Boney C M, Gruppuso P A, Faris R A, Frackelton A R
Department of Pediatrics Rhode Island Hospital, Providence 02903, USA.
Mol Endocrinol. 2000 Jun;14(6):805-13. doi: 10.1210/mend.14.6.0487.
Insulin-like growth factor-I (IGF-I) stimulates mitogenesis in proliferating preadipocytes, but when cells reach confluence and become growth arrested, IGF-I stimulates differentiation into adipocytes. IGF-I induces signaling pathways that involve IGF-I receptor-mediated tyrosine phosphorylation of Shc and insulin receptor substrate 1 (IRS-1). Either of these adaptor proteins can lead to activation of the three-kinase cascade ending in activation of the extracellular signal-regulated kinase 1 and -2 (ERK-1 and -2) mitogen-activated protein kinases (MAPKs). Several lines of evidence suggest that activation of MAPK inhibits 3T3-L1 preadipocyte differentiation. We have shown that IGF-I stimulation of MAPK activity is lost as 3T3-L1 preadipocytes begin to differentiate. This change in MAPK signaling coincides with loss of IGF-I-mediated Shc, but not IRS-1, tyrosine phosphorylation. We hypothesized that down-regulation of MAPK via loss of proximal signaling through Shc is an early component in the IGF-I switch from mitogenesis to differentiation in 3T3-L1 preadipocytes. Treatment of subconfluent cells with the MEK inhibitor PD098059 inhibited both IGF-I-activation of MAPK as well as 3H-thymidine incorporation. PD098059, in the presence of differentiation-inducing media, accelerated differentiation in subconfluent cells as measured by expression of adipocyte protein-2 (aP-2), peroxisome proliferator-activated receptor gamma (PPARgamma) and lipoprotein lipase (LPL). Transient transfection of subconfluent cells with Shc-Y317F, a dominant-negative mutant, attenuated IGF-I-mediated MAPK activation, inhibited DNA synthesis, and accelerated expression of differentiation markers aP-2, PPARgamma, and LPL. We conclude that signaling through Shc to MAPK plays a critical role in mediating IGF-I-stimulated 3T3-L1 mitogenesis. Our results suggest that loss of the ability of IGF-I to activate Shc signaling to MAPK may be an early component of adipogenesis in 3T3-L1 cells.
胰岛素样生长因子-I（IGF-I）可刺激增殖中的前脂肪细胞发生有丝分裂，但当细胞达到汇合状态并停止生长时，IGF-I会刺激其分化为脂肪细胞。IGF-I诱导的信号通路涉及IGF-I受体介导的Shc和胰岛素受体底物1（IRS-1）的酪氨酸磷酸化。这两种衔接蛋白中的任何一种都可导致三激酶级联反应的激活，最终激活细胞外信号调节激酶1和-2（ERK-1和-2）丝裂原活化蛋白激酶（MAPK）。多项证据表明，MAPK的激活会抑制3T3-L1前脂肪细胞的分化。我们已经表明，随着3T3-L1前脂肪细胞开始分化，IGF-I对MAPK活性的刺激作用丧失。MAPK信号的这种变化与IGF-I介导的Shc酪氨酸磷酸化丧失一致，但IRS-1的酪氨酸磷酸化未丧失。我们推测，通过Shc近端信号丧失导致的MAPK下调是3T3-L1前脂肪细胞中IGF-I从有丝分裂向分化转变的早期组成部分。用MEK抑制剂PD098059处理亚汇合细胞，可抑制IGF-I对MAPK的激活以及3H-胸腺嘧啶核苷掺入。在存在分化诱导培养基的情况下，PD098059可加速亚汇合细胞的分化，这通过脂肪细胞蛋白-2（aP-2）、过氧化物酶体增殖物激活受体γ（PPARγ）和脂蛋白脂肪酶（LPL）的表达来衡量。用显性负性突变体Shc-Y317F瞬时转染亚汇合细胞，可减弱IGF-I介导的MAPK激活，抑制DNA合成，并加速分化标志物aP-2、PPARγ和LPL的表达。我们得出结论，通过Shc向MAPK的信号传导在介导IGF-I刺激的3T3-L1有丝分裂中起关键作用。我们的结果表明，IGF-I激活Shc信号至MAPK的能力丧失可能是3T3-L1细胞脂肪生成的早期组成部分。
