The Regulating Role And Mechanism Of PP2A Catalyzed Subunit Isomers In Autophagy

Objective: Protein phosphatase 2A catalyzes the subunit ?-isomer(PP2Ac?2),a recently discovered cleavage isomer encoded by the protein phosphatase 2A catalytic subunit alpha gene.Protein phosphatase 2A as an important protein phosphatase almost runs through all aspects of life activities,but because of its structure and function is very complex at present its research there are still many blind spots.The discovery of protein phosphatase 2A catalytic subunit ? isomers has become an opportunity to uncover blind spots.In this study,the cell lines expressing PP2Ac?2 were used as the research platform to explore the function of PP2Ac?2 and the shear triggering conditions of isomer gene,and its related life activities.Methods:(1)The mammalian expression vector of PP2Ac?2 was constructed by molecular cloning techniques.(2)HEK293 cells expressing PP2Ac?2 were constructed using liposome transfection.(3)Cell cycle and cell apoptosis were measured by flow cytometry,and cell growth curves were plotted using cell counting method.(4)The expression levels of PP2 Ac,IGBP1,mTOR and autophagic biomarker LC3 protein were detected by immunoblotting assay.Results:(1)successfully constructed the PP2Ac?2 mammalian expression vector.(2)successfully constructed the HEK293 cells expressing PP2Ac?2.(3)The constructed cell did not change significantly in cell growth curve,cell cycle and apoptosis.(4)The constructed cell did not change significantly in phosphorylation and demethylation of PP2 Ac protein,IGBP1 protein expression increased the difference was statistically significant.(5)The constructed cell phosphorylation level of mTOR protein was increased,the ratio of LC3? / ?of LC3 protein was decreasedConclusion:(1)The mammalian expression vector of PP2Ac?2 was successfully constructed.(2)The HEK293 cell line expressing PP2Ac?2 was successfully constructed.(3)This study found that the expression of PP2Ac?2HEK293 cell basal autophagy levels decreased.