臺灣博碩士論文加值系統

癌轉移是指惡性癌細胞在體內從啟始發生的位置移動到遠方第二個位置出現的過程。此過程涵蓋許多階段，需要許多不同的基因累積表現變異所致。為了發掘出和癌轉移相關的新穎基因，我們使用一系列具不同侵襲能力的肺腺癌模式細胞株和cDNA微陣列技術來找出哪些基因的表現和癌侵襲或轉移功能具有關連性。在此研究中，我們發現在非小細胞肺癌(NSCLC)病人身上，低表現量的HLJ1和手術後早期復發及較短的生存週期有顯著相關。此外，為了探討在肺癌中是否HLJ1基因會參與抑制腫瘤生成及癌侵襲的過程，我們將人類HLJ1 cDNA轉殖到具高侵襲能力的肺腺癌細胞CL1-5中，此細胞本身僅有少量的內生性HLJ1表現。結果顯示在肺癌細胞中恢復HLJ1基因的表現，可以抑制癌細胞的增生速率、非依賴固著生長(anchorage independent growth)、腫瘤生成、癌細胞侵襲和移動(migration)能力。此外，為瞭解HLJ1的基因調控機制，我們針對人類HLJ1基因可能的啟動子(promoter)及增強子(enhancer)調控區域做進一步的定性研究，並探討其轉錄調節作用。實驗結果顯示，位於-232到+176的區域片段可以啟動HLJ1基因的基礎轉錄能力。透過將YY1和HLJ1基礎啟動子區域一起進行共轉殖(cotransfection)試驗，結合位定點突變試驗(site-directed mutagenesis)，以及電泳移動能力(electrophoretic mobility shift assay)分析試驗，發現YY1會增強HLJ1基礎啟動子的轉錄能力。除此之外，我們發現在轉錄啟始位前端-2,025到-1,039的位置，存在著一個正向調控的轉錄元素，可以持續表現出最大量的相對luciferase基因活性。利用電泳移動能力分析以及結合位定點突變分析，對最小增強子(minimum enhancer)片段進行定性研究後發現，位於-1,457到-1,451位置的AP-1結合位，對於完整的最小增強子活性是必要的。而由抗體supershift分析結果得知，FosB, JunB和JunD蛋白可以作用在此AP-1結合位上。釐清調控HLJ1基因轉錄作用的分子機制，對於發展增加HLJ1基因表現，進而抑制癌細胞生長以及其侵襲作用的治療方法，深具意義及重要性。

Metastasis, the malignant cancer cells migrating from their primary sites of origin to distant secondary sites within the body, is the multiple-step process requiring the accumulation of altered expression of many different genes. To discover novel genes associated with metastasis, a lung adenocarcinoma model cell line and cDNA microarray technology were used to identify the genes whose expression correlated with a function of invasive and metastatic competence. In this study, the results reported that reduced expression of HLJ1 was statistically significantly associated with early postoperative relapse and shorter survival time in NSCLC (non-small cell lung cancer) patients. In addition, to determine whether HLJ1 expression is responsible for the tumor suppression or invasion suppression in lung cancer, the human HLJ1 cDNA was transfected into highly invasive lung adenocarcinoma cells (CL1-5), which have low levels of endogenous HLJ1 expression. Restoration of HLJ1 in lung cancer cells exhibited marked inhibition in the proliferation rate, anchorage independent growth, tumorigenesis, cell invasion and migration. To study the regulatory mechanisms in HLJ1 expression, I further characterized the putative promoter and enhancer regions and investigated the transcriptional regulations of human HLJ1 gene. The results indicated that the region from –232 to +176 could drive the basal transcriptional activity of the HLJ1 gene. Co-transfection of the YY1 and HLJ1 basal promoter regions, the site-directed mutagenesis analysis, and electrophoretic mobility shift assay confirmed that YY1 could up regulate HLJ1 basal promoter activity. In addition, it is also demonstrated that a positive transcriptional element located at position –2,025 to –1,039 base pair upstream of the transcription start site consistently exhibited the highest level of relative luciferase activity. Characterization of the minimum enhancer by electrophoretic mobility shift assay and mutational analyses revealed that AP-1 site (-1,457 to –1,451) is essential for most minimum enhancer activity. Antibody supershift assays determined that FosB, JunB, and JunD could bind to this site. Verifying the molecular mechanisms of HLJ1 transcriptional regulation is important to develop the therapeutic methods with increasing HLJ1 gene expression and inhibiting cancer cell growth and invasion.

正 文 目 錄

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第一章 抑癌基因HLJ1之功能定性分析
第一節 緒言………………………………………………………………...1
第二節 材料與方法………………………………………………………...5
壹、 細胞培養方法…………………………………………………...5
貳、 細胞株RNA之製備…………………………………………….8
參、 反轉錄聚合酶連鎖反應 ……………………………………..…9
肆、 微陣列雜合反應……………………………………………….11
伍、 北方墨點法…………………………………………………….15
陸、 HLJ1基因專一性多株抗體的製備…………………………...17
柒、 蛋白質萃取與西方墨點法…………………………………….18
捌、 即時定量反轉錄聚合酶連鎖反應…………………………….23
玖、 細胞基因組及質體之抽取方法……………………………….25
壹拾、 核酸處理方法(DNA treatment methods)………………....30
壹拾壹、 構築HLJ1基因表現載體………………………………...34
壹拾貳、 建立可大量表現HLJ1基因活性的細胞株……………...35
壹拾參、 HLJ1基因蛋白在細胞內的表現座落位置分析…………37
壹拾肆、 細胞增生試驗(MTT assay)……………………………….37
壹拾伍、 瓊脂膠體株化生長試驗 (Soft agar clonogenic assay)……38
壹拾陸、 試管內侵襲能力試驗 (In vitro invasion assay)…………..38
壹拾柒、 細胞移動性試驗(In vitro wounding migration assay)……39
壹拾捌、 動物體內腫瘤生成試驗 (Animal tumorigenesis)...………40
壹拾玖、 臨床肺癌病人組織檢體樣本之檢測………………...…...40
貳拾、 流動細胞測量計數方法……………………………….….43
貳拾壹、 小片段干擾RNA轉殖實驗(siRNA Transfection)………45
第三節 結果………………………………………………………………47
壹、 HLJ1基因的表現和肺癌細胞株的侵襲能力降低有關……..47
貳、 HLJ1基因在不同侵襲能力肺癌細胞內的熱休克效應……..51
參、 HLJ1蛋白在細胞內的座落位置分佈情形…………………..53
肆、 HLJ1基因持續或誘發表現系統之建立……………………..56
伍、 HLJ1基因表現會降低癌細胞的增生能力和非依賴固著生長(anchorage-independent growth)能力…………………………60
陸、 HLJ1基因表現會在體外情況下抑制癌細胞的侵襲和移動能力………………………………………………………………64
柒、 HLJ1基因表現抑制小鼠體內腫瘤的生長…………………..67
捌、 HLJ1基因表現可預測NSCLC病人的生存時間長短………70
玖、 HLJ1基因表現和細胞週期暫停(arrest)之關連性…………...79
壹拾、 以Affymetrix寡核苷酸微陣列分析實驗找出受HLJ1基因調控的下游基因……………………………………………….84
壹拾壹、 HLJ1基因可以正向調節p21WAF1及負向調節cyclin D1基因的表現……………………………………………………….87
第四節 討論……………………………………………………………….91
第二章 抑癌基因HLJ1啟動子轉錄調節機制之探討
第一節 緒言……………………………………………………………..101
第二節 材料與方法……………………………………………………..104
壹、 HLJ1基因5’端相鄰區域的選殖與定序……………………104
貳、 Luciferase報導基因之構築…………………………………105
參、 5’-RACE (rapid amplification of cDNA ends)………….…...106
肆、 轉殖試驗和luciferase 報導基因分析實驗…………………107
伍、 北方及西方墨點法分析…………………………………….109
陸、 即時定量反轉錄聚合酶連鎖反應………………………….109
柒、 Matrigel侵襲能力試驗……………………………………...110
捌、 膠體訊號移動分析(Elecphoretic mobility shift assay)……...111
玖、 組織免疫染色試驗 (IHC, immunohistochemical stain)……..112
壹拾、 統計分析方法……………………………………………113
第三節 結果……………………………………………………………...115
壹、 人類HLJ1基因啟動子的選殖及序列分析…………………115
貳、 人類HLJ1基因具有細胞專一性的啟動子能力……………118
參、 大量表現YY1可以增加HLJ1的表現並降低CL1-0細胞的侵襲能力………………………………………………………..122
肆、 完整的YY1 DNA結合區域對於HLJ1啟動子的transactivation是必要的……………………………………..127
伍、 HLJ1基因的表現可以一部份透過調升E-cadherin的表現而抑制細胞的侵襲能力…………………………………………...133
第四節 討論……………………………………………………………...141
第三章 抑癌基因HLJ1增強子轉錄調節機制之探討
第一節 緒言………………………………………………………….…..149
第二節 材料與方法…………………………..………………………….152
壹、 細胞培養方法 ………………………………………………..152
貳、 HLJ1基因增強子區域的選殖及定序……………………….152
參、 Luciferase報導基因之構築………………………………….153
肆、 轉殖試驗和luciferase 報導基因分析實驗………………….156
伍、 膠體訊號移動分析(Elecphoretic mobility shift assay)………156
陸、 統計分析方法………………………………………………...158
第三節 結果……………………………………………………………...159
壹、 HLJ1基因啟動子之功能分析……………………………….159
貳、 人類HLJ1基因增強子活性分析……………………………163
參、 基因基礎啟動子和5’端增強子的交互作用………………...166
肆、 確認位於HLJ1基因5’端相鄰區域中的最小增強子元素…168
伍、 剖析最小增強子元素以找出表現增強子活性時所需的可能正向調節motif………………………………………………….170
陸、 人類HLJ1基因最小增強子元素在CL1-0細胞中的特殊DNA-protein交互作用……………………………………….174
第四節 討論……………………………………………………………...177
第四章 結論………………………………………………………………….183
第五章 參考文獻…………………………………………………………….185

1.Parkin DM, Bray FI and Devesa SS: Cancer burden in the year 2000. The global picture. Eur. J. Cancer 37: S4-S66, 2001.
2.Department of Health, Executive Yuan, Taiwan, R. O. C. Taiwan area main causes of death, 2003. Health and Vital Statistics (http://www.doh.gov.tw)
3.Hoffman PC, Mauer AM and Vokes EE: Lung cancer. Lancet 355: 479-485, 2000.
4.Jemal A, Thomas A, Murray T and Thun M: Cancer statistics, 2002. CA Cancer J. Clin. 52: 23-47, 2002.
5.Jaklitsch MT, Strauss GM, Healey EA, DeCamp MM Jr and Sugarbaker DJ: An historical perspective of multi-modality treatment for respectable non-small cell lung cancer. Lung Cancer 12: S17-32, 1995.
6.Niklinski J, Niklinska W, Chyczewski L, Becker HD and Pluygers E: Molecular genetic abnormalities in premalignant lung lesions: biological and clinical implications. Eur. J. Cancer Prev. 10: 213-226, 2001.
7.Mountain CF: Revisions in the International System for Staging Lung Cancer. Chest 111: 1710-1717, 1997.
8.Meyer T and Hart IR: Mechanisms of tumor metastasis. Eur. J. Cancer 34: 214-221, 1998.
9.Yoneda T: Cellular and molecular mechanisms of breast and prostate cancer metastasis to bone. Eur. J. Cancer 34:240-245, 1998.
10. Zhao H, Jhanwar-Uniyal M, Datta PK, Yemul S, Ho L, Khitrov G, Kupershmidt I, Pasinetti GM, Ray T, Athwal RS and Achary MP: Expression profile of genes associated with antimetastatic gene: nm23-mediated metastasis inhibition in breast carcinoma cells. Int. J. Cancer 109: 65-70, 2004.
11. Gao AC, Lou W, Dong JT and Isaacs JT: CD44 is a metastasis suppressor gene for prostatic cancer located on human chromosome 11p13. Cancer Res. 57: 846-849, 1997.
12. Nicolson GL, Nawa A, Toh Y, Taniguchi S, Nishimori K and Moustafa A: Tumor metastasis-associated human MTA-1 gene and its MTA1 protein product: role in epithelial cancer cell invasion, proliferation and nuclear regulation. Clin. Exp. Metastasis. 20: 19-24, 2003.
13. Shiomi T and Okada Y: MT1-MMP and MMP-7 in invasion and metastasis of human cancers. Cancer Metastasis Rev. 22:145-152, 2003.
14. Lee JH, Seo YW, Park SR, Kim YJ and Kim KK: Expression of a splice variant of KAI1, a tumor metastasis suppressor gene, influences tumor invasion and progression. Cancer Res. 63: 7247-7255, 2003.
15. Jiang WG: E-cadherin and its associated protein catenins, cancer invasion and metastasis. Br. J. Surg. 83: 437-446, 1996.
16. Bremnes RM, Veve R, Hirsch FR and Franklin WA: The E-cadherin cell-cell adhesion complex and lung cancer invasion, metastasis, and prognosis. Lung Cancer 36: 115-124, 2002.
17. Harms JF, Welch DR and Miele ME: KISS1 metastasis suppression and emergent pathways. Clin. Exp. Metastasis 20: 11-18, 2003.
18. Steeg PS: Metastasis suppressors alter the signal transduction of cancer cells. Nat. Rev. Cancer 3: 55-63, 2003.
19. Chen JJW, Peck K, Hong TM, Yang SC, Sher YP, Shih JY, Wu R, Cheng JL, Rofler SR, Wu CW and Yang PC: Global analysis of gene expression in invasion by a lung cancer model. Cancer Res. 61: 5223-5230, 2001.
20. Shih JY, Yang SC, Hong TM, Yuan A, Chen JJ, Yu CJ, Chang YL, Lee YC, Peck K, Wu CW and Yang PC: Coolapsin response mediator protein-1 and the invasion and metastasis of cancer cells. J. Natl. Cancer Inst. 93: 1392-1400, 2001.
21. Hoe KL, Won M, Chung KS, Jang YJ, Lee SB, Kim DU, Lee JW, Tun JH and Yoo HS: Isolation of a new member of Dnaj-like heat shock protein 40 (Hsp40) from human liver. Biochem. Biophys. Acta. 1383: 4-8, 1998.
22. Ritossa F: A new puffing pattern induced by temperature shock and DNP in drosophila. Experientia. 18: 571-573, 1962.
23. Ritossa F: Discovery of the heat shock response. Cell Stress Chaperones 1: 97-98, 1996.
24. Morimoto RI, Kline MP, Bimston DN and Cotto JJ: The heat-shock response: regulation and function of heat-shock proteins and molecular chaperones. Essays Biochem. 32: 17-29, 1997.
25. Hartl FU and Hayer-Hartl M: Molecular chaperones in the cytosol: from nascent chain to folded protein. Science 295: 1852-1858, 2002.
26. Csermeley P, Schnaider T, Soti C, Prohaszka Z and Nardai G: The 90-kDa molecular chaperone family: structure, function, and clinical applications. A comprehensive review. Pharmacol. Ther. 79: 129–168, 1998.
27. Lindquist S and Craig EA: The heat shock proteins. Annu. Rev. Genet. 22: 631–677, 1988.
28. Kiang J and Tsokos G: Heat shock protein 70 kDa: molecular biology, biochemistry, and physiology. Pharmacol. Ther. 80: 183–201, 1998.
29. Robert J: Evolution of heat shock protein and immunity. Dev. Comp. Immunol. 27: 449-464, 2003.
30. Helmbrecht K, Zeise E and Rensing L: Chaperones in cell cycle regulation and mitogenic signal transduction: a review. Cell Prolif. 33: 341-365, 2000.
31. Neininger A and Gaestel M: Evidence for a hsp25-specific mechanism involved in transcriptional activation by heat shock. Exp. Cell Res. 242: 285-293, 1998.
32. Jolly C and Morimoto RI: Role of the heat shock response and molecular chaperones in oncogenesis and cell death. J. Natl. Cancer Inst. 92: 1564-1572, 2000.
33. Ohtsuka K: Cloning of cDNA for heat-shock protein hsp40, a human homologue of bacterial DnaJ. Biochem. Biophys. Res. Commun. 197: 235-240, 1993.
34. Cheetham ME and Caplan AJ: Structure, function and evolution of DnaJ: conservation and adaptation of chaperon function. Cell Stress Chaperones 3: 28-36, 1998.
35. Sullivan CS and Pipas JM: T antigens of simian virus 40: molecular chaperones for viral replication and tumorigenesis. Microbiol. Mol. Biol. Rev. 66: 179-202, 2002.
36. Schilling B, De-Medina T, Syken J, Vidal M and Munger K: A novel human DnaJ protein, hTid-1, a homolog of the Drosophila tumor suppressor protein Tid56, can interact with the human papillomavirus type 16 E7 oncoprotein. Virology 247: 74-85, 1998.
37. Canamasas I, Debes A, Natali PG and Kurzik-Dumke U: Understanding human cancer using Drosophila: Tid47, a cytosolic product of the DnaJ-like tumor suppressor gene I2Tid, is a novel molecular partner of patched related to skin cancer. J. Biol. Chem. 278: 30952-30960, 2003.
38. Kurzik-Dumke U, Gundacker D, Renthrop M and Gateff E: Tumor suppression in Drosophila is causally related to the function of the lethal (2) tumorous imaginal discs gene, a dnaJ homolog. Dev. Genet. 16: 64-76, 1995.
39. Chu YW, Yang PC, Yang SC, Shyu YC, Hendrix MJ, Wu R and Wu CW: Selection of invasive and metastatic subpopulations from a human lung adenocarcinoma cell line. Am. J. Respir. Cell Mol. Biol. 17: 353-360, 1997.
40. Hong TM, Yang PC, Peck K, Chen JJW, Yang SC, Chen YC and Wu CW: Profiling the downstream genes of tumor suppressor PTEN in lung cancer cells by complementary DNA microarray. Am. J. Respir. Cell Mol. Biol. 23: 355-363, 2000.
41. Andrews NC and Faller DV: A rapid micropreparation technique for extraction of DNA-binding proteins from limiting numbers of mammalian cells. Nucleic Acid Res. 19: 2499, 1991.
42. Bieche I, Onody P, Laurendeau I, Olivi M, Vidaud D, Lidereau R and Vidaud M: Real-time reverse transcription-PCR assay for future management of ERBB2-based clinical applications. Clin. Chem. 45: 1148-56, 1999.
43. Sambrook J, Fritsch EF and Maniatis T: Molecular cloning. A Laboratory Manual, 2nd edition. Cold Spring Harbor, New York. 1989
44. World Health Organization. The World Health Organization: Histological typing of lung tumors, 2nd edition. Am. J. Clin. Pathol. 77: 123-136, 1982.
45. Al-Mohanna MA, Al-Khodairy FM, Krezolek Z, Bertilsson PA, Al-Houssein KA and Aboussekhra A: A p53 is dispensable for UV-induced cell cycle arrest at late G1 in mammalian cells. Carcinogenesis 22: 573-578, 2001.
46. Nakai K and Horton P: PSORT: a program for detecting sorting signals in proteins and predicting theirsubcellular localization. Trends Biochem. Sci. 24: 34-36, 1999.
47. Wang S, Yan-Neale Y, Fischer D, Zeremski M, Cai R, Zhu J, Asselbergs F, Hampton G and Cohen D: Histone deacetylase 1 represses the small GTPase RhoB expression in human non-small lung carcinoma cell line. Oncogene 22: 6204-6213, 2003.
48. Ikoma T, Takahashi T, Nagano S, Li YM, Ohno Y, Ando K, Fujiwara T, Fujiwara H and Kosai K: A definitive role of RhoC in metastasis of orthotopic lung cancer in mice. Clin. Cancer Res. 10: 1192-1200, 2004.
49. Hirata T, Fukuse T, Naiki H and Wada H: Expression of E-cadherin and lymph node metastasis in resected non-small cell lung cancer. Clin. Lung Cancer 3: 134-140, 2001.
50. Higashiyama M, Kodama K, Yokouchi H, Takami K, Adachi M, Taki T, Ishiguro S, Nakamori S, Yoshie O and Miyake M: KAI1/CD82 expression in non-small cell lung carcinoma is a novel, favorable prognostic factor: an immunohistochemical analysis. Cancer 83: 466-474, 1998.
51. Lamb RF, Roy C, Diefenbach TJ, Vinters HV, Johnson MW, Jay DG, and Hall A: The TSC1 tumour suppressor hamartin regulates cell adhesion through ERM proteins and the GTPase Rho. Nat. Cell Biol. 2: 281-287, 2000.
52. Kim S, Han J, Kim J and Park C: Maspin expression is transactivated by p63 and is critical for the modulation of lung cancer progression. Cancer Res. 64: 6900-6905, 2004.
53. Sanchez-Cespedes M: Dissecting the genetic alterations involved in lung carcinogenesis. Lung Cancer 40: 111-121, 2003.
54. Girard L, Zochbauer-Muller S, Virmani AK, Gazdar AF and Minna JD: Genome-wide allelotyping of lung cancer identifies new regions of allelic loss, differences between small cell lung cancer and non-small cell lung cancer, and loci clustering. Cancer Res. 60: 4894-4906, 2000.
55. Ragnarsson G, Eiriksdottir G, Johannsdottir J, Jonasson JG, Egilsson V and Ingvarsson S: Loss of heterozygosity at chromosome 1p in different solid human tumours: association with survival. Br. J. Cancer 79: 1468-1474, 1999.
56. Fearon ER: Human cancer syndromes: clues to the origin and nature of cancer. Science 278: 1043-1050, 1997.
57. Esteller M: CpG island hypermethylation and tumor suppressor genes: a booming present, a brighter future. Oncogene 21: 5427-5440, 2002.
58. Takise A, Kodama T, Shimosato Y, Watanabe S and Suemasu K: Histopathologic prognostic factors in adenocarcinomas of the peripheral lung less than 2 cm in diameter. Cancer 61: 2083-2088, 1988.
59. Harpole DH, Herndon JE, Wolfe WG, Iglehart JD and Marks JR: A prognostic model of recurrence and death in stage I non-small cell lung cancer utilizing presentation, histopathology, and oncoprotein expression. Cancer Res. 55: 51-56, 1995.
60. Roth JA, Fossella F, Komaki R, Ryan MB, Putnam JB, Lee JS, Dhingra H, De Caro L, Chasen M and McGavran M: A randomized trial comparing perioperative chemotherapy and surgery with surgery alone in resectable stage IIIA non-small-cell lung cancer. J. Natl. Cancer Inst. 86: 673-680, 1994.
61. Rosell R, Gomez-Codina J, Camps C, Maestre J, Padille J, Canto A, Mate JL, Li S, Roig J, Olazabal A, Canela M, Ariza A, Skacel Z, Morera-Prat J and Abad A: A randomized trial comparing preoperative chemotherapy plus surgery with surgery alone in patients with non-small-cell lung cancer. N. Engl. J. Med. 330: 153-158, 1994.
62. Moldvay J, Scheid P, Wild P, Nabil K, Siat J, Borrelly J, Marie B, Farré G, Labib T, Pottier G, Sesboüé R, Bronner C, Vignaud J-M, Martinet Y and Martinet N: Predictive survival markers in patients with surgically resected non-small cell lung carcinoma. Clin. Cancer Res. 6: 1125-1134, 2000.
63. Beer DG, Kardia SLR, Huang CC, Giordano TJ, Levin AM, Misek DE, Lin L, Chen G, Gharib TG, Thomas DG, Lizyness ML, Kuick R, Hayasaka S, Taylor JMG, Iannettoni MD, Orringer MB and Hanash S: Gene-expression profiles predict survival of patients with lung adenocarcinoma. Nat. Med. 8: 816-824, 2002. Nelson HH, Christiani DC, Mark EJ, Wiencke JK, Wain JC and Kelsey KT: Implications and prognostic value of K-ras mutation for early-stage lung cancer in women. J. Natl. Cancer Inst. 91: 2032-2038, 1999.
65. Kelly WL: The J-domain family and the recruitment of chaperone power. Trends Biochem. Sci. 23: 222-227, 1998.
66. Ohtsuka K and Hata M: Mammalian HSP40/DNAJ homologues: cloning of novel cDNAs and a proposal for their classification and nomenclature. Cell Stress Chaperones 5: 98-112, 2000.
67. Minami Y, Hohfeld J, Ohtsuka K and Hartl FU: Regulation of the heat-shock protein 70 reaction cycle by the mammalian DnaJ homologue, Hsp40. J. Biol. Chem. 271: 19617-19624, 1996.
68. Meacham GC, Lu Z, King S, Sorscher E, Tousson A and Cyr DM: The Hdj-2/Hsc70 chaperone pair facilitates early steps in CFTR biogenesis. EMBO J. 18: 1492-1505, 1999.
69. Rassow J, Voos W and Pfanner N: Partner proteins determine multiple functions of Hsp70. Trends Cell Biol. 5: 207-212, 1995.
70. Izawa I, Nishizawa M, Ohtakara K, Ohtsuka K, Inada H and Inagaki M: Identification of Mrj, a DnaJ/Hsp40 family protein, as a keratin 8/18 filament regulatory protein. J. Biol. Chem. 275: 34521-34527, 2000.
71. Sedbrook JC, Chen R and Masson PH: ARG1 (altered response to gravity) encodes a DnaJ-like protein that potentially interacts with the cytoskeleton. Proc. Natl. Acad. Sci. 96: 1140-1145, 1999.
72. Baldin V, Lukas J, Marcote MJ, Pagano M and Draetta G: Cyclin D1 is a nuclear protein required for cell cycle progression in G1. Genes Dev. 7: 812-821, 1993.
73. Yang ZY, Perkins ND, Ohno T, Nabel EG and Nabel GJ: The p21 cyclin-dependent kinase inhibitor suppresses tumorigenicity in vivo. Nat. Med. 1: 1052-1056, 1995.
74. Kumar A, Commane M, Flickinger TW, Horvath CM and Stark GR: Defective TNF-alpha-induced apoptosis in STAT1-null cells due to low constitutive levels of caspases. Science 278: 1630-1632, 1997.
75. Chin YE, Kitagawa M, Su WCS, You ZH, Iwamoto Y and Fu XY: Cell growth arrest and induction of cyclin-dependent kinase inhibitor p21 WAF1/CIP1 mediated by STAT1. Science 272: 719-722, 1996.
76. Webb DJ, Donais K, Whitmore LA, Thomas SM, Turner CE, Parsons JT and Horwitz AF: FAK-Src signaling through paxillin, ERK and MLCK regulates adhesion disassembly. Nat. Cell Biol. 6: 154-161, 2004.
77. Crean JK, Furlong F, Finlay D, Mitchell D, Conway B, Brady HR, Godson C and Martin F: Connective tissue growth factor [CTGF]/CCN2 stimulates mesangial cell migration through integrated dissolution of focal adhesion complexes and activation of cell polarization. FASEB J. 18: 1541-1543, 2004.
78. Welch WJ: Mammalian stress response: cell physiology, structure/function of stress proteins, and implications for medicine and disease. Physiol. Rev. 72: 1063-1081, 1992.
79. Hendric JP and Hartl FU: Molecular chaperone functions of heat-shock proteins. Annu. Rev. Biochem. 62: 349-384, 1993.
80. Craig EA, Weissman JS and Horwich AL: Heat shock proteins and molecular chaperones: mediators of protein conformation and turnover in the cell. Cell 78: 365-372, 1994.
81. Caroline J and Richard IM: Role of the heat shock response and molecular chaperons in oncogenesis and cell death. J. Natl. Cancer Inst. 92: 1564-1572, 2000.
82. Hartl FU: Molecular chaperones in cellular protein folding. Nature 381: 571-580, 1996.
83. Gottesman S, Wickner S and Maurizi MR: Protein quality control: triage by chaperones and proteases. Genes Dev. 11: 815-823, 1997.
84. Fry CJ and Farnham PJ: Context-dependent transcriptional regulation. J. Biol. Chem. 274: 29583-29586, 1999.
85. Kim J and Shapiro DJ: In simple synthetic promoters YY1-induced DNA bending is important in transcription activation and repression. Nucleic Acids Res. 24:4341-4348, 1996.
86. Khachigian LM, Williams AJ and Collins T: Interplay of Sp1 and Egr-1 in the Proximal Platelet-derived Growth Factor A-Chain Promoter in Cultured Vascular Endothelial Cells. J. Biol. Chem. 270: 27679-27686, 1995.
87. Perkins ND, Edwards NL, Duckett CS, Agranoff AB, Schmid RM and Nabel GJ: A cooperative interaction between NF-kappa B and Sp1 is required for HIV-1 enhancer activation. EMBO J. 12: 3551-3558, 1993.
88. Quandt K, Frech K, Karas H, Wingender E and Werner T: MatInd and MatInspector: new fast and versatile tools for detection of consensus matches in nucleotide sequence data. Nucleic Acids Res. 23: 4878-4884, 1995
89. Prestridge DS: SIGNAL SCAN: A computer program that scans DNA sequences for eukaryotic transcriptional elements. CABIOS 7: 203-206, 1991.
90. Heinemeyer T, Chen X, Karas H, Kel AE, Kel OV, Liebich I, Meinhardt T, Reuter I, Schacherer F and Wingender E: Expanding the TRANSFAC database towards an expert system of regulatory molecular mechanisms. Nucleic Acids Res. 27: 318-322, 1999.
91. Matz M, Shagin D, Bogdanova E, Britanova O, Lukyanov S, Diatchenko L and Chenchik A: Amplification of cDNA ends based on template-switching effect and step-out PCR. Nucleic Acids Res. 27: 1558-1560, 1999.
92. Yao YL, Yang WM and Seto E: Regulation of transcription factor YY1 by acetylation and deacetylation. Mol. Cell Biol. 21: 5979-5991, 2001.
93. Sreenath T, Matrisian LM, Stetler-Stevenson W, Gattoni-Celli S and Pozzatti RO: Expression of matrix metalloproteinase genes in transformed rat cell lines of high and low metastatic potential. Cancer Res. 52: 4942-4947, 1992.
94. Birch M, Mitchell S and Hart IR: Isolation and characterization of human melanoma cell variants expressing high and low levels of CD44. Cancer Res. 51: 6660-6667, 1991.
95. Uleminckx K, Vackat L, Mareel M, Fiers W and Von Roy F: Genetic manipulation of E cadherin expression by epithelial tumor cells reveals an invasion suppressor role. Cell 66: 107-119, 1991.
96. Liotta LA, Steeg PS and Stetler-Stevenson WG: Cancer metastatis and angiogenesis: an imbalance of positive and negative regulation. Cell 64: 327-336, 1991.
97. Mantovani R: A survey of 178 NF-Y binding CCAAT boxes. Nucleic Acids Res. 26: 1135-1143, 1998.
98. Bird AP: CpG rich islands and the function of DNA methylation. Nature 321: 209-213, 1986.
99. Li WW, Hsiung Y, Wong V, Galvin K, Zhou Y, Shi Y and Lee AS: Suppression of grp78 core promoter element-mediated stress induction by the dbpA and dbpB (YB-1) cold shock domain proteins. Mol. Cell Biol. 17: 61-68, 1997.
100.Shi Y, Seto E, Chang LS and Shenk T: Transcriptional repression by YY1, a human GLI-Kruppel-related protein, and relief of repression by adenovirus E1A protein. Cell 67: 377-388, 1991.
101.Galvin KM and Shi Y: Multiple mechanisms of transcriptional repression by YY1. Mol. Cell Biol. 17: 3723-3732, 1997.
102.Ficzycz A and Ovsenek N: The Ying Yang 1 transcription factor associates with ribonucleoprotein (mRNP) complexes in the cytoplasm of Xenopus oocytes. J. Biol. Chem. 277: 8382-8387, 2002.
103.Shi Y, Lee JS and Galvin KM: Everything you have ever wanted to know about Ying Yang 1. Biochim. Biophys. Acta. 1332: F49-F66, 1997.
104.Usheva A and Shenk T: TATA-binding protein-independent initiation: YY1, TFIIB and RNA polymerase II direct basal transcription on supercoiled template DNA. Cell 76: 1115-1121, 1994.
105.Seto E, Shi Y and Shenk T: YY1 is an initiator sequence-binding protein that directs and activates transcription in vitro. Nature 354: 241-245, 1991.
106.Natesan S and Gilman MZ: DNA bending and orientation-dependent function of YY1 in the c-fos promoter. Genes Dev. 7: 2497-2509, 1993.
107.Lu SY, Rodriguez M and Liao WSL: YY1 represses rat serum amyloid A1 transcription and is antagonized by NF-kappaB during acute-phase response. Mol. Cell Biol. 14: 6253-6263, 1994.
108.Meier VS and Groner B: The nuclear factor YY1 participates in repression of the beta-casein gene promoter in mammary epithelial cells and is counteracted by mammary gland factor during lactogenic hormone induction. Mol. Cell Biol. 14: 128-137, 1994.
109.Zhou Q and Engel DA: Transcriptional repression of the c-fos gene by YY1 is mediated by a direct interaction with ATF/CREB. J. Virol. 69: 7402-7409, 1995.
110.Guo B, Aslam F, van Wijnen AJ, Roberts SGE, Frenkel B, Green MR, Deluca H, Lian JB, Stein GS and Stein JL: YY1 regulates VDR/RXR mediated transactivation of the vitamin D responsive osteocalcin gene. Proc. Natl. Acad. Sci. USA 94: 121-126, 1997.
111.Hariharan N, Kelley DE and Perry RP: Delta, a transcription factor that binds to downstream elements in several polymerase II promoters, is a functionally versatile zinc finger protein. Proc. Natl. Acad. Sci. USA 88: 9799-9803, 1991.
112.Azizkhan JC, Jensen DE, Pierce AJ and Wade M: Transcription from TATA-less promoters: Dihydrofolate reductase as a model. Crit. Rev. Eukartotic Gene Expr. 3: 229-254, 1993.
113.Riggs KJ, Saleque S, Wong KK, Merrell KT, Lee JS, Shi Y and Calame K: Yin-yang 1 activates the c-myc promoter. Mol. Cell Biol. 13: 7487-7495, 1993.
114.Hehlgans T and Strominger JL: Activation of transcription by binding of NF-E1 (YY1) to a newly identified element in the first exon of the human DR alpha gene J. Immunol. 154: 5181-5187, 1995.
115.Takeichi M: Cadherin cell adhesion receptors, as morphogenetic regulator. Science 251: 1451-1455, 1991.
116.Knudsen KA, Soler AP, Johnson KR and Wheelock MJ: Interaction of alpha-actinin with the cadherin/catenin cell-cell adhesion complex via alpha-catenin. J. Cell Biol. 130: 67-77, 1995.
117.Angst BD, Marcozzi C and Magee AI: The cadherin superfamily: diversity in form and function. J. Cell Sci. 114: 629-641, 2001.
118.Berx G, Becker KF, Hofler H and Van Roy F: Mutations of the human E-cadherin (CDH1) gene. Hum. Mutat. 12: 226-237, 1998.
119.Hirohashi S: Inactivation of the E-Cadherin-Mediated Cell Adhesion System in Human Cancers. Am. J. Pathol. 153: 333-339, 1998.
120.Bremnes RM, Veve R, Hirsch FR and Franklin WA: The E-cadherin cell-cell adhesion complex and lung cancer invasion, metastasis, and prognosis. Lung Cancer 36: 115-124, 2002.
121.Bukholm IK, Nesland JM, Karesen R, Jacobsen U and Borresen-Dale AL: E-cadherin and a-, b-, and g-catenin protein expression in relation to metastasis in human breast carcinoma. J. Pathol. 185: 262-266, 1998.
122.Hajra KM, Chen DY and Fearon ER: The SLUG zinc-finger protein represses E-cadherin in breast cancer. Cancer Res. 62: 1613-1618, 2002.
123.Thomas MJ and Seto E: Unlocking the mechanisms of transcription factor YY1: are chromatin modifying enzymes the key? Gene 236: 197-208, 1999.
124.Lee JS, Galvin KM, See RH, Eckner R, Livingston D, Moran E and Shi Y: Relief of YY1 transcriptional repression by adenovirus E1A is mediated by E1A-associated protein p300. Genes Dev. 9: 1188-1198, 1995.
125.Li WW, Hsiung Y, Zhou Y, Roy B and Lee AS: Induction of the mammalian GRP78/BiP gene by Ca2+ depletion and formation of aberrant proteins: activation of the conserved stress- inducible grp core promoter element by the human nuclear factor YY1. Mol. Cell Biol. 17: 54-60, 1997.
126.Austen M, Cerni C, Luscher-Firzlaff JM and Luscher B: YY1 can inhibit c-Myc function through a mechanism requiring DNA binding of YY1 but neither its transactivation domain nor direct interaction with c-Myc. Oncogene 17: 511-520, 1998.
127.Sistonen L, Sarge KD, Phillips B, Abravaya K and Morimoto R: Activation of heat shock factor 2 during hemin-induced differentiation of human erythroleukemia cells. Mol. Cell Biol. 12: 4104-4111, 1992.
128.Miernyk JA: The J-domain proteins of Arabidopsis thaliana: An unexpectedly large and diverse family of chaperones. Cell Stress Chaperones 6: 209-218, 2001.
129.Borges JC, Peroto MC and Ramos CHI: Molecular chaperone genes in the sugarcane expressed sequence database (SUCEST). Gen. Mol. Biol. 24: 85-92, 2001.
130.Venter JC, Adams MD, Myers EW, et al. The Sequence of the Human Genome. Science 291: 1304-1351, 2001.
131.Jack J, Dorsett D, Delotto Y and Liu S: Expression of the cut locus in the Drosophila wing margin is required for cell type specification and is regulated by a distant enhancer. Development 113: 735-747, 1991.
132.Khoury G and Gruss P: Enhancer elements. Cell 33:313-314, 1983.
133.Bondarenko VA, Liu YV, Jiang YI and Studitsky VM: Communication over a large distance: enhancers and insulators. Biochem. Cell Biol. 81: 241-251, 2003.
134.Tjian R and Maniatis T: Transcriptional activation: a complex puzzle with few easy pieces. Cell 77: 5-8, 1994.
135.Hill CS and Treisman R: Transcriptional regulation by extracellular signals: mechanisms and specificity. Cell 80: 199-211, 1995.
136.Grosschedl R: Higher-order nucleoprotein complexes in transcription: analogies with site-specific recombination. Curr. Opin. Cell Biol. 7: 362-370, 1995.
137.Thanos D and Maniatis T: Virus induction of human IFN-b gene expression requires the assembly of an enhanceosome. Cell 83: 1091-1100, 1995.
138.Falvo JV, Thanos D and Maniatis T: Reversal of intrinsic DNA bends in the IFN-b gene enhancer by transcription factors and the architectural protein HMG I(Y). Cell 83: 1101-1111, 1995.
139.Cosma MP: Ordered recruitment: gene-specific mechanism of transcription activation. Mol. Cell 10: 227-236, 2002.
140.Ptashne M: Gene regulation by proteins acting nearby and at a distance. Nature 322: 697-701, 1986.
141.Matthews KS: DNA looping. Microbiol. Rev. 56: 123-136, 1992.
142.Dorsett D: Distant liaisions: long-range enhancer-promoter interactions in Drodophila. Curr. Opin. Genet. Dev. 9: 505-514, 1999.
143.Li JJ, Westergaard C, Ghosh P and Colburn NH: Inhibitors of both nuclear factor-kappaB and activator protein-1 activation block the neoplastic transformation response. Cancer Res. 57: 3569-3576, 1997.
144.Li JJ, Rhim JS, Schlegel R, Vousden KH and Colburn NH: Expression of dominant negative Jun inhibits elevated AP-1 and NF- B transactivation and suppresses anchorage independent growth of HPV immortalized human keratinocytes. Oncogene 16: 2711-2721, 1998.
145.Li J, Cho Y, Young MR and Colburn NH: Induced expression of dominant-negative c-jun downregulates NF B and AP-1 target genes and suppresses tumor phenotype in human keratinocytes. Mol. Carcinog. 29: 159-169, 2000.
146.Barton MC, Madani N and Emerson BM: Distal enhancer regulation by promoter derepression in topologically constrained DNA in vitro. Proc. Natl. Acad. Sci. USA 94: 7257-7262, 1997.
147.Willis SD and Seyfred MA: Pituitary-specific chromatin structure of the rat prolactin distal enhancer element. Nucleic Acids Res. 24: 1065-1072, 1996
148.Yang X, Taylor L and Polgar P: Mechanisms in the transcriptional regulation of Bradykinin B1 receptor gene expression. J. Biol. Chem. 273: 10763-10770. 1998.
149.Wang YN and Chang WC: Induction of disease-associated keratin 16 gene expression by epidermal growth factor is regulated through cooperation of transcriptio factors Sp1 and c-Jun. J. Biol. Chem. 278: 45848-45857, 2003.
150.Chen BK and Chang WC: Functional interaction between c-Jun and promoter factor Sp1 in epidermal growth factor-induced gene expression of human 12(S)-lipoxygenase. Proc. Natl. Acad. Sci. USA 97: 10406-10411, 2000.
151.Welter JF, Crish JF, Agarwal C and Eckert RL: Fos-related Antigen (Fra-1), junB, and junD Activate Human Involucrin Promoter Transcription by Binding to Proximal and Distal AP1 Sites to Mediate Phorbol Ester Effects on Promoter Activity. J. Biol. Chem. 270: 12614-12622, 1995.
152.Angel P and Karin M: The role of Jun, Fos and the AP-1 complex in cell-proliferation and transformation. Biochim. Biophys. Acta. 1072: 129-157, 1991.
153.Crowe DL and Brown TN: Transcriptional inhibition of matrix metalloproteinase 9 (MMP-9) activity by a c-fos/estrogen receptor fusion protein is mediated by the proximal AP-1 site of the MMP-9 promoter and correlates with reduced tumor cell invasion. Neoplasia 1: 368-372, 1999.
154.Ryseck RP and Bravo R: c-Jun, JunB, and JunD differ in their binding affinities to AP-1 and CRE consensus sequences: effect of FOS proteins. Oncogene 6: 533-542, 1991.
155.Kaminska B, Pyrzynska B, Ciechomska I and Wisniewska M: Modulation of the composition of AP-1 complex and its impact on transcriptional activity. Acta. Neurobiol. Exp. 60: 395-402, 2000.
156.Grundker C, Schlotawa L, Viereck V and Emons G: Protein kinase C-independent stimulation of activator protein-1 and c-Jun N-terminal kinase activity in human endometrial cancer cells by the LHRH agonist triptorelin. Eur. J. Endocrinol. 145: 651-658, 2001.
157.Song MS, Park YK, Lee JH and Park K: Induction of Glucose-regulated Protein 78 by Chronic Hypoxia in Human Gastric Tumor Cells through a Protein Kinase C-{epsilon}/ERK/AP-1 Signaling Cascade. Cancer Res. 61: 8322-8330, 2001.
158.Simon C, Simon M, Vucelic G, Hicks MJ, Plinkert PK, Koitschev A and Zenner HP: The p38 SAPK Pathway Regulates the Expression of the MMP-9 Collagenase via AP-1-Dependent Promoter Activation. Exp. Cell Res. 271: 344-355, 2001.