Next Generation Sequencing Facilitates Quantitative Analysis of Wild Type and Hrd1-/- T regulatory cells

Purpose: Natural T regulatory cells (nTreg) and induced Tregs (iTreg) were isolated from hrd1 WT and knockout mice. The purpose of this project is to look at the transcriptional profiles changes of genes specifically involved in T cell receptor signaling, TGF-beta signaling, and metabolic genes in Hrd1 deficient Tregs. Methods: natural Tregs were sorted according to CD4+GFP+, and RNA was harvested using QIAGEN Rneasy Mini Kit. For induced iTreg, CD4 T cells were isolated and polarized with IL2/TGFbeta for 5 days to induce FoxP3 induction. Day 5, GFP positive cells were isolated and RNA were harvested using QIAGEN RNeasy mini Kit. Results and Conclusions: Our study represents the first detailed analysis of transcriptional profiles in Hrd1 deficient T regulatory cells, with biologic replicates, generated by RNA-seq technology. The optimized data analysis workflows reported here should provide a framework for comparative investigations of expression profiles. natural and induced T regulatory cells mRNA profiles of 8 weeks old wild type (WT) and Treg specific deletion of Hrd1 mice were generated by deep sequencing, in triplicate, using Illumina NextSeq 500