Protein kinase C-λ knockout in embryonic stem cells and adipocytes impairs insulin-stimulated glucose transport
Protein kinase C-λ knockout in embryonic stem cells and adipocytes impairs insulin-stimulated glucose transport
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DOI：
10.1210/me.2003-0087
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发表时间：
2004-02-01
影响因子：
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通讯作者：
Farese, RV
中科院分区：
文献类型：
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作者：
Bandyopadhyay, G;Standaert, ML;Farese, RV
非典型蛋白激酶C（aPKC）亚型已被认为介导胰岛素对脂肪细胞和其他细胞中葡萄糖转运的影响。为了更严格地测试这一假设，我们产生了小鼠胚胎干细胞（ES）和ES衍生的脂肪细胞，其中aPKC-λ等位基因通过重组方法被敲除。胰岛素激活PKC-λ并刺激野生型（WT）PKC-λ（+/+）中的葡萄糖转运，但在敲除PKC-λ（-/-）的ES细胞中则不然。然而，在PKC-λ（-/-）ES细胞中，WT PKC-λ的表达挽救了胰岛素刺激的葡萄糖转运。令人惊讶的是，胰岛素诱导的PKC-λ活性和葡萄糖转运的增加依赖于富含脯氨酸的酪氨酸蛋白激酶2、ERK途径和磷脂酶D（PLD）的激活，但不依赖于PKC-λ（+/+）ES细胞中的磷脂酰肌醇3-激酶（PI 3 K）。有趣的是，这种依赖性在ES细胞分化为脂肪细胞后完全逆转，即胰岛素对PKC-λ和葡萄糖转运的作用依赖于PI 3 K，而不是富含脯氨酸的酪氨酸蛋白激酶2/ERK/PLD。在ES细胞中，胰岛素对葡萄糖转运的影响在PKC-λ（-/-）脂肪细胞中不存在，但在这些脂肪细胞中通过表达WT PKC-λ而被拯救。我们的研究结果表明，胰岛素激活aPKCs和葡萄糖转运在ES细胞通过一个新认识的PI 3 K非依赖性ERK/PLD依赖性途径，并提供了一个令人信服的证据表明，aPKCs是胰岛素刺激的葡萄糖转运所必需的，无论aPKCs是由PI 3 K依赖性或PI 3 K非依赖性机制激活。
Atypical protein kinase C (aPKC) isoforms have been suggested to mediate insulin effects on glucose transport in adipocytes and other cells. To more rigorously test this hypothesis, we generated mouse embryonic stem (ES) cells and ES-derived adipocytes in which both aPKC-lambda alleles were knocked out by recombinant methods. Insulin activated PKC-lambda and stimulated glucose transport in wild-type (WT) PKC-lambda(+/+), but not in knockout PKC-lambda(-/-), ES cells. However, insulin-stimulated glucose transport was rescued by expression of WT PKC-lambda in PKC-lambda(-/-) ES cells. Surprisingly, insulin-induced increases in both PKC-lambda activity and glucose transport were dependent on activation of proline-rich tyrosine protein kinase 2, the ERK pathway, and phospholipase D (PLD) but were independent of phosphatidylinositol 3-kinase (PI3K) in PKC-lambda(+/+) ES cells. Interestingly, this dependency was completely reversed after differentiation of ES cells to adipocytes, i.e. insulin effects on PKC-lambda and glucose transport were dependent on PI3K, rather than proline-rich tyrosine protein kinase 2/ERK/PLD. As in ES cells, insulin effects on glucose transport were absent in PKC-lambda(-/-) adipocytes but were rescued by expression of WT PKC-lambda in these adipocytes. Our findings suggest that insulin activates aPKCs and glucose transport in ES cells by a newly recognized PI3K-independent ERK/PLD-dependent pathway and provide a compelling line of evidence suggesting that aPKCs are required for insulin-stimulated glucose transport, regardless of whether aPKCs are activated by PI3K-dependent or PI3K-independent mechanisms.