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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 張?仁(Ching-Jin Chang) | |
| dc.contributor.author | Ching-Han Kao | en |
| dc.contributor.author | 高靖涵 | zh_TW |
| dc.date.accessioned | 2021-06-14T17:12:08Z | - |
| dc.date.available | 2008-08-05 | |
| dc.date.copyright | 2008-08-05 | |
| dc.date.issued | 2008 | |
| dc.date.submitted | 2008-07-25 | |
| dc.identifier.citation | 1. Fenger-Gron, M., et al., Multiple processing body factors and the ARE binding protein TTP activate mRNA decapping. Mol Cell, 2005. 20(6): p. 905-915.
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Dunckley, T. and R. Parker, The DCP2 protein is required for mRNA decapping in Saccharomyces cerevisiae and contains a functional MutT motif. Embo J, 1999. 18(19): p. 5411-5422. 10. Cunningham, K.S., et al., Vigilin binding selectively inhibits cleavage of the vitellogenin mRNA 3'-untranslated region by the mRNA endonuclease polysomal ribonuclease 1. Proc Natl Acad Sci U S A, 2000. 97(23): p. 12498-12502. 11. Daugeron, M.C., F. Mauxion, and B. Seraphin, The yeast POP2 gene encodes a nuclease involved in mRNA deadenylation. Nucleic Acids Res, 2001. 29(12): p. 2448-2455. 12. Eulalio, A., et al., P-body formation is a consequence, not the cause, of RNA-mediated gene silencing. Mol Cell Biol, 2007. 27(11): p. 3970-3981. 13. Tucker, M., et al., Ccr4p is the catalytic subunit of a Ccr4p/Pop2p/Notp mRNA deadenylase complex in Saccharomyces cerevisiae. Embo J, 2002. 21(6): p. 1427-1436. 14. Decker, C.J. and R. Parker, A turnover pathway for both stable and unstable mRNAs in yeast: evidence for a requirement for deadenylation. Genes Dev, 1993. 7(8): p. 1632-1643. 15. Wang, Z. and M. Kiledjian, Functional link between the mammalian exosome and mRNA decapping. Cell, 2001. 107(6): p. 751-762. 16. Liu, S.W., et al., Functional analysis of mRNA scavenger decapping enzymes. Rna, 2004. 10(9): p. 1412-1422. 17. Chen, C.Y., et al., AU binding proteins recruit the exosome to degrade ARE-containing mRNAs. Cell, 2001. 107(4): p. 451-464. 18. LaGrandeur, T.E. and R. Parker, Isolation and characterization of Dcp1p, the yeast mRNA decapping enzyme. Embo J, 1998. 17(5): p. 1487-1496. 19. Callebaut, I., An EVH1/WH1 domain as a key actor in TGFbeta signalling. FEBS Lett, 2002. 519(1-3): p. 178-180. 20. Bai, R.Y., et al., SMIF, a Smad4-interacting protein that functions as a co-activator in TGFbeta signalling. 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Kiledjian, Functional characterization of the mammalian mRNA decapping enzyme hDcp2. Rna, 2003. 9(9): p. 1138-1147. 27. Coller, J.M., et al., The DEAD box helicase, Dhh1p, functions in mRNA decapping and interacts with both the decapping and deadenylase complexes. Rna, 2001. 7(12): p. 1717-1727. 28. Tharun, S., et al., Yeast Sm-like proteins function in mRNA decapping and decay. Nature, 2000. 404(6777): p. 515-518. 29. Fillman, C. and J. Lykke-Andersen, RNA decapping inside and outside of processing bodies. Curr Opin Cell Biol, 2005. 17(3): p. 326-331. 30. Schwartz, D.C. and R. Parker, Mutations in translation initiation factors lead to increased rates of deadenylation and decapping of mRNAs in Saccharomyces cerevisiae. Mol Cell Biol, 1999. 19(8): p. 5247-5256. 31. LaGrandeur, T. and R. Parker, The cis acting sequences responsible for the differential decay of the unstable MFA2 and stable PGK1 transcripts in yeast include the context of the translational start codon. 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Lykke-Andersen, mRNA surveillance: the perfect persist. J Cell Sci, 2002. 115(Pt 15): p. 3033-3038. 39. Lykke-Andersen, J. and E. Wagner, Recruitment and activation of mRNA decay enzymes by two ARE-mediated decay activation domains in the proteins TTP and BRF-1. Genes Dev, 2005. 19(3): p. 351-361. 40. Ntambi, J.M. and K. Young-Cheul, Adipocyte differentiation and gene expression. J Nutr, 2000. 130(12): p. 3122S-3126S. 41. Patel, Y.M. and M.D. Lane, Mitotic clonal expansion during preadipocyte differentiation: calpain-mediated turnover of p27. J Biol Chem, 2000. 275(23): p. 17653-17660. 42. Lin, N.Y., C.T. Lin, and C.J. Chang, Modulation of immediate early gene expression by tristetraprolin in the differentiation of 3T3-L1 cells. Biochem Biophys Res Commun, 2008. 365(1): p. 69-74. 43. Tang, Q.Q., T.C. Otto, and M.D. Lane, Mitotic clonal expansion: a synchronous process required for adipogenesis. Proc Natl Acad Sci U S A, 2003. 100(1): p. 44-49. 44. Zhang, J.W., et al., Dominant-negative C/EBP disrupts mitotic clonal expansion and differentiation of 3T3-L1 preadipocytes. Proc Natl Acad Sci U S A, 2004. 101(1): p. 43-47. 45. Villen, J., et al., Large-scale phosphorylation analysis of mouse liver. Proc Natl Acad Sci U S A, 2007. 104(5): p. 1488-1493. 46. She, M., et al., Structural basis of dcp2 recognition and activation by dcp1. Mol Cell, 2008. 29(3): p. 337-349. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/41018 | - |
| dc.description.abstract | 訊息核醣核酸降解的調控在基因的表現中是非常重要的一環。由於在去除3’端的長poly(A)序列 (deadenylation)後，去頭蓋的動作會將標的訊息核醣核酸5’端的頭蓋移除，製造出m7GDP和在5’端帶有單一個磷酸，而會受到Xrn1進行5’到3’端的exonucleolytic降解的受質，所以，去頭蓋在訊息核醣核酸代謝過程扮演著相當重要的角色。去頭蓋蛋白質1 (Dcp1)和去頭蓋蛋白質2 (Dcp2)會形成去頭蓋複合物，當中並包含其他和降解相關的因子，像是Tristetraprolin (TTP)。TTP是在3T3-L1前脂肪細胞在受到誘發之後，所表現的立即早期基因，並會與人類去頭蓋蛋白質1a形成在ARE傳達降解路徑中作用的複合物。因此，本研究想要了解在前脂肪細胞的分化過程中，老鼠去頭蓋蛋白1a參與在有絲分裂細胞株落擴張的訊息核醣核酸降解(mitotic clonal expansion)的可能性。去頭蓋蛋白1a在3T3-L1早期分化時期會受到磷酸化，而這活性很可能是受到ERK訊息路徑的調控。老鼠去頭蓋蛋白1a主要會存在於細胞質中，並且它在細胞中的位置是不會受到磷酸化的影響的。而大量表現的去頭蓋蛋白1a和去頭蓋蛋白2 也被發現會聚集在細胞質中的特定裂解體中。在in vitro 去頭蓋的實驗，去頭蓋蛋白2展現強烈的去頭蓋活性，另一方面，去頭蓋蛋白1a表現了些微的去頭蓋活性。進一步的實驗發現，處於高度磷酸化狀態的去頭蓋蛋白1a在高劑量時，可以增強去頭蓋蛋白2的去頭蓋活性。這方面在之後會需要更多的分析來了解磷酸化在對於去頭蓋蛋白1a在3T3-L1細胞分化早期進行去頭蓋的動作或增進Dcp2之活性時所扮演的角色。 | zh_TW |
| dc.description.abstract | Regulation of mRNA decay is a critical step in gene expression. Decapping plays a major role in mRNA turnover as it removes the 5’ cap of target mRNA after deadenylation to generate m7GDP and a 5’ monophosphate substrate that is susceptible for 5’ to 3’exonucleolytic degradation by Xrn1. Dcp1p and Dcp2p were noted to form a decapping complex and associate with other mRNA decay regulators such as Tristetraprolin (TTP). TTP is an immediate early gene stimulated upon induction in 3T3-L1 preadipocytes, and was reported to precipitate with hDcp1a and the complex functioned in the ARE-mediated decay pathway. The potential role of mouse Dcp1a during mitotic clonal expansion in preadipocyte differentiation was thus studied. Dcp1a was phosphorylated during the early differentiation of 3T3-L1 cells, and was possibly regulated by the ERK signaling pathway. Mouse Dcp1a was localized mostly in the cytoplasm, and its position was not affected by its phosphorylation status. The colocalization of overexpressed Dcp1a and Dcp2 was also observed in cytoplasmic processing bodies. In the in vitro decapping assay, mouse Dcp2 displayed strong decapping activity while Dcp1a expressed partial decapping activity. In addition, highly phosphorylated Dcp1a appeared to enhance the catalytic activity of Dcp2 in a dosage dependent manner. Further analysis would be required to reveal the effect of phosphorylation on Dcp1a’s role in decapping and enhancing the activity of Dcp2 during early differentiation of 3T3-L1 cells. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-14T17:12:08Z (GMT). No. of bitstreams: 1 ntu-97-R95b46034-1.pdf: 932423 bytes, checksum: c56e8f6b22a23595ceeb7e2d9f386fdb (MD5) Previous issue date: 2008 | en |
| dc.description.tableofcontents | 中文摘要 i
Abstract iii Contents v Abbreviations vi I. INTRODUCTION 1 Overview of mRNA Decay Pathways 1 The mRNA Decapping 2 The mRNA Decapping Machinery 4 The Activation or Inhibition of Decapping by Various Protein Factors 7 Decapping Can Occur in Processing Bodies 10 Decapping is Involved in mRNA Surveillance Decay Pathways 11 II. MATERIALS AND METHODS 15 Plasmid Constructs 15 Cell Culture 16 Cell Extracts Preparation 17 SDS-PAGE and Western Blotting 18 Indirect immunofluorescence and confocal microscopy 19 In vitro transcription 20 In vitro decapping assay 20 III. Results 22 Dcp1a is phosphorylated during early differentiation of 3T3-L1 preadipocytes. 22 Phosphorylation of Dcp1a is possibly regulated by the ERK signaling pathway. 24 The localization of Mouse Dcp1a is not affected by its phosphorylation state. 26 Dcp1a is mostly distributed in the cytoplasm and it colocalizes with Dcp2 in specific cellular structures. 27 Mouse Dcp1a possess partial decapping activity which is independent of its phosphorylation status. 28 IV. Discussion 32 V. Figures 42 Figure 1. The expression profile of endogenous Dcp1a during 3T3-L1 early differentiation time course. 43 Figure 2. The phosphorylation state of Dcp1a is affected by ERK activity. 45 Figure 3. Subcellular localization of Dcp1a during 3T3-L1 early differentiation time course. 47 Figure 4. Dcp1a is concentrated in the processing bodies located in the cytoplasm 48 Figure 5. In vitro decapping assay performed with immunopurified recombinant Dcp1a and Dcp2. 50 VI. References 53 VII. Appendix 57 Appendix I. Endogenous mouse Dcp1a starts translation at second AUG initiation site 58 Appendix II. Coimmunoprecipitation of overexpressed Dcp1a and Dcp2. 64 Appendix III. In vitro decay assay with S100 extracts. 66 | |
| dc.language.iso | en | |
| dc.subject | 前脂肪細胞分化 | zh_TW |
| dc.subject | 去頭蓋 | zh_TW |
| dc.subject | 訊息核醣核酸降解 | zh_TW |
| dc.subject | 去頭蓋蛋白1a | zh_TW |
| dc.subject | 特定裂解體 | zh_TW |
| dc.subject | mRNA decay | en |
| dc.subject | preadipocyte | en |
| dc.subject | processing bodies | en |
| dc.subject | Dcp1a | en |
| dc.subject | decapping | en |
| dc.title | 訊息RNA去頭蓋蛋白質1a磷酸化之功能分析 | zh_TW |
| dc.title | Functional Characterization of Phosphorylation of Decapping Protein 1a (Dcp1a) | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 96-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 李芳仁,呂勝春,陳瑞華 | |
| dc.subject.keyword | 去頭蓋,訊息核醣核酸降解,去頭蓋蛋白1a,特定裂解體,前脂肪細胞分化, | zh_TW |
| dc.subject.keyword | decapping,mRNA decay,Dcp1a,processing bodies,preadipocyte, | en |
| dc.relation.page | 55 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2008-07-28 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 生化科學研究所 | zh_TW |
| 顯示於系所單位： | 生化科學研究所 | |
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