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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 呂勝春 | |
| dc.contributor.author | Chia-Wei Lee | en |
| dc.contributor.author | 李家瑋 | zh_TW |
| dc.date.accessioned | 2021-06-17T00:40:43Z | - |
| dc.date.available | 2017-03-02 | |
| dc.date.copyright | 2012-03-02 | |
| dc.date.issued | 2012 | |
| dc.date.submitted | 2012-01-18 | |
| dc.identifier.citation | Ahmed, A.A., Lu, Z., Jennings, N.B., Etemadmoghadam, D., Capalbo, L., Jacamo, R.O., Barbosa-Morais, N., Le, X.F., Vivas-Mejia, P., Lopez-Berestein, G., et al. (2010). SIK2 is a centrosome kinase required for bipolar mitotic spindle formation that provides a potential target for therapy in ovarian cancer. Cancer cell 18, 109-121.
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Nature 449, 366-369. Horike, N., Kumagai, A., Shimono, Y., Onishi, T., Itoh, Y., Sasaki, T., Kitagawa, K., Hatano, O., Takagi, H., Susumu, T., et al. (2010). Downregulation of SIK2 expression promotes the melanogenic program in mice. Pigment cell & melanoma research 23, 809-819. Horike, N., Takemori, H., Katoh, Y., Doi, J., Min, L., Asano, T., Sun, X.J., Yamamoto, H., Kasayama, S., Muraoka, M., et al. (2003). Adipose-specific expression, phosphorylation of Ser794 in insulin receptor substrate-1, and activation in diabetic animals of salt-inducible kinase-2. The Journal of biological chemistry 278, 18440-18447. Janssens, V., and Goris, J. (2001). Protein phosphatase 2A: a highly regulated family of serine/threonine phosphatases implicated in cell growth and signalling. The Biochemical journal 353, 417-439. Kowluru, A. (2005). Novel regulatory roles for protein phosphatase-2A in the islet beta cell. Biochemical pharmacology 69, 1681-1691. Leulliot, N., Quevillon-Cheruel, S., Sorel, I., Li de La Sierra-Gallay, I., Collinet, B., Graille, M., Blondeau, K., Bettache, N., Poupon, A., Janin, J., et al. (2004). Structure of protein phosphatase methyltransferase 1 (PPM1), a leucine carboxyl methyltransferase involved in the regulation of protein phosphatase 2A activity. The Journal of biological chemistry 279, 8351-8358. Li, X., Scuderi, A., Letsou, A., and Virshup, D.M. (2002). B56-associated protein phosphatase 2A is required for survival and protects from apoptosis in Drosophila melanogaster. Molecular and cellular biology 22, 3674-3684. Li, X., and Virshup, D.M. (2002). Two conserved domains in regulatory B subunits mediate binding to the A subunit of protein phosphatase 2A. European journal of biochemistry / FEBS 269, 546-552. Li, Y., Wei, H., Hsieh, T.C., and Pallas, D.C. (2008). Cdc55p-mediated E4orf4 growth inhibition in Saccharomyces cerevisiae is mediated only in part via the catalytic subunit of protein phosphatase 2A. Journal of virology 82, 3612-3623. Longin, S., Zwaenepoel, K., Louis, J.V., Dilworth, S., Goris, J., and Janssens, V. (2007). Selection of protein phosphatase 2A regulatory subunits is mediated by the C terminus of the catalytic Subunit. The Journal of biological chemistry 282, 26971-26980. Muraoka, M., Fukushima, A., Viengchareun, S., Lombes, M., Kishi, F., Miyauchi, A., Kanematsu, M., Doi, J., Kajimura, J., Nakai, R., et al. (2009). Involvement of SIK2/TORC2 signaling cascade in the regulation of insulin-induced PGC-1alpha and UCP-1 gene expression in brown adipocytes. American journal of physiology Endocrinology and metabolism 296, E1430-1439. Ogris, E., Du, X., Nelson, K.C., Mak, E.K., Yu, X.X., Lane, W.S., and Pallas, D.C. (1999). A protein phosphatase methylesterase (PME-1) is one of several novel proteins stably associating with two inactive mutants of protein phosphatase 2A. The Journal of biological chemistry 274, 14382-14391. Ortega-Gutierrez, S., Leung, D., Ficarro, S., Peters, E.C., and Cravatt, B.F. (2008). Targeted disruption of the PME-1 gene causes loss of demethylated PP2A and perinatal lethality in mice. PloS one 3, e2486. Palanivel, R., Veluthakal, R., and Kowluru, A. (2004). Regulation by glucose and calcium of the carboxylmethylation of the catalytic subunit of protein phosphatase 2A in insulin-secreting INS-1 cells. American journal of physiology Endocrinology and metabolism 286, E1032-1041. Puustinen, P., Junttila, M.R., Vanhatupa, S., Sablina, A.A., Hector, M.E., Teittinen, K., Raheem, O., Ketola, K., Lin, S., Kast, J., et al. (2009). PME-1 protects extracellular signal-regulated kinase pathway activity from protein phosphatase 2A-mediated inactivation in human malignant glioma. Cancer research 69, 2870-2877. Sablina, A.A., Chen, W., Arroyo, J.D., Corral, L., Hector, M., Bulmer, S.E., DeCaprio, J.A., and Hahn, W.C. (2007). The tumor suppressor PP2A Abeta regulates the RalA GTPase. Cell 129, 969-982. Sasaki, T., Takemori, H., Yagita, Y., Terasaki, Y., Uebi, T., Horike, N., Takagi, H., Susumu, T., Teraoka, H., Kusano, K., et al. (2011). SIK2 is a key regulator for neuronal survival after ischemia via TORC1-CREB. Neuron 69, 106-119. Schonthal, A.H. (1998). Role of PP2A in intracellular signal transduction pathways. Frontiers in bioscience : a journal and virtual library 3, D1262-1273. Screaton, R.A., Conkright, M.D., Katoh, Y., Best, J.L., Canettieri, G., Jeffries, S., Guzman, E., Niessen, S., Yates, J.R., 3rd, Takemori, H., et al. (2004). The CREB coactivator TORC2 functions as a calcium- and cAMP-sensitive coincidence detector. Cell 119, 61-74. Silverstein, A.M., Barrow, C.A., Davis, A.J., and Mumby, M.C. (2002). Actions of PP2A on the MAP kinase pathway and apoptosis are mediated by distinct regulatory subunits. Proceedings of the National Academy of Sciences of the United States of America 99, 4221-4226. Sjostrom, M., Stenstrom, K., Eneling, K., Zwiller, J., Katz, A.I., Takemori, H., and Bertorello, A.M. (2007). SIK1 is part of a cell sodium-sensing network that regulates active sodium transport through a calcium-dependent process. Proceedings of the National Academy of Sciences of the United States of America 104, 16922-16927. Sontag, E. (2001). Protein phosphatase 2A: the Trojan Horse of cellular signaling. Cellular signalling 13, 7-16. Sontag, E., Hladik, C., Montgomery, L., Luangpirom, A., Mudrak, I., Ogris, E., and White, C.L., 3rd (2004). Downregulation of protein phosphatase 2A carboxyl methylation and methyltransferase may contribute to Alzheimer disease pathogenesis. Journal of neuropathology and experimental neurology 63, 1080-1091. Stanevich, V., Jiang, L., Satyshur, K.A., Li, Y., Jeffrey, P.D., Li, Z., Menden, P., Semmelhack, M.F., and Xing, Y. (2011). The structural basis for tight control of PP2A methylation and function by LCMT-1. Molecular cell 41, 331-342. Tolstykh, T., Lee, J., Vafai, S., and Stock, J.B. (2000). Carboxyl methylation regulates phosphoprotein phosphatase 2A by controlling the association of regulatory B subunits. The EMBO journal 19, 5682-5691. Vafai, S.B., and Stock, J.B. (2002). Protein phosphatase 2A methylation: a link between elevated plasma homocysteine and Alzheimer's Disease. FEBS letters 518, 1-4. Wang, Z., Takemori, H., Halder, S.K., Nonaka, Y., and Okamoto, M. (1999). Cloning of a novel kinase (SIK) of the SNF1/AMPK family from high salt diet-treated rat adrenal. FEBS letters 453, 135-139. Wu, J., Tolstykh, T., Lee, J., Boyd, K., Stock, J.B., and Broach, J.R. (2000). Carboxyl methylation of the phosphoprotein phosphatase 2A catalytic subunit promotes its functional association with regulatory subunits in vivo. The EMBO journal 19, 5672-5681. Xing, Y., Li, Z., Chen, Y., Stock, J.B., Jeffrey, P.D., and Shi, Y. (2008). Structural mechanism of demethylation and inactivation of protein phosphatase 2A. Cell 133, 154-163. Zhou, X.W., Gustafsson, J.A., Tanila, H., Bjorkdahl, C., Liu, R., Winblad, B., and Pei, J.J. (2008). Tau hyperphosphorylation correlates with reduced methylation of protein phosphatase 2A. Neurobiology of disease 31, 386-394. | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/66528 | - |
| dc.description.abstract | SIK2是屬於AMPK家族，SIK次家族的一員。AMPK家族的成員主要功能在於偵測細胞能量狀態，對於細胞凋亡以及自噬作用的逆境反應。在AMPK家族中，SIK2是唯一可以與PP2A及VCP/P97有交互作用的成員。然而，對於SIK2-PP2A複合體的功能仍然不清楚。在癌細胞中，SIK2的降解在泛素蛋白質體降解系統及自噬作用中扮演重要角色。SIK2 knockdown會造成PP2Ac量的下降，並且會增加Bcl-2/S70的磷酸化。我發現PME-1/S15的磷酸化在SIK2 knockdown時會上升。在之前的研究中，調節PP2A次單元B的招募對於 PP2A受質特異性及活性是相當重要的步驟。LCMT-1對於PP2Ac次單元的羧基末端白胺酸的羧基甲基化和PP2A之活化是必需的酵素。PME-1可和PP2A結和並抑制其活性，是移除PP2Ac/L309甲基的羧基甲基化酯酶。我發現PME-1會被排除於SIK2-PP2A複合體之外，進而能保存PP2A的活性。同時在SIK2 knockdown下CaMK1會被活化並且直接磷酸化PME-1/S15。正常狀態下，SIK2會藉由PP2A負調控 CaMK1的活性。意外的是，SIK2-KD同樣可以形成SIK2-PP2A複合體，因此可以保存PP2A的活性。綜合以上結果，SIK2可能扮演adaptor的角色，藉由排除PME-1，因而保存PP2A的活性。在泛素蛋白質體降解系統及自噬作用進行的過程中，鈣離子依賴活化的CaMK1以及磷酸化PME-1/S15會受到PP2A的回饋抑制。以上結果顯示，SIK2透過維持PP2A活性及Bcl-2的量在ER恆定的過程中扮演重要的角色。 | zh_TW |
| dc.description.abstract | Salt-inducible kinase 2 (SIK2) is a member of AMPK family, SIK subfamily. Members of AMPK family play wide array of functions ranging from sensing energy status, stress response to apoptosis and autophagy. SIK2 is the only member of AMPK family that can interact with PP2A and VCP/P97. However, the function of this SIK2-PP2A complex is largely unknown. It was demonstrated in cancer cells that SIK2 is important protein degradation by both UPS and autophagy. SIK2 knockdown resulted in decreasing both levels of PP2Ac and carboxymethylated PP2Ac (PP2Ac/L309Me). SIK2 knockdown also resulted in increasing the phosphorylation of Bcl-2/S70. I identified the phosphorylation of PME-1/S15 was positively regulated by knockdown of SIK2. The enzymatic activity of PP2A may be regulated by heterotrimer formation in which recruiting the substrate-specific regulatory B subunit is believed to be the crucial step. A novel PP2Ac subunit C-terminal Leucine modification by LCMT1-mediated carboxymethylation is also essential for the catalytic activity of PP2A. PME-1, a carboxymethylesterase specific for the removal of PP2Ac/L309Me could complex with PP2Ac and inhibits its activity. I have also uncovered that SIK2-PP2A complex preserves the phosphatase activity. PME-1 is excluded from that complex. I also showed that SIK2 knockdown resulted in the activation of CaMK1 and phosphorylation of PME-1/S15. It turned out that the activity of CaMK1 is negatively regulated by PP2A in a SIK2-dependent manner. I have demonstrated that PME-1/S15 is the target of CaMK1. Unexpectedly, kinase-dead mutant of SIK2 (i.e., SIK2/K49M) could also form SIK2/K49M-PP2A complex and preserve the PP2A activity. Together, these results suggest that one of the functions of SIK2 may be an adaptor. PP2A activity is protected in the SIK2-PP2A complex from the inhibition of PME-1. The Ca2+-dependent activation of CaMK1 during UPS- or autophagosome-induction and phosphorylation of PME-1/S15 may be regulated by PP2A-dependent feedback inhibition. Taken together, these findings suggest that SIK2 is an important regulator for ER homeostasis through maintaining PP2A activity and the level of antiapoptotic protein Bcl-2 in the ER membrane. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-17T00:40:43Z (GMT). No. of bitstreams: 1 ntu-101-R98448013-1.pdf: 1711848 bytes, checksum: 568a56a66191943ea718514c272f4fa5 (MD5) Previous issue date: 2012 | en |
| dc.description.tableofcontents | Master Thesis ……………………………………………………………….…………..i
中文摘要 ……………………………………………………..…….....……………....ii ABSTRACT …………………………………………………...…...…………….…..iii CONTENTS ………………………………………………………...………...………..v INTRODUCTION ……………………………………………………...…...………....1 MATERIALS AND METHODS ……………………………….……………………..6 Plasmids and Mutagenesis ……………………………………………….…....6 Cell Culture and Transfection ………………………………………………....6 Drugs and Antibodies ………………………………………………….……....7 Immunoprecipitation and Western Blotting ………………….………………7 Protein purification and CaMK1 in vitro kinase assay ………………………8 Preparation of microsomal fraction …………………………..………...…….9 In vitro PP2A phosphatase assay ………………..………………….…………9 RESULTS …………………………………………………………………..………….11 Characterization of antibodies …………..………………………….………..11 SIK2-PP2A complex formation and phosphatase activity …………..…..…11 SIK2 is required for ER homeostasis of cancer cells ……………...………..12 SIK2-dependent negative regulation of CaMK1 …………..………………..13 Feedback regulation of CaMK1 and PME-1/S15 phosphorylation by SIK2-PP2A complex ………………………………………...………………..15 DISCUSSION ………………………………………………………………..………..16 REFERENCES …………………………………………………..……………….…..21 FIGURES AND TABLES …………………………………………….……………..29 Fig. 1. Characterization of phosphor-specific antibodies for PME-1/S15 and PME-1/S243. ……………………………………………………….…………29 Fig. 2. Formation of SIK2-PP2A complex was independent of SIK2 activity. .…………………………………………………...………………..…30 Fig. 3. Depletion of SIK2 resulted in a reduction of both PP2Ac and carboxymethylated PP2Ac (PP2Ac/L309Me) levels. .………………….…...31 Fig. 4. SIK2 interacts with p97/VCP and PP2A, but not PME-1. …….…...32 Fig. 5. PP2A activity is preserved in SIK2-PP2A complex. ………...….…...33 Fig. 6. PP2A activity is required for SIK2 interaction. …………………..…35 Fig. 7. PP2A activity is required for preserving SIK2 activity. .………....…36 Fig. 8. Reduced PP2A activity and SIK2 level resulted in decreased BCL-2 in the microsomal fraction. ……………………………….………….………37 Fig. 9. SIK2 regulates the phosphorylation of PME-1/S15, but not PME-1/S243. ………………………………………….………………….……39 Fig. 10. CaMK1 activity is negatively regulated by PP2A. ………….......…40 Fig. 11. Phosphorylated PME-1/S15 is dephosphorylated by PP2A. ...…....41 Fig. 12. CaMK1, but not CaMK2, is responsible for phosphorylation of PME-1/S15. ……………………...……………….…………………......…..…42 Fig. 13. Reduced SIK2 level resulted in CaMK1 activation. .……….……...44 Fig. 14. Feedback regulation of CaMK1/T177 and PME-1/S15 phosphorylation by PP2A. .………………………………………………..…45 Fig. 15. Phosphorylation of PME-1/S15 enhances its association with PP2A. ………………………………………………………………..……....…47 Fig. 16. SIK2 level is negatively regulated by CaMK1. .………….…...……48 Fig. 17. Cross-regulation of SIK2-PP2A complex and CaMK1-PME-1 axis. .……………...………………………………………….……....…………49 Table.1. Supporting Primer List ……………………..………….………...…50 | |
| dc.language.iso | en | |
| dc.subject | 第二型鹽誘導激酶 | zh_TW |
| dc.subject | 內質網逆境 | zh_TW |
| dc.subject | 細胞凋亡 | zh_TW |
| dc.subject | 蛋白質磷酸酶 | zh_TW |
| dc.subject | 2A | zh_TW |
| dc.subject | 磷酸酶 | zh_TW |
| dc.subject | B細胞淋巴瘤-2 | zh_TW |
| dc.subject | 甲酯酶 | zh_TW |
| dc.subject | PP2A | en |
| dc.subject | SIK2 | en |
| dc.subject | apoptosis | en |
| dc.subject | BcL-2 | en |
| dc.subject | CaMK1 | en |
| dc.subject | ER-stress | en |
| dc.subject | PME-1 | en |
| dc.title | SIK2與PP2A功能性交互作用之探討 | zh_TW |
| dc.title | Functional interaction between SIK2 and PP2A | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 100-1 | |
| dc.description.degree | 碩士 | |
| dc.contributor.oralexamcommittee | 陳瑞華,李芳仁 | |
| dc.subject.keyword | 第二型鹽誘導激酶,蛋白質磷酸酶,2A,磷酸酶,甲酯酶,-1,內質網逆境,B細胞淋巴瘤-2,細胞凋亡, | zh_TW |
| dc.subject.keyword | SIK2,PP2A,PME-1,ER-stress,CaMK1,BcL-2,apoptosis, | en |
| dc.relation.page | 52 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2012-01-19 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 分子醫學研究所 | zh_TW |
| 顯示於系所單位： | 分子醫學研究所 | |
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