Federici M, Hribal M L, Ranalli M, Marselli L, Porzio O, Lauro D, Borboni P, Lauro R, Marchetti P, Melino G, Sesti G
Laboratory of Molecular Medicine, Department of Internal Medicine, University of Rome-'Tor Vergata', 00133 Rome, Italy.
FASEB J. 2001 Jan;15(1):22-24. doi: 10.1096/fj.00-0414fje. Epub 2000 Nov 14.
Molecular scanning of human IRS-1 gene revealed a common polymorphism causing Gly-->Arg972 change. Diabetic and pre-diabetic carriers of Arg972 IRS-1 are characterized by low fasting levels of insulin and C-peptide. To investigate directly whether the Arg 972 IRS-1 affects human islet cells survival, we took advantage of the unique opportunity to analyze pancreatic islets isolated from three donors heterozygous for the Arg972 and six donors carrying wild-type IRS-1. Islets from carriers of Arg972 IRS-1 showed a two-fold increase in the number of apoptotic cells as compared with wild-type. IRS-1-associated PI3-kinase activity was decreased in islets from carriers of Arg972 IRS-1. Same results were reproduced in RIN rat b-cell lines stably expressing wild-type IRS-1 or Arg972 IRS-1. Using these cells, we characterized the downstream pathway by which Arg972 IRS-1 impairs b-cell survival. RIN-Arg972 cells exhibited a marked impairment in the sequential activation of PI3-kinase, Akt, and BAD as compared with RI N-WT. Impaired BAD phosphorylation resulted in increased binding to Bcl-XL instead of 14-3-3 protein, thus sequestering the Bcl-XL antiapoptotic protein to promote survival. Both caspase-9 and caspase-3 activities were increased in RIN-Arg972 cells. The results show that the common Arg972 polymorphism in IRS-1 impairs human b-cell survival and causes resistance to antiapoptotic effects of insulin by affecting the PI3-kinase/Akt survival pathway. These findings establish an important role for the insulin signaling in human b-cell survival and suggest that genetic defects in early steps of insulin signaling may contribute to b-cell failure.
对人类胰岛素受体底物-1（IRS-1）基因进行分子扫描发现，一种常见的多态性导致了甘氨酸（Gly）向精氨酸（Arg）972位点的改变。携带精氨酸972位点IRS-1的糖尿病患者和糖尿病前期患者的空腹胰岛素和C肽水平较低。为了直接研究精氨酸972位点的IRS-1是否影响人类胰岛细胞的存活，我们利用了一个独特的机会，分析了从三名携带精氨酸972位点杂合子的供体和六名携带野生型IRS-1的供体中分离出的胰岛。与野生型相比，携带精氨酸972位点IRS-1的供体的胰岛凋亡细胞数量增加了两倍。携带精氨酸972位点IRS-1的供体的胰岛中，与IRS-1相关的磷脂酰肌醇-3激酶（PI3-激酶）活性降低。在稳定表达野生型IRS-1或精氨酸972位点IRS-1的RIN大鼠β细胞系中也得到了相同的结果。利用这些细胞，我们确定了精氨酸972位点的IRS-1损害β细胞存活的下游途径。与表达野生型IRS-1的RIN细胞（RIN-WT）相比，表达精氨酸972位点IRS-1的RIN细胞（RIN-Arg972）在PI3-激酶、蛋白激酶B（Akt）和BAD的顺序激活方面表现出明显的损害。BAD磷酸化受损导致其与Bcl-XL的结合增加，而不是与14-3-3蛋白结合，从而隔离了Bcl-XL抗凋亡蛋白以促进存活。RIN-Arg972细胞中的半胱天冬酶-9（caspase-9）和半胱天冬酶-3（caspase-3）活性均增加。结果表明，IRS-1中常见的精氨酸972位点多态性损害了人类β细胞的存活，并通过影响PI3-激酶/Akt存活途径导致对胰岛素抗凋亡作用的抵抗。这些发现确立了胰岛素信号在人类β细胞存活中的重要作用，并表明胰岛素信号早期步骤中的遗传缺陷可能导致β细胞功能衰竭。
