Plasma membrane subdomain compartmentalization contributes to distinct mechanisms of ceramide action on insulin signaling.
Plasma membrane subdomain compartmentalization contributes to distinct mechanisms of ceramide action on insulin signaling.
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作者：
Blouin CM;Prado C;Takane KK;Lasnier F;Garcia-Ocana A;Ferré P;Dugail I;Hajduch E
神经酰胺现在被认为是胰岛素信号的负调节因子，通过损害蛋白激酶B (PKB)/Akt的激活。在不同的细胞中，已经提出了两种不同的机制来介导神经酰胺对PKB/Akt的抑制：一种涉及非典型蛋白激酶Cζ (PKCζ)，另一种涉及蛋白磷酸酶-2 （PP2A）。我们假设神经酰胺通过PKCζ或PP2A的作用可能取决于质膜（PM）的结构组织，特别是小泡蛋白富集域（CEM）的丰度。我们使用了不同的PKCζ突变体结构或PP2A抑制剂冈田酸（OKA），在表达不同小窝蛋白-1水平的细胞中选择性地抑制PKCζ和PP2A依赖通路，并评估了胰岛素和神经酰胺对不同PM亚结构域PKB/Akt活性的影响。尽管神经酰胺对胰岛素刺激的PKB/Akt的负面影响在脂肪细胞中占主导地位，但神经酰胺通过CEMs外的PP2A作用，被OKA阻止，也被揭示。为了测试CEM通过PKCζ通路直接神经酰胺作用的重要性，我们用神经酰胺处理缺乏CEM的3T3-L1前脂肪细胞，我们发现PKB/Akt的脂质阴性作用转变为pp2a介导的机制。在CEM丰度较低的成纤维细胞中，神经酰胺激活的PP2A途径占主导地位，但在caveolin-1过表达后，可以转移到神经酰胺激活的PKCζ途径。我们的研究结果表明，神经酰胺可以从pkc - ζ依赖机制转变为PP2A途径，对PKB/Akt起负作用，从而揭示了PM的CEMs在这一过程中的关键作用。
Ceramide is now recognized as a negative regulator of insulin signaling by impairing protein kinase B (PKB)/Akt activation. In different cells, two distinct mechanisms have been proposed to mediate ceramide inhibition of PKB/Akt: one involving atypical protein kinase C zeta (PKCζ) and the other the protein phosphatase-2 (PP2A). We hypothesized that ceramide action through PKCζ or PP2A might depend on plasma membrane (PM) structural organization and especially on caveolin-enriched domain (CEM) abundance. We have used different PKCζ mutant constructs or the PP2A inhibitor, okadaic acid (OKA), to selectively inhibit PKCζ- and PP2A-dependent pathways in cells expressing different caveolin-1 levels and evaluated the impact of insulin and ceramide on PKB/Akt activity in different PM subdomains. Although the PKCζ-mediated negative effect of ceramide on insulin-stimulated PKB/Akt was dominant in adipocytes, a ceramide action through PP2A outside CEMs, prevented by OKA, was also unraveled. To test the importance of CEM to direct ceramide action through the PKCζ pathway, we treated 3T3-L1 preadipocytes devoid of CEMs with ceramide and we saw a shift of the lipid-negative action on PKB/Akt to a PP2A-mediated mechanism. In fibroblasts with low CEM abundance, the ceramide-activated PP2A pathway dominated, but could be shifted to a ceramide-activated PKCζ pathway after caveolin-1 overexpression. Our results show that ceramide can switch from a PKCζ-dependent mechanism to a PP2A pathway, acting negatively on PKB/Akt, and hence revealing a critical role of CEMs of the PM in this process.
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DOI：
10.1073/pnas.92.19.8655
发表时间：
1995-09-12
影响因子：
11.1
作者：
FRA, AM;WILLIAMSON, E;PARTON, RG
通讯作者：
PARTON, RG
影响因子：
12.8
作者：
Ha, H;Pak, Y
通讯作者：
Pak, Y
影响因子：
3.1
作者：
GUILLOT, FL;AUDUS, KL;RAUB, TJ
通讯作者：
RAUB, TJ
影响因子：
7.8
作者：
KANDROR, KV;STEPHENS, JM;PILCH, PF
通讯作者：
PILCH, PF
影响因子：
4.8
作者：
Bourbon, NA;Yun, J;Kester, M
通讯作者：
Kester, M