Poster abstracts
Poster number 8 submitted by Deepak Kumar Lakshmipathi
Alternative splicing profile comparison of differentiating T-helper cells to dissect the splicing signatures of Th1, Th2, Th17 and Treg cells
Deepak Kumar Lakshmipathi (Department of BioHealth Informatics, School of Informatics and Computing, Indiana University Purdue University), Mir Quoseena (Department of BioHealth Informatics, School of Informatics and Computing, Indiana University Purdue University), Benjamin Ulrich (Department of Microbiology and Immunolgy, Indiana University School of Medicine, Cancer Research Institute, Indianapolis), Mark Kaplan (Department of Microbiology and Immunolgy, Indiana University School of Medicine, Cancer Research Institute, Indianapolis, Indiana), Sarath Chandra Janga (Department of BioHealth Informatics, School of Informatics and Computing, Indiana University Purdue University)
Abstract:
T cell differentiation is regulated through complex changes in cellular transcription. This study focuses on the contribution of Alternative Splicing (AS) events in the differentiation and post differentiation functions of T-helper cells, specifically in Th1, Th2, Th9, Th17 and Treg cells. T cell RNA-seq data from 72hr and 2week post differentiation time points was analyzed using (r-MATS) for alternative splicing events. We observed majority of the significant events are Skipped Exon (SE) events originating from a total of 1,556 genes and Intron Retention (RI) events were the second most abundant event occurring in 1,254 genes at 72 hours post differentiation. These numbers were significantly lower at 2 weeks post differentiation. Data was further validated using PCR and qPCR techniques. Results showed several skipped exonic events in KTN1, IL4RA IL27, Hnrmpd, CREM and Arid4b show different mRNA isoforms across multiple naïve vs differentiated T cell combinations. Overall, RI event associated genes were more prevalent (3,239 genes) than those exhibiting SE (2810 genes). SE events were associated with 10.8% (Th17), 11.2% (Treg), 12.1% (Th2) and 13.9% (Th1) of the genes, a similar trend was observed with RI events with a prevalence of 12.2% (Th17), 12.5% (Treg), 14.2% (Th1) and 14.4% (Th2) of the genes. Gene ontology results showed most of the genes showing SE and RI events are involved in process like ‘Regulation of mRNA splicing and alternative mRNA splicing via spliceosome’, ‘RNA splicing’, ‘mRNA Processing’, ‘RNA Processing’ and ‘RNA Binding’. It was also observed that Introns consistently favored retention at the 3’ end of the gene than the 5’, with upper bound showing 430 genes with introns retained at the 3’ end and 21 genes showing introns retained at the 5’ for the 72 hour time point. Enriched functional ontologies were consistently seen across all cell types to be exclusive for the genes showing RI in the 5’ end vs the 3’ end.
Keywords: T-helper cells, Alternative Splicing , Next Generation Sequencing