The Association Study And Preliminary Functional Exploration Of CCKAR, DRD2, DAT And SNAPIN Genes In Schizophrenia

Schizophrenia is a common yet severe psychiatric condition that affects about1%of the population. The disorder always suffers the youth, and brings heavy economical and social burdens to families and societies.Schizophrenia is a typical complex disorder with the heredity over80%. The genetic heterogeneity, inter/intra-gene interactions and gene-environment interactions exacerbate its complexity. The dopamine hypothesis of schizophrenia is one of the generally accepted hypothesis and dopamine system dysfunction is considered to be the final common pathway of schizophrenia. In a previous study, our group screened the dopamine signaling genes and found that the cholecystokinin type A receptor (CCKAR) gene is associated with schizophrenics with auditory hallucination. In another work, we detected an association between the SNAP-associated protein (SNAPIN) gene and schizophrenia in a small population. Based on the dopamine hypothesis and our previous results, we conducted the association study of CCKAR, DRD1-5, DAT and SNAPIN genes with paranoid schizophrenia in Han Chinese, and explored the function of associated genes in gene expression, miRNA regulation and protein interaction levels.Five single nucleotide polymorphisms (SNPs) including rs1800857located in the5'end of the CCKAR gene were checked in522cases and527controls. Although no association between these single SNPs and this disorder, cis-phase interactions were detected between rs1800857and other4SNPs, respectively. Moreover,3risky5-SNP haplotypes of G-A-G-C-T, T-G-G-G-T and T-G-G-C-C were found [p=2.2x10-5,p1.1×10-6, and p=1.9×10-6), indicating allelic heterogeneity of the CCKAR gene is involved in schizophrenia. Quantitative trait analysis found that the5-SNP haplotypes associated with hallucination, suspicion, hostile and the total positive scores of PANSS. In addition, we used real-time PCR to detect CCKAR gene expression in antipsychotic drugs treated human CHP212cell lines and found that within36h, haloperidol rather than clozapine made the CCKAR mRNA increased gradually and then returned to the initial expression level.By screening functional SNPs of the DRD1-5and DAT genes, the DRD2and DAT genes showed associated trend with paranoid schizophrenia. Thereafter, we validated that rs1076560-T in the DRD2gene confers risk of schizophrenia in1351patients and1640controls (p=0.022). We also found that rs2455391in the DAT gene is associated with the disease (allelic p=0.015, genotypic p=0.018) and the frequency of rs2975223(G)-rs2455391(C) haplotype in the patient group is significantly higher than that in the control group (p=0.0012).In this study, we firstly report that rs7345located in SNAPIN3'UTR is associated with paranoid schizophrenia in516patients and532controls (allelic p=0.014, genotypic p=0.003). However, this SNP did not directly influence gene expression by reporter gene assay. Also by the dual luciferases reporter gene assay, we discovered that miR-30e can bind SNAPIN-3'UTR to inhibit expression of the reporter gene. Furthermore, we tested plasma miR-30e concentration in113patients and127controls and found that the miR-30e in cases was significantly lower (p=0.001). At the same time,57candidate proteins which may interact with Snapin in the SH-SY5Y cells were initially identified with tandem affinity purification (TAP)-mass spectrometry (MS) method.In summary, we not only verified the association of CCKAR, DRD2, DAT and SNAPIN genes with paranoid schizophrenia in Han Chinese population, but also conducted preliminary functional exploration of the susceptibility genes in expression, miRNA regulation and protein interaction levels.