Genetic Variants And Functional Analyses Of The CTSB Gene Promoter In Acute Myocardial Infarction

Objective: In this paper,we studied the CTSB gene promoter’s single nucleotide sequence variation in the AMI group and the control group,and explored whether the onset of AMI is related to the sub-genetic variation initiated by CTSB gene and the onset of AMI.Methods: In this study,we use case-control researc method,analyze the clinical data of AMI and control groups,extract DNA from them,collect the peripheral blood of all subjects,separate mononuclear cells,extract DNA,after PCR amplification,we sequenced DNA and compared the sequences for data statistics,functional analysis was carried out on DNA sequence variation through dual luciferase reporter gene experiment,predicted the transcription factor binding sites affected by SNPs.Finally,we analyzed the association between SNPs and the onset of AMIby multiple statistical analysis methods.Results:1.In this study,a total of 7 SNPs were identified in 405 subjects.4SNPs [g.4482 T>C(rs1369930956),g.4567 C>T(rs891018567),g.4953G>T(rs1369930956)and g.4954 T>A(rs942670850)] were found in 9AMI patients but not in controls.1 SNP [g.4803 T>C(rs1293312)] was present in both AMI and control groups,and the frequency of distribution was similar between the two groups(P=0.926).2 SNPs [g.5061C>T(rs965406972),g.5130 A>G(rs1817431932)] were found only in the control group and not in the AMI group.2.The HEK-293 and NRCMs cells transfection results showed that the transcription activity of CTSB gene promoter was significan tly increased by the four SNPs[g.4482 T>C(183062389)、g.4803 T>C(rs1293312)、g.4953 G>T(rs1369930956)and g.4954 T>A(rs942670850)] detected in AMI group.SNP[g.4567 C>T(rs891018567)] did not affect the transcription activity(P > 0.05).Two SNPs [g.5061C>T(rs965406972)and g.5130 A>G(rs1817431932)] detected in the control group did not affect the transcription activity(P > 0.05).3.The results predicted by TRANSFAC database showd that SNP g.4482 T>C(rs183062389)can eliminate the binding site of transcription factor FOXO1A;SNP g.4803 T>C(rs1293312)can create the binding sites of transcription factors CBPB and DPF2,and eliminate the binding sites of Spi-B;SNP g.4953 G>T(rs1369930956)can create the binding sites of transcription factors ZEB1 and TTF-1,and eliminate the binding site of HIF1.SNP g.4954 T > A(rs942670850)can also eliminate the binding site of HIF1.Conclusion: In this study,we found that four SNPs[g.4482T>C(rs183062389),g.4803 T>C(rs1293312),g.4953 G>T(rs1369930956)and g.4954 T>A(rs942670850)] changed the transcription activity of the CTSB gene promoter by affecting its binding to transcription factors in HEK-293 cells and NRCMs cells,thereby affecting the expression level of CTSB gene and participating in the occurrence and development of AMI.