Luo Moulun, Langlais Paul, Yi Zhengping, Lefort Natalie, De Filippis Elena A, Hwang Hyonson, Christ-Roberts Christine Y, Mandarino Lawrence J
Center for Metabolic Biology, College of Liberal Arts and Sciences, Tempe, AZ 85287-3704, USA.
Endocrinology. 2007 Oct;148(10):4895-905. doi: 10.1210/en.2007-0049. Epub 2007 Jul 19.
The function of insulin receptor substrate-1 (IRS-1) is regulated by both tyrosine and serine/threonine phosphorylation. Phosphorylation of some serine/threonine residues in IRS-1 dampens insulin signaling, whereas phosphorylation of other serine/threonine residues enhances insulin signaling. Phosphorylation of human IRS-1 at Ser(629) was increased by insulin in Chinese hamster ovary cells expressing the insulin receptor (1.26 +/- 0.09-fold; P < 0.05) and L6 cells (1.35 +/- 0.29-fold; P < 0.05) expressing human IRS-1. Sequence analysis surrounding Ser(629) revealed conformity to the consensus phosphorylation sequence recognized by Akt. Phosphorylation of IRS-1 at Ser(629) in cells was decreased upon treatment with either an Akt inhibitor or by coexpression with kinase dead Akt, whereas Ser(629) phosphorylation was increased by coexpression with constitutively active Akt. In addition, Ser(629) of IRS-1 is directly phosphorylated by Akt in vitro. In cells, preventing phosphorylation of Ser(629) by a Ser(629)Ala mutation resulted in increased phosphorylation of Ser(636), a known negative regulator of IRS-1, without affecting phosphorylation of Tyr(632) or Ser(616). Cells expressing the Ser(629)Ala mutation, along with increased Ser(636) phosphorylation, had decreased insulin-stimulated association of the p85 regulatory subunit of phosphatidylinositol 3'-kinase with IRS-1 and decreased phosphorylation of Akt at Ser(473). Finally, in vitro phosphorylation of a Ser(629)-containing IRS-1 fragment with Akt reduces the subsequent ability of ERK to phosphorylate Ser(636/639). These results suggest that a feed-forward mechanism may exist whereby insulin activation of Akt leads to phosphorylation of IRS-1 at Ser(629), resulting in decreased phosphorylation of IRS-1 at Ser(636) and enhanced downstream signaling. Understanding the complex phosphorylation patterns of IRS-1 is crucial to elucidating the factors contributing to insulin resistance and, ultimately, the pathogenesis of type 2 diabetes.
胰岛素受体底物-1（IRS-1）的功能受酪氨酸和丝氨酸/苏氨酸磷酸化的调节。IRS-1中某些丝氨酸/苏氨酸残基的磷酸化会减弱胰岛素信号传导，而其他丝氨酸/苏氨酸残基的磷酸化则会增强胰岛素信号传导。在表达胰岛素受体的中国仓鼠卵巢细胞（1.26±0.09倍；P<0.05）和表达人IRS-1的L6细胞（1.35±0.29倍；P<0.05）中，胰岛素可增加人IRS-1在Ser(629)位点的磷酸化。围绕Ser(629)的序列分析显示其符合Akt识别的共有磷酸化序列。用Akt抑制剂处理细胞或与激酶失活的Akt共表达时，细胞中IRS-1在Ser(629)位点的磷酸化会降低，而与组成型活性Akt共表达则会增加Ser(629)的磷酸化。此外，IRS-1的Ser(629)在体外可被Akt直接磷酸化。在细胞中，通过Ser(629)Ala突变阻止Ser(629)的磷酸化会导致IRS-1的已知负调节因子Ser(636)的磷酸化增加，而不影响Tyr(632)或Ser(616)的磷酸化。表达Ser(629)Ala突变的细胞，随着Ser(636)磷酸化的增加，胰岛素刺激的磷脂酰肌醇3'-激酶p85调节亚基与IRS-1的结合减少，Akt在Ser(473)位点的磷酸化也减少。最后，用Akt对含Ser(629)的IRS-1片段进行体外磷酸化会降低ERK随后对Ser(636/639)磷酸化的能力。这些结果表明可能存在一种前馈机制，即胰岛素激活Akt导致IRS-1在Ser(629)位点磷酸化，从而导致IRS-1在Ser(636)位点磷酸化减少并增强下游信号传导。了解IRS-1复杂的磷酸化模式对于阐明导致胰岛素抵抗以及最终2型糖尿病发病机制的因素至关重要。
