Exploring the role of NF-kB in TNF mediated repression of gene expression

Abstract

Inflammation is one of the first symptomatic events in an immune response. A controlled inflammatory response is beneficial. However, it can have certain detrimental effects if unchecked (Takeuchi and Akira, 2010). Tumour Necrosis Factor (TNF) is an important mediator of inflammation, which induces NF-B during this inflammatory response. NF-B plays diverse roles including induction of genes involved in inflammation and producing pro-survival factors (Medzhitov and Horng, 2009). However, a plausible role of TNF-activated NF-B factors in transcriptional repression has not been systematically investigated at the genome scale.

Here, I obtained transcriptomic data of TNF-stimulated mouse embryonic fibroblasts (MEFs) genetically devoid of one or more NF-B monomers or monomer precursors. I then interrogated this data to dissect the relative contributions of different NF-B heterodimers in the repression of gene expression. First, I included genes that exhibited NF-B-dependent transcriptional downregulation in the analyses utilising appropriate controls in this data set. Using a clustering algorithm, I catalogued these genes into four groups, based on their repression in different genetic backgrounds. Next, in collaboration, I verified the repression of representative genes from each gene-groups by performing qRT-PCR experiments. Further, I carried out Gene Ontology analysis to explore the biological processes and pathways which are enriched in the NF-B repressed gene sets. I evaluated binding of specific NF-B factors at proximal locations of the downregulated genes by analysing the ChIP-Seq data that I received, generated using antibodies against specific NF-B factors in TNF-stimulated MEFs. I finally examined the involvement of secondary transcription factors in repression via motif de novo enrichment and transcriptomic analysis.