Harashima Shin-ichi, Clark Anne, Christie Michael R, Notkins Abner Louis
Experimental Medicine Section, Oral Infection and Immunity Branch, National Institute of Dental and Craniofacial Research, National Institutes of Health, Bethesda, MD 20892, USA.
Proc Natl Acad Sci U S A. 2005 Jun 14;102(24):8704-9. doi: 10.1073/pnas.0408887102. Epub 2005 Jun 6.
IA-2 is an enzymatically inactive member of the transmembrane protein tyrosine phosphate family located in dense core secretory vesicles and a major autoantigen in type 1 diabetes. Recent studies showed that targeted disruption of the IA-2 gene in mice resulted in impairment of insulin secretion and glucose intolerance. Insulin homeostasis, however, is a complex process involving a cascade of regulatory factors, and IA-2 is widely expressed in neuroendocrine cells throughout the body. Consequently, it is uncertain whether the impairment of insulin secretion in IA-2 knockout mice is a direct result of the knockout of IA-2 in beta cells or to counter regulatory alterations resulting from IA-2 knockout in other neuroendocrine cells. To define the function of IA-2, we studied the secretion of insulin in a single cell type, MIN-6, by overexpressing and knocking down IA-2. Our experiments showed that overexpression of IA-2 resulted in a 6-fold increase in glucose- or K+-induced insulin secretion and a approximately 3-fold increase in the number of secretory vesicles and the insulin content of cells. In contrast, knockdown of endogenous IA-2 by short interfering RNA resulted in nearly a complete loss of glucose-induced insulin secretion and a 50% decrease in basal insulin release. The half-life of insulin in cells overexpressing IA-2 was nearly twice as great as that in mock-transfected cells, suggesting that IA-2 was stabilizing the insulin-containing vesicles. From these results we conclude that in beta cells, IA-2 is an important regulator of dense core vesicle number and glucose-induced and basal insulin secretion.
IA-2是跨膜蛋白酪氨酸磷酸酶家族中一种无酶活性的成员，位于致密核心分泌囊泡中，是1型糖尿病中的主要自身抗原。最近的研究表明，小鼠IA-2基因的靶向破坏导致胰岛素分泌受损和葡萄糖不耐受。然而，胰岛素稳态是一个涉及一系列调节因子的复杂过程，并且IA-2在全身的神经内分泌细胞中广泛表达。因此，尚不确定IA-2基因敲除小鼠中胰岛素分泌的受损是β细胞中IA-2基因敲除的直接结果，还是其他神经内分泌细胞中IA-2基因敲除导致的反调节改变的结果。为了确定IA-2的功能，我们通过过表达和敲低IA-2来研究单一细胞类型MIN-6中的胰岛素分泌。我们的实验表明，IA-2的过表达导致葡萄糖或钾离子诱导的胰岛素分泌增加6倍，分泌囊泡数量和细胞胰岛素含量增加约3倍。相反，通过短干扰RNA敲低内源性IA-2导致葡萄糖诱导的胰岛素分泌几乎完全丧失，基础胰岛素释放减少50%。过表达IA-2的细胞中胰岛素的半衰期几乎是 mock 转染细胞中的两倍，这表明IA-2正在稳定含胰岛素的囊泡。从这些结果我们得出结论，在β细胞中，IA-2是致密核心囊泡数量以及葡萄糖诱导和基础胰岛素分泌的重要调节因子。
