Elucidation of the role of microRNA in innate immune response and adaptive immune response.
Chew, Chun Cheong.
Date of Issue2009
School of Biological Sciences
A*STAR Bioprocessing Technology Institute
MicroRNAs (miRNAs) are non-coding RNA sequences involved in post-transcriptional gene regulation of various biological processes in organisms. miRNA targets mRNA though miRNA recognition sites along 3’ untranslated region or even in the CDS region. In recent years, the discovery of miRNAs regulation in immunity has contributed to increasing focus on its role in immune response modulation. Our first objective was to clone 3’ UTR X Box Paired 1. Using common molecular techniques, the XBP1 3’UTR was amplified and ligated in pCR2.1® vector. The sequence had been validated using automated sequencing technology. In Baltimore et al study of miRNA-146a in THP-1 cells, he showed that miR-146a may regulate secretion of inflammatory mediator in innate immune respone. This led us to the next objective: to attempt to elucidate miRNA-146a (miR-146a) role in Toll-like receptor signaling in primary macrophages. miR-146a was overexpressed or knocked-down in primary macrophages via transfection of miR-146a precursor and miR-146a inhibitor. Interestingly, we observed that TRAF6, one of the potential targets of miR-146a, was negatively regulated when miR-146a was being knocked-down. Expression level of cytokines measured using Quantitative Real Time Polymerase Chain Reaction showed no effects by miR-146a on expression level of cytokine. Also secreted level of cytokines measured using Enzyme-linked Immunosorbent Assay (ELISA) showed that IL6 secretion was also not regulated via miRNA146a.
Final Year Project (FYP)
Nanyang Technological University