CTLA-4 gene polymorphisms result in inefficient transcription in myasthenia gravis
Objective To study the expression of cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) in patients with myasthenia gravis (MG) and molecular immunology pathogenesis of MG caused by CTLA-4 gene promoter polymorphism. Methods ELISA assay was used to determine the expression level of serum sCTLA-4. Two independent single nucleotide polymorphisms (SNP) of CTLA-4 at position -1772 and position -1661 in the promoter were analyzed by RFLP. Transcription factor NF-1 and c/EBPβ binding site were confirmed by CHIP assay. Results sCTLA-4 levels in patients serum are correlated with the haplotype and genotype. Patients with CT -1772 genotype or CT -1772 AA -1661 haplotypes have higher levels of sCTLA-4 than patients with TT -1772 or TT -1772 AA -1661. The frequency of the CT -1772 genotype is significantly higher in MG patients and especially, in MG with thymoma as comparing with controls. Meanwhile, the frequency of the G -1661 allele and GG -1661 genotype is lower in MG patients. These SNPs change the sequence of transcription factor binding sites: NF-1 and c/EBPβ. T→C -1772 mutation deletes NF-1 binding site while A→G -1661 mutation creates c/EBPβ binding site, which is confirmed by CHIP assay. DNA variants lose site-specific binding activity of transcription factor regulated by lectin. Conclusion Patients with MG have an aberrant expression of the CTLA-4 products and C/T -1772 and A/G -1661 polymorphisms which result in inefficient transcription of CTLA-4 gene. T→C -1772 mutation also affect gene splicing. The two SNPs may function as an etiological factor to disease susceptibility.