鈣離子結合蛋-1對調控興奮性細胞刺激-分泌耦合機轉之研究

Modulation of Stimulus-secretion Coupling by Calcium Binding protein-1 in Excitable Cells

陳明靈(Ming-Ling Chen)

指導教授 ： 潘建源

國立臺灣大學/生命科學院/動物學研究研究所/碩士(2006年)

https://doi.org/10.6342/NTU.2006.01934

摘要

鈣調素(calmodulin)是細胞中常見的鈣離子結合蛋白，調控細胞內許多重要的生理機制。而近幾年發現有一鈣離子結合蛋白家族，在結構上與鈣調素類似，且主要表現在神經細胞中。CaBP1因為alternative splicing，而有L-CaBP1和S-CaBP1之分。在本實驗中我們以電生理的技術，探討鈣離子結合蛋白-1 (CaBP1)在興奮性細胞中所扮演的功能。首先由大鼠E14.5胚胎大腦中選殖出CaBP1在其C端接上EYFP，再轉殖入初級培養的牛嗜鉻細胞及胎鼠大腦神經細胞中。結果顯示，在嗜鉻細胞中CaBP1主要分布在細胞膜上，且會抑制鈣離子電流及胞吐作用。而無法進行荳蔻酸化(myristoylation)修飾的G2A變異型，則主要分布在細胞質中，且無此抑制作用。而在神經細胞中，CaBP1對鈣離子電流，並無顯著的抑制作用。但當以高鉀溶液刺激這兩種細胞，鈣離子濃度上升都會因CaBP1大量表現而被抑制，但G2A變異型則無此作用。這些結果顯示，CaBP1需要在膜上才能發揮其對鈣離子通道及胞吐作用的抑制功能。

關鍵字

鈣離子通道 ； 胞吐作用 ； 鈣離子結合蛋白1 ； 膜片箝制

並列摘要

Calmodulin is a calcium binding protein ubiquitously expressed in cells and modulates many physiological activities. Recently, a new family of calcium binding proteins was found to have similar structure to calmodulin. In this report, the functions of a member of this family, calcium binding protein-1 (CaBP1), in modulating the stimulus-secretion coupling in excitable cells was characterized. Alternative splicing L- and S-CaBP1 were cloned from rat E14.5 embryonic cortical neurons. When expressed in primary cultured bovine chromaffin and embryonic neurons, both L- and S-CaBP1-EYFP fusion proteins were found to be localized on the plasma membrane but the G2A mutants, deficient in myristoylation, was in cytosol. The changes in calcium currents and capacitance evoked by membrane depolarization were inhibited by the expression of wild type L- and S-CaBP1 but not G2A mutants. The Ca2+ imaging recorded form cells stimulated by high K+ buffer also showed comparable results. These suggest that overexpression of CaBP1s in chromaffin cells and neurons inhibit calcium currents and exocytosis. These suggest the importance of CaBP1 in modulating the stimulus-secretion coupling in excitable cells.

並列關鍵字

calcium channel ； exocytosis ； CaBP1 ； patch clamp

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