Grønning Line M, Cederberg Anna, Miura Naoyuki, Enerbäck Sven, Taskén Kjetil
Department of Medical Biochemistry, Institute of Basic Medical Sciences, University of Oslo, N-0317 Oslo, Norway.
Mol Endocrinol. 2002 Apr;16(4):873-83. doi: 10.1210/mend.16.4.0803.
We have recently identified the winged helix/forkhead gene Foxc2 as a key regulator of adipocyte metabolism that counteracts obesity and diet-induced insulin resistance. This study was performed to elucidate the hormonal regulation of Foxc2 in adipocytes. We find that TNF alpha and insulin induce Foxc2 mRNA in differentiated 3T3-L1 cells with the kinetics of an immediate early response (1-2 h with 100 ng/ml insulin or 5 ng/ml TNF alpha). This induction is, in both cases, attenuated by the PI3K inhibitor wortmannin as well as the MAPK kinase inhibitor PD98059. Furthermore, we show that stimulation of 3T3-L1 adipocytes with phorbol-12-myristate-13-acetate or 8-(4-chlorophenyl)thio-cAMP induces the expression of Foxc2. Interestingly, we find that the basal level of Foxc2 mRNA is down-regulated whereas hormonal responsiveness increases during differentiation of 3T3-L1 from preadipocytes to adipocytes. At the protein level, immunoblots with Foxc2 antibody demonstrated an induction of Foxc2 by insulin and TNF alpha in nuclear extracts of 3T3-L1 adipocytes. EMSA of nuclear proteins from phorbol-12-myristate-13-acetate- and TNF alpha-treated 3T3-L1 adipocytes using a forkhead consensus oligonucleotide revealed specific binding of a Foxc2/DNA complex. In conclusion, our data suggest that insulin and TNF alpha regulate the expression of Foxc2 via a PI3K- and ERK 1/2-dependent pathway in 3T3-L1 adipocytes. Also, signaling pathways downstream of PKA and PKC induce the expression of Foxc2 mRNA.
我们最近确定了翼状螺旋/叉头基因Foxc2是脂肪细胞代谢的关键调节因子，它可对抗肥胖和饮食诱导的胰岛素抵抗。进行这项研究是为了阐明脂肪细胞中Foxc2的激素调节机制。我们发现，TNFα和胰岛素可在分化的3T3-L1细胞中诱导Foxc2 mRNA表达，其动力学表现为即时早期反应（100 ng/ml胰岛素或5 ng/ml TNFα作用1-2小时）。在这两种情况下，PI3K抑制剂渥曼青霉素以及MAPK激酶抑制剂PD98059均可减弱这种诱导作用。此外，我们还发现，用佛波醇-12-肉豆蔻酸酯-13-乙酸酯或8-（4-氯苯基）硫代-cAMP刺激3T3-L1脂肪细胞可诱导Foxc2的表达。有趣的是，我们发现，在3T3-L1从前脂肪细胞分化为脂肪细胞的过程中，Foxc2 mRNA的基础水平下调，而激素反应性增加。在蛋白质水平上，用Foxc2抗体进行的免疫印迹显示，胰岛素和TNFα可在3T3-L1脂肪细胞核提取物中诱导Foxc2表达。使用叉头共有寡核苷酸对经佛波醇-12-肉豆蔻酸酯-13-乙酸酯和TNFα处理的3T3-L1脂肪细胞核蛋白进行EMSA分析，结果显示存在Foxc2/DNA复合物的特异性结合。总之，我们的数据表明，胰岛素和TNFα通过PI3K和ERK 1/2依赖性途径调节3T3-L1脂肪细胞中Foxc2的表达。此外，PKA和PKC下游的信号通路可诱导Foxc2 mRNA的表达。
