Glucagon-like peptide 1 activates protein kinase C through Ca2+-dependent activation of phospholipase C in insulin-secreting cells
Glucagon-like peptide 1 activates protein kinase C through Ca2+-dependent activation of phospholipase C in insulin-secreting cells
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DOI：
10.1074/jbc.m604291200
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发表时间：
2006-09-29
影响因子：
4.8
通讯作者：
Mogami, Hideo
中科院分区：
文献类型：
--
作者：
Suzuki, Yuko;Zhang, Hui;Mogami, Hideo
虽然胰高血糖素样肽1（GLP 1），cAMP生成激动剂，对Ca 2+信号和胰岛素分泌的刺激作用是公认的，但其潜在的机制仍有待充分阐明。我们最近发现，Ca ~（2+）内流可以单独激活胰岛素分泌细胞（INS-1）中的常规蛋白激酶C（PKC）以及新型PKC。基于这一早期发现，在此我们研究了GLP-1诱导的Ca 2+信号传导是否可以在INS-1细胞中在亚刺激浓度的葡萄糖（3 mM）下激活PKC α和PKC β。我们首先发现GLP-1将内源性PKC α和PKC β从细胞质转运到质膜。接下来，我们通过使用MARCKS-GFP评估PKC底物，肉豆蔻酰化富含丙氨酸的C激酶底物（MARCKS）的磷酸化状态。GLP-1以Ca 2+依赖性方式将MARCKS-GFP转运至胞质溶胶，并且GLP-1诱发的MARCKS-GFP转运被PKC抑制剂阻断，所述PKC抑制剂为广泛的PKC抑制剂双吲哚马来酰亚胺I或PKC β抑制剂肽、与触角足肽融合的假底物PKC β-（149-164）（antp-PKC β）和常规PKC抑制剂Go-6976。此外，毛喉素诱导的MARCKS-GFP易位几乎完全被磷脂酶C抑制剂U 73122抑制。这些观察结果以两种不同的方式得到验证，证明1）毛喉素诱导的PKC γ的GFP标记的C1结构域的易位和2）PKC α-DsRed和PKC α-GFP的易位。此外，PKC抑制剂减少毛喉素诱导的INS-1细胞和大鼠胰岛的胰岛素分泌。因此，GLP-1可以激活PKC α和PKC β，这些GLP-1激活的PKC可能在亚刺激浓度的葡萄糖下显著促进胰岛素分泌。
Although the stimulatory effect of glucagon-like peptide 1(GLP1), a cAMP-generating agonist, on Ca2+ signal and insulin secretion is well established, the underlying mechanisms remain to be fully elucidated. We recently discovered that Ca2+ influx alone can activate conventional protein kinase C (PKC) as well as novel PKC in insulin-secreting (INS-1) cells. Building on this earlier finding, here we examined whether GLP-1-evoked Ca2+ signaling can activate PKC alpha and PKC epsilon at a substimulatory concentration of glucose (3 mM) in INS-1 cells. We first showed that GLP-1 translocated endogenous PKC alpha and PKC epsilon from the cytosol to the plasma membrane. Next, we assessed the phosphorylation state of the PKC substrate, myristoylated alanine-rich C kinase substrate ( MARCKS), by using MARCKS-GFP. GLP-1 translocated MARCKS-GFP to the cytosol in a Ca2+-dependent manner, and the GLP-1-evoked translocation of MARCKS-GFP was blocked by PKC inhibitors, either a broad PKC inhibitor, bisindolylmaleimide I, or a PKC epsilon inhibitor peptide, antennapedia peptide-fused pseudosubstrate PKC epsilon-(149-164) (antp-PKC epsilon) and a conventional PKC inhibitor, Go-6976. Furthermore, forskolin-induced translocation of MARCKS-GFP was almost completely inhibited by U73122, a putative inhibitor of phospholipase C. These observations were verified in two different ways by demonstrating 1) forskolin-induced translocation of the GFP-tagged C1 domain of PKC gamma and 2) translocation of PKC alpha-DsRed and PKC alpha-GFP. In addition, PKC inhibitors reduced forskolin-induced insulin secretion in both INS-1 cells and rat islets. Thus, GLP-1 can activate PKC alpha and PKC epsilon, and these GLP-1-activated PKCs may contribute considerably to insulin secretion at a substimulatory concentration of glucose.