磷酸化對維生素D受體轉錄功能的影響

Effect of phosphorylation on the transcriptional activity of vitamin D3 receptor

鄭雪岑(Hsueh-Tsen Cheng)

指導教授 ： 洪文俊 ； 黃友利

高雄醫學大學/健康科學院/公共衛生學研究所/碩士(2005年)

摘要

蛋白質的磷酸化，是經由蛋白質激酶(protein kinases)，在tyrosine、serine或threonine作用。根據文獻指出，荷爾蒙接受器的磷酸化，與調控DNA結合、蛋白質間作用、核內位置、基因活化有關。在核接受器(nuclear receptor) superfamily中，像progesterone receptor、glucocorticoid receptor、thyroid hormone receptor、estrogen receptor、androgen receptor和vitamin D receptor都被認為是phosphoprotein。
荷爾蒙 1α,25-dihydroxyvitamin D3 (vitamin D3)對於細胞生理機能的調控佔有重要的地位，如血液與骨頭中鈣和磷濃度的恆定、細胞生長及分化、細胞凋亡等。Vitamin D3藉由與VDR (vitamin D3 receptor)的結合，促進VDR與RXR (retinoid X receptor)形成heterodimer複合物，進而去活化或抑制目標基因的表現，影響目標基因轉錄、轉譯的活性。
Osteocalcin(OC)被造骨細胞(osteoblast)合成並分泌至血清中，可用來當作是骨骼合成的指標。文獻指出OC可被維生素D藉著VDR-RXR複合體來調控，而且VDR是一個phosphoprotein，故本論文藉著單點突變法將VDR上可被磷酸化的位置突變後進一步探討VDR上的磷酸化作用是如何影響VDR-RXR複合體及其他co-factor而對OC基因進行調控。另外也將探討AF2 domain對於osteocalcin基因轉錄調控的重要性。

關鍵字

磷酸化 ； 維生素D受體

並列摘要

Proteins can be phosphorylated at serine ,tyrosine ,threonine residues by protein kinases. Hormone dependent phosphorylation of nuclear receptor has been reported and may be involved in regulation of DNA binding, protein-protein interaction, nuclear localization and gene transactivation. Most members of the steroid hormone receptor superfamily, including progesterone receptor、glucocorticoid receptor、thyroid hormone receptor、estrogen receptor、androgen receptor and vitamin D receptor, have been shown to be phosphoprotein.
The steroid hormone 1α,25-dihydroxyvitamin D3 (vitamin D3) plays an important role in the regulation of numerous physiological and cellular processes including calcium and phosphorous homeostasis, cell growth, differentiation and apoptosis. The biological effect of vitamin D3 is mainly mediated via the vitamin D3 receptor (VDR). Binding of vitamin D3 to VDR promotes the interaction between VDR and retinoid X receptor (RXR) and enhances the formation of VDR/RXR heterodimer. The heterodimer then attaches to the vitamin D3 response element (VDRE) in the promoter of target genes to trigger or repress gene transcription.
Osteocalcin is synthesized and secreted by the osteoblast and whose concentration in serum has been increasingly used as an index of bone formation. Previous report shows that vitamin D could regulate osteocalcin by VDR-RXR complex and vitamin D receptor was a phophoprotein. In this thesis ,we use site-mutagenesis to create serine-mutated VDR constructs. Further, we define how the phosphorylation of VDR affect VDR-RXR complex or other co-factors to regulate the expression of osteocalcin. Besides, we will study the importance of AF2 domain in the transcriptional regulation of Osteocalcin gene.

並列關鍵字

phosphorylation ； VDR

參考文獻

Reference List

1. R.Lin and J.H.White, The pleiotropic actions of vitamin D, Bioessays 26 (2004) 21.

2. L.L.Issa, G.M.Leong, and J.A.Eisman, Molecular mechanism of vitamin D receptor action, Inflamm. Res. 47 (1998) 451.

3. T.C.Chen and M.F.Holick, Vitamin D and prostate cancer prevention and treatment, Trends Endocrinol. Metab 14 (2003) 423.

4. J.C.Hsieh, P.W.Jurutka, M.A.Galligan, C.M.Terpening, C.A.Haussler, D.S.Samuels, Y.Shimizu, N.Shimizu, and M.R.Haussler, Human vitamin D receptor is selectively phosphorylated by protein kinase C on serine 51, a residue crucial to its trans-activation function, Proc. Natl. Acad. Sci. U. S. A 88 (1991) 9315.