STUDIES ON REGULATING MECHANISMS OF INTRACELLULAR MOVEMENT OF THE INSULIN GRANULES

在胞吐作用之前，分泌颗粒被合成，携带在细胞质中，并分布在质膜附近。在胰腺β细胞中，胞吐上游的这些调节步骤主要是基于测量释放到细胞外空间的胰岛素来讨论的，并且很少有论文发表在这些胞吐前步骤的直接测量和定量上。我们的研究得到了该基金的支持，证明了前胞吐步骤对胰岛素释放的调节至关重要，并证明了胰岛素释放的一些控制机制; 1）胰岛素颗粒的细胞内运动不具有如对于神经元细胞中的突触颗粒所报道的任何固定方向，2）肌球蛋白II（最近被其他人报道为肌球蛋白IIA）是胰岛素颗粒的运动蛋白，3）胰岛素颗粒的细胞内运动被来自内部储存的动员的Ca 2+激活（4）通过肌球蛋白轻链激酶对肌球蛋白轻链的磷酸化和蛋白激酶A对未知底物的磷酸化激活颗粒。此外，我们发现TPA反应性蛋白激酶C负责颗粒与质膜的对接。我们的研究表明，第二信使和由此产生的蛋白质磷酸化在胰岛素分泌级联反应中起着分工作用。这有助于研究胰岛素通过葡萄糖的阶段性释放，这在2型糖尿病中受损。

Prior to exocytosis, the secretory granules are synthesized, carried in the cytoplasm, and distributed in the vicinity of the plasma membrane. In the pancreatic beta cell, these regulatory steps upstream exocytosis have been mainly discussed on the basis of the measurement of insulin released into the extracellular space, and very few papers have been published on the direct measurement and quantification of these pre-exocytotic steps. Our study supported by this grant, demonstrated that the preexocytotic steps are crucial for the regulation of insulin release and, we demonstrated some controlling mechanisms characteristic for insulin release ; 1) intracellular movement of the insulin granules do not have any fixed direction as reported for the synaptic granules in neuronal cells, 2 ) myosin II (very recently reported to be myosin IIA by others) is a motor protein of the insulin granules, 3) intracellular movement of the insulin granule is activated by mobilized Ca2+ from internal stores (not Ca2+ influx) and cAMP, 4) the activation is via phosphorylation of myosin light chain by myosin light chain kinase, and via that of unknown substrate by protein kinase A.Furthermore, we found TPA-responsive protein kinase C is responsible for the granule docking to the plasma membrane. Our study demonstrated that second messengers and resultant protein phosphorylation are dividing works in the insulin secretory cascade. This contributes to investigation of phasic release of insulin by glucose, which is impaired in type 2 diabetes mellitus.