| AimsSchizophrenia(SZ)is a devastating mental disorder that seriously affects human health and life quality.The clinical symptoms of schizophrenia are complex,with the underlying pathogenetic mechanism of schizophrenia being unknown.However,a large body of studies indicate,genetic factors,especially the polygenic effects,play a crucial part in the pathogenicity of schizophrenia.Based on the microarray analysis from our lab,two differentially expressed genes between schizophrenia and control group: RNF4(ring finger protein 4)and SART3(squamous cell carcinoma antigen recognized by T cells 3)were identified and were investigated for their underlying pathogenetic roles in schizophrenia in this study.MethodsWe studied the expression of RNF4 and SART3 genes in patients with schizophrenia,as well as the association study between these two genes with schizophrenia,and explored the biological interaction between RNF4 and SART3 proteins in this work.The methods used in the three parts of this study are as follows:(1)Correlation analysis was performed between the transcriptome level of RNF4 and SART3 from Brain Span database.Besides,real-time PCR was applied to identify the m RNA level of RNF4 and SART3 in 30 patients with schizophrenia and 30 healthy controls.(2)Genotyping was implemented to detect 6 single nucleotide polymorphisms in RNF4(rs1203849,rs1203860,rs2282765,rs3849666,rs4974695,rs704352)and 3single nucleotide polymorphisms in SART3(rs2287550,rs2374998,rs73189225)between 392 schizophrenia and 572 control samples.What’s more,the association study in single locus between RNF4 and SART3 genes with schizophrenia was conducted.Meanwhile,we analyzed the interaction of RNF4 and SART3 on schizophrenia susceptibility with potential risk by MDR(multifactor-dimensionality reduction)software.(3)Immunofluorescence and co-immunoprecipitation assays were carried out to examine the location of RNF4 and SART3 protein as well as the protein-protein interaction,and the biological relationship of the two proteins was further studied by overexpression and si RNA silencing the target protein SART3 to observe the alteration of expression about RNF4 protein.Results(1)Correlation analysis of Brain Span database demonstrated that there exists a co-expression relationship about transcriptome between RNF4 and SART3 genes with a high correlation coefficient.In addition,as compared with controls,the m RNA level of RNF4 and SART3 up-regulated in patients with schizophrenia.(2)Six single nucleotide polymorphisms in RNF4(rs1203849,rs1203860,rs2282765,rs3849666,rs4974695,rs704352)and three single nucleotide polymorphisms in SART3(rs2287550,rs2374998,rs73189225)were calculated with Hardy-Weinberg equilibrium,followed by in-depth analysis.The results are as following:Single locis analysis revealed there is no statistically significant differences in the genotype or allele distribution or the frequency of the haplotype and the analysis about gender stratification between schizophrenia and controls.Gene-gene interaction analysis indicated there exists a positive association between the interaction of RNF4 and SART3 genes with schizophrenia,with the combination of rs1203860,rs2282765(RNF4)and rs2287550(SART3)being the best model for predicting the risk of schizophrenia,which contributes to increase vulnerability for schizophrenia.(3)Immunofluorescence and co-immunoprecipitation experiments showed both RNF4 and SART3 protein expressed in the nucleus,and there existed protein-protein interaction between SART3 and RNF4 protein.Furthermore,the results of that overexpression SART3 displayed RNF4 protein expression decreased,and that silencing SART3 showed the expression level of RNF4 protein increased,revealed that SART3 directly regulated RNF4 protein.Moreover,western blotting assay uncovered SART3 protein can modulate RNF4 protein by ubiquitination.ConclusionsThere exists a biological interaction between RNF4 and SART3 proteins,and our findings suggested the interaction of RNF4 and SART3 confers susceptibility on patients with schizophrenia,and both genes may be involved in schizophrenia. |
