Generation of NOD mice carrying R619W mutation in PTPN22 gene by CRISPR/Cas9-mediated genome engineering (BA4P.134)

Abstract

Abstract An allelic variant of protein tyrosine phosphatase nonreceptor 22 (PTPN22), PTPN22R620W, is strongly associated with type 1 diabetes (T1D) and several other autoimmune diseases in human, and increases the risk of developing T1D by 2-4 fold. Non-obese diabetic (NOD) mouse is an ideal spontaneous T1D model that shares with human many genetic pathways contributing to T1D; however, it has not previously been possible to introduce targeted mutations into NOD mice due to a lack of germline competent NOD embryonic stem cells. To overcome this technological hindrance, we used CRISPR/Cas9 technology to precisely mutate PTPN22 gene in its endogenous locus in NOD zygotes, and successfully produced 2 independent lines of NOD mice expressing PTPN22R619W variant, a murine ortholog of the human autoimmune-associated allele. We have also obtained a line carrying PTPN22 null allele. We have demonstrated the mutated PTPN22 genes in all lines have successfully germline transmitted. We have found a significant increase of T1D incidence in the heterozygous NOD PTPN22R619W mice, suggesting its strong effect in heterozygosity, which is very relevant to human T1D since PTPN22R620W is most commonly carried in heterozygosity. This is the first time the autoimmune-associated PTPN22 allele has been introduced into a spontaneous model of T1D and, to the best of our knowledge, the first time CRISPR/Cas9 technology has been used to genetically modify NOD mice.