ChIP-seq study of sigma factors in Pseudomonas aeruginosa

Sigma factors are master regulators of bacterial transcription which direct gene expression of specific subsets of genes. In particular, alternative sigma factors are well-known to be key players of bacterial adaptation to changing environments. To elucidate the regulatory network of sigma factors in P. aeruginosa, an integrative approach including ChIP-seq of 11 polyhistidine-tag sigma factors was performed to define the primary regulon of each sigma factor. Overall design: Sigma factor genes were fused to a polyhistidine-tag and provided in trans. Under optimal conditions regarding sigma factor activity and induction of sigma factor expression, DNA-sigma factor interactions were conserved by formaldehyde treatment. Upon DNA fragmentation by sonication, the complexes were specifically immunoprecipitated by polyclonal anti-6X His-tag antibodies and the purified DNA was analyzed by Illumina sequencing. DNA enrichment to a control strain was calculated and used for peak calling within promoter region (-500,+100) to identify directly regulated genes/operons.