Hormonal induction of adipogenesis induces Skp2 expression through PI3K and MAPK pathways
Hormonal induction of adipogenesis induces Skp2 expression through PI3K and MAPK pathways
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DOI：
10.1002/jcb.21063
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发表时间：
2007-01-01
影响因子：
4
通讯作者：
Morrison, Ron F.
中科院分区：
文献类型：
--
作者：
Auld, Corinth A.;Caccia, Carla D.;Morrison, Ron F.
我们以前已经表明，F-box蛋白，S-相激酶相关蛋白（Skp 2）在靶向细胞周期抑制剂，p27降解3 T3-L1脂肪细胞分化的早期阶段的26 S蛋白酶体中发挥了机械作用。在这里，我们证明了Skp 2及其辅助蛋白，Cks 1的蛋白水平增加密度逮捕前脂肪细胞重新进入细胞周期在克隆扩增，减少分化诱导的生长停滞，并成为难治性激素刺激后发病的终端脂肪细胞分化。成分分析显示，虽然最大的Skp 2/Cks 1蛋白积累需要完全分化的鸡尾酒，胰岛素主要参与。Skp 2 mRNA的积累先于Skp 2蛋白的增加，并继Akt和Erk 1/2的激活，Akt和Erk 1/2分别是磷脂酰肌醇-3激酶（PI 3 K）和丝裂原活化蛋白激酶（MAPK）信号转导通路的介质。使用特定的抑制剂，我们发现，虽然这两种途径的激活是最大表达所必需的，但PI 3 K信号转导主要负责Skp 2/Cks 1积累的增加。Skp 2 mRNA的增加在激素刺激后4小时是显著的，在中期G期进展期间12小时达到平台，并且发生在mRNA稳定性没有变化的情况下。我们进一步证明，荧光素酶活性，从含有2.4 kb的Skp 2启动子的pGL 3载体，增加2.5倍，激素刺激。这种启动子活性的增加在PI 3 K和MAPK阻断后被显著抑制。缺失研究表明响应元件位于Skp 2启动子近端。这些数据表明，Skp 2的转录调节PI 3 K和MAPK途径作为3 T3-L1前脂肪细胞从静止到增殖的脂肪细胞增生过程中的过渡。
We have previously shown that the F-box protein, S-phase kinase-associated protein (Skp2) plays a mechanistic role in targeting the cell-cycle inhibitor, p27 for degradation by the 26S proteasome during early stages of 3T3-L1 adipocyte differentiation. Here, we demonstrate that protein levels of Skp2 and its accessory protein, Cks1 increased as density-arrested preadipocytes re-entered the cell cycle during clonal expansion, decreased with differentiation-induced growth arrest, and became refractory to hormonal stimulation following the onset of terminal adipocyte differentiation. Component analysis revealed that while maximal Skp2/Cks1 protein accumulation required the complete differentiation cocktail, that insulin was principally involved. Skp2 mRNA accumulation was found to precede the increase in Skp2 protein and succeed the activation of Akt and Erk1/2, mediators of phosphatidylinositol-3 kinase (PI3K) and mitogen-activated protein kinase(MAPK) signal transduction pathways, respectively. Using specific inhibitors, we found that while activation of both pathways was required for maximal expression, PI3K signaling was primarily responsible for the increase in Skp2/Cks1 accumulation. The increase in Skp2 mRNA was notable 4 h following hormonal stimulation, plateaued by 12 h during mid-G, phase progression, and occurred without change to mRNA stability. We further demonstrate that luciferase activity, originating from a pGL3 vector containing 2.4 kb of the Skp2 promoter, increased 2.5-fold with hormonal stimulation. This increase in promoter activity was markedly suppressed following PI3K and MAPK blockade. Deletion studies indicate that responsive elements were located within the proximal Skp2 promoter. These data demonstrate that Skp2 is transcriptionally regulated by PI3K and MAPK pathways as 3T3-L1 preadipocytes transition from quiescence to proliferation during adipocyte hyperplasia.