Correlation Between BBS7 Gene And SIRT2 Gene And The Effect Of BBS7 Gene On Cell Function

Objective: To explore the interaction between BBS7 and SIRT2;and to study the effect of BBS7 on cell proliferation,apoptosis and cell cycle.To explore the effect of BBS7 gene on lysine acetylation and ubiquitination.Through the above study,to further understand the pathogenesis of Bardet-Biedl Syndrome(BBS)..Method: To construct BBS7 high expression cell model,p ENTER BBS7 recombinant plasmid was transient transfected to 293 T cell.To construct BBS7 knock down cell model,p X458-574 and p X458-593 recombinant plasmids targeting to BBS7 were constructed by using Crispr/Cas9 system and were co-transfected to 293 T cell.To construct stabilized BBS7 low expressing cell model,BBS7sh2 lentiviral vector was infected to 293 T cells.Proteins,RNAs and DNAs were extracted from the above cell models,and Western Blot,q RT-PCR and DNA digestion were used to verify whether the cell model was successfully constructed.Under the premise of successfully constructing the above cell models,q RT-PCR technology and Western Blot technique were used to study the effect of different expression levels of BBS7 on SIRT2.The co-immunoprecipitation technique was used to further determine whether BBS7 and SIRT2 have a mutual binding effect.And immunofluorescence technique was performed to explore the intracellular colocalization of BBS7 and SIRT2.The effects of BBS7 on cell function were explored by detecting cell cycle,apoptosis and cell proliferation.Western Blot was used to study the effect of BBS7 on lysine acetylation and ubiquitination.Result: In this experiment,cell models with different expression levels of BBS7 were successfully constructed.It was found that there was a negative correlation between BBS7 and SIRT2;and BBS7 and SIRT2 were interacting proteins;BBS7 and SIRT2 were co-localized in cell nucleus.The correlation coefficient was strong;BBS7low expression or high expression inhibited cell proliferation;BBS7 low expression promoted apoptosis,BBS7 high expression inhibited apoptosis;BBS7 different expression levels led to cell cycle changes;Cell fuction was rescued by wild BBS7.Ubiquitylation level of lysine residues was higher than control when BBS7 was knockdown.Conclusion:In this experiment,BBS7 can regulate the expression of SIRT2,the potential regulatory mechanism still needs to be further studied.The effect of BBS7 on cell proliferation,apoptosis,cell cycle and ubiquitylated lysine residue provide a new clue to interpret the pathogenic mechanism of BBS,and to apply a theoretical basis for the treatment of BBS patients.