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系統識別號 U0026-2801201412595700
論文名稱(中文) 研究hRAD9蛋白於癌症中扮演之功能性角色
論文名稱(英文) Study on the Functional Role of hRAD9 in Cancer
校院名稱 成功大學
系所名稱(中) 基礎醫學研究所
系所名稱(英) Institute of Basic Medical Sciences
學年度 102
學期 1
出版年 103
研究生(中文) 溫凡志
研究生(英文) Fan-Chih Wen
學號 s58961604
學位類別 博士
語文別 英文
論文頁數 118頁
口試委員 指導教授-張敏政
召集委員-賴明德
口試委員-張文粲
口試委員-鄭宏祺
口試委員-戴明泓
口試委員-洪文俊
中文關鍵字 hRAD9  細胞老化  上皮間質轉化  Slug  磷酸化  缺氧 
英文關鍵字 hRAD9  senescence  EMT  Slug  phosphorylation  hypoxia 
學科別分類
中文摘要 細胞老化與上皮間質轉化在癌症進程中扮演著相反的角色,前者為防止腫瘤生成的障壁,後者為增強癌細胞侵犯性與惡性化的作用。hRAD9為一DNA損傷反應蛋白參與在DNA修復、細胞週期檢查點調控、細胞凋亡、基因調控與維持基因體穩定等功能中,而hRAD9在癌症進程中扮演的角色卻是有爭議性的。在本研究中,我們在乳癌、肺癌與大腸癌檢體中進行了hRAD9的臨床分析以及對hRAD9超表現與抑制細胞中進行了功能性分析,結果說明hRAD9在癌細胞中具有促進細胞老化與抑制上皮間質轉化等兩項功能。首先,以西方墨點法分析乳癌與肺癌病人檢體的結果中顯示,在同一病人中,相較其正常組織而言,hRAD9的蛋白表現量於腫瘤組織中明顯降低;而相較於癌症前期病人而言,hRAD9的相對蛋白表現量在癌症後期病人中亦明顯降低,而低hRAD9的蛋白表現量與細胞侵犯能力呈負相關。將hRAD9超表現於具高侵犯性且低hRAD9蛋白表現量的肺癌細胞H1299與乳癌細胞MDA-MB 231時,會增強p21的蛋白表現量且此增強的p21會累積於細胞核內,進而促使癌細胞產生細胞老化,此外,超表現hRAD9同時產生抑制癌細胞的爬行、侵犯與小鼠皮下腫瘤的生長能力。相反地,在具低侵犯性且高hRAD9蛋白表現量的A549與MCF7中以shRNA抑制hRAD9後,則大幅提升癌細胞之爬行與侵犯的能力,並導致癌細胞產生上皮間質轉化的發生。利用超表現hRAD9或抑制hRAD9後發現,hRAD9的超表現可抑制上皮間質轉化關鍵因子Slug的蛋白表現量,相反地,hRAD9表現抑制後則引起Slug的蛋白表現量上升,進一步的實驗結果顯示,hRAD9能夠直接結合在Slug啟動子上並抑制其轉錄活性。總結以上所說,這些實驗結果說明hRAD9是一個在乳癌與肺癌中有潛力的腫瘤抑制蛋白,其具有透過轉錄活化p21的表現促使癌細胞產生細胞老化,並透過轉錄抑制Slug的表現抑制癌細胞產生上皮間質轉化的功能。除此之外,在大腸癌中,相較於同一病人的正常組織來說,高表現的高度磷酸化hRAD9出現在57.1%的病人癌症檢體中,並且此高度磷酸化的現象會伴隨著兩個hRAD9的蛋白質激酶CK2A與CDK1的高度表現。我們證實CK2A與CDK1能夠和hRAD9產生交互作用並磷酸化hRAD9。而高度磷酸化的hRAD9會失去其抑制細胞生長與促進細胞老化的能力。透過抑制CK2A與CDK1的表現後可降低hRAD9的磷酸化情形,並強化其對染色質的結合能力及增強其下游標的基因p21的活化而促使細胞老化發生。這些實驗結果說明,CK2A與CDK1對hRAD9的磷酸化作用可能是癌細胞抑制hRAD9抑癌作用的可能機制。最後,我們在研究對於hRAD9的轉錄調控研究中發現,hRAD9的表現量在缺氧環境中會顯著性降低,且此降低來自於缺氧誘導因子HIF-1α的轉錄抑制。總結以上所述,我們的結果顯示hRAD9是一個具有促進細胞老化與抑制上皮間質轉化的腫瘤抑制蛋白,且磷酸化修飾作用對於hRAD9促進細胞老化的能力具有負面的影響。對於hRAD9在癌症進程中表現量降低的可能調控機制是來自於HIF-1α在癌症缺氧中的抑制作用。透過研究hRAD9的功能性角色、磷酸化修飾作用於轉錄調控機制,我們最終希望能夠藉此評估將hRAD9作為癌症治療標的之可能性。
英文摘要 Senescence and epithelial-mesenchymal transition (EMT) have opposing roles in tumor progression in that one is a barrier against tumorigenesis, while the other is required for invasive malignancies. hRAD9 is a DNA damage response (DDR) protein, participating in DNA repair, cell-cycle checkpoint regulation, apoptosis, gene transcription and maintenance of genomic stability, but the role of hRAD9 is controversial in tumor progression. In this study, we performed clinical investigations of hRAD9 in breast, lung, and colon cancers and functional analyses of stable cells with hRad9 overexpression or knockdown, and found that hRAD9 contributes to induction of senescence and repression of EMT. Our results showed that hRAD9 was downregulated in most of breast and lung cancer tumor tissues. The decreased hRAD9 expression was associated with tumor stage in breast and lung cancers, as well as with the invasive phenotype. By overexpressing hRAD9 in highly invasive cancer cell lines, H1299 and MDA-MB 231, with low endogenous hRAD9 induced senescence by upregulation of nuclear p21, independent of the p53 status. In addition, overexpression of hRAD9 attenuated cellular migration and invasion in vitro and tumor growth in a xenograft mouse model in vivo. In contrast, silencing hRAD9 in lower invasive cancer cell lines, A549 and MCF7, with high endogenous hRAD9 significantly enhanced their migration and invasion abilities, and simultaneously activated EMT. Knockdown of hRAD9 increased, whereas overexpression of hRAD9 decreased, the expression of an EMT inducer, Slug. Moreover, we demonstrated that hRAD9 directly bound to the promoter region of slug gene and repressed its expression in transcription. Altogether, these results suggest that hRAD9 is a potential tumor suppressor in breast and lung cancers and that it is likely to function by upregulating p21 and inhibiting Slug to regulate tumorigenesis. Besides, in colon cancer specimens, hyper-phosphorylated hRAD9 was seen in 57.1% of cancerous tissues and high levels of hyper-phosphorylated hRAD9 were associated with high expression levels of two protein kinases of hRAD9, CK2A and CDK1. CK2A and CDK1 interacted with and phosphorylated hRAD9 in vivo. The hyper-phosphorylation of hRAD9 abolished its ability of growth inhibition and senescence induction. Silencing CK2A and CDK1 resulted in decreasing phosphorylation of hRAD9, enhancing its binding capacity to chromatin and thereby activated its target, p21, to trigger senescence. These results provide evidence for a mechanism suggesting the CK2A and CDK1 phosphorylation of hRAD9 attenuates its tumor suppression capacity of inducing senescence. Finally, our investigation of transcriptional regulation of hRAD9 in cancer showed hRAD9 was markedly downregulated in hypoxic condition through the hypoxia inducible factor-1 alpha (HIF-1α)-dependent repression. Taken together, our data indicates hRAD9 is a tumor suppressor with potentials to trigger senescence and inhibit EMT, and phosphorylation modification of hRAD9 confers a negative impact on its ability of senescence induction. A possible regulatory mechanism that leads to the downregulation of hRAD9 in cancer cells may be caused by HIF-1α in tumor hypoxia. By exploring the functional role, modification, and transcriptional regulation of hRAD9, we eventually hope to evaluate the potency of hRAD9 as a therapeutic target for cancer treatment.
論文目次 中文摘要 I
Abstract III
Acknowledgement V
Contents VII
Table contents XII
Figure contents XIII
List of abbreviations XVI
Chapter 1 Introduction 1
1-1 Premature senescence and tumor suppression 1
1-2 Epithelial-mesenchymal transition and tumor progression 2
1-3 Crossed path: senescence and EMT 3
1-4 Rad9: from yeast to human 4
1-5 Functional domains and phosphorylation sites of hRAD9 5
1-6 Two-faced hRAD9: oncoprotein or tumor suppressor 6
1-7 Specific aim 8
Chapter 2 Materials and methods 9
2-1 Bacteria strains, expression vectors and growth medium 9
2-1-1 Bacteria strains 9
2-1-2 Expression vector (map of lentiviral plasmids shown in Appendix 5) 9
2-1-3 Antibiotic and growth medium preparation 10
2-2 Gene cloning 11
2-2-1 Plasmid extraction 11
2-2-2 Polymerase chain reaction (PCR) 12
2-2-3 Restriction enzyme digestion 12
2-2-4 DNA purification 13
2-2-5 Ligation 13
2-2-6 Competent cells preparation and transformation 14
2-3 Cell cultivation techniques 15
2-3-1 Cell lines properties and culture conditions 15
2-3-2 Unfreezing cells 16
2-3-3 Subculture 16
2-3-4 Storage of cells 17
2-3-5 Cell counts 17
2-4 Transfection and lentivirus infection 18
2-4-1 Transient transfection (plasmid DNA) 18
2-4-2 Transient transfection (siRNA) 18
2-4-3 Stably transfection 19
2-4-4 Lentivirus preparation 20
2-4-5 Stably lentiviurs infection 21
2-5 Protein analysis 22
2-5-1 Protein extraction 22
2-5-2 Protein quantification 23
2-5-3 Western blot analysis 23
2-5-4 Immunofluorescence staining 27
2-6 Functional analysis of cell behavior 28
2-6-1 Cell proliferation 28
2-6-1-1 Cell counts 28
2-6-1-2 Anchorage dependent colony formation assay 29
2-6-1-3 Ki67 staining 29
2-6-2 Cell migration, invasion, and adhesion analysis 30
2-6-2-1 Transwell assay (migration and invasion assay) 30
2-5-2-2 Adhesion assay 32
2-6-3 Senescence analysis 32
2-6-3-1 Senescence associated β-Galactosidase staining 32
2-6-3-2 F-Actin staining 33
2-7 Animal model and IHC staining 34
2-8 Promoter assay 35
2-9 ChIP assay 36
2-10 Patients and tissue samples 38
2-11 Statistical analysis 38
Chapter 3 Results 40
3-1 Part. Ⅰ The functional role of hRAD9 in tumor progression 40
3-1-1 hRAD9 expression is significantly downregulated in breast and lung cancer tissues and cell lines 40
3-1-2 Ectopic hRAD9 expression induces senescence in highly invasive cancer cells 41
3-1-3 Ectopic hRAD9 expression induces senescence by nuclear upregulation of p21, independent of the p53 status 41
3-1-4 hRAD9 suppresses tumorigenicity of invasive cancer cells in vitro and in vivo 42
3-1-5 Silencing hRAD9 in cancer cells with lower invasive capacity activates EMT and increases migratory and invasive potential 43
3-1-6 hRAD9 inhibits EMT through suppression of Slug 44
3-1-7 hRAD9 directly binds the slug promoter and represses its transcriptional activity 45
3-2 Part. Ⅱ Phosphorylation of hRAD9 modulates its function of inducing senescence 46
3-2-1 Levels of hyper-phosphorylated hRAD9 are significantly elevated in colon cancer tissues 46
3-2-2 CK2A and CDK1 interacts with and phosphorylates hRAD9 in vivo 47
3-2-3 Hyper-phosphorylated hRAD9 is unable to trigger senescence in cancer cells 48
3-2-4 hRAD9 mutants with deficiency of CK2A- or CDK1-dependent phosphorylation can trigger senescence 49
3-2-5 Decreased phosphorylation of hRAD9 facilitates senescence induction and increases its binding capacity to chromatin 50
3-3 Part. Ⅲ Transcriptional regulation of hRAD9 in hypoxia 51
3-3-1 HIF-1α-mediated downregulation of hRAD9 in hypoxia 51
3-3-2 HIF-1α downregulates hRAD9 by directly transcriptional repression 52
3-3-3 Re-expression of hRAD9 inhibits Slug expression and upregulates E-cadherin in the presence of hypoxia-mimetic agent 53
Chapter 4 Conclusion 54
Chapter 5 Discussion 55
Figures and tables 61
References 103
Appendix 111
Appendix 1 111
Appendix 2 112
Appendix 3 113
Appendix 4 114
Appendix 5 115
參考文獻 1. Adams, P.D. Healing and hurting: molecular mechanisms, functions, and pathologies of cellular senescence. Mol Cell 36, 2-14 (2009).
2. Hayflick, L. & Moorhead, P.S. The serial cultivation of human diploid cell strains. Exp Cell Res 25, 585-621 (1961).
3. Ohtani, N., Mann, D.J. & Hara, E. Cellular senescence: its role in tumor suppression and aging. Cancer Sci 100, 792-797 (2009).
4. Di Micco, R., et al. Oncogene-induced senescence is a DNA damage response triggered by DNA hyper-replication. Nature 444, 638-642 (2006).
5. Aliouat-Denis, C.M., et al. p53-independent regulation of p21Waf1/Cip1 expression and senescence by Chk2. Mol Cancer Res 3, 627-634 (2005).
6. Chen, C.R., et al. Dual induction of apoptosis and senescence in cancer cells by Chk2 activation: checkpoint activation as a strategy against cancer. Cancer Res 65, 6017-6021 (2005).
7. Nardella, C., Clohessy, J.G., Alimonti, A. & Pandolfi, P.P. Pro-senescence therapy for cancer treatment. Nat Rev Cancer 11, 503-511 (2011).
8. Kagawa, S., et al. Overexpression of the p21 sdi1 gene induces senescence-like state in human cancer cells: implication for senescence-directed molecular therapy for cancer. Cell Death Differ 6, 765-772 (1999).
9. Chen, Q.M., et al. Involvement of Rb family proteins, focal adhesion proteins and protein synthesis in senescent morphogenesis induced by hydrogen peroxide. Journal of cell science 113 ( Pt 22), 4087-4097 (2000).
10. Campisi, J. & d'Adda di Fagagna, F. Cellular senescence: when bad things happen to good cells. Nature reviews. Molecular cell biology 8, 729-740 (2007).
11. Romagosa, C., et al. p16(Ink4a) overexpression in cancer: a tumor suppressor gene associated with senescence and high-grade tumors. Oncogene 30, 2087-2097 (2011).
12. Meek, D.W. Tumour suppression by p53: a role for the DNA damage response? Nat Rev Cancer 9, 714-723 (2009).
13. Ferbeyre, G., et al. Oncogenic ras and p53 cooperate to induce cellular senescence. Mol Cell Biol 22, 3497-3508 (2002).
14. Wang, Y., Blandino, G. & Givol, D. Induced p21waf expression in H1299 cell line promotes cell senescence and protects against cytotoxic effect of radiation and doxorubicin. Oncogene 18, 2643-2649 (1999).
15. Chen, X., Zhang, W., Gao, Y.F., Su, X.Q. & Zhai, Z.H. Senescence-like changes induced by expression of p21(waf1/Cip1) in NIH3T3 cell line. Cell Res 12, 229-233 (2002).
16. Campisi, J. Senescent cells, tumor suppression, and organismal aging: good citizens, bad neighbors. Cell 120, 513-522 (2005).
17. Burgess, D.J. Senescence: Tumorigenesis under surveillance. Nat Rev Cancer (2011).
18. Collado, M., et al. Tumour biology: senescence in premalignant tumours. Nature 436, 642 (2005).
19. Collado, M. & Serrano, M. Senescence in tumours: evidence from mice and humans. Nat Rev Cancer 10, 51-57 (2010).
20. d'Adda di Fagagna, F. Living on a break: cellular senescence as a DNA-damage response. Nature reviews. Cancer 8, 512-522 (2008).
21. Lleonart, M.E., Artero-Castro, A. & Kondoh, H. Senescence induction; a possible cancer therapy. Mol Cancer 8, 3 (2009).
22. Kalluri, R. EMT: when epithelial cells decide to become mesenchymal-like cells. The Journal of clinical investigation 119, 1417-1419 (2009).
23. Floor, S., van Staveren, W.C., Larsimont, D., Dumont, J.E. & Maenhaut, C. Cancer cells in epithelial-to-mesenchymal transition and tumor-propagating-cancer stem cells: distinct, overlapping or same populations. Oncogene 30, 4609-4621 (2011).
24. Sanchez-Tillo, E., et al. ZEB1 represses E-cadherin and induces an EMT by recruiting the SWI/SNF chromatin-remodeling protein BRG1. Oncogene 29, 3490-3500 (2010).
25. Martin, A. & Cano, A. Tumorigenesis: Twist1 links EMT to self-renewal. Nature cell biology 12, 924-925 (2010).
26. Berx, G. & van Roy, F. Involvement of members of the cadherin superfamily in cancer. Cold Spring Harbor perspectives in biology 1, a003129 (2009).
27. Peinado, H., Olmeda, D. & Cano, A. Snail, Zeb and bHLH factors in tumour progression: an alliance against the epithelial phenotype? Nat Rev Cancer 7, 415-428 (2007).
28. Vuoriluoto, K., et al. Vimentin regulates EMT induction by Slug and oncogenic H-Ras and migration by governing Axl expression in breast cancer. Oncogene 30, 1436-1448 (2011).
29. Zeisberg, M. & Neilson, E.G. Biomarkers for epithelial-mesenchymal transitions. J Clin Invest 119, 1429-1437 (2009).
30. Singh, A. & Settleman, J. EMT, cancer stem cells and drug resistance: an emerging axis of evil in the war on cancer. Oncogene 29, 4741-4751 (2010).
31. Bartkova, J., et al. Oncogene-induced senescence is part of the tumorigenesis barrier imposed by DNA damage checkpoints. Nature 444, 633-637 (2006).
32. Mooi, W.J. & Peeper, D.S. Oncogene-induced cell senescence--halting on the road to cancer. N Engl J Med 355, 1037-1046 (2006).
33. Smit, M.A. & Peeper, D.S. Epithelial-mesenchymal transition and senescence: two cancer-related processes are crossing paths. Aging 2, 735-741 (2010).
34. Chang, C.J., et al. p53 regulates epithelial-mesenchymal transition and stem cell properties through modulating miRNAs. Nat Cell Biol 13, 1467 (2011).
35. Liu, M., et al. p21CIP1 attenuates Ras- and c-Myc-dependent breast tumor epithelial mesenchymal transition and cancer stem cell-like gene expression in vivo. Proc Natl Acad Sci U S A 106, 19035-19039 (2009).
36. Arima, Y., et al. Induction of ZEB proteins by inactivation of RB protein is key determinant of mesenchymal phenotype of breast cancer. The Journal of biological chemistry 287, 7896-7906 (2012).
37. Emadi Baygi, M., Soheili, Z.S., Schmitz, I., Sameie, S. & Schulz, W.A. Snail regulates cell survival and inhibits cellular senescence in human metastatic prostate cancer cell lines. Cell Biol Toxicol 26, 553-567 (2010).
38. Liu, Y., El-Naggar, S., Darling, D.S., Higashi, Y. & Dean, D.C. Zeb1 links epithelial-mesenchymal transition and cellular senescence. Development 135, 579-588 (2008).
39. Ansieau, S., et al. Induction of EMT by twist proteins as a collateral effect of tumor-promoting inactivation of premature senescence. Cancer Cell 14, 79-89 (2008).
40. Tsai, C.C., et al. Hypoxia inhibits senescence and maintains mesenchymal stem cell properties through down-regulation of E2A-p21 by HIF-TWIST. Blood 117, 459-469 (2011).
41. Prakash, L. Repair of pyrimidine dimers in radiation-sensitive mutants rad3, rad4, rad6 and rad9 of Saccharomyces cerevisiae. Mutat Res 45, 13-20 (1977).
42. Smolinska, U. Mitochondrial mutagenesis in yeast: mutagenic specificity of EMS and the effects of RAD9 and REV3 gene products. Mutat Res 179, 167-174 (1987).
43. Weinert, T.A. & Hartwell, L.H. The RAD9 gene controls the cell cycle response to DNA damage in Saccharomyces cerevisiae. Science 241, 317-322 (1988).
44. Schiestl, R.H., Reynolds, P., Prakash, S. & Prakash, L. Cloning and sequence analysis of the Saccharomyces cerevisiae RAD9 gene and further evidence that its product is required for cell cycle arrest induced by DNA damage. Mol Cell Biol 9, 1882-1896 (1989).
45. Weinert, T. & Hartwell, L. Control of G2 delay by the rad9 gene of Saccharomyces cerevisiae. J Cell Sci Suppl 12, 145-148 (1989).
46. Weinert, T.A. & Hartwell, L.H. Characterization of RAD9 of Saccharomyces cerevisiae and evidence that its function acts posttranslationally in cell cycle arrest after DNA damage. Mol Cell Biol 10, 6554-6564 (1990).
47. Lieberman, H.B., Hopkins, K.M., Laverty, M. & Chu, H.M. Molecular cloning and analysis of Schizosaccharomyces pombe rad9, a gene involved in DNA repair and mutagenesis. Mol Gen Genet 232, 367-376 (1992).
48. Murray, J.M., Carr, A.M., Lehmann, A.R. & Watts, F.Z. Cloning and characterisation of the rad9 DNA repair gene from Schizosaccharomyces pombe. Nucleic Acids Res 19, 3525-3531 (1991).
49. Lieberman, H.B. & Hopkins, K.M. Schizosaccharomyces malidevorans and Sz. octosporus homologues of Sz. pombe rad9, a gene that mediates radioresistance and cell-cycle progression. Gene 150, 281-286 (1994).
50. Lieberman, H.B., Hopkins, K.M., Nass, M., Demetrick, D. & Davey, S. A human homolog of the Schizosaccharomyces pombe rad9+ checkpoint control gene. Proceedings of the National Academy of Sciences of the United States of America 93, 13890-13895 (1996).
51. Bessho, T. & Sancar, A. Human DNA damage checkpoint protein hRAD9 is a 3' to 5' exonuclease. J Biol Chem 275, 7451-7454 (2000).
52. St Onge, R.P., Udell, C.M., Casselman, R. & Davey, S. The human G2 checkpoint control protein hRAD9 is a nuclear phosphoprotein that forms complexes with hRAD1 and hHUS1. Mol Biol Cell 10, 1985-1995 (1999).
53. Bao, S., et al. Disruption of the Rad9/Rad1/Hus1 (9-1-1) complex leads to checkpoint signaling and replication defects. Oncogene 23, 5586-5593 (2004).
54. Pandita, R.K., et al. Mammalian Rad9 plays a role in telomere stability, S- and G2-phase-specific cell survival, and homologous recombinational repair. Mol Cell Biol 26, 1850-1864 (2006).
55. Lieberman, H.B. & Yin, Y. A novel function for human Rad9 protein as a transcriptional activator of gene expression. Cell Cycle 3, 1008-1010 (2004).
56. Komatsu, K., et al. Human homologue of S. pombe Rad9 interacts with BCL-2/BCL-xL and promotes apoptosis. Nat Cell Biol 2, 1-6 (2000).
57. Wang, L., et al. Human checkpoint protein hRad9 functions as a negative coregulator to repress androgen receptor transactivation in prostate cancer cells. Mol Cell Biol 24, 2202-2213 (2004).
58. Yin, Y., et al. Human RAD9 checkpoint control/proapoptotic protein can activate transcription of p21. Proceedings of the National Academy of Sciences of the United States of America 101, 8864-8869 (2004).
59. Ishikawa, K., Ishii, H., Saito, T. & Ichimura, K. Multiple functions of rad9 for preserving genomic integrity. Curr Genomics 7, 477-480 (2006).
60. Lee, M.W., Hirai, I. & Wang, H.G. Caspase-3-mediated cleavage of Rad9 during apoptosis. Oncogene 22, 6340-6346 (2003).
61. Ishikawa, K., et al. Rad9 modulates the P21WAF1 pathway by direct association with p53. BMC Mol Biol 8, 37 (2007).
62. Hopkins, K.M., et al. Deletion of mouse rad9 causes abnormal cellular responses to DNA damage, genomic instability, and embryonic lethality. Mol Cell Biol 24, 7235-7248 (2004).
63. Komatsu, K., Hopkins, K.M., Lieberman, H.B. & Wang, H. Schizosaccharomyces pombe Rad9 contains a BH3-like region and interacts with the anti-apoptotic protein Bcl-2. FEBS Lett 481, 122-126 (2000).
64. Broustas, C.G. & Lieberman, H.B. Contributions of RAD9 to tumorigenesis. Journal of cellular biochemistry (2011).
65. Hirai, I. & Wang, H.G. A role of the C-terminal region of human Rad9 (hRad9) in nuclear transport of the hRad9 checkpoint complex. J Biol Chem 277, 25722-25727 (2002).
66. Roos-Mattjus, P., et al. Phosphorylation of human Rad9 is required for genotoxin-activated checkpoint signaling. J Biol Chem 278, 24428-24437 (2003).
67. St Onge, R.P., Besley, B.D., Pelley, J.L. & Davey, S. A role for the phosphorylation of hRad9 in checkpoint signaling. J Biol Chem 278, 26620-26628 (2003).
68. Chen, M.J., Lin, Y.T., Lieberman, H.B., Chen, G. & Lee, E.Y. ATM-dependent phosphorylation of human Rad9 is required for ionizing radiation-induced checkpoint activation. The Journal of biological chemistry 276, 16580-16586 (2001).
69. Yoshida, K., Wang, H.G., Miki, Y. & Kufe, D. Protein kinase Cdelta is responsible for constitutive and DNA damage-induced phosphorylation of Rad9. The EMBO journal 22, 1431-1441 (2003).
70. Zhan, Z., et al. Phosphorylation of Rad9 at serine 328 by cyclin A-Cdk2 triggers apoptosis via interfering Bcl-xL. PLoS One 7, e44923 (2012).
71. Takeishi, Y., et al. Casein kinase 2-dependent phosphorylation of human Rad9 mediates the interaction between human Rad9-Hus1-Rad1 complex and TopBP1. Genes to cells : devoted to molecular & cellular mechanisms 15, 761-771 (2010).
72. Chan, V., et al. Localization of hRad9 in breast cancer. BMC Cancer 8, 196 (2008).
73. Maniwa, Y., et al. Accumulation of hRad9 protein in the nuclei of nonsmall cell lung carcinoma cells. Cancer 103, 126-132 (2005).
74. Cheng, C.K., Chow, L.W., Loo, W.T., Chan, T.K. & Chan, V. The cell cycle checkpoint gene Rad9 is a novel oncogene activated by 11q13 amplification and DNA methylation in breast cancer. Cancer Res 65, 8646-8654 (2005).
75. Zhu, A., Zhang, C.X. & Lieberman, H.B. Rad9 has a functional role in human prostate carcinogenesis. Cancer Res 68, 1267-1274 (2008).
76. Kebebew, E., et al. Diagnostic and prognostic value of cell-cycle regulatory genes in malignant thyroid neoplasms. World J Surg 30, 767-774 (2006).
77. Hopkins, K.M., et al. Expression of mammalian paralogues of HRAD9 and Mrad9 checkpoint control genes in normal and cancerous testicular tissue. Cancer Res 63, 5291-5298 (2003).
78. Hayashi, K., et al. Induction of hRAD9 is required for G2/M checkpoint signal transduction in gastric cancer cells. Pathobiology 70, 40-46 (2002).
79. Broustas, C.G., Zhu, A. & Lieberman, H.B. Rad9 protein contributes to prostate tumor progression by promoting cell migration and anoikis resistance. J Biol Chem 287, 41324-41333 (2012).
80. Lee, E., et al. Inhibition of androgen receptor and beta-catenin activity in prostate cancer. Proceedings of the National Academy of Sciences of the United States of America 110, 15710-15715 (2013).
81. Hu, Z., et al. Targeted deletion of Rad9 in mouse skin keratinocytes enhances genotoxin-induced tumor development. Cancer Res 68, 5552-5561 (2008).
82. Gyorffy, B., Lanczky, A. & Szallasi, Z. Implementing an online tool for genome-wide validation of survival-associated biomarkers in ovarian-cancer using microarray data from 1287 patients. Endocr Relat Cancer 19, 197-208 (2012).
83. Yuki, T., et al. DNA damage sensor protein hRad9, a novel molecular target for lung cancer treatment. Oncol Rep 20, 1047-1052 (2008).
84. Lambertini, E., et al. SLUG: a new target of lymphoid enhancer factor-1 in human osteoblasts. BMC Mol Biol 11, 13 (2010).
85. Wen, F.C., Chang, T.W., Tseng, Y.L., Lee, J.C. & Chang, M.C. hRAD9 functions as a tumor suppressor by inducing p21-dependent senescence and suppressing epithelial-mesenchymal transition through inhibition of Slug transcription. Carcinogenesis (2014).
86. Zou, J., et al. Protein kinase CK2alpha is overexpressed in colorectal cancer and modulates cell proliferation and invasion via regulating EMT-related genes. Journal of translational medicine 9, 97 (2011).
87. Roberson, R.S., Kussick, S.J., Vallieres, E., Chen, S.Y. & Wu, D.Y. Escape from therapy-induced accelerated cellular senescence in p53-null lung cancer cells and in human lung cancers. Cancer research 65, 2795-2803 (2005).
88. Richter, K.H., et al. Down-regulation of cdc2 in senescent human and hamster cells. Cancer research 51, 6010-6013 (1991).
89. Elmore, L.W., Di, X., Dumur, C., Holt, S.E. & Gewirtz, D.A. Evasion of a single-step, chemotherapy-induced senescence in breast cancer cells: implications for treatment response. Clinical cancer research : an official journal of the American Association for Cancer Research 11, 2637-2643 (2005).
90. Mirzayans, R., Scott, A., Andrais, B., Pollock, S. & Murray, D. Ultraviolet light exposure triggers nuclear accumulation of p21(WAF1) and accelerated senescence in human normal and nucleotide excision repair-deficient fibroblast strains. J Cell Physiol 215, 55-67 (2008).
91. Polyak, K. & Weinberg, R.A. Transitions between epithelial and mesenchymal states: acquisition of malignant and stem cell traits. Nature reviews. Cancer 9, 265-273 (2009).
92. Bolos, V., et al. The transcription factor Slug represses E-cadherin expression and induces epithelial to mesenchymal transitions: a comparison with Snail and E47 repressors. J Cell Sci 116, 499-511 (2003).
93. Hajra, K.M., Chen, D.Y. & Fearon, E.R. The SLUG zinc-finger protein represses E-cadherin in breast cancer. Cancer Res 62, 1613-1618 (2002).
94. Shih, J.Y. & Yang, P.C. The EMT regulator slug and lung carcinogenesis. Carcinogenesis 32, 1299-1304 (2011).
95. Maniwa, Y., et al. His239Arg SNP of HRAD9 is associated with lung adenocarcinoma. Cancer 106, 1117-1122 (2006).
96. Meng, A.X., et al. Hypoxia down-regulates DNA double strand break repair gene expression in prostate cancer cells. Radiother Oncol 76, 168-176 (2005).
97. Bristow, R.G. & Hill, R.P. Hypoxia and metabolism. Hypoxia, DNA repair and genetic instability. Nat Rev Cancer 8, 180-192 (2008).
98. Sullivan, R. & Graham, C.H. Hypoxia prevents etoposide-induced DNA damage in cancer cells through a mechanism involving hypoxia-inducible factor 1. Mol Cancer Ther 8, 1702-1713 (2009).
99. Welford, S.M., et al. HIF1alpha delays premature senescence through the activation of MIF. Genes & development 20, 3366-3371 (2006).
100. Huang, C.H., et al. Regulation of membrane-type 4 matrix metalloproteinase by SLUG contributes to hypoxia-mediated metastasis. Neoplasia 11, 1371-1382 (2009).
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