Investigation of the mechanisms underlying the regulation of macrophage activation and glomerular inflammation by JunD

Abstract

Macrophages are the predominant immune effector cell found in one of the most severe causes of renal failure in humans, crescentic glomerulonephritis (Crgn). The Wistar Kyoto (WLY) rat is uniquely susceptible among rat strains to Crgn and susceptibility genes have previously been mapped to seven quantitative trait loci (Crgn1-7). The AP-1 transcription factor JunD is markedly overexpressed in WKY macrophages and was identified as a major determinant of macrophage activation associated with Crgn susceptibility at Crgn2. The work presented in this thesis aimed to investigate mechanisms underlying the regulation of macrophage activation and glomerular inflammation by JunD in the NTN-susceptible WKY rat strain and in patients with Crgn. Genomic-based approaches, including microarray analysis of bone marrow-derived macrophage transcriptomes, RNA interference and cistrome analysis using ChIP-Seq, along with histological techniques and genotyping studies, were used to identify key genes and pathways underlying JunD-mediated activation of macrophages. Transcriptome analyses showed that the Crgn2 locus regulates expression of over 800 genes in WKY macrophage responses to lipopolysaccharide (LPS). Acute changes in Jund expression by RNA interference also modulated gene expression in WKY macrophages, including effector genes for the macrophage LPS response. Cistrome analysis revealed that genetically determined differences in Jund expression alter its genome binding pattern and the increased numbers of JunD-bound genes in WKY macrophages were functionally linked with signalling pathways and cell activation. Integration of the data identified 47 genes, that were associated with JunD binding peaks and whose expression was both regulated by Crgn2 and altered by siRNA knockdown of Jund, as the key candidates through which JunD determines macrophage activation. Overall, this work provides the basis for understanding genes and pathways through which JunD regulates macrophage activation and has identified novel gene targets for modulation of macrophage phenotypes in WKY rat and human macrophages.