Construction And Studies Of Trex1 Autoimmune Disease Mouse Models

Autoimmune diseases are chronic diseases that are caused by autoimmune reactions due to immune dysfunction.Currently,tens of millions of people around the world are suffering from these diseases,but the etiology remains unclear.Nucleases play an important role in DNA replication,injury,repair and maintenance of genomic stability.Trex1 is the most abundant exonuclease in mammalian cells which can degrade DNA editing and repairing of aberrant or mismatched DNA.Mutation of the Trex1 gene has been associated with a variety of autoimmune diseases.Thereby,to explore the role of Trex1 in the immune dysfunction is important for better understanding the pathogenesis of the related autoimmune diseases.In this study,we studied the relationship of Trex1 and its mutation to autoimmune diseases by means of CRISPR/Cas9 technology,the expression of recombinant protein,Western blotting,qRT-PCR,Flow Cytometry etc.We first established the Trex1 gene knockout cell model by the CRISPR/Cas9 technology,and found that the deletion of Trexl gene could cause the immune response mediated by the cGAS-STING DNA-sensing pathway.In Trex1 multiple loci mutations,D18N mutation occurs in the DNA enzymatic domain,which is related to autoimmune diseases,but no further research has been reported.Therefore,we studied the effect of Trexl D18N mutation on the autoimmune reaction.We first constructed and expressed the recombinant protein of Trex1 D18N in E.coli.In vitro studies found that the Trex1 D18N mutation led to the loss of DNA enzymatic activity.Then,the Trexl D18N mouse model was constructed by the CRISPR/Cas9 technology.Compared with the wild type,the MEF cells derived from the Trex1 D18N mouse model had a higher level of type I interferon expression in the absence of stimulation,and the expression of inflammatory cytokines in plasma and CD4+T cells also increased significantly.These data suggest that the Trex1 D18N mutation can cause autoimmune reaction and the Trex1 D18N mouse model can be used for autoimmune disease research.These studies lay a solid foundation for the mechanistic and therapeutic target research of Trex1 in autoimmune diseases.