Study of the ubiquitination of RNF6 on insulin receptor substrate-1 / 国际检验医学杂志

ABSTRACT

Objective To construct an eukaryotic expression vetor of ring finger protein 6 (RNF6)and to investigate its ubiqu-itylation on insulin receptor substrate-1(IRS-1).Methods Human RNF6 coding sequence was amplified by polymerase chain reac-tion (PCR)method with human cDNA as template.The pcDNA3.1-CHA-RNF6 was constructed with routine molecular methods and transfected into hepatocarcinoma cell HepG2.Relative quantification of IRS-1 mRNA was preformed by real-time reverse tran-scription PCR.Western blot was applied to detect the expression levels of IRS-1.Results 48 h after the plasmid carring RNF6 gene transfected HepG2 cells,the mRNA level of ISR-1 gene reduced to 69% of control group.The expression of IRS-1 was markedly lower than control group (P <0.01).Conclusion The expression of IRS-1 is markedly down-regulated in HepG2 and enhancement of the ubiquitylation level of IRS-1 contribute to the disorder in insulin signal transferring.