EPIGENETIC REGULATION IN BREVUNDIMONAS SUBVIBRIOIDES AND COMPARISON WITH THE CAULOBACTER CRESCENTUS MODEL

Epigenetics is a form of DNA methylation-based gene regulation. The CcrM methyltransferase and transcriptional regulator GcrA, constitute an epigenetic regulatory system well-conserved throughout the Alphaproteobacteria. The regulatory targets of GcrA and CcrM were identified in the Alphaproteobacterium Brevundimonas subvibrioides, and then compared to those of its close relative Caulobacter crescentus. Both these bacteria belong to the same family and inhabit the same environment. Although the regulators themselves are highly conserved, the genes they regulate are vastly different. GcrA directly regulates 204 genes in C. crescentus, and though B. subvibrioides has orthologs to 147 of those genes, only 48 genes retained GcrA binding in their promoter regions. Only 12 of those 48 genes demonstrated significant transcriptional change in a gcrA mutant, suggesting extensive transcriptional rewiring between these organisms. Similarly, out of hundreds of genes CcrM regulates in each of these organisms, only 2 genes were found in common. One of the few genes regulated by GcrA in both bacteria is a conserved hypothetical gene, bresu_2828 in B. subvibrioides (cc_0493 in C. crescentus). Bresu_2828 likely belongs to the highly conserved YbjN-like family of proteins. Deletion of bresu_2828 led to defects in surface adhesion but had no effect on holdfast production. The adhesion defect could partially be explained by compromised pilus synthesis or function, as bresu_2828 showed increased resistance to pilitropic bacteriophage. Using co-IP and bacterial two hybrid approaches, several proteins were found to interact with Bresu_2828 including GcrA, PopA and SciP. SciP is a known regulator of pilus biosynthesis in C. crescentus. Deletion of sciP in B. subvibrioides recapitulated the adhesion defect, and also resulted in complete resistance to pilitropic phage, suggesting this mutant does not produce pili. Deletion of bresu_2828 also lead to a ~2.5-fold increase in GcrA protein levels in bulk culture, suggesting that Bresu_2828 also has a role in protein stability. The results suggest Bresu_2828 has a role in regulating pilus production, potentially by influencing SciP stability. This work begins to describe a specific cellular function for a protein belonging to a broadly conserved but largely uncharacterized family