Iida Y, Senda T, Matsukawa Y, Onoda K, Miyazaki J I, Sakaguchi H, Nimura Y, Hidaka H, Niki I
Department of Pharmacology, Nagoya University School of Medicine, Japan.
Am J Physiol. 1997 Oct;273(4):E782-9. doi: 10.1152/ajpendo.1997.273.4.E782.
The aim of this study was to investigate how insulin secretion is controlled by phosphorylation of the myosin light chain (MLC). Ca2+-evoked insulin release from pancreatic islets permeabilized with streptolysin O was inhibited by different monoclonal antibodies against myosin light-chain kinase (MLCK) to an extent parallel to their inhibition of purified MLCK. Anti-MLCK antibody also inhibited insulin release caused by the stable GTP analog guanosine 5'-O-(3-thiodiphosphate), even at a substimulatory concentration (0.1 microM) of Ca2+. Free Ca2+ increased MLC peptide phosphorylation by beta-cell extracts in vitro. In contrast to the phosphorylation by purified MLCK or by calmodulin (CaM) kinase II, the activity partially remained with the beta-cell under nonstimulatory Ca2+ (0.1 microM) conditions. The MLCK inhibitor ML-9 inhibited the activity in the beta-cell with both substimulatory and stimulatory Ca2+, whereas KN-62, an inhibitor of CaM kinase II, only exerted an influence in the latter case. ML-9 decreased intracellular granule movement in MIN6 cells under basal and acetylcholine-stimulated conditions. We propose that MLC phosphorylation may modulate translocation of secretory granules, resulting in enhanced insulin secretion.
本研究的目的是探究肌球蛋白轻链（MLC）磷酸化如何控制胰岛素分泌。用链球菌溶血素O通透处理的胰岛中，Ca2+诱发的胰岛素释放受到不同抗肌球蛋白轻链激酶（MLCK）单克隆抗体的抑制，其抑制程度与它们对纯化的MLCK的抑制程度平行。抗MLCK抗体也抑制了由稳定的GTP类似物鸟苷5'-O-（3-硫代二磷酸）引起的胰岛素释放，即使在亚刺激浓度（0.1微摩尔）的Ca2+条件下也是如此。游离Ca2+在体外增加了β细胞提取物对MLC肽的磷酸化。与纯化的MLCK或钙调蛋白（CaM）激酶II的磷酸化作用不同，在非刺激的Ca2+（0.1微摩尔）条件下，该活性部分保留在β细胞中。MLCK抑制剂ML-9在亚刺激和刺激的Ca2+条件下均抑制β细胞中的活性，而CaM激酶II的抑制剂KN-62仅在后者情况下发挥作用。在基础和乙酰胆碱刺激条件下，ML-9减少了MIN6细胞内颗粒的移动。我们提出，MLC磷酸化可能调节分泌颗粒的转运，从而导致胰岛素分泌增加。
