muscle before and after exercise. The mRNA level of peroxisome proliferator-activated receptor co-activator 1α (PGC-1α, citrate synthase, cytochome c and FOXO1 was elevated at rest following ACID (P0.05; the difference in PGC-1α mRNA content 2 h post-exercise between ACID and PLA was not significant (P = 0.08. Thus, metabolic acidosis abolished the early post-exercise increase of PGC-1α mRNA and the mRNA of downstream mitochondrial and glucose-regulating proteins. These findings indicate that metabolic acidosis may affect mitochondrial biogenesis, with divergent responses in resting and post-exercise skeletal muscle.

  11. Conventional and unconventional mechanisms for capping viral mRNA.

    Science.gov (United States)

    Decroly, Etienne; Ferron, François; Lescar, Julien; Canard, Bruno

    2011-12-05

    In the eukaryotic cell, capping of mRNA 5' ends is an essential structural modification that allows efficient mRNA translation, directs pre-mRNA splicing and mRNA export from the nucleus, limits mRNA degradation by cellular 5'-3' exonucleases and allows recognition of foreign RNAs (including viral transcripts) as 'non-self'. However, viruses have evolved mechanisms to protect their RNA 5' ends with either a covalently attached peptide or a cap moiety (7-methyl-Gppp, in which p is a phosphate group) that is indistinguishable from cellular mRNA cap structures. Viral RNA caps can be stolen from cellular mRNAs or synthesized using either a host- or virus-encoded capping apparatus, and these capping assemblies exhibit a wide diversity in organization, structure and mechanism. Here, we review the strategies used by viruses of eukaryotic cells to produce functional mRNA 5'-caps and escape innate immunity.

  12. The functional half-life of an mRNA depends on the ribosome spacing in an early coding region

    DEFF Research Database (Denmark)

    Pedersen, Margit; Nissen, Søren; Mitarai, Namiko

    2011-01-01

    Bacterial mRNAs are translated by closely spaced ribosomes and degraded from the 5'-end, with half-lives of around 2 min at 37 °C in most cases. Ribosome-free or "naked" mRNA is known to be readily degraded, but the initial event that inactivates the mRNA functionally has not been fully described....... Here, we characterize a determinant of the functional stability of an mRNA, which is located in the early coding region. Using literature values for the mRNA half-lives of variant lacZ mRNAs in Escherichia coli, we modeled how the ribosome spacing is affected by the translation rate of the individual...... codons. When comparing the ribosome spacing at various segments of the mRNA to its functional half-life, we found a clear correlation between the functional mRNA half-life and the ribosome spacing in the mRNA region approximately between codon 20 and codon 45. From this finding, we predicted that inserts...

  13. mRNA vaccines - a new era in vaccinology.

    Science.gov (United States)

    Pardi, Norbert; Hogan, Michael J; Porter, Frederick W; Weissman, Drew

    2018-01-12

    mRNA vaccines represent a promising alternative to conventional vaccine approaches because of their high potency, capacity for rapid development and potential for low-cost manufacture and safe administration. However, their application has until recently been restricted by the instability and inefficient in vivo delivery of mRNA. Recent technological advances have now largely overcome these issues, and multiple mRNA vaccine platforms against infectious diseases and several types of cancer have demonstrated encouraging results in both animal models and humans. This Review provides a detailed overview of mRNA vaccines and considers future directions and challenges in advancing this promising vaccine platform to widespread therapeutic use.

  14. The yeast POP2 gene encodes a nuclease involved in mRNA deadenylation.

    Science.gov (United States)

    Daugeron, M C; Mauxion, F; Séraphin, B

    2001-06-15

    The major mRNA degradation pathway involves deadenylation of the target molecule followed by decapping and, finally, 5'-->3' exonuclease digestion of the mRNA body. While yeast factors involved in the decapping and exonuclease degradation steps have been identified, the nature of the factor(s) involved in the deadenylation step remained elusive. Database searches for yeast proteins related to the mammalian deadenylase PARN identified the Pop2 protein (Pop2p) as a potential deadenylase. While Pop2p was previously identified as a factor affecting transcription, we identified a non-canonical RNase D sequence signature in its sequence. Analysis of the fate of a reporter mRNA in a pop2 mutant demonstrates that Pop2p is required for efficient mRNA degradation in vivo. Characterisation of mRNA degradation intermediates accumulating in this mutant supports the involvement of Pop2p in mRNA deadenylation in vivo. Similar phenotypes are observed in yeast strains lacking the Ccr4 protein, which is known to be associated with Pop2p. A recombinant Pop2p fragment encompassing the putative catalytic domain degrades poly(A) in vitro demonstrating that Pop2p is a nuclease. We also demonstrate that poly(A) is a better competitor than poly(G) or poly(C) of the Pop2p nuclease activity. Altogether, our study indicates that Pop2p is a nuclease subunit of the yeast deadenylase and suggests that Pop2p homologues in other species may have similar functions.

  15. Novel RNA-binding activity of NQO1 promotes SERPINA1 mRNA translation.

    Science.gov (United States)

    Di Francesco, Andrea; Di Germanio, Clara; Panda, Amaresh C; Huynh, Phu; Peaden, Robert; Navas-Enamorado, Ignacio; Bastian, Paul; Lehrmann, Elin; Diaz-Ruiz, Alberto; Ross, David; Siegel, David; Martindale, Jennifer L; Bernier, Michel; Gorospe, Myriam; Abdelmohsen, Kotb; de Cabo, Rafael

    2016-10-01

    NAD(P)H: quinone oxidoreductase (NQO1) is essential for cell defense against reactive oxidative species, cancer, and metabolic stress. Recently, NQO1 was found in ribonucleoprotein (RNP) complexes, but NQO1-interacting mRNAs and the functional impact of such interactions are not known. Here, we used ribonucleoprotein immunoprecipitation (RIP) and microarray analysis to identify comprehensively the subset of NQO1 target mRNAs in human hepatoma HepG2 cells. One of its main targets, SERPINA1 mRNA, encodes the serine protease inhibitor α-1-antitrypsin, A1AT, which is associated with disorders including obesity-related metabolic inflammation, chronic obstructive pulmonary disease (COPD), liver cirrhosis and hepatocellular carcinoma. Biotin pulldown analysis indicated that NQO1 can bind the 3' untranslated region (UTR) and the coding region (CR) of SERPINA1 mRNA. NQO1 did not affect SERPINA1 mRNA levels; instead, it enhanced the translation of SERPINA1 mRNA, as NQO1 silencing decreased the size of polysomes forming on SERPINA1 mRNA and lowered the abundance of A1AT. Luciferase reporter analysis further indicated that NQO1 regulates SERPINA1 mRNA translation through the SERPINA1 3'UTR. Accordingly, NQO1-KO mice had reduced hepatic and serum levels of A1AT and increased activity of neutrophil elastase (NE), one of the main targets of A1AT. We propose that this novel mechanism of action of NQO1 as an RNA-binding protein may help to explain its pleiotropic biological effects. Published by Elsevier Inc.

  16. Occupational health hazards of trichloroethylene among workers in relation to altered mRNA expression of cell cycle regulating genes (p53, p21, bax and bcl-2 and PPARA

    Directory of Open Access Journals (Sweden)

    Meenu Varshney

    2015-01-01

    Full Text Available Trichloroethylene (TCE is widely used as a metal degreaser in industrial processes. The present study reports on the effects of TCE exposure on workers employed in the lock industries. To ensure exposure of the workers to TCE, its toxic metabolites, trichloroacetic acid (TCA, dichloroacetic acid (DCA and trichloroethanol (TCEOH were detected in the plasma of the subjects through solid phase microextraction-gas chromatography-electron capture detection. TCA, DCA and TCEOH were detected in the range of 0.004–2.494 μg/mL, 0.01–3.612 μg/mL and 0.002–0.617 μg/mL, respectively. Quantitative reverse transcription polymerase chain reaction analysis revealed up-regulated expression of p53 (2.4-fold; p < 0.05, p21 (2-fold; p < 0.01, bax (2.9-fold; p < 0.01 mRNAs and down-regulated expression of bcl-2 (67%; p < 0.05 mRNAs, indicating DNA damaging potential of these metabolites. No effects were observed on the levels of p16 and c-myc mRNAs. Further, as TCA and DCA, the ligand of peroxisome proliferator activated receptor alpha (PPARA, are involved in the process of hepatocarcinogenesis in rodents, we examined expression of PPARA mRNA and let-7c miRNA in the workers. No statistically significant differences in expression of PPARA mRNA and let-7c miRNA in patients were observed as compared to values in controls. Dehydroepiandosterone sulfate (DHEAS is a reported endogenous ligand of PPARA so its competitive role was also studied. We observed decreased levels of DHEAS hormone in the subjects. Hence, its involvement in mediation of the observed changes in the levels of various mRNAs analyzed in this study appears unlikely.

  17. ErbB3 mRNA leukocyte levels as a biomarker for major depressive disorder

    Directory of Open Access Journals (Sweden)

    Milanesi Elena

    2012-09-01

    Full Text Available Abstract Background In recent years, the identification of peripheral biomarkers that are associated with psychiatric diseases, such as Major Depressive Disorder (MDD, has become relevant because these biomarkers may improve the efficiency of the differential diagnosis process and indicate targets for new antidepressant drugs. Two recent candidate genes, ErbB3 and Fgfr1, are growth factors whose mRNA levels have been found to be altered in the leukocytes of patients that are affected by bipolar disorder in a depressive state. On this basis, the aim of the study was to determine if ErbB3 and Fgfr1 mRNA levels could be a biomarkers of MDD. Methods We measured by Real Time PCR ErbB3 and Fgfr1 mRNA expression levels in leukocytes of MDD patients compared with controls. Successively, to assess whether ErbB3 mRNA levels were influenced by previous antidepressant treatment we stratified our patients sample in two cohorts, comparing drug-naive versus drug-free patients. Moreover, we evaluated the levels of the transcript in MDD patients after 12 weeks of antidepressant treatment, and in prefrontal cortex of rats stressed and treated with an antidepressant drug of the same class. Results These results showed that ErbB3 but not Fgfr1 mRNA levels were reduced in leukocytes of MDD patients compared to healthy subjects. Furthermore, ErbB3 levels were not affected by antidepressant treatment in either human or animal models Conclusions Our data suggest that ErbB3 might be considered as a biomarker for MDD and that its deficit may underlie the pathopsysiology of the disease and is not a consequence of treatment. Moreover the study supports the usefulness of leukocytes as a peripheral system for identifying biomarkers in psychiatric diseases.

  18. Integrin beta 4 MRNA expression levels in bronchial asthma patients ...

    African Journals Online (AJOL)

    Serum total IgE was measured by ELISA and mRNA expression of ITGβ4 was assessed by reverse transcriptase PCR (RT-PCR) using real time PCR.. ITGβ4 mRNA expression was significantly down regulated with increased serum total IgE in patients with asthma compared to controls. Moreover, ITGβ4 expression was ...

  19. Functional Integration of mRNA Translational Control Programs

    Directory of Open Access Journals (Sweden)

    Melanie C. MacNicol

    2015-07-01

    Full Text Available Regulated mRNA translation plays a key role in control of cell cycle progression in a variety of physiological and pathological processes, including in the self-renewal and survival of stem cells and cancer stem cells. While targeting mRNA translation presents an attractive strategy for control of aberrant cell cycle progression, mRNA translation is an underdeveloped therapeutic target. Regulated mRNAs are typically controlled through interaction with multiple RNA binding proteins (RBPs but the mechanisms by which the functions of distinct RBPs bound to a common target mRNA are coordinated are poorly understood. The challenge now is to gain insight into these mechanisms of coordination and to identify the molecular mediators that integrate multiple, often conflicting, inputs. A first step includes the identification of altered mRNA ribonucleoprotein complex components that assemble on mRNAs bound by multiple, distinct RBPs compared to those recruited by individual RBPs. This review builds upon our knowledge of combinatorial control of mRNA translation during the maturation of oocytes from Xenopus laevis, to address molecular strategies that may mediate RBP diplomacy and conflict resolution for coordinated control of mRNA translational output. Continued study of regulated ribonucleoprotein complex dynamics promises valuable new insights into mRNA translational control and may suggest novel therapeutic strategies for the treatment of disease.

  20. Phytochrome B mRNA expression enhances biomass yield and ...

    African Journals Online (AJOL)

    The present study shows successful transformation and mRNA expression in Phytochrome B transformed CIM 482 cotton plants. Transgenic cotton plants expressing Phytochrome B mRNA have showed more than two times increase in relative leaf growth rate (RLGR) and photosynthetic rate, more than one time increase in ...

  1. mRNA pseudoknot structures can act as ribosomal roadblocks

    DEFF Research Database (Denmark)

    Hansen, Jesper Tholstrup; Oddershede, Lene Broeng; Sørensen, Michael Askvad

    2012-01-01

    Several viruses utilize programmed ribosomal frameshifting mediated by mRNA pseudoknots in combination with a slippery sequence to produce a well defined stochiometric ratio of the upstream encoded to the downstream-encoded protein. A correlation between the mechanical strength of mRNA pseudoknots...

  2. E1a promotes c-Myc-dependent replicative stress: Implications in glioblastoma radiosensitization

    OpenAIRE

    Valero, María Llanos; Cimas, Francisco Jose; Arias, Laura; Melgar-Rojas, Pedro; García, Elena; Callejas-Valera, Juan Luis; García-Cano, Jesús; Serrano-Oviedo, Leticia; Ángel de la Cruz-Morcillo, Miguel; Sánchez-Pérez, Isabel; Sánchez-Prieto, Ricardo

    2013-01-01

    The E1a gene from adenovirus is known to be a potent inducer of chemo/radiosensitivity in a wide range of tumors. However, the molecular bases of its radiosensitizer properties are still poorly understood. In an attempt to study this effect, U87MG cells, derived from a radio-resistant tumor as glioblastoma, where infected with lentivirus carrying E1a gene developing an acute sensitivity to ionizing radiation. The induction of radiosensitivity correlated with a marked G2/M phase accumulation a...

  3. Treatment of Multiple Myeloma with VLA4-targeted Nanoparticles Delivering Novel c-MYC Inhibitor Prodrug

    Science.gov (United States)

    2012-09-01

    Bagnasco, L., Malacarne, D., Melchiori, A., Valente, P., Millo, E., Bruno, S., Basso, S., and Parodi, S. A retro-inverso peptide homologous to...81. 25. Ruoslahti, E. RGD and other recognition sequences for integrins. Annu Rev Cell Dev Bioi. 1996; 12697-715. 26. Ria, R., Vacca, A., Ribatti...Haematologica. 2002; 87(8):836-845. 27. Temming, K., Schiffelers, R.M., Molema, G., and Kok, R.J. RGD -based strategies for selective delivery of

  4. Treatment of Endocrine-Resistant Breast Cancer with a Small Molecule c-Myc Inhibitor

    Science.gov (United States)

    2016-08-01

    myeloid leukemia [21-23]. Based on our results, we tested whether JQ1 can suppress ERα expression. As shown in Figure 2G, treatment of MCF7 cells with...al. Small-molecule inhibition of BRD4 as a new potent approach to eliminate leukemic stem- and progenitor cells in acute myeloid leukemia AML...tamoxifen, bromodomain, resistance 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT 18. NUMBER OF PAGES 19a. NAME OF RESPONSIBLE PERSON

  5. Cdk2 suppresses cellular senescence induced by the c-myc oncogene.

    Science.gov (United States)

    Campaner, Stefano; Doni, Mirko; Hydbring, Per; Verrecchia, Alessandro; Bianchi, Lucia; Sardella, Domenico; Schleker, Thomas; Perna, Daniele; Tronnersjö, Susanna; Murga, Matilde; Fernandez-Capetillo, Oscar; Barbacid, Mariano; Larsson, Lars-Gunnar; Amati, Bruno

    2010-01-01

    Activated oncogenes induce compensatory tumour-suppressive responses, such as cellular senescence or apoptosis, but the signals determining the main outcome remain to be fully understood. Here, we uncover a role for Cdk2 (cyclin-dependent kinase 2) in suppressing Myc-induced senescence. Short-term activation of Myc promoted cell-cycle progression in either wild-type or Cdk2 knockout mouse embryo fibroblasts (MEFs). In the knockout MEFs, however, the initial hyper-proliferative response was followed by cellular senescence. Loss of Cdk2 also caused sensitization to Myc-induced senescence in pancreatic beta-cells or splenic B-cells in vivo, correlating with delayed lymphoma onset in the latter. Cdk2-/- MEFs also senesced upon ectopic Wnt signalling or, without an oncogene, upon oxygen-induced culture shock. Myc also causes senescence in cells lacking the DNA repair protein Wrn. However, unlike loss of Wrn, loss of Cdk2 did not enhance Myc-induced replication stress, implying that these proteins suppress senescence through different routes. In MEFs, Myc-induced senescence was genetically dependent on the ARF-p53-p21Cip1 and p16INK4a-pRb pathways, p21Cip1 and p16INK4a being selectively induced in Cdk2-/- cells. Thus, although redundant for cell-cycle progression and development, Cdk2 has a unique role in suppressing oncogene- and/or stress-induced senescence. Pharmacological inhibition of Cdk2 induced Myc-dependent senescence in various cell types, including a p53-null human cancer cell line. Our data warrant re-assessment of Cdk2 as a therapeutic target in Myc- or Wnt-driven tumours.

  6. Overexpression of the c-myc oncogene inhibits nonsense-mediated RNA decay in B lymphocytes.

    Science.gov (United States)

    Wang, Ding; Wengrod, Jordan; Gardner, Lawrence B

    2011-11-18

    The Myc transcription factor plays a vital role in both normal cellular physiology and in many human cancers. We have recently demonstrated that nonsense-mediated RNA decay (NMD), a mechanism that rapidly degrades select mRNAs, is inhibited by the stress-induced phosphorylation of translation initiation factor eIF2α, and this inhibition stabilizes many transcripts necessary for tumorigenesis. Here, we demonstrate that NMD is inhibited by high Myc expression. We show that the phosphorylation of eIF2α, likely due to the ability of Myc to generate reactive oxygen species and augment endoplasmic reticulum stress, is necessary for the inhibition of NMD by Myc. The inhibition of NMD both stabilizes and up-regulates multiple Myc targets, suggesting that the inhibition of NMD may play an important role in the dynamic regulation of genes by Myc.

  7. C-MYC Involvement in Chronic Lymphocytic Leukemia (CLL): A Molecular and Cytogenetic Update.

    Science.gov (United States)

    Fonseka, Lakshan N; Tirado, Carlos A

    2015-01-01

    Chronic lymphocytic leukemia (CLL) is a disorder entailing the slow proliferation of B-cell lymphocytes in the bone marrow and blood. In 2015, it is estimated that 14,620 patients will be diagnosed with CLL, and approximately 4,650 patients will die due to disease progression. CLL typically presents in patients about 71 years of age. Initially, the patients exhibit leukocytosis; however, as the disease progresses, they experience splenomegaly, lymphadenopathy, hepatomegaly, anemia, and infections. Although about 84% of CLL patients will survive for five years or more, CLL cases that report MYC (8q24) translocations with IGH, IGK, IGL, and TCR genes have poor prognoses and low survival rates. Recent studies have shown data supporting both a positive correlation and no correlation between disease progression and MYC expression. Nonetheless, other studies have revealed new information on multiple MYC-dependent pathways responsible for leukemogenesis and tumorigenesis. Herein, we summarize the current molecular nd cytogenetic findings in MYC-associated CLL, with focus on the underlying MYC-dependent mechanisms of leukemogenesis and MYC-associated CLL progression and treatment regimen.

  8. Effects of codon optimization on the mRNA levels of heterologous genes in filamentous fungi.

    Science.gov (United States)

    Tanaka, Mizuki; Tokuoka, Masafumi; Gomi, Katsuya

    2014-05-01

    Filamentous fungi, particularly Aspergillus species, have recently attracted attention as host organisms for recombinant protein production. Because the secretory yields of heterologous proteins are generally low compared with those of homologous proteins or proteins from closely related fungal species, several strategies to produce substantial amounts of recombinant proteins have been conducted. Codon optimization is a powerful tool for improving the production levels of heterologous proteins. Although codon optimization is generally believed to improve the translation efficiency of heterologous genes without affecting their mRNA levels, several studies have indicated that codon optimization causes an increase in the steady-state mRNA levels of heterologous genes in filamentous fungi. However, the mechanism that determines the low mRNA levels when native heterologous genes are expressed was poorly understood. We recently showed that the transcripts of heterologous genes are polyadenylated prematurely within the coding region and that the heterologous gene transcripts can be stabilized significantly by codon optimization, which is probably attributable to the prevention of premature polyadenylation in Aspergillus oryzae. In this review, we describe the detailed mechanism of premature polyadenylation and the rapid degradation of mRNA transcripts derived from heterologous genes in filamentous fungi.

  9. Developmental changes in hypothalamic oxytocin and oxytocin receptor mRNA expression and their sensitivity to fasting in male and female rats.

    Science.gov (United States)

    Matsuzaki, Toshiya; Iwasa, Takeshi; Munkhzaya, Munkhsaikhan; Tungalagsuvd, Altankhuu; Kawami, Takako; Murakami, Masahiro; Yamasaki, Mikio; Yamamoto, Yuri; Kato, Takeshi; Kuwahara, Akira; Yasui, Toshiyuki; Irahara, Minoru

    2015-04-01

    Oxytocin (OT) affects the central nervous system and is involved in a variety of social and non-social behaviors. Recently, the role played by OT in energy metabolism and its organizational effects on estrogen receptor alpha (ER-α) during the neonatal period have gained attention. In this study, the developmental changes in the hypothalamic mRNA levels of OT, the OT receptor (OTR), and ER-α were evaluated in male and female rats. In addition, the fasting-induced changes in the hypothalamic mRNA levels of OT and the OTR were evaluated. Hypothalamic explants were taken from postnatal day (PND) 10, 20, and 30 rats, and the mRNA level of each molecule was measured. Hypothalamic OT mRNA expression increased throughout the developmental period in both sexes. The rats' hypothalamic OTR mRNA levels were highest on PND 10 and decreased throughout the developmental period. In the male rats, the hypothalamic mRNA levels of ER-α were higher on PND 30 than on PND 10. On the other hand, no significant differences in hypothalamic ER-α mRNA expression were detected among the examined time points in the female rats, although hypothalamic ER-α mRNA expression tended to be higher on PND 30 than on PND 10. Significant positive correlations were detected between hypothalamic OT and ER-α mRNA expression in both the male and female rats. Hypothalamic OT mRNA expression was not affected by fasting at any of the examined time points in either sex. These results indicate that hypothalamic OT expression is not sensitive to fasting during the developmental period. In addition, as a positive correlation was detected between hypothalamic OT and ER-α mRNA expression, these two molecules might interact with each other to induce appropriate neuronal development. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Cellular retinoic-acid-binding-protein and retinol-binding-protein mRNA expression in the cells of the rat seminiferous tubules and their regulation by retinoids.

    Science.gov (United States)

    Faraonio, R; Galdieri, M; Colantuoni, V

    1993-02-01

    The levels of the mRNA corresponding to the intracellular binding proteins for retinoic acid and retinol (CRABP1 and CRBP1, respectively) were studied in primary cultures of somatic and germ cells of the rat seminiferous tubules. We show that the CRABP1 mRNA is expressed in Sertoli and germ cells and a single molecular species of mRNA is detected. CRBP1 mRNA is detected in Sertoli and peritubular cells. The regulation of the expression of both genes by retinoids was studied in Sertoli cells. CRABP1 mRNA levels are not affected by either retinoic acid or retinol, whereas both compounds positively regulate CRBP1 mRNA synthesis in a dose-dependent manner. A fivefold increase in CRBP1 mRNA levels was observed 32-48 h after addition of either agent. These results demonstrate that in Sertoli cells the expression of CRABP1 is not affected by retinoids, similar to the situation observed in vivo and in other in-vitro cultures. CRBP1-gene expression is, instead, induced and the variations in CRBP1-mRNA levels may regulate the intracellular concentrations of retinoids, as a response to changes in the vitamin-A nutritional status.

  11. Role of mRNA Methylation in Prostate Cancer

    Science.gov (United States)

    2015-02-01

    regulation of mRNA. Recent technological advances have made it possible to detect mRNA methylation . The m6A was found near regulatory regions and...TERMS mRNA methylation , FTO, MeRIP-seq, RNA-seq, m6A 16. SECURITY CLASSIFICATION OF: 17. LIMITATION OF ABSTRACT 18. NUMBER OF PAGES 19a. NAME OF...10 9. Appendices……………………………………………………………10 1. INTRODUCTION: Methylation at the N6 position of adenosine ( m6A ) is a post-transcriptional modification of

  12. Optimization of preservation and storage time of sponge tissues to obtain quality mRNA for next-generation sequencing.

    Science.gov (United States)

    Riesgo, Ana; Pérez-Porro, Alicia R; Carmona, Susana; Leys, Sally P; Giribet, Gonzalo

    2012-03-01

    Transcriptome sequencing with next-generation sequencing technologies has the potential for addressing many long-standing questions about the biology of sponges. Transcriptome sequence quality depends on good cDNA libraries, which requires high-quality mRNA. Standard protocols for preserving and isolating mRNA often require optimization for unusual tissue types. Our aim was assessing the efficiency of two preservation modes, (i) flash freezing with liquid nitrogen (LN₂) and (ii) immersion in RNAlater, for the recovery of high-quality mRNA from sponge tissues. We also tested whether the long-term storage of samples at -80 °C affects the quantity and quality of mRNA. We extracted mRNA from nine sponge species and analysed the quantity and quality (A260/230 and A260/280 ratios) of mRNA according to preservation method, storage time, and taxonomy. The quantity and quality of mRNA depended significantly on the preservation method used (LN₂) outperforming RNAlater), the sponge species, and the interaction between them. When the preservation was analysed in combination with either storage time or species, the quantity and A260/230 ratio were both significantly higher for LN₂-preserved samples. Interestingly, individual comparisons for each preservation method over time indicated that both methods performed equally efficiently during the first month, but RNAlater lost efficiency in storage times longer than 2 months compared with flash-frozen samples. In summary, we find that for long-term preservation of samples, flash freezing is the preferred method. If LN₂ is not available, RNAlater can be used, but mRNA extraction during the first month of storage is advised. © 2011 Blackwell Publishing Ltd.

  13. Effect of vitrification on the mRNA transcriptome of bovine oocytes.

    Science.gov (United States)

    Wang, N; Li, C-Y; Zhu, H-B; Hao, H-S; Wang, H-Y; Yan, C-L; Zhao, S-J; Du, W-H; Wang, D; Liu, Y; Pang, Y-W; Zhao, X-M

    2017-08-01

    Vitrification has been shown to decrease the developmental capacity of mammalian oocytes, and this is closely associated with the abnormal mRNA expressions of vitrified oocytes. However, the effect of vitrification on transcriptional machinery of oocytes examined by RNA sequencing (RNA-seq) has yet to be defined. In the present study, the mRNA transcriptomes of fresh and vitrified bovine oocytes were analysed by Smart-seq2 with the differently expressed genes determined by DEseq2 (an adjusted p-value of .05 and a minimum fold change of 2). The differentially expressed mRNAs were then searched against the Gene Ontology (GO) and Genomes (KEGG) database. Finally, the mRNA expressions of 10 candidate genes were validated using quantitative real-time PCR (qRT-PCR). Approximately 12,000 genes were detected in each sample of fresh or vitrified oocytes. Of these, the expression levels of 102 genes differed significantly in vitrified groups: 12 genes mainly involved in cell cycle, fertilization and glucose metabolism were upregulated, and 90 genes mainly involved in mitochondria, ribosomal protein, cytoskeleton, transmembrane protein, cell cycle and calcium ions were downregulated. GO analysis showed that these genes were mainly enriched in terms of membrane-bounded organelles, macromolecular complex, and intracellular part. The mRNA expression levels of 10 candidate genes selected randomly were in agreement with the results of the RNA-seq. In conclusion, our results showed that vitrification affected the mRNA transcriptome of bovine oocytes by downregulating genes, which contributed to the decreased developmental capacity of vitrified oocytes. Our findings will be useful in determining approaches to improve the efficiency of vitrified oocytes. © 2017 Blackwell Verlag GmbH.

  14. Regulation and dysregulation of vitellogenin mRNA accumulation in daphnids (Daphnia magna)

    Energy Technology Data Exchange (ETDEWEB)

    Hannas, Bethany R.; Wang, Ying H.; Thomson, Susanne; Kwon, Gwijun; Li Hong [Department of Environmental and Molecular Toxicology, North Carolina State University, Raleigh, NC 27695-7633 (United States); LeBlanc, Gerald A., E-mail: Gerald_LeBlanc@ncsu.edu [Department of Environmental and Molecular Toxicology, North Carolina State University, Raleigh, NC 27695-7633 (United States)

    2011-01-25

    The induction of vitellogenin in oviparous vertebrates has become the gold standard biomarker of exposure to estrogenic chemicals in the environment. This biomarker of estrogen exposure also has been used in arthropods, however, little is known of the factors that regulate the expression of vitellogenin in these organisms. We investigated changes in accumulation of mRNA products of the vitellogenin gene Vtg2 in daphnids (Daphnia magna) exposed to a diverse array of chemicals. We further evaluated the involvement of hormonal factors in the regulation of vitellogenin expression that may be targets of xenobiotic chemicals. Expression of the Vtg2 gene was highly responsive to exposure to various chemicals with an expression range spanning approximately four orders of magnitude. Chemicals causing the greatest induction were piperonyl butoxide, chlordane, 4-nonylphenol, cadmium, and chloroform. Among these, only 4-nonylphenol is recognized to be estrogenic. Exposure to several chemicals also suppressed Vtg2 mRNA levels, as much as 100-fold. Suppressive chemicals included cyproterone acetate, acetone, triclosan, and atrazine. Exposure to the estrogens diethylstilbestrol and bisphenol A had little effect on vitellogenin mRNA levels further substantiating that these genes are not induced by estrogen exposure. Exposure to the potent ecdysteroids 20-hydroxyecdysone and ponasterone A revealed that Vtg2 was subject to strong suppressive control by these hormones. Vtg2 mRNA levels were not significantly affected from exposure to several juvenoid hormones. Results indicate that ecdysteroids are suppressors of vitellogenin gene expression and that vitellogenin mRNA levels can be elevated or suppressed in daphnids by xenobiotics that elicit antiecdysteroidal or ecdysteroidal activity, respectively. Importantly, daphnid Vtg2 is not elevated in response to estrogenic activity.

  15. Viral and Cellular mRNA Translation in Coronavirus-Infected Cells.

    Science.gov (United States)

    Nakagawa, K; Lokugamage, K G; Makino, S

    2016-01-01

    Coronaviruses have large positive-strand RNA genomes that are 5' capped and 3' polyadenylated. The 5'-terminal two-thirds of the genome contain two open reading frames (ORFs), 1a and 1b, that together make up the viral replicase gene and encode two large polyproteins that are processed by viral proteases into 15-16 nonstructural proteins, most of them being involved in viral RNA synthesis. ORFs located in the 3'-terminal one-third of the genome encode structural and accessory proteins and are expressed from a set of 5' leader-containing subgenomic mRNAs that are synthesized by a process called discontinuous transcription. Coronavirus protein synthesis not only involves cap-dependent translation mechanisms but also employs regulatory mechanisms, such as ribosomal frameshifting. Coronavirus replication is known to affect cellular translation, involving activation of stress-induced signaling pathways, and employing viral proteins that affect cellular mRNA translation and RNA stability. This chapter describes our current understanding of the mechanisms involved in coronavirus mRNA translation and changes in host mRNA translation observed in coronavirus-infected cells. © 2016 Elsevier Inc. All rights reserved.

  16. Different loci and mRNA copy number of the increased serum survival gene of Escherichia coli.

    Science.gov (United States)

    Xu, Wang-Ye; Li, Yi-Jing; Fan, Chen

    2018-02-01

    The increased serum survival gene (iss) has been identified as a virulence trait associated with the virulence of Escherichia coli, causing colibacillosis in poultry. However, it remains unclear as to whether iss mRNA copy number and sequence affect virulence. To examine these influences, we assessed the presence of iss, sequence analysis, iss mRNA copy number, and serum resistance. The iss gene was detected in 88 (all) E. coli isolates from different sources, and sequencing identified 16 alleles (32 different loci) and 10 amino acid sequences (10 different loci). Nested polymerase chain reaction improved iss detection. The isolates from sick chickens had >68% livability in serum resistance tests and higher iss mRNA copy number. The iss mRNA copy number highly correlated with mortality and E. coli livability. Student's t tests confirmed the relationship between the different loci to iss transcription, serum resistance, and virulence. These data suggest that iss mRNA copy number and different loci affect the virulence and serum resistance. These findings could be useful in further studies on the prevalence of iss among E. coli isolates and other virulence factors.

  17. REDUCED NITRIC OXIDE PRODUCTION AND INOS MRNA EXPRESSION IN IFN-G STIMULATED CHICKEN MACROPHAGES TRANSFECTED WITH INOS SIRNAS

    Science.gov (United States)

    Utilizing RNA interference technology with siRNA in the HD-11 macrophage cell line, we determined how the inhibition or knock-down of the iNOS (inducible nitric oxide synthase) gene affected IFN-y' induced macrophage production of nitric oxide (NO) and mRNA expression of genes involved in this biolo...

  18. Long Open Reading Frame Transcripts Escape Nonsense-Mediated mRNA Decay in Yeast

    Directory of Open Access Journals (Sweden)

    Laurence Decourty

    2014-02-01

    Full Text Available Nonsense-mediated mRNA decay (NMD destabilizes eukaryotic transcripts with long 3′ UTRs. To investigate whether other transcript features affect NMD, we generated yeast strains expressing chromosomal-derived mRNAs with 979 different promoter and open reading frame (ORF regions and with the same long, destabilizing 3′ UTR. We developed a barcode-based DNA microarray strategy to compare the levels of each reporter mRNA in strains with or without active NMD. The size of the coding region had a significant negative effect on NMD efficiency. This effect was not specific to the tested 3′ UTR because two other different NMD reporters became less sensitive to NMD when ORF length was increased. Inefficient NMD was not due to a lack of association of Upf1 to long ORF transcripts. In conclusion, in addition to a long 3′ UTR, short translation length is an important feature of NMD substrates in yeast.

  19. Regulation of Egr-1, VIP, and Shh mRNA and Egr-1 protein in the mouse retina by light and image quality.

    Science.gov (United States)

    Brand, Christine; Burkhardt, Eva; Schaeffel, Frank; Choi, Jeong Won; Feldkaemper, Marita Pauline

    2005-04-28

    To analyze mRNA expression changes of Egr-1, VIP, and Shh under different light and treatment conditions in mice. The mRNA expression levels of the three genes and additionally the Egr-1 protein expression were compared in form deprived eyes and eyes with normal vision. Moreover, the influence of dark to light and light to dark transitions and of changes in retinal illumination on mRNA levels was investigated. Form deprivation of mice was induced by fitting frosted diffusers over one eye and an attentuation matched neutral density (ND) filter over the other eye. To measure the effects of retinal illumination changes on mRNA expression, animals were bilaterally fitted with different ND filters. Semiquantitative real-time RT-PCR was used to measure the mRNA levels and immunohistochemistry was applied to localize and detect Egr-1 protein. The expression levels of both Egr-1 mRNA and protein were reduced in form deprived eyes compared to their fellow eyes after 30 min and 1 h, respectively. Egr-1 mRNA was strikingly upregulated both after dark to light and light to dark transitions, whereas minor changes in retinal illumination by covering the eyes with neutral density filters did not alter Egr-1 mRNA expression. In mice, the mRNA levels of VIP and Shh were not affected by form deprivation, but they were found to be regulated depending on the time of day. Both Egr-1 mRNA and protein expression levels were strongly regulated by light, especially by transitions between light and darkness. Image contrast may exert an additional influence on mRNA and protein expression of Egr-1, particularly in the cells in the ganglion cell layer and in bipolar cells.

  20. Relationship Between the DPD and TS mRNA Expression and the Response to S-1-Based Chemotherapy and Prognosis in Patients with Advanced Gastric Cancer.

    Science.gov (United States)

    Shen, Xiao-Ming; Zhou, Chong; Lian, Lian; Li, Li-Qun; Li, Wei; Tao, Min

    2015-04-01

    The aim was to determine changes in dihydropyrimidine dehydrogenase (DPD) and thymidylate synthase (TS) mRNAs in the blood of advanced gastric cancer (AGC) patients to see whether these enzymes affected the patients' response to S-1-based chemotherapy and prognosis. For this purpose, pretreatment DPD/TS mRNA expressions were determined in 40 AGC patients using RT-PCR. The patients were then administered with S-1-based regimen (S-1 + cisplatin) and toxicities were recorded. The relationship between the DPD/TS mRNA expressions and the chemotherapy response, drug resistance, and prognosis was analyzed. The data show that DPD mRNA expression correlated significantly with Lauren type while TS mRNA expression correlated with distant metastasis. Patients with higher DPD and/or TS mRNA expression(s) showed poor response, while those with low DPD mRNA expression showed better response to the chemotherapy. Pooled analysis showed that the patients with low DPD/TS mRNA expressions had better therapeutic response. The incidence of bone marrow suppression, diarrhea, and oral mucositis was high in patients with low DPD mRNA expression. Median overall survival (OS) in 40 patients was 13.5 months. It was 17 months for low and 10 months for high DPD (P = 0.044) and TS mRNA expression (P = 0.047). Pooled analysis showed that the patients with both low DPD/TS mRNA expressions had longer OS (P = 0.001). In conclusion, the detection of DPD and/or TS mRNA expression can be used to predict the response to S-1-based chemotherapy, drug resistance, and prognosis in AGC patients as well as to help guide the individualized treatment of gastric cancer.

  1. Tissue-specific mRNA expression profiling in grape berry tissues

    Directory of Open Access Journals (Sweden)

    Cramer Grant R

    2007-06-01

    wall function and transport processes. Seeds, which supply essential resources for embryo development, showed higher mRNA abundance of genes encoding phenylpropanoid biosynthetic enzymes, seed storage proteins, and late embryogenesis abundant proteins. Water-deficit stress affected the mRNA abundance of 13% of the genes with differential expression patterns occurring mainly in the pulp and skin. In pulp and seed tissues transcript abundance in most functional categories declined in water-deficit stressed vines relative to well-watered vines with transcripts for storage proteins and novel (no-hit functional assignments being over represented. In the skin of berries from water-deficit stressed vines, however, transcripts from several functional categories including general phenypropanoid and ethylene metabolism, pathogenesis-related responses, energy, and interaction with the environment were significantly over-represented. Conclusion These results revealed novel insights into the tissue-specific expression mRNA expression patterns of an extensive repertoire of genes expressed in berry tissues. This work also establishes an extensive catalogue of gene expression patterns for future investigations aimed at the dissection of the transcriptional regulatory hierarchies that govern tissue-specific expression patterns associated with tissue differentiation within berries. These results also confirmed that water-deficit stress has a profound effect on mRNA expression patterns particularly associated with the biosynthesis of aroma and color metabolites within skin and pulp tissues that ultimately impact wine quality.

  2. Demonstration that a mRNA Binding Protein is Responsible for GADD45 mRNA Destabilization

    National Research Council Canada - National Science Library

    Abcouwer, Steve

    2003-01-01

    ...) Using regions of the GADD45 mRNA 3'-untranslated region (UTR) in KNA gel shift assays, we have observed that glutamine causes distinct changes in RBP activities in cytoplasmic and nuclear protein extracts...

  3. Alkaline decontamination of sputum specimens adversely affects stability of mycobacterial mRNA.

    OpenAIRE

    Desjardin, L E; Perkins, M D; Teixeira, L; Cave, M D; Eisenach, K D

    1996-01-01

    Reverse transcriptase PCR (RT-PCR) is an important tool for Mycobacterium tuberculosis research and diagnostics. A standard procedure using N-acetyl-L-cysteine (NALC) and NaOH has been widely adopted for digestion and decontamination of sputum specimens for mycobacterial culture. The objective of this study was to determine the compatibility of this method with the recovery of RNA for RT-PCR assays. Nineteen sputum specimens were collected from smear-positive, pretreatment tuberculosis patien...

  4. Protein targeting to subcellular organelles via MRNA localization.

    Science.gov (United States)

    Weis, Benjamin L; Schleiff, Enrico; Zerges, William

    2013-02-01

    Cells have complex membranous organelles for the compartmentalization and the regulation of most intracellular processes. Organelle biogenesis and maintenance requires newly synthesized proteins, each of which needs to go from the ribosome translating its mRNA to the correct membrane for insertion or transclocation to an a organellar subcompartment. Decades of research have revealed how proteins are targeted to the correct organelle and translocated across one or more organelle membranes ro the compartment where they function. The paradigm examples involve interactions between a peptide sequence in the protein, localization factors, and various membrane embedded translocation machineries. Membrane translocation is either cotranslational or posttranslational depending on the protein and target organelle. Meanwhile research in embryos, neurons and yeast revealed an alternative targeting mechanism in which the mRNA is localized and only then translated to synthesize the protein in the correct location. In these cases, the targeting information is coded by the cis-acting sequences in the mRNA ("Zipcodes") that interact with localization factors and, in many cases, are transported by the molecular motors on the cytoskeletal filaments. Recently, evidence has been found for this "mRNA based" mechanism in organelle protein targeting to endoplasmic reticulum, mitochondria, and the photosynthetic membranes within chloroplasts. Here we review known and potential roles of mRNA localization in protein targeting to and within organelles. This article is part of a Special Issue entitled: Protein Import and Quality Control in Mitochondria and Plastids.

  5. Effect of atrophy and contractions on myogenin mRNA concentration in chick and rat myoblast omega muscle cells

    Science.gov (United States)

    Krebs, J. M.; Denney, R. M.

    1997-01-01

    The skeletal rat myoblast omega (RMo) cell line forms myotubes that exhibit spontaneous contractions under appropriate conditions in culture. We examined if the RMo cells would provide a model for studying atrophy and muscle contraction. To better understand how to obtain contractile cultures, we examined levels of contraction under different growing conditions. The proliferation medium and density of plating affected the subsequent proportion of spontaneously contracting myotubes. Using a ribonuclease protection assay, we found that exponentially growing RMo myoblasts contained no detectable myogenin or herculin mRNA, while differentiating myoblasts contained high levels of myogenin mRNA but no herculin mRNA. There was no increase in myogenin mRNA concentration in either primary chick or RMo myotubes whose contractions were inhibited by depolarizing concentrations of potassium (K+). Thus, altered myogenin mRNA concentrations are not involved in atrophy of chick myotubes. Depolarizing concentrations of potassium inhibited spontaneous contractions in both RMo cultures and primary chick myotube cultures. However, we found that the myosin concentration of 6-d-old contracting RMo cells fed medium plus AraC was 11 +/- 3 micrograms myosin/microgram DNA, not significantly different from 12 +/- 4 micrograms myosin/microgram DNA (n = 3), the myosin concentration of noncontracting RMo cells (treated with 12 mM K+ for 6 d). Resolving how RMo cells maintained their myosin content when contraction is inhibited may be important for understanding atrophy.

  6. Binding of DEAD-box helicase Dhh1 to the 5'-untranslated region of ASH1 mRNA represses localized translation of ASH1 in yeast cells.

    Science.gov (United States)

    Zhang, Qianjun; Meng, Xiuhua; Li, Delin; Chen, Shaoyin; Luo, Jianmin; Zhu, Linjie; Singer, Robert H; Gu, Wei

    2017-06-09

    Local translation of specific mRNAs is regulated by dynamic changes in their subcellular localization, and these changes are due to complex mechanisms controlling cytoplasmic mRNA transport. The budding yeast Saccharomyces cerevisiae is well suited to studying these mechanisms because many of its transcripts are transported from the mother cell to the budding daughter cell. Here, we investigated the translational control of ASH1 mRNA after transport and localization. We show that although ASH1 transcripts were translated after they reached the bud tip, some mRNAs were bound by the RNA-binding protein Puf6 and were non-polysomal. We also found that the DEAD-box helicase Dhh1 complexed with the untranslated ASH1 mRNA and Puf6. Loss of Dhh1 affected local translation of ASH1 mRNA and resulted in delocalization of ASH1 transcript in the bud. Forcibly shifting the non-polysomal ASH1 mRNA into polysomes was associated with Dhh1 dissociation. We further demonstrated that Dhh1 is not recruited to ASH1 mRNA co-transcriptionally, suggesting that it could bind to ASH1 mRNA within the cytoplasm. Of note, Dhh1 bound to the 5'-UTR of ASH1 mRNA and inhibited its translation in vitro These results suggest that after localization to the bud tip, a portion of the localized ASH1 mRNA becomes translationally inactive because of binding of Dhh1 and Puf6 to the 5'- and 3'-UTRs of ASH1 mRNA. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  7. Recognition of nonsense mRNA: towards a unified model.

    Science.gov (United States)

    Mühlemann, Oliver

    2008-06-01

    Among the different cellular surveillance mechanisms that ensure accurate gene expression, nonsense-mediated mRNA decay rapidly degrades mRNAs harbouring PTCs (premature translation-termination codons) and thereby prevents the accumulation of potentially deleterious proteins with C-terminal truncations. In the present article, I review recent data from yeast, fluitflies, nematode worms and human cells and endeavour to merge these results into a unified model for recognition of nonsense mRNA. According to this model, the distinction between translation termination at PTCs and at 'normal' termination codons relies on the physical distance between the terminating ribosome and PABP [poly(A)-binding protein]. Correct translation termination is promoted by a PABP-mediated signal to the terminating ribosome, whereas the absence of this signal leads to the assembly of an mRNA decay-promoting protein complex including the conserved NMD factors UPF (up-frameshift) 1-3.

  8. Post-transcriptional gene regulation by mRNA modifications

    Science.gov (United States)

    Zhao, Boxuan Simen; Roundtree, Ian A.; He, Chuan

    2016-01-01

    The recent discovery of reversible mRNA methylation has opened a new realm of post-transcriptional gene regulation in eukaryotes. The identification and functional characterization of proteins that specifically recognize RNA N6-methyladenosine (m6A) unveiled it as a modification that cells utilize to accelerate mRNA metabolism and translation. N6-adenosine methylation directs mRNAs to distinct fates by grouping them for differential processing, translation and decay in processes such as cell differentiation, embryonic development and stress responses. Other mRNA modifications, including N1-methyladenosine (m1A), 5-methylcytosine (m5C) and pseudouridine, together with m6A form the epitranscriptome and collectively code a new layer of information that controls protein synthesis. PMID:27808276

  9. Increased IL-10 mRNA and IL-23 mRNA expression in multiple sclerosis: interferon-beta treatment increases IL-10 mRNA expression while reducing IL-23 mRNA expression

    DEFF Research Database (Denmark)

    Krakauer, M.; Sorensen, P.; Khademi, M.

    2008-01-01

    volunteers served to confirm initial findings. mRNA was analyzed by real-time reverse transcriptase polymerase chain reaction (PCR). RESULTS: We found elevated expression of interleukin (IL)-23 and IL-10 in untreated MS patients. IFN-beta therapy increased IL-10 and decreased IL-23 expression independently...... of the regulatory cytokine IL-10. The elevated IL-23 mRNA levels in MS patients are noteworthy in view of the newly discovered IL-23-driven Th17 T-cell subset, which is crucial in animal models of MS. Since IFN-beta therapy resulted in decreased IL-23 mRNA levels, the Th17 axis could be another target of IFN...

  10. Simultaneous detection of mRNA and protein stem cell markers in live cells

    Directory of Open Access Journals (Sweden)

    Bao Gang

    2009-04-01

    Full Text Available Abstract Background Biological studies and medical application of stem cells often require the isolation of stem cells from a mixed cell population, including the detection of cancer stem cells in tumor tissue, and isolation of induced pluripotent stem cells after eliciting the expression of specific genes in adult cells. Here we report the detection of Oct-4 mRNA and SSEA-1 protein in live carcinoma stem cells using respectively molecular beacon and dye-labeled antibody, aiming to establish a new method for stem cells detection and isolation. Results Quantification of Oct-4 mRNA and protein in P19 mouse carcinoma stem cells using respectively RT-PCR and immunocytochemistry confirmed that their levels drastically decreased after differentiation. To visualize Oct-4 mRNA in live stem cells, molecular beacons were designed, synthesized and validated, and the detection specificity was confirmed using control studies. We found that the fluorescence signal from Oct-4-targeting molecular beacons provides a clear discrimination between undifferentiated and retinoic acid-induced differentiated cells. Using deconvolution fluorescence microscopy, Oct-4 mRNAs were found to reside on one side of the cytosol. We demonstrated that, using a combination of Oct-4 mRNA-targeting molecular beacon with SSEA-1 antibody in flow cytometric analysis, undifferentiated stem cells can be clearly distinguished from differentiated cells. We revealed that Oct-4 targeting molecular beacons do not seem to affect stem cell biology. Conclusion Molecular beacons have the potential to provide a powerful tool for highly specific detection and isolation of stem cells, including cancer stem cells and induced pluripotent stem (iPS cells without disturbing cell physiology. It is advantageous to perform simultaneous detection of intracellular (mRNA and cell-surface (protein stem cell markers in flow cytometric analysis, which may lead to high detection sensitivity and efficiency.

  11. Complexity on Acute Myeloid Leukemia mRNA Transcript Variant

    Directory of Open Access Journals (Sweden)

    Carlo Cattani

    2011-01-01

    Full Text Available This paper deals with the sequence analysis of acute myeloid leukemia mRNA. Six transcript variants of mlf1 mRNA, with more than 2000 bps, are analyzed by focusing on the autocorrelation of each distribution. Through the correlation matrix, some patches and similarities are singled out and commented, with respect to similar distributions. The comparison of Kolmogorov fractal dimension will be also given in order to classify the six variants. The existence of a fractal shape, patterns, and symmetries are discussed as well.

  12. Aging in Rats Differentially Affects Markers of Transcriptional and Translational Capacity in Soleus and Plantaris Muscle

    Directory of Open Access Journals (Sweden)

    Christopher B. Mobley

    2017-07-01

    Full Text Available Alterations in transcriptional and translational mechanisms occur during skeletal muscle aging and such changes may contribute to age-related atrophy. Herein, we examined markers related to global transcriptional output (i.e., myonuclear number, total mRNA and RNA pol II levels, translational efficiency [i.e., eukaryotic initiation and elongation factor levels and muscle protein synthesis (MPS levels] and translational capacity (ribosome density in the slow-twitch soleus and fast-twitch plantaris muscles of male Fischer 344 rats aged 3, 6, 12, 18, and 24 months (n = 9–10 per group. We also examined alterations in markers of proteolysis and oxidative stress in these muscles (i.e., 20S proteasome activity, poly-ubiquinated protein levels and 4-HNE levels. Notable plantaris muscle observations included: (a fiber cross sectional area (CSA was 59% (p < 0.05 and 48% (p < 0.05 greater in 12 month vs. 3 month and 24 month rats, respectively, suggesting a peak lifetime value near 12 months and age-related atrophy by 24 months, (b MPS levels were greatest in 18 month rats (p < 0.05 despite the onset of atrophy, (c while regulators of ribosome biogenesis [c-Myc and upstream binding factor (UBF protein levels] generally increased with age, ribosome density linearly decreased from 3 months of age and RNA polymerase (Pol I protein levels were lowest in 24 month rats, and d 20S proteasome activity was robustly up-regulated in 6 and 24 month rats (p < 0.05. Notable soleus muscle observations included: (a fiber CSA was greatest in 6 month rats and was maintained in older age groups, and (b 20S proteasome activity was modestly but significantly greater in 24 month vs. 3/12/18 month rats (p < 0.05, and (c total mRNA levels (suggestive of transcriptional output trended downward in older rats despite non-significant between-group differences in myonuclear number and/or RNA Pol II protein levels. Collectively, these findings suggest that plantaris, not soleus

  13. Developmental changes in the hypothalamic mRNA levels of prepro-orexin and orexin receptors and their sensitivity to fasting in male and female rats.

    Science.gov (United States)

    Iwasa, Takeshi; Matsuzaki, Toshiya; Munkhzaya, Munkhsaikhan; Tungalagsuvd, Altankhuu; Kuwahara, Akira; Yasui, Toshiyuki; Irahara, Minoru

    2015-11-01

    Orexin, which is also called as hypocretin (Hcrt), a product of the prepro-orexin (pp-orexin//Hcrt) gene, affects various physiological and behavioral functions, such as the sleep-wake cycle and appetite. The developmental changes in the hypothalamic mRNA levels of pp-prexin and the orexin receptors OX1R and OX2R and their sensitivity to fasting were evaluated in both male and female rats. During development, hypothalamic pp-orexin/Hcrt mRNA expression increased in both male and female rats, whereas hypothalamic OX1R mRNA expression decreased in both sexes. In addition, hypothalamic OX2R mRNA expression increased in male rats, but did not change in female rats. Fasting did not affect hypothalamic pp-orexin/Hcrt mRNA expression in either sex. Hypothalamic OX1R mRNA expression was increased by fasting in the prepubertal period (postnatal days 20 and 30) in female rats, but was not affected by fasting in males. In male rats, hypothalamic OX2R mRNA expression was decreased by fasting during the neonatal period (postnatal day 10), but not the prepubertal period (postnatal days 20 and 30). In females, hypothalamic OX2R mRNA expression was also decreased by fasting; however, the fasting-induced downregulation of hypothalamic OX2R expression persisted until postnatal day 20. These results indicate that the developmental patterns of components of the orexin system and their sensitivity to fasting during the neonatal and prepubertal periods only differ slightly between the sexes. These differences might be involved in the development of some physiological and behavioral functions. Copyright © 2015 Elsevier Ltd. All rights reserved.

  14. mRNA expression of genes regulating lipid metabolism in ringed seals (Pusa hispida) from differently polluted areas

    Energy Technology Data Exchange (ETDEWEB)

    Castelli, Martina Galatea [Norwegian Polar Institute, Fram Centre, 9296 Tromsø (Norway); University of Bergen, Department of Biology, 5020 Bergen (Norway); Rusten, Marte; Goksøyr, Anders [University of Bergen, Department of Biology, 5020 Bergen (Norway); Routti, Heli, E-mail: heli.routti@npolar.no [Norwegian Polar Institute, Fram Centre, 9296 Tromsø (Norway)

    2014-01-15

    Highlights: •Genes regulating lipid metabolism were studied in ringed seals. •We compared highly contaminated Baltic seals and less contaminated Svalbard seals. •mRNA expression of hepatic PPARγ was higher in the Baltic seals. •mRNA expression of adipose PPARγ target genes was higher in the Baltic seals. •Contaminant exposure may affect lipid metabolism in the Baltic ringed seals. -- Abstract: There is a growing concern about the ability of persistent organic pollutants (POPs) to influence lipid metabolism. Although POPs are found at high concentrations in some populations of marine mammals, for example in the ringed seal (Pusa hispida) from the Baltic Sea, little is known about the effects of POPs on their lipid metabolism. An optimal regulation of lipid metabolism is crucial for ringed seals during the fasting/molting season. This is a physiologically stressful period, during which they rely on the energy stored in their fat reserves. The mRNA expression levels for seven genes involved in lipid metabolism were analyzed in liver and/or blubber tissue from molting ringed seals from the polluted Baltic Sea and a less polluted reference location, Svalbard (Norway). mRNA expression of genes encoding peroxisome proliferator-activated receptors (PPAR) α and γ and their target genes acyl-coenzyme A oxidase 1 (ACOX1) and cluster of differentiation 36 (CD36) were analyzed in liver. mRNA expression level of genes encoding PPARβ, PPARγ and their target genes encoding fatty acid binding protein 4 (FABP4) and adiponectin (ADIPOQ) were measured in inner and middle blubber layers. In addition, we evaluated the influence of molting status on hepatic mRNA expression of genes encoding PPARs and their target genes in ringed seals from Svalbard. Our results show higher mRNA expression of genes encoding hepatic PPARγ and adipose PPARβ, FABP4, and ADIPOQ in the Baltic seals compared to the Svalbard seals. A positive relationship between mRNA expressions of genes

  15. Xplore mRNA assays for the quantification of IL-1 beta and TNF-alpha mRNA in lipopolysaccharide-induced mouse macrophages

    National Research Council Canada - National Science Library

    Van Arsdell, S W; Murphy, K P; Pazmany, C; Erickson, D; Burns, C; Moody, M D

    2000-01-01

    Because the accurate measurement of a number of cytokine mRNA transcripts provides valuable knowledge about cytokine gene regulation, we have developed the Xplore assay for the quantification of cytokine mRNA...

  16. Analysis of mRNA recognition by human thymidylate synthase.

    Science.gov (United States)

    Brunn, Nicholas D; Dibrov, Sergey M; Kao, Melody B; Ghassemian, Majid; Hermann, Thomas

    2014-12-23

    Expression of hTS (human thymidylate synthase), a key enzyme in thymidine biosynthesis, is regulated on the translational level through a feedback mechanism that is rarely found in eukaryotes. At low substrate concentrations, the ligand-free enzyme binds to its own mRNA and stabilizes a hairpin structure that sequesters the start codon. When in complex with dUMP (2'-deoxyuridine-5'-monophosphate) and a THF (tetrahydrofolate) cofactor, the enzyme adopts a conformation that is unable to bind and repress expression of mRNA. Here, we have used a combination of X-ray crystallography, RNA mutagenesis and site-specific cross-linking studies to investigate the molecular recognition of TS mRNA by the hTS enzyme. The interacting mRNA region was narrowed to the start codon and immediately flanking sequences. In the hTS enzyme, a helix-loop-helix domain on the protein surface was identified as the putative RNA-binding site.

  17. Association between VDAC1 mRNA expression and intracellular ...

    African Journals Online (AJOL)

    One way in which xenobiotics induce apoptotic cell death is to alter the selective permeability of the intracellular voltage-dependent anion channel (VDAC1) in the mitochondrial membrane. In this study, we explored the association between VDAC1 mRNA expression and mitochondrial function during hexavalent chromium ...

  18. Cytokine mRNA expression during experimental corneal allograft rejection

    NARCIS (Netherlands)

    Torres, P. F.; de Vos, A. F.; van der Gaag, R.; Martins, B.; Kijlstra, A.

    1996-01-01

    Allograft rejection is the main cause of corneal graft failure. T lymphocytes and macrophages have been implied to be involved in corneal rejection, but little is known about the molecular mechanism in this process. In this study, cytokine mRNA expression in the cornea was analysed during

  19. Human mRNA response to exercise and temperature.

    Science.gov (United States)

    Slivka, D R; Dumke, C L; Tucker, T J; Cuddy, J S; Ruby, B

    2012-02-01

    The purpose of this research was to determine the mRNA response to exercise in different environmental temperatures. 9 recreationally active males (27±1 years, 77.4±2.7  kg, 13.5±1.5% fat, 4.49±0.15  L · min (-1) VO2 max) completed 3 trials consisting of 1 h cycling exercise at 60% Wmax followed by a 3 h recovery in the cold (7°C), room temperature (20°C), and hot (33°C) environments. Muscle biopsies were obtained pre, post, and 3 h post exercise for the analysis of glycogen and mRNA. Expired gases were collected to calculate substrate use. PGC-1α increased to a greater degree in the cold trial than in the room temperature trial (p=0.036) and the hot trial (p=0.006). PGC1-α mRNA was also higher after the room temperature trial than the hot trial (p=0.050). UCP3 and MFN2 mRNA increased with exercise (pcold than exercise in the heat. However, VO2 was higher during recovery in the cold trial than in the room temperature and hot trials (p<0.05). This study presents evidence of PGC-1α temperature sensitivity in human skeletal muscle. © Georg Thieme Verlag KG Stuttgart · New York.

  20. Cloning and mRNA expression pattern analysis under low ...

    African Journals Online (AJOL)

    This research cloned endochitinase-antifreeze protein precursor (EAPP) gene of Dong-mu 70 rye (Secale cereale) by designing special primers according to Genbank's EAPP gene sequence, and analyzing the influence of low temperature stress on the expression of mRNA with RT-PCR. The results indicated that the ...

  1. Cloning and mRNA expression pattern analysis under low ...

    African Journals Online (AJOL)

    Jane

    2011-07-13

    Jul 13, 2011 ... This research cloned endochitinase-antifreeze protein precursor (EAPP) gene of Dong-mu 70 rye. (Secale cereale) by designing special primers according to Genbank's EAPP gene sequence, and analyzing the influence of low temperature stress on the expression of mRNA with RT-PCR. The results.

  2. Regulation of mRNA Trafficking by Nuclear Pore Complexes

    Directory of Open Access Journals (Sweden)

    Amandine Bonnet

    2014-09-01

    Full Text Available Over the last two decades, multiple studies have explored the mechanisms governing mRNA export out of the nucleus, a crucial step in eukaryotic gene expression. During transcription and processing, mRNAs are assembled into messenger ribonucleoparticles (mRNPs. mRNPs are then exported through nuclear pore complexes (NPCs, which are large multiprotein assemblies made of several copies of a limited number of nucleoporins. A considerable effort has been put into the dissection of mRNA export through NPCs at both cellular and molecular levels, revealing the conserved contributions of a subset of nucleoporins in this process, from yeast to vertebrates. Several reports have also demonstrated the ability of NPCs to sort out properly-processed mRNPs for entry into the nuclear export pathway. Importantly, changes in mRNA export have been associated with post-translational modifications of nucleoporins or changes in NPC composition, depending on cell cycle progression, development or exposure to stress. How NPC modifications also impact on cellular mRNA export in disease situations, notably upon viral infection, is discussed.

  3. MRNA-based skin identification for forensic applications

    NARCIS (Netherlands)

    M. Visser (Mijke); D. Zubakov (Dmitry); K. Ballantyne (Kaye); M.H. Kayser (Manfred)

    2011-01-01

    textabstractAlthough the identification of human skin cells is of important relevance in many forensic cases, there is currently no reliable method available. Here, we present a highly specific and sensitive messenger RNA (mRNA) approach for skin identification, meeting the key requirements in

  4. Reduced mRNA expression of PTGDS in peripheral blood mononuclear cells of rapid-cycling bipolar disorder patients compared with healthy control subjects

    DEFF Research Database (Denmark)

    Munkholm, Klaus; Peijs, Lone; Kessing, Lars Vedel

    2015-01-01

    that mRNA expression of PTGDS and AKR1C3 is deregulated in rapid-cycling disorder patients in a euthymic or current affective state compared with healthy control subjects, and that expression alters with affective states. METHODS: PTGDS and AKR1C3 mRNA expression in peripheral blood mononuclear cells......BACKGROUND: Disturbances related to the arachidonic acid cascade and prostaglandin metabolism may be involved in the pathophysiology of bipolar disorder, as supported by a recent genome-wide association study meta-analysis; however, evidence from clinical studies on a transcriptional level...... was measured in 37 rapid-cycling bipolar disorder patients and 40 age- and gender-matched healthy control subjects using reverse transcription quantitative real-time polymerase chain reaction. Repeated measurements of PTGDS and AKR1C3 mRNA expression were obtained in various affective states during 6-12 months...

  5. Dinaciclib potently suppresses MCL-1 and selectively induces the cell death in human iPS cells without affecting the viability of cardiac tissue.

    Science.gov (United States)

    Alsayegh, Khaled; Matsuura, Katsuhisa; Sekine, Hidekazu; Shimizu, Tatsuya

    2017-03-31

    Induced pluripotent stem (iPS) cells hold great potential for being a major source of cells for regenerative medicine. One major issue that hinders their advancement to clinic is the persistence of undifferentiated iPS cells in iPS-derived tissue. In this report, we show that the CDKs inhibitor, Dinaciclib, selectively eliminates iPS cells without affecting the viability of cardiac cells. We found that low nanomolar concentration of dinaciclib increased DNA damage and p53 protein levels in iPSCs. This was accompanied by negative regulation of the anti-apoptotic protein MCL-1. Gene knockdown experiments revealed that p53 downregulation only increased the threshold of dinaciclib induced apoptosis in iPS cells. Dinaciclib also inhibited the phosphorylation of Serine 2 of the C-terminal domain of RNA Polyemrase II through CDK9 inhibition. This resulted in the inhibition of transcription of MCL-1 and the pluripotency genes, NANOG and c-MYC. Even though dinaciclib caused a slight downregulation of MCL-1 in iPS-derived cardiac cells, the viability of the cells was not significantly affected, and beating iPS-derived cardiac cell sheet could still be fabricated. These findings suggest a difference in tolerance of MCL-1 downregulation between iPSCs and iPS-derived cardiac cells which could be exploited to eliminate remaining iPS cells in bioengineered cell sheet tissues.

  6. Reliable PCR quantitation of estrogen, progesterone and ERBB2 receptor mRNA from formalin-fixed, paraffin-embedded tissue is independent of prior macro-dissection

    DEFF Research Database (Denmark)

    Tramm, Trine; Hennig, Guido; Kyndi, Marianne

    2013-01-01

    Gene expression analysis on messenger RNA (mRNA) purified from formalin-fixed, paraffin-embedded tissue is increasingly used for research purposes. Tissue heterogeneity may question specificity and interpretation of results from mRNA isolated from a whole slide section, and thresholds for minimal...... tumor content in the paraffin block or macrodissection are used to avoid contamination from non-neoplastic tissue. The aim was to test if mRNA from tissue surrounding breast cancer affected quantification of estrogen receptor α (ESR1), progesterone receptor (PGR) and human epidermal growth factor...... receptor 2 (ERBB2), by comparing gene expression from whole slide and tumor-enriched sections, and correlating gene expression from whole slide sections with corresponding immunohistochemistry. Gene expression, based on mRNA extracted from a training set (36 paraffin blocks) and two validation sets (133...

  7. Reliable PCR quantitation of estrogen, progesterone and ERBB2 receptor mRNA from formalin-fixed, paraffin-embedded tissue is independent of prior macro-dissection

    DEFF Research Database (Denmark)

    Tramm, Trine; Hennig, Guido; Kyndi, Marianne

    2013-01-01

    Gene expression analysis on messenger RNA (mRNA) purified from formalin-fixed, paraffin-embedded tissue is increasingly used for research purposes. Tissue heterogeneity may question specificity and interpretation of results from mRNA isolated from a whole slide section, and thresholds for minimal...... tumor content in the paraffin block or macrodissection are used to avoid contamination from non-neoplastic tissue. The aim was to test if mRNA from tissue surrounding breast cancer affected quantification of estrogen receptor α (ESR1), progesterone receptor (PGR) and human epidermal growth factor...... and ERBB2, and 83 % for PGR. Overall agreements, when comparing mRNA expression to immunohistochemistry, were 100 % (ERBB2), 89 % (ESR1) and 83 % (PGR), which was confirmed in the validation sets. Percentage of tumor in the sections did not influence the results. In conclusion, reliable quantification...

  8. DBIRD complex integrates alternative mRNA splicing with RNA polymerase II transcript elongation

    DEFF Research Database (Denmark)

    Close, Pierre; East, Philip; Dirac-Svejstrup, A Barbara

    2012-01-01

    and help to integrate transcript elongation with mRNA splicing remain unclear. Here we characterize the human interactome of chromatin-associated mRNP particles. This led us to identify deleted in breast cancer 1 (DBC1) and ZNF326 (which we call ZNF-protein interacting with nuclear mRNPs and DBC1 (ZIRD......)) as subunits of a novel protein complex--named DBIRD--that binds directly to RNAPII. DBIRD regulates alternative splicing of a large set of exons embedded in (A + T)-rich DNA, and is present at the affected exons. RNA-interference-mediated DBIRD depletion results in region-specific decreases in transcript...... elongation, particularly across areas encompassing affected exons. Together, these data indicate that the DBIRD complex acts at the interface between mRNP particles and RNAPII, integrating transcript elongation with the regulation of alternative splicing....

  9. Alternative mRNA Splicing from the Glial Fibrillary Acidic Protein (GFAP) Gene Generates Isoforms with Distinct Subcellular mRNA Localization Patterns in Astrocytes

    DEFF Research Database (Denmark)

    Thomsen, Rune; Daugaard, Tina Fuglsang; Holm, Ida E

    2013-01-01

    The intermediate filament network of astrocytes includes Glial fibrillary acidic protein (Gfap) as a major component. Gfap mRNA is alternatively spliced resulting in generation of different protein isoforms where Gfapa is the most predominant isoform. The Gfapd isoform is expressed in proliferating......RNA localization patterns were dependent on the different 39-exon sequences included in Gfapd and Gfapa mRNA. The presented results show that alternative Gfap mRNA splicing results in isoform-specific mRNA localization patterns with resulting different local mRNA concentration ratios which have potential...

  10. The Impact of Ramadan Fasting on SIRT1 mRNA Expression in Peripheral Blood Mononuclear Cells

    OpenAIRE

    Mostafa Haji Molahoseini; kanaan Gorjipour; Farshid Yeganeh

    2016-01-01

    Background:The aim of this study was to evaluate the effect of Ramadan fasting on SIRT1 mRNA expression in healthy men.Islamic Ramadan fasting is a holy religious ceremony that has many spiritual benefits. Additionally, it can be considered as the equivalent of calorie restriction that may affect physical health. The results of previous studies revealed that calorie restriction increases the lifespan in laboratory rodents via increasing the expression of a histone deacetylase named SIRT1. Add...

  11. Interleukin-21 mRNA expression during virus infections

    DEFF Research Database (Denmark)

    Holm, Christian; Nyvold, Charlotte Guldborg; Paludan, Søren Riis

    2006-01-01

    and activational effects of IL-21 on different leukocytes come into play in vivo in an immune response has so far not been fully investigated. We show here for the first time in vivo, that IL-21 mRNA is produced in the spleen when mice are challenged with herpes simplex virus type 2 (HSV-2) or lymphocytic...... choriomeningitis virus (LCMV). We show in HSV-2 challenged mice that this production takes place in CD4+ T cell fractions and is absent in CD4+ T cell-depleted fractions. We also show that the peak of IL-21 mRNA production in both the HSV-2 and LCMV-challenged mice coincides with the onset of the adaptive immune...... response. Thus, our data suggest a role for IL-21 in the early stages of adaptive immune response against virus infections....

  12. Translation with frameshifting of ribosome along mRNA transcript

    CERN Document Server

    Li, Jingwei

    2015-01-01

    Translation is an important process for prokaryotic and eukaryotic cells to produce necessary proteins for cell growth. Numerious experiments have been performed to explore the translational properties. Diverse models have also been developed to determine the biochemical mechanism of translation. However, to simplify the majority of the existing models, the frameshifting of ribosome along the mRNA transcript is neglected, which actually occurs in real cells and has been extensively experimentally studied. The frameshifting of ribosome evidently influences the efficiency and speed of translation, considering that the peptide chains synthesized by shifted ribosomes will not fold into functional proteins and will degrade rapidly. In this study, a theoretical model is presented to describe the translational process based on the model for totally asymmetric simple exclusion process. In this model, the frameshifting of the ribosome along the mRNA transcript and the attachment/detachment of the ribosome to/from the ...

  13. Translatability of rat kidney mRNA after mercury administration.

    Science.gov (United States)

    Samji, S; Kuliszewski, M J; Girgis, G R; Nicholls, D M

    1985-09-01

    Young male rats received an intraperitoneal injection of 0.5 mg HgCl2/kg body weight and 16 h later the kidneys were removed and homogenized to prepare the polysomal fraction from which the poly(A)+ RNA was obtained. The activity of this fraction was assessed by translating the poly(A)+ RNA in a mRNA-dependent rabbit reticulocyte lysate and the activity was markedly elevated relative to preparations from control rat kidneys. The incorporation of labelled leucine and cysteine, but not phenylalanine, into a low molecular weight protein (approximately 10 000 as judged by denaturing polyacrylamide gel electrophoresis) accounted for the increased mRNA activity. The mobility during electrophoresis of the denatured labelled product and carboxymethylated product, as well as their acidic isoelectric points, provided evidence that it is metallothionein mRNA which exhibits increased translatability in preparations derived from mercury-treated rats.

  14. Affect Regulation

    DEFF Research Database (Denmark)

    Pedersen, Signe Holm; Poulsen, Stig Bernt; Lunn, Susanne

    2014-01-01

    Gergely and colleagues’ state that their Social Biofeedback Theory of Parental Affect Mirroring” can be seen as a kind of operationalization of the classical psychoanalytic concepts of holding, containing and mirroring. This article examines to what extent the social biofeedback theory of parental...... affect mirroring may be understood as a specification of these concepts. It is argued that despite similarities at a descriptive level the concepts are embedded in theories with different ideas of subjectivity. Hence an understanding of the concept of affect regulation as a concretizisation...... and specification of the classical concepts dilutes the complexity of both the concept of affect regulation and of the classical concepts....

  15. Sequence and structure-specific elements of HERG mRNA determine channel synthesis and trafficking efficiency

    Science.gov (United States)

    Sroubek, Jakub; Krishnan, Yamini; McDonald, Thomas V.

    2013-01-01

    Human ether-á-gogo-related gene (HERG) encodes a potassium channel that is highly susceptible to deleterious mutations resulting in susceptibility to fatal cardiac arrhythmias. Most mutations adversely affect HERG channel assembly and trafficking. Why the channel is so vulnerable to missense mutations is not well understood. Since nothing is known of how mRNA structural elements factor in channel processing, we synthesized a codon-modified HERG cDNA (HERG-CM) where the codons were synonymously changed to reduce GC content, secondary structure, and rare codon usage. HERG-CM produced typical IKr-like currents; however, channel synthesis and processing were markedly different. Translation efficiency was reduced for HERG-CM, as determined by heterologous expression, in vitro translation, and polysomal profiling. Trafficking efficiency to the cell surface was greatly enhanced, as assayed by immunofluorescence, subcellular fractionation, and surface labeling. Chimeras of HERG-NT/CM indicated that trafficking efficiency was largely dependent on 5′ sequences, while translation efficiency involved multiple areas. These results suggest that HERG translation and trafficking rates are independently governed by noncoding information in various regions of the mRNA molecule. Noncoding information embedded within the mRNA may play a role in the pathogenesis of hereditary arrhythmia syndromes and could provide an avenue for targeted therapeutics.—Sroubek, J., Krishnan, Y., McDonald, T V. Sequence- and structure-specific elements of HERG mRNA determine channel synthesis and trafficking efficiency. PMID:23608144

  16. In silico evaluation of miRNA binding site in mutated 3'UTR mRNA of G6PD

    Science.gov (United States)

    Azmi, Syarifah Anis Wafa Binti Syed Mohd; Noorden, Mohd Shihabudin; Yusof, Nurul Yuziana Mohd; Ismail, Endom

    2015-09-01

    MicroRNAs (miRNAs) are small non coding RNA sized 21-25 nucleotide. It has the ability to bind to the 3'- untranslated regions (3'UTR) of their target genes. Consequently, the binding of miRNA in the 3'UTR of targeted mRNA will regulate the expression of this gene. Thus, changes in 3'UTR may affect miRNA binding to mRNA of their target gene, leading to aberrations in mRNA regulations or expression and likely contribute to the various phenotypic changes or clinical risk for certain diseases in man. Therefore, the aim of this study is to evaluate candidate miRNAs species involved during the regulation of glucose-6-phosphate dehydrogenase (G6PD) mRNA with and without a specific 3'UTR nucleotide change that was previously shown to be responsible for G6PD deficiency in a Negrito sub-group of the Malaysian Orang Asli. We have conducted in silico analysis using TargetScan, PITA, RegRNA 2.0 and miRanda platform. Our results indicate that three potential miRNAs may have a functional role towards the regulated expression of those bearing the 3'UTR mutation. The role of these eleven miRNA can be investigated in future in vitro expression studies in order to verify its miRNA:mRNA relationship.

  17. Polysome Fractionation to Analyze mRNA Distribution Profiles.

    Science.gov (United States)

    Panda, Amaresh C; Martindale, Jennifer L; Gorospe, Myriam

    2017-02-05

    Eukaryotic cells adapt to changes in external or internal signals by precisely modulating the expression of specific gene products. The expression of protein-coding genes is controlled at the transcriptional and post-transcriptional levels. Among the latter steps, the regulation of translation is particularly important in cellular processes that require rapid changes in protein expression patterns. The translational efficiency of mRNAs is altered by RNA-binding proteins (RBPs) and noncoding (nc)RNAs such as microRNAs (Panda et al., 2014a and 2014b; Abdelmohsen et al., 2014). The impact of factors that regulate selective mRNA translation is a critical question in RNA biology. Polyribosome (polysome) fractionation analysis is a powerful method to assess the association of ribosomes with a given mRNA. It provides valuable information about the translational status of that mRNA, depending on the number of ribosomes with which they are associated, and identifies mRNAs that are not translated (Panda et al., 2016). mRNAs associated with many ribosomes form large polysomes that are predicted to be actively translated, while mRNAs associated with few or no ribosomes are expected to be translated poorly if at all. In sum, polysome fractionation analysis allows the direct determination of translation efficiencies at the level of the whole transcriptome as well as individual mRNAs.

  18. Cup regulates oskar mRNA stability during oogenesis.

    Science.gov (United States)

    Broyer, Risa M; Monfort, Elena; Wilhelm, James E

    2017-01-01

    The proper regulation of the localization, translation, and stability of maternally deposited transcripts is essential for embryonic development in many organisms. These different forms of regulation are mediated by the various protein subunits of the ribonucleoprotein (RNP) complexes that assemble on maternal mRNAs. However, while many of the subunits that regulate the localization and translation of maternal transcripts have been identified, relatively little is known about how maternal mRNAs are stockpiled and stored in a stable form to support early development. One of the best characterized regulators of maternal transcripts is Cup - a broadly conserved component of the maternal RNP complex that in Drosophila acts as a translational repressor of the localized message oskar. In this study, we have found that loss of cup disrupts the localization of both the oskar mRNA and its associated proteins to the posterior pole of the developing oocyte. This defect is not due to a failure to specify the oocyte or to disruption of RNP transport. Rather, the localization defects are due to a drop in oskar mRNA levels in cup mutant egg chambers. Thus, in addition to its role in regulating oskar mRNA translation, Cup also plays a critical role in controlling the stability of the oskar transcript. This suggests that Cup is ideally positioned to coordinate the translational control function of the maternal RNP complex with its role in storing maternal transcripts in a stable form. Published by Elsevier Inc.

  19. Final report: FASEB Summer Research Conference on ''Post-transcriptional control of gene expression: Effectors of mRNA decay'' [agenda and attendees list

    Energy Technology Data Exchange (ETDEWEB)

    Maquat, Lynne

    2002-12-01

    The goal of this meeting was to provide an interactive forum for scientists working on prokaryotic and eukaryotic mRNA decay. A special seminar presented by a leader in the field of mRNA decay in S. cerevisiae focused on what is known and what needs to be determined, not only for yeast but for other organisms. The large attendance (110 participants) reflects the awareness that mRNA decay is a key player in gene regulation in a way that is affected by the many steps that precede mRNA formation. Sessions were held on the following topics: mRNA transport and mRNP; multicomponent eukaryotic nucleases; nonsense-mediated mRNA decay and nonsense-associated altered splicing; Cis-acting sequences/Trans-acting factors of mRNA decay; translational accuracy; multicomponent bacterial nucleases; interplay between mRNA polyadenylation, translation and decay in prokaryotes and prokaryotic organelles; and RNA interference and other RNA mediators of gene expression. In addition to the talks and two poster sessions, there were three round tables: (1) Does translation occur in the nucleus? (2) Differences and similarities in the mechanisms of mRNA decay in different eukaryotes, and (3) RNA surveillance in bacteria?

  20. Genetic effect of CysLTR2 polymorphisms on its mRNA synthesis and stabilization

    Directory of Open Access Journals (Sweden)

    Chung Il

    2009-10-01

    Full Text Available Abstract Background We previously demonstrated that single nucleotide polymorphism (SNP and haplotypes were associated with aspirin hypersensitivity in asthmatics. We investigated the genetic effects of the SNPs and haplotypes on the expression of the CysLTR2 gene. Methods We measured CysLTR2 protein and mRNA expression in EB virus-infected B cell lines from asthmatics having ht1+/+ and ht2+/+. A gel retardation assay was used to identify nuclear protein binding to the c.-819 promoter site. The function of promoter and 3'-UTR were assessed using pGL3 luciferase and pEGFP reporter system, respectively. Results We found that the expression of CysLTR2 protein was higher in B cell lines of asthmatics having ht2+/+ than in those having ht1+/+. PMA/ionomycin induced higher mRNA expression of CysLTR2 in B cell lines from ht2+/+ asthmatics than those from ht1+/+ asthmatics. A nuclear protein from the B cell lines showed stronger DNA binding affinity with a probe containing c.-819T than one containing c.-819G. The luciferase activity of the c.-819T type of CysLTR2 promoter was higher than that of the c.-819G type. EGFP expression was higher in the EGFP-c.2078T 3'-UTR fusion construct than in the c.2078C construct. Conclusion The sequence variants of CysLTR2 may affect its transcription and the stability of its mRNA, resulting in altered expression of CysLTR2 protein, which in turn causes some asthmatics to be susceptible to aspirin hypersensitivity.

  1. The Substrates of Nonsense-Mediated mRNA Decay in Caenorhabditis elegans

    Directory of Open Access Journals (Sweden)

    Virginia S. Muir

    2018-01-01

    Full Text Available Nonsense-mediated mRNA decay (NMD is a conserved pathway that strongly influences eukaryotic gene expression. Inactivating or inhibiting NMD affects the abundance of a substantial fraction of the transcriptome in numerous species. Transcripts whose abundance is altered in NMD-deficient cells may represent either direct substrates of NMD or indirect effects of inhibiting NMD. We present a genome-wide investigation of the direct substrates of NMD in Caenorhabditis elegans. Our goals were (i to identify mRNA substrates of NMD and (ii to distinguish those mRNAs from others whose abundance is indirectly influenced by the absence of NMD. We previously demonstrated that Upf1p/SMG-2, the central effector of NMD in all studied eukaryotes, preferentially associates with mRNAs that contain premature translation termination codons. We used this preferential association to distinguish direct from indirect effects by coupling immunopurification of Upf1/SMG-2 with high-throughput mRNA sequencing of NMD-deficient mutants and NMD-proficient controls. We identify 680 substrates of NMD, 171 of which contain novel spliced forms that (i include sequences of annotated introns and (ii have not been previously documented in the C. elegans transcriptome. NMD degrades unproductively spliced mRNAs with sufficient efficiency in NMD-proficient strains that such mRNAs were not previously known. Two classes of genes are enriched among the identified NMD substrates: (i mRNAs of expressed pseudogenes and (ii mRNAs of gene families whose gene number has recently expanded in the C. elegans genome. Our results identify novel NMD substrates and provide a context for understanding NMD’s role in normal gene expression and genome evolution.

  2. Real-time RT-PCR analysis of mRNA decay: half-life of Beta-actin mRNA in human leukemia CCRF-CEM and Nalm-6 cell lines

    Directory of Open Access Journals (Sweden)

    Barredo Julio C

    2002-03-01

    Full Text Available Abstract Background We describe an alternative method to determine mRNA half-life (t1/2 based on the Real-Time RT-PCR procedure. This approach was evaluated by using the β-actin gene as a reference molecule for measuring of mRNA stability. Results Human leukemia Nalm-6 and CCRF-CEM cells were treated with various concentrations of Actinomycin D to block transcription and aliquots were removed periodically. Total RNA was isolated and quantified using the RiboGreen® fluorescent dye with the VersaFluor Fluorometer System. One μg of total RNA was reverse transcribed and used as template for the amplification of a region of the β-actin gene (231 bp. To generate the standard curve, serial ten-fold dilutions of the pBactin-231 vector containing the cDNA amplified fragment were employed, β-actin mRNAs were quantified by Real-Time RT-PCR using the SYBR® Green I fluorogenic dye and data analyzed using the iCycle iQ system software. Using this method, the β-actin mRNA exhibited a half-life of 6.6 h and 13.5 h in Nalm-6 and CCRF-CEM cells, respectively. The t1/2 value obtained for Nalm-6 is comparable to those estimated from Northern blot studies, using normal human leukocytes (5.5 h. Conclusions We have developed a rapid, sensitive, and reliable method based on Real-Time RT-PCR for measuring mRNA half-life. Our results confirm that β-actin mRNA half-life can be affected by the cellular growth rate.

  3. Survivin inhibitor YM155 suppresses gastric cancer xenograft growth in mice without affecting normal tissues.

    Science.gov (United States)

    Cheng, Xiao Jiao; Lin, Jia Cheng; Ding, Yan Fei; Zhu, Liming; Ye, Jing; Tu, Shui Ping

    2016-02-09

    Survivin overexpression is associated with poor prognosis of human gastric cancer, and is a target for gastric cancer therapy. YM155 is originally identified as a specific inhibitor of survivin. In this study, we investigated the antitumor effect of YM155 on human gastric cancer. Our results showed that YM155 treatment significantly inhibited cell proliferation, reduced colony formation and induced apoptosis of gastric cancer cells in a dose-dependent manner. Accordingly, YM155 treatment significantly decreased survivin expression without affecting XIAP expression and increased the cleavage of apoptosis-associated proteins caspase 3, 7, 8, 9. YM155 significantly inhibited sphere formation of gastric cancer cells, suppressed expansion and growth of the formed spheres (cancer stem cell-like cells, CSCs) and downregulated the protein levels of β-catenin, c-Myc, Cyclin D1 and CD44 in gastric cancer cells. YM155 infusion at 5 mg/kg/day for 7 days markedly inhibited growth of gastric cancer xenograft in a nude mouse model. Immunohistochemistry staining and Western Blot showed that YM155 treatment inhibited expression of survivin and CD44, induced apoptosis and reduced CD44+ CSCs in xenograft tumor tissues in vivo. No obvious pathological changes were observed in organs (e.g. heart, liver, lung and kidney) in YM155-treated mice. Our results demonstrated that YM155 inhibits cell proliferation, induces cell apoptosis, reduces cancer stem cell expansion, and inhibits xenograft tumor growth in gastric cancer cells. Our results elucidate a new mechanism by which YM155 inhibits gastric cancer growth by inhibition of CSCs. YM155 may be a promising agent for gastric cancer treatment.

  4. Correlation of thyroid stimulating hormone receptor mRNA expression levels in peripheral blood with undesirable clinicopathological features in papillary thyroid carcinoma patients

    Science.gov (United States)

    Liu, Riming; Hao, Shaolong; Zhang, Hua; Ma, Jihong; Liu, Xincheng; Xu, Jie; Liu, Xin; Ning, Jinyao; Sun, Yan; Jiang, Lixin; Li, Guojun; Song, Xicheng; Zheng, Haitao

    2017-01-01

    To determine the extent to which thyroid stimulating hormone receptor (TSHR) mRNA in peripheral blood (PB) has diagnostic value for papillary thyroid carcinoma (PTC). We obtained pre- and postoperative PB samples from 104 thyroid disease patients and collected 11 healthy volunteers’ PB samples twice apiece at different times. We used reverse transcription polymerase chain reaction (RT-PCR) to quantify TSHR mRNA expression levels in the samples. T-test and chi-square test were used to compare quantitative data and rates. The mean preoperative PB TSHR mRNA expression level of the PTC patients was significantly higher than that of the healthy volunteers. However, on the postoperative day 1, PB TSHR mRNA level of PTC patients significantly decreased but not for healthy controls. Preoperative PB TSHR mRNA expression levels were significantly associated with patient age, capsular invasion status, lymph node metastasis status, and BRAFV600E mutation status (P cancer foci, or Hashimoto thyroiditis status. Preoperative assessment of the PB TSHR mRNA expression level combined with ultrasonography of the thyroid had better accuracy in the diagnosis of PTC than either method alone did. Moreover, TSHR mRNA expression significantly affected recurrence of PTC patients. Our findings suggest that PB TSHR mRNA expression level is a promising novel biomarker for the early detection, diagnosis, and treatment of PTC. It may serve as a noninvasive means of PTC detection and a prognostic biomarker of residual tumor and help guide further treatment. PMID:29088773

  5. Processing bodies are not required for mammalian nonsense-mediated mRNA decay.

    Science.gov (United States)

    Stalder, Lukas; Mühlemann, Oliver

    2009-07-01

    Nonsense-mediated mRNA decay (NMD) is a eukaryotic quality-control mechanism that recognizes and degrades mRNAs with premature termination codons (PTCs). In yeast, PTC-containing mRNAs are targeted to processing bodies (P-bodies), and yeast strains expressing an ATPase defective Upf1p mutant accumulate P-bodies. Here we show that in human cells, an ATPase-deficient UPF1 mutant and a fraction of UPF2 and UPF3b accumulate in cytoplasmic foci that co-localize with P-bodies. Depletion of the P-body component Ge-1, which prevents formation of microscopically detectable P-bodies, also impairs the localization of mutant UPF1, UPF2, and UPF3b in cytoplasmic foci. However, the accumulation of the ATPase-deficient UPF1 mutant in P-bodies is independent of UPF2, UPF3b, or SMG1, and the ATPase-deficient UPF1 mutant can localize into the P-bodies independent of its phosphorylation status. Most importantly, disruption of P-bodies by depletion of Ge-1 affects neither the mRNA levels of PTC-containing reporter genes nor endogenous NMD substrates. Consistent with the recently reported decapping-independent SMG6-mediated endonucleolytic decay of human nonsense mRNAs, our results imply that microscopically detectable P-bodies are not required for mammalian NMD.

  6. Antiinflammatory Effect of Rosiglitazone via Modulation of mRNA Stability of Interleukin 10 and Cyclooxygenase 2 in Astrocytes.

    Science.gov (United States)

    Pankevich, E V; Astakhova, A A; Chistyakov, D V; Sergeeva, M G

    2017-11-01

    Investigation of molecular mechanisms of proinflammatory stimuli signaling in astrocytes is important for understanding their role in pathogenesis of central nervous system diseases as well as in functioning of the innate immunity system in non-immune cells. Here we show that lipopolysaccharide (LPS) stimulation of primary rat astrocytes led to conventional inflammatory response: increase in both proinflammatory (tumor necrosis factor, TNFα; prostaglandin E 2 , PGE 2 ) and antiinflammatory marker (interleukin 10, IL-10) levels. The protein level of cyclooxygenase 2 (COX-2) was also increased. Rosiglitazone strengthened LPS-induced mRNA expression of COX-2 and IL-10 but not TNFα. Rosiglitazone is an agonist of nuclear receptor PPARγ, but its impact on IL-10 expression was not influenced by a PPARγ antagonist, GW9662, suggesting PPARγ-independent effect of rosiglitazone. The degradation of mRNA is one of the steps of inflammation regulation and might be affected by small molecules. In experiments with actinomycin D, we found that mRNA half-lives of IL-10, COX-2, and TNFα in naive astrocytes were 70, 44, and 19 min, respectively. LPS stimulation caused 2-fold increase in IL-10 and COX-2 mRNA decay rates, whereas addition of rosiglitazone restored them to the initial level. TNFα decay rate was not changed by these stimulations. This suggests that mRNA decay rate could be regulated by small molecules. Moreover, rosiglitazone could be used as a substance stimulating the resolution of inflammation without influence on proinflammatory signals. These results open new perspectives in the search for inflammation resolution modulators.

  7. Influence of progesterone on GAD65 and GAD67 mRNA expression in the dorsolateral striatum and prefrontal cortex of female rats repeatedly treated with cocaine

    Directory of Open Access Journals (Sweden)

    M.F. Souza

    2009-11-01

    Full Text Available Female rats are intensely affected by cocaine, with estrogen probably playing an important role in this effect. Progesterone modulates the GABA system and attenuates the effects of cocaine; however, there is no information about its relevance in changing GABA synthesis pathways after cocaine administration to female rats. Our objective was to investigate the influence of progesterone on the effects of repeated cocaine administration on the isoenzymes of glutamic acid decarboxylase (GAD65 and GAD67 mRNA in brain areas involved in the addiction circuitry. Ovariectomized, intact and progesterone replacement-treated female rats received saline or cocaine (30 mg/kg, ip acutely or repeatedly. GAD isoenzyme mRNA levels were determined in the dorsolateral striatum (dSTR and prefrontal cortex (PFC by RT-PCR, showing that repeated, but not acute, cocaine decreased GADs/β-actin mRNA ratio in the dSTR irrespective of the hormonal condition (GAD65: P < 0.001; and GAD67: P = 0.004. In the PFC, repeated cocaine decreased GAD65 and increased GAD67 mRNA ratio (P < 0.05. Progesterone replacement decreased both GAD isoenzymes mRNA ratio after acute cocaine in the PFC (P < 0.001 and repeated cocaine treatment reversed this decrease (P < 0.001. These results suggest that cocaine does not immediately affect GAD mRNA expression, while repeated cocaine decreases both GAD65 and GAD67 mRNA in the dSTR of female rats, independently of their hormonal conditions. In the PFC, repeated cocaine increases the expression of GAD isoenzymes, which were decreased due to progesterone replacement.

  8. Sodium arsenite-induced inhibition of cell proliferation is related to inhibition of IL-2 mRNA expression in mouse activated T cells

    Energy Technology Data Exchange (ETDEWEB)

    Conde, Patricia; Acosta-Saavedra, Leonor C.; Calderon-Aranda, Emma S. [Centro de Investigacion y de Estudios Avanzados, CINVESTAV, Seccion Toxicologia, P.O. Box 14-740, Mexico, D.F. (Mexico); Goytia-Acevedo, Raquel C. [Universidad Juarez del Estado de Durango, Facultad de Medicina, Gomez Palacio, Durango (Mexico)

    2007-04-15

    A proposed mechanism for the As-induced inhibition of cell proliferation is the inhibition of IL-2 secretion. However, the effects of arsenite on IL-2 mRNA expression or on the ERK pathway in activated-T cells have not yet been described. We examined the effect of arsenite on IL-2 mRNA expression, cell activation and proliferation in PHA-stimulated murine lymphocytes. Arsenite (1 and 10 {mu}M) decreased IL-2 mRNA expression, IL-2 secretion and cell proliferation. Arsenite (10 {mu}M) strongly inhibited ERK-phosphorylation. However, the partial inhibition (50%) of IL-2 mRNA produced by 1 {mu}M, consistent with the effects on IL-2 secretion and cell proliferation, could not be explained by the inhibition of ERK-phosphorylation, which was not affected at this concentration. The inhibition of IL-2 mRNA expression caused by 1 {mu}M could be associated to effects on pathways located downstream or parallel to ERK. Arsenite also decreased early activation (surface CD69{sup +} expression) in both CD4{sup +} and CD8{sup +}, and decreased total CD8{sup +} count without significantly affecting CD4{sup +}, supporting that the cellular immune response mediated by cytotoxic T cells is an arsenic target. Thus, our results suggest that arsenite decreases IL-2 mRNA levels and T-cell activation and proliferation. However, further studies on the effects of arsenite on IL-2 gene transcription and IL-2 mRNA stability are needed. (orig.)

  9. Ethylmalonic acid modulates Na+, K(+)-ATPase activity and mRNA levels in rat cerebral cortex.

    Science.gov (United States)

    Schuck, Patrícia Fernanda; De Assis, Dênis Reis; Viegas, Carolina Maso; Pereira, Talita Carneiro Brandão; Machado, Jéssica Luca; Furlanetto, Camila Brulezi; Bogo, Mauricio Reis; Streck, Emilio Luiz; Ferreira, Gustavo Costa

    2013-03-01

    Ethylmalonic acid (EMA) accumulates in tissues of patients affected by short-chain acyl-CoA dehydrogenase deficiency and ethylmalonic encephalopathy, illnesses characterized by variable neurological symptoms. In this work, we investigated the in vitro and in vivo EMA effects on Na(+), K(+)-ATPase (NAK) activity and mRNA levels in cerebral cortex from 30-day-old rats. For in vitro studies, cerebral cortex homogenates were incubated in the presence of EMA at 0.5, 1, or 2.5 mM concentrations for 1 h. For in vivo experiments, animals received three subcutaneous EMA injections (6 μmol g(-1); 90-min interval) and were killed 60 min after the last injection. After that, NAK activity and its mRNA expression were measured. We observed that EMA did not affect this enzyme activity in vitro. In contrast, EMA administration significantly increased NAK activity and decreased mRNA NAK expression as assessed by semiquantitative reverse transcriptase polymerase chain reaction when compared with control group. Considering the high score of residues prone to phosphorylation on NAK, this profile can be associated with a possible regulation by specific phosphorylation sites of the enzyme. Altogether, the present results suggest that NAK alterations may be involved in the pathophysiology of brain damage found in patients in which EMA accumulates. Copyright © 2012 Wiley Periodicals, Inc.

  10. IGF-I mRNA levels in bovine satellite cell cultures: effects of fusion and anabolic steroid treatment.

    Science.gov (United States)

    Kamanga-Sollo, E; Pampusch, M S; Xi, G; White, M E; Hathaway, M R; Dayton, W R

    2004-11-01

    Androgenic and estrogenic steroids enhance muscle growth in a number of species; however, the mechanism by which anabolic steroids enhance muscle growth is not known. Castrated male cattle (steers) provide a particularly good model system in which to study the effects of anabolic steroids on muscle growth because they respond dramatically to treatment with both estrogens and androgens. The goal of this study was to determine if treatment of bovine satellite cell (BSC) cultures with 17beta-estradiol (E(2)) or trenbolone (a synthetic androgen) directly affects proliferation rate or level of mRNA for estrogen receptor (ER)-alpha, androgen receptor, and growth factors that have been shown to affect muscle growth (insulin-like growth factor (IGF)-I, IGF binding protein (IGFBP)-3, and myostatin). BSC cultures were established from the semimembranosus muscles of steers and then treated for 48 h with various concentrations of E(2) or trenbolone ranging from 0.001 to 10 nM. IGF-I mRNA levels in proliferating BSC cultures were significantly increased at 0.01 (1.9-times control values, P steroids. Although, levels of IGF-I mRNA were 10-times greater (P steroids have direct anabolic effects on cells present in the BSC culture. Copyright 2004 Wiley-Liss, Inc.

  11. Live Imaging of mRNA Synthesis in Drosophila.

    Science.gov (United States)

    Garcia, Hernan G; Gregor, Thomas

    2018-01-01

    mRNA synthesis is one of the earliest readouts of the activity of a transcribed gene, which is of particular interest in the context of metazoan cell fate specification. These processes are intrinsically dynamic and stochastic, which makes in vivo single-cell measurements inevitable. Here, we present the application of a technology that has been widely used in single celled organisms to measure transcriptional activity in developing embryos of the fruit fly Drosophila melanogaster. The method allows for quantification of instantaneous polymerase occupancy of active gene loci and thereby enables the development and testing of models of gene regulation in development.

  12. NITRIC OXIDE PRODUCTION AND iNOS mRNA EXPRESSION IN IFN-8-STIMULATED CHICKEN MACROPHAGES TRANSFECTED WITH iNOS siRNAs

    Science.gov (United States)

    Utilizing RNA interference technology with siRNA in the HD-11 macrophage cell line, we determined how the knock-down of the iNOS (inducible nitric oxide synthase) gene affected IFN-' induced macrophage production of nitric oxide (NO) and mRNA expression of genes involved in this biological pathway i...

  13. Association of a PIT1 gene polymorphism with growth hormone mRNA levels in pig pituitary glands

    Directory of Open Access Journals (Sweden)

    Maurício Machaim Franco

    2005-03-01

    Full Text Available Fourty-six non-castrated, halothane-free, male Landrace pigs were genotyped by PCR-RFLP for the Rsa I polymorphism in the PIT1 gene and classified into AA and AB genotypes. Total RNA was extracted from the pituitaries and the relative quantities of growth hormone (GH mRNA were determined by semi-quantitative RT-PCR. Pigs with the AB genotype had higher levels of GH mRNA than those with the AA genotype (p = 0.034; Kruskal-Wallis test. This result suggests that the Rsa I polymorphism may be involved in Pit-1 protein expression or function, which in turn may influence GH transcription and expression. Thus, the Rsa I PIT1 gene polymorphism in this pig line may be used as a molecular marker to identify higher GH expression and possibly select for carcass and performance traits affected by GH.

  14. Naturally occurring BRCA2 alternative mRNA splicing events in clinically relevant samples

    DEFF Research Database (Denmark)

    Fackenthal, James D; Yoshimatsu, Toshio; Zhang, Bifeng

    2016-01-01

    BACKGROUND: BRCA1 and BRCA2 are the two principal tumour suppressor genes associated with inherited high risk of breast and ovarian cancer. Genetic testing of BRCA1/2 will often reveal one or more sequence variants of uncertain clinical significance, some of which may affect normal splicing...... patterns and thereby disrupt gene function. mRNA analyses are therefore among the tests used to interpret the clinical significance of some genetic variants. However, these could be confounded by the appearance of naturally occurring alternative transcripts unrelated to germline sequence variation...... or defects in gene function. To understand which novel splicing events are associated with splicing mutations and which are part of the normal BRCA2 splicing repertoire, a study was undertaken by members of the Evidence-based Network for the Interpretation of Germline Mutant Alleles (ENIGMA) consortium...

  15. Matrin 3 binds and stabilizes mRNA.

    Directory of Open Access Journals (Sweden)

    Maayan Salton

    Full Text Available Matrin 3 (MATR3 is a highly conserved, inner nuclear matrix protein with two zinc finger domains and two RNA recognition motifs (RRM, whose function is largely unknown. Recently we found MATR3 to be phosphorylated by the protein kinase ATM, which activates the cellular response to double strand breaks in the DNA. Here, we show that MATR3 interacts in an RNA-dependent manner with several proteins with established roles in RNA processing, and maintains its interaction with RNA via its RRM2 domain. Deep sequencing of the bound RNA (RIP-seq identified several small noncoding RNA species. Using microarray analysis to explore MATR3's role in transcription, we identified 77 transcripts whose amounts depended on the presence of MATR3. We validated this finding with nine transcripts which were also bound to the MATR3 complex. Finally, we demonstrated the importance of MATR3 for maintaining the stability of several of these mRNA species and conclude that it has a role in mRNA stabilization. The data suggest that the cellular level of MATR3, known to be highly regulated, modulates the stability of a group of gene transcripts.

  16. Expression of type XXIII collagen mRNA and protein.

    Science.gov (United States)

    Koch, Manuel; Veit, Guido; Stricker, Sigmar; Bhatt, Pinaki; Kutsch, Stefanie; Zhou, Peihong; Reinders, Elina; Hahn, Rita A; Song, Rich; Burgeson, Robert E; Gerecke, Donald R; Mundlos, Stefan; Gordon, Marion K

    2006-07-28

    Collagen XXIII is a member of the transmembranous subfamily of collagens containing a cytoplasmic domain, a membrane-spanning hydrophobic domain, and three extracellular triple helical collagenous domains interspersed with non-collagenous domains. We cloned mouse, chicken, and humanalpha1(XXIII) collagen cDNAs and showed that this non-abundant collagen has a limited tissue distribution in non-tumor tissues. Lung, cornea, brain, skin, tendon, and kidney are the major sites of expression. In contrast, five transformed cell lines were tested for collagen XXIII expression, and all expressed the mRNA. In vivo the alpha1(XXIII) mRNA is found in mature and developing organs, the latter demonstrated using stages of embryonic chick cornea and mouse embryos. Polyclonal antibodies were generated in guinea pig and rabbit and showed that collagen XXIII has a transmembranous form and a shed form. Comparison of collagen XXIII with its closest relatives in the transmembranous subfamily of collagens, types XIII and XXV, which have the same number of triple helical and non-collagenous regions, showed that there is a discontinuity in the alignment of domains but that striking similarities remain despite this.

  17. Tissue-specific expression and regulation by 1,25(OH)2D3 of chick protein kinase inhibitor (PKI) mRNA.

    Science.gov (United States)

    Marchetto, G S; Henry, H L

    1997-02-01

    The heat-stable protein kinase inhibitor (PKI) protein is a specific and potent competitive inhibitor of the catalytic subunit of cAMP-dependent protein kinase (PKA). Previously, it has been shown that vitamin D status affects chick kidney PKI activity: a 5- to 10-fold increase in PKI activity was observed in kidneys of chronically vitamin D-deficient chicks and treatment with 1,25-dihydroxyvitamin D3 (1,25[OH]2D3) in cultured kidney cells resulted in a 95% decrease in PKI activity. The authors have recently cloned the cDNA for chick kidney PKI and have used the coding sequence to study the regulation of PKI mRNA. Northern analysis showed the expression of two PKI messages, which are 2.7 and 3.3 kb in size. These mRNAs are expressed in brain, muscle, testis, and kidney, but not in pancreas, liver, or intestine. PKI mRNA steady-state levels are downregulated by 47% in kidneys from vitamin D-replete chicks as compared to vitamin D-deficient chicks. PKI mRNA levels in brain, muscle, and testis are not affected by vitamin D status. Treatment of primary chick kidney cultures treated with 10(-7) M 1,25(OH)2D3 for 24h resulted in a 20-30% decrease in PKI mRNA. 1,25(OH)2D3 treatment does not affect the stability of PKI mRNA as determined by treatment of cell cultures with actinomycin D. This study shows that 1,25(OH)2D3 directly and tissue-specifically downregulates PKI mRNA in the chick kidney.

  18. Affective Networks

    OpenAIRE

    Jodi Dean

    2010-01-01

    This article sets out the idea of affective networks as a constitutive feature of communicative capitalism. It explores the circulation of intensities in contemporary information and communication networks, arguing that this circulation should be theorized in terms of the psychoanalytic notion of the drive. The article includes critical engagements with theorists such as Guy Debord, Jacques Lacan, Tiziana Terranova, and Slavoj Zizek.

  19. Affective Urbanism

    DEFF Research Database (Denmark)

    Samson, Kristine

    Urban design and architecture are increasingly used as material and affective strategies for setting the scene, for manipulation and the production of urban life: The orchestration of atmospheres, the framing and staging of urban actions, the programming for contemplation, involvement, play...

  20. The Role of CYP3A4 mRNA Transcript with Shortened 3′-Untranslated Region in Hepatocyte Differentiation, Liver Development, and Response to Drug InductionS⃞

    OpenAIRE

    Li, Dan; Gaedigk, Roger; Hart, Steven N.; Leeder, J Steven; Zhong, Xiao-bo

    2012-01-01

    Cytochrome P450 3A4 (CYP3A4) metabolizes more than 50% of prescribed drugs. The expression of CYP3A4 changes during liver development and may be affected by the administration of some drugs. Alternative mRNA transcripts occur in more than 90% of human genes and are frequently observed in cells responding to developmental and environmental signals. Different mRNA transcripts may encode functionally distinct proteins or contribute to variability of mRNA stability or protein translation efficien...

  1. Affective Networks

    Directory of Open Access Journals (Sweden)

    Jodi Dean

    2010-02-01

    Full Text Available This article sets out the idea of affective networks as a constitutive feature of communicative capitalism. It explores the circulation of intensities in contemporary information and communication networks, arguing that this circulation should be theorized in terms of the psychoanalytic notion of the drive. The article includes critical engagements with theorists such as Guy Debord, Jacques Lacan, Tiziana Terranova, and Slavoj Zizek.

  2. Integrative genomics analysis of genes with biallelic loss and its relation to the expression of mRNA and micro-RNA in esophageal squamous cell carcinoma.

    Science.gov (United States)

    Hu, Nan; Wang, Chaoyu; Clifford, Robert J; Yang, Howard H; Su, Hua; Wang, Lemin; Wang, Yuan; Xu, Yi; Tang, Ze-Zhong; Ding, Ti; Zhang, Tongwu; Goldstein, Alisa M; Giffen, Carol; Lee, Maxwell P; Taylor, Philip R

    2015-09-26

    Genomic instability plays an important role in human cancers. We previously characterized genomic instability in esophageal squamous cell carcinomas (ESCC) in terms of loss of heterozygosity (LOH) and copy number (CN) changes in tumors. In the current study we focus on biallelic loss and its relation to expression of mRNA and miRNA in ESCC using results from 500 K SNP, mRNA, and miRNA arrays in 30 cases from a high-risk region of China. (i) Biallelic loss was uncommon but when it occurred it exhibited a consistent pattern: only 77 genes (RNA expression data, and 41 (79%) showed lower expression levels in cases with biallelic loss compared to those without. (iii) The relation of biallelic loss to miRNA expression was less clear but appeared to favor higher miRNA levels: of 60 miRNA-target gene pairs, 34 pairs (57%) had higher miRNA expression with biallelic loss than without, while 26 pairs (43%) had lower miRNA expression. (iv) Finally, the effect of biallelic loss on the relation between miRNA and mRNA expression was complex. Biallelic loss was most commonly associated with a pattern of elevated miRNA and reduced mRNA (43%), but a pattern of both reduced miRNA and mRNA was also common (35%). Our results indicate that biallelic loss in ESCC is uncommon, but when it occurs it is localized to a few specific chromosome regions and is associated with reduced mRNA expression of affected genes. The effect of biallelic loss on miRNA expression and on the relation between miRNA and mRNA expressions was complex.

  3. The Effects of Exercise on Expression of CYP19 and StAR mRNA in Steroid-Induced Polycystic Ovaries of Female Rats.

    Science.gov (United States)

    Aghaie, Fatemeh; Khazali, Homayoun; Hedayati, Mehdi; Akbarnejad, Ali

    2018-01-01

    Polycystic ovarian syndrome (PCOS) is the most frequent female endocrine disorder that affects 5-10% of women. PCOS is characterized by hyperandrogenism, oligo-/anovulation, and polycystic ovaries. The aim of the present research is to evaluate the expression of steroidogenic acute regulatory protein (StAR) and aromatase (CYP19) mRNA in the ovaries of an estradiol valerate (EV)-induced PCOS rat model, and the effect of treadmill and running wheel (voluntary) exercise on these parameters. In this experimental study, we divided adult female Wistar rats that weighed approximately 220 ± 20 g initially into control (n=10) and PCOS (n=30). Subsequently, PCOS group were divided to PCOS, PCOS with treadmill exercise (P-ExT), and PCOS with running wheel exercise (P-ExR) groups (n=10 per group). The expressions of StAR and CYP19 mRNA in the ovaries were determined by quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR). Data were analyzed by one-way ANOVA using SPSS software, version 16. The data were assessed at α=0.05. There was significantly lower mRNA expression of CYP19 in the EV-induced PCOS, running wheel and treadmill exercise rats compared to the control group (PStAR in the ovaries of the PCOS group indicated an increasing trend compared to the control group, however this was not statistically significant (P=0.810). We observed that 8 weeks of running wheel and treadmill exercises could not statistically decrease StAR mRNA expression compared to the PCOS group (P=0.632). EV-induced PCOS in rats decreased CYP19 mRNA expression, but had no effect on StAR mRNA expression. We demonstrated that running wheel and moderate treadmill exercise could not modify CYP19 and StAR mRNA expressions.

  4. MDR1 mRNA EXPRESSION BY REAL TIME - PCR AND RELATIONSHIP WITH CLINICAL RESPONSE BEFORE AND AFTER GIVING NEOADJUVANT CHEMOTHERAPY REGIMEN CYCLOPHOSPHAMIDDOXORUBICIN- 5FU CDF) IN LOCAL ADVANCED BREAST CANCER LABC)

    OpenAIRE

    Rosdiana Natzir, Rosdiana Natzir

    2017-01-01

    Background: Chemoresistanceinvolving P-gp / MDR1 as uncertain factors affect the clinical response after the administration of neoadjuvant chemotherapy NAC) regimen Cyclophosphamid- Doxorubicin-5FU CDF). Response to chemotherapy can be assessed by changes in mRNA expression of MDR1 gene due to DNA damage in the cancer cell DNA synthesis by examination of quantitative Real Time PCR qRT-PCR). Objective: Knowing the direction of change in MDR1 mRNA expression before and after admi...

  5. Synergistic Effect of Caffeine and Glucocorticoids on Expression of Surfactant Protein B (SP-B) mRNA

    Science.gov (United States)

    Fehrholz, Markus; Bersani, Iliana; Kramer, Boris W.; Speer, Christian P.; Kunzmann, Steffen

    2012-01-01

    Administration of glucocorticoids and caffeine is a common therapeutic intervention in the neonatal period, but possible interactions between these substances are still unclear. The present study investigated the effect of caffeine and different glucocorticoids on expression of surfactant protein (SP)-B, crucial for the physiological function of pulmonary surfactant. We measured expression levels of SP-B, various SP-B transcription factors including erythroblastic leukemia viral oncogene homolog 4 (ErbB4) and thyroid transcription factor-1 (TTF-1), as well as the glucocorticoid receptor (GR) after administering different doses of glucocorticoids, caffeine, cAMP, or the phosphodiesterase-4 inhibitor rolipram in the human airway epithelial cell line NCI-H441. Administration of dexamethasone (1 µM) or caffeine (5 mM) stimulated SP-B mRNA expression with a maximal of 38.8±11.1-fold and 5.2±1.4-fold increase, respectively. Synergistic induction was achieved after co-administration of dexamethasone (1 mM) in combination with caffeine (10 mM) (206±59.7-fold increase, pcaffeine with hydrocortisone (87.9±39.0), prednisolone (154±66.8), and betamethasone (123±6.4). Rolipram also induced SP-B mRNA (64.9±21.0-fold increase). We detected a higher expression of ErbB4 and GR mRNA (7.0- and 1.7-fold increase, respectively), whereas TTF-1, Jun B, c-Jun, SP1, SP3, and HNF-3α mRNA expression was predominantly unchanged. In accordance with mRNA data, mature SP-B was induced significantly by dexamethasone with caffeine (13.8±9.0-fold increase, p = 0.0134). We found a synergistic upregulation of SP-B mRNA expression induced by co-administration of various glucocorticoids and caffeine, achieved by accumulation of intracellular cAMP. This effect was mediated by a caffeine-dependent phosphodiesterase inhibition and by upregulation of both ErbB4 and the GR. These results suggested that caffeine is able to induce the expression of SP-transcription factors and affects the

  6. Effects of extremely low frequency electromagnetic field and cisplatin on mRNA levels of some DNA repair genes.

    Science.gov (United States)

    Sanie-Jahromi, Fatemeh; Saadat, Iraj; Saadat, Mostafa

    2016-12-01

    It has been shown that exposure to extremely-low frequency (˂300Hz) oscillating electromagnetic field (EMF) can affect gene expression. The effects of different exposure patterns of 50-Hz EMF and co-treatment of EMF plus cisplatin (CDDP) on mRNA levels of seven genes involved in DNA repair pathways (GADD45A, XRCC1, XRCC4, Ku70, Ku80, DNA-PKcs and LIG4) were evaluated. Two 50-Hz EMF intensities (0.25 and 0.50mT), three exposure patterns (5min field-on/5min field-off, 15min field-on/15min field-off, 30min field-on continuously) and two cell lines (MCF-7 and SH-SY5Y) were used. The mRNA levels were measured using quantitative real-time PCR. The examined genes had tendency to be down-regulated in MCF-7 cells treated with EMF. In the pattern of 15min field-on/15min field-off of the 0.50mT EMF, no increase in mRNA levels were observed, but the mRNA levels of GADD45A, XRCC1, XRCC4, Ku80, Ku70, and LIG4 were down-regulated. A significant elevation in IC50 of CDDP was observed when MCF-7 and SH-SY5Y cells were co-treated with CDDP+EMF in comparison with the cells treated with CDDP alone. GADD45A mRNA levels in MCF-7 and SH-SY5Y cells co-treated with CDDP+EMF were increased and at the same time the mRNA levels of XRCC4, Ku80, Ku70 and DNA-PKcs were down-regulated. Present study provides evidence that co-treatment of CDDP+EMF can enhance down-regulation of the genes involved in non-homologous end-joining pathway. It might be suggested that co-treatment of CDDP+EMF could be more promising for sensitizing cancer cells to DNA double strand breaks. Copyright © 2016 Elsevier Inc. All rights reserved.

  7. Cytokine mRNA expressions after racing at a high altitude and at sea level in horses with exercise-induced pulmonary hemorrhage.

    Science.gov (United States)

    Saulez, Montague N; Godfroid, Jacques; Bosman, Anamarie; Stiltner, Jackie L; Breathnach, Cormac C; Horohov, David W

    2010-04-01

    To determine concentrations of cytokine mRNA in horses with exercise-induced pulmonary hemorrhage (EIPH) after racing. 97 Thoroughbreds. Following tracheobronchoscopy, the severity of EIPH was graded (scale of 0 to 4), and venous blood samples were collected from 10 horses in each grade. After RNA isolation and cDNA synthesis, real-time PCR assay was conducted to detect cytokinespecific mRNA for interleukin (IL)-1, IL-6, and IL-10; interferon (INF)-gamma; and tumor necrosis factor (TNF)-alpha. Neither location nor grade of EIPH affected the expression of IL-1 and INF-gamma. There was significantly greater overall expression of IL-6 mRNA at sea level, with significantly more IL-6 expressed in horses with grade 4 EIPH than in horses with grade 0, 1, or 2 EIPH. At a high altitude, no difference was detected for IL-6 expression among the various EIPH grades. There was significantly greater overall expression of TNF-alpha mRNA at a high altitude; however, there was no difference within the various grades of EIPH. Expression of IL-10 was significantly affected by grade of EIPH because horses with grade 3 EIPH expressed significantly more IL-10 mRNA than did horses with grade 0 or 2 EIPH; this expression was not affected by location. At sea level, increased IL-6 expression was associated with more severe EIPH, and altitude may affect gene expressions of the proinflammatory cytokine TNF-alpha and anti-inflammatory cytokine IL-6. Studies on protein concentrations of cytokine expression are needed. The pathophysiologic importance of these findings remains to be explained.

  8. Ccr4-not complex mRNA deadenylase activity contributes to DNA damage responses in Saccharomyces cerevisiae.

    Science.gov (United States)

    Traven, Ana; Hammet, Andrew; Tenis, Nora; Denis, Clyde L; Heierhorst, Jörg

    2005-01-01

    DNA damage checkpoints regulate gene expression at the transcriptional and post-transcriptional level. Some components of the yeast Ccr4-Not complex, which regulates transcription as well as transcript turnover, have previously been linked to DNA damage responses, but it is unclear if this involves transcriptional or post-transcriptional functions. Here we show that CCR4 and CAF1, which together encode the major cytoplasmic mRNA deadenylase complex, have complex genetic interactions with the checkpoint genes DUN1, MRC1, RAD9, and RAD17 in response to DNA-damaging agents hydroxyurea (HU) and methylmethane sulfonate (MMS). The exonuclease-inactivating ccr4-1 point mutation mimics ccr4Delta phenotypes, including synthetic HU hypersensitivity with dun1Delta, demonstrating that Ccr4-Not mRNA deadenylase activity is required for DNA damage responses. However, ccr4Delta and caf1Delta DNA damage phenotypes and genetic interactions with checkpoint genes are not identical, and deletions of some Not components that are believed to predominantly function at the transcriptional level rather than mRNA turnover, e.g., not5Delta, also lead to increased DNA damage sensitivity and synthetic HU hypersensitivity with dun1Delta. Taken together, our data thus suggest that both transcriptional and post-transcriptional functions of the Ccr4-Not complex contribute to the DNA damage response affecting gene expression in a complex manner.

  9. The Impact of Ramadan Fasting on SIRT1 mRNA Expression in Peripheral Blood Mononuclear Cells

    Directory of Open Access Journals (Sweden)

    Mostafa Haji Molahoseini

    2016-11-01

    Full Text Available Background:The aim of this study was to evaluate the effect of Ramadan fasting on SIRT1 mRNA expression in healthy men.Islamic Ramadan fasting is a holy religious ceremony that has many spiritual benefits. Additionally, it can be considered as the equivalent of calorie restriction that may affect physical health. The results of previous studies revealed that calorie restriction increases the lifespan in laboratory rodents via increasing the expression of a histone deacetylase named SIRT1. Additionally, SIRT1 is known for its anti-inflammatory properties. Materials and Methods: Overall, 43 men volunteered for participating in this one-group before and after (self-controlled study. Two mL blood samples were taken prior to fasting and at the end of the 30th day of fasting. Routine biochemical tests and SIRT1 mRNA expression analysis were performed. Results: Cholesterol and low-density lipoproteins increase, however, high-density lipoproteins level decreased after Ramadan fasting. The analysis of real-time PCR results revealed that SIRT1 mRNA expression in human peripheral blood mononuclear cells increased 4.63 fold in fasting state in comparison with non-fasting state. Conclusion: Ramadan fasting has a significant effect on SIRT1 gene expression. Considering the immunosuppressive and anti-inflammatory properties of SIRT1, further studies are needed to evaluate the effects of SIRT1 up-regulation on the autoimmune and inflammatory diseases during Ramadan fasting.

  10. Increased mRNA Levels of TCF7L2 and MYC of the Wnt Pathway in Tg-ArcSwe Mice and Alzheimer's Disease Brain

    Directory of Open Access Journals (Sweden)

    Elin S. Blom

    2011-01-01

    Full Text Available Several components in the Wnt pathway, including β-catenin and glycogen synthase kinase 3 beta, have been implied in AD pathogenesis. Here, mRNA brain levels from five-month-old tg-ArcSwe and nontransgenic mice were compared using Affymetrix microarray analysis. With surprisingly small overall changes, Wnt signaling was the most affected pathway with altered expression of nine genes in tg-ArcSwe mice. When analyzing mRNA levels of these genes in human brain, transcription factor 7-like 2 (TCF7L2 and v-myc myelocytomatosis viral oncogene homolog (MYC, were increased in Alzheimer's disease (AD (P<.05. Furthermore, no clear differences in TCF7L2 and MYC mRNA were found in brains with frontotemporal lobar degeneration, suggesting that altered regulation of these Wnt-related genes could be specific to AD. Finally, mRNA levels of three neurogenesis markers were analyzed. Increased mRNA levels of dihydropyrimidinase-like 3 were observed in AD brain, suggesting that altered Wnt pathway regulation may signify synaptic rearrangement or neurogenesis.

  11. Transcriptome kinetics is governed by a genome-wide coupling of mRNA production and degradation: a role for RNA Pol II.

    Directory of Open Access Journals (Sweden)

    Ophir Shalem

    2011-09-01

    Full Text Available Transcriptome dynamics is governed by two opposing processes, mRNA production and degradation. Recent studies found that changes in these processes are frequently coordinated and that the relationship between them shapes transcriptome kinetics. Specifically, when transcription changes are counter-acted with changes in mRNA stability, transient fast-relaxing transcriptome kinetics is observed. A possible molecular mechanism underlying such coordinated regulation might lay in two RNA polymerase (Pol II subunits, Rpb4 and Rpb7, which are recruited to mRNAs during transcription and later affect their degradation in the cytoplasm. Here we used a yeast strain carrying a mutant Pol II which poorly recruits these subunits. We show that this mutant strain is impaired in its ability to modulate mRNA stability in response to stress. The normal negative coordinated regulation is lost in the mutant, resulting in abnormal transcriptome profiles both with respect to magnitude and kinetics of responses. These results reveal an important role for Pol II, in regulation of both mRNA synthesis and degradation, and also in coordinating between them. We propose a simple model for production-degradation coupling that accounts for our observations. The model shows how a simple manipulation of the rates of co-transcriptional mRNA imprinting by Pol II may govern genome-wide transcriptome kinetics in response to environmental changes.

  12. Effects of the pesticides prochloraz and methiocarb on human estrogen receptor alpha and beta mRNA levels analyzed by on-line RT-PCR.

    Science.gov (United States)

    Hofmeister, M V; Bonefeld-Jørgensen, E C

    2004-08-01

    Exposure to endocrine disrupters such as dioxins, PCBs and certain pesticides are suspected to affect human reproductive health. We have analyzed the effect of the currently used pesticides prochloraz and methiocarb on the estrogen receptor (ER)alpha and beta mRNA levels in parallel with the natural ligand, 17beta-estradiol (E2). Using the highly sensitive on-line RT-PCR technique we were able to quantify the ERalpha and ERbeta mRNA levels in the human breast cancer cell line, MCF7-BUS. Upon exposure with E2 or prochloraz a down regulation of ERalpha and ERbeta mRNAs was observed after 48 h of treatment. Co-treatment with the ER antagonist ICI 182,780 abolished these mRNA down regulations. Western blot analyses elicited a decreased ER protein level after 3 h of exposure with prochloraz but after 24 h the ERalpha protein level had recovered to basal level. Methiocarb exposure had no effect on the ERalpha mRNA level, whereas an increase in the ERbeta mRNA level was observed after 3 h of exposure. Our study demonstrates that like E2, prochloraz had the potential to down regulate the expression of ERalpha and ERbeta mRNAs as well as the ERalpha protein level in MCF7-BUS cells.

  13. Identification of an RNase J ortholog in Sulfolobus solfataricus: implications for 5'-to-3' directional decay and 5'-end protection of mRNA in Crenarchaeota.

    Science.gov (United States)

    Hasenöhrl, David; Konrat, Robert; Bläsi, Udo

    2011-01-01

    In both Bacteria and Eukaryotes, degradation is known to start at the 5' and at the 3' extremities of mRNAs. Until the recent discovery of 5'-to-3' exoribonucleases in hyperthermophilic Euryarchaeota, the exosome was assumed to be the key enzyme in mRNA degradation in Archaea. By means of zymogram assays and bioinformatics, we have identified a 5'-to-3' exoribonuclea