Role of type Ialpha phosphatidylinositol-4-phosphate 5-kinase in insulin secretion, glucose metabolism, and membrane potential in INS-1 beta-cells.
Insulin secretion from beta-cells is regulated by a complex signaling network. Our earlier study has reported that Rac1 participates in glucose- and cAMP-induced insulin secretion probably via maintaining a functional actin structure for recruitment of insulin granules. Type Ialpha phosphatidylinositol-4-phosphate 5-kinase (PIP5K-Ialpha) is a downstream effector of Rac1 and a critical enzyme for synthesis of phosphatidylinositol-4,5-bisphosphate (PIP(2)). By using an RNA interference technique, PIP5K-Ialpha in INS-1 beta-cells could be specifically knocked down by 70-75%. PIP5K-Ialpha knockdown disrupted filamentous actin structure and caused changes in cell morphology. In addition, PIP(2) content in the plasma membrane was reduced and the glucose effect on PIP(2) was abolished but without affecting glucose-induced formation of inositol 1,4,5-trisphosphate. At basal conditions (2.8 mM glucose), PIP5K-Ialpha knockdown doubled insulin secretion, elevated glucose metabolic rate, depolarized resting membrane potential, and raised cytoplasmic free Ca(2+) levels ([Ca(2+)](i)). The total insulin release at high glucose was increased upon PIP5K-Ialpha knockdown. However, the percent increment of insulin secretion by high glucose and forskolin over the basal release was significantly reduced, an effect more apparent on the late phase of insulin secretion. Metabolism and [Ca(2+)](i) rises at high glucose were also attenuated in cells after PIP5K-Ialpha knockdown. In contrast, PIP5K-Ialpha knockdown had no effect on cell growth and viability. Taken together, our data suggest that PIP5K-Ialpha may play an important role in both the proximal and distal steps of signaling cascade for insulin secretion in beta-cells.
Zhang
Jiping
J
Cardiovascular Research Institute, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597.
Luo
Ruihua
R
Wu
Heqing
H
Wei
Shunhui
S
Han
Weiping
W
Li
Guodong
G
D000199:Actins / Q000378:metabolism; D000818:Animals; D016923:Cell Death / Q000187:drug effects / Q000235:genetics; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D048430:Cell Shape; D055785:Gene Knockdown Techniques; D005947:Glucose / Q000378:metabolism* / Q000494:pharmacology; D007328:Insulin / Q000378:metabolism*; D000078790:Insulin Secretion; D050417:Insulin-Secreting Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism / Q000502:physiology*; D008564:Membrane Potentials / Q000235:genetics*; D051379:Mice; D019269:Phosphatidylinositol 4,5-Diphosphate / Q000378:metabolism; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000037:antagonists & inhibitors / Q000235:genetics / Q000502:physiology*; D034741:RNA, Small Interfering / Q000235:genetics / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects / Q000235:genetics; D014162:Transfection
D000199:Actins; D007328:Insulin; D019269:Phosphatidylinositol 4,5-Diphosphate; D034741:RNA, Small Interfering; D017853:Phosphotransferases (Alcohol Group Acceptor); C020077:1-phosphatidylinositol-4-phosphate 5-kinase; D005947:Glucose