AMPK对Rab-GAP的调节作用研究

Study on the regulation of Rab-GAP by AMPK

目的:检测激活型AMPK对Rab-GAP的调节作用,明确它们之间的上下游关系。方法:激活型AMPK腺病毒(AdAMPK-CA)感染C2C12小鼠骨骼肌细胞,通过细胞成像系统测定细胞中的荧光强度,明确AMPK-CA在C2C12细胞中表达。MTS试验确定没有细胞毒性的腺病毒浓度,Western blot检测AMPK底物ACC和两个Rab-GAP(TCB1D1和TCB1D4)的磷酸化。结果:AMPK-CA显著升高ACC的磷酸化,并磷酸化TBC1D1 Thr590和TBC1D4 Ser318。结论:AMPK直接磷酸化TCB1D1和TCB1D4。

Objective:To examine the effect of constitutive active AMPK on Rab-GAP phosphorylaiton to elucidate the signal relationship.Methods:Mouse C2C12skeletal muscle cells were infected with constitutive active AMPK adenovirus(Ad-AMPK-CA).The expression of AMPK in C2C12cells was observed by cell fluorescence imaging system.Cytotoxicity of the adenovirus was determined by MTS.Phosphorylations of AMPK substrate ACC and two Rab-GAPs(TCB1D1and TCB1D4)were detected by Western blot.Results:AMPK-CA significantly elevated the phosphorylations of ACC and TBC1D1Thr590and TBC1D4Ser318.Conclusion:AMPK directly could regulate TBC1D1and TBC1D4.