Comparing the chromosome binding profiles of nucleoid structuring protein H-NS in the presence or absence of conditions eliciting the depletion of transcription termination protein NusG in Salmonella
Enterobacterial protein H-NS forms oligomers along DNA by binding to high affinity AT-rich nucleation sites and spreading cooperatively to adjacent sequences. In doing so, H-NS blocks RNA polymerase access to promoters and silences gene expression over extended regions. In Salmonella, H-NS silences the vast majority of virulence genes when bacteria grow outside the host. Some evidence suggests that although blocking transcription initiation, H-NS cannot stop the progression of spurious transcripts that initiate outside the H-NS-bound region. These transcripts can dislodge H-NS and render the DNA accessible to RNA polymerase and regulatory proteins. Spurious transcription is normally suppressed by the activity of termination factor Rho recruited by its cofactor NusG. In order to assess the effect of spurious transcription on H-NS binding to Salmonella pathogenicity island 1 (SPI-1), CHIP-Seq experiments were performed in cells grown in the presence or absence of treatments eliciting NusG depletion. This was achieved using strains carrying the nusG gene under the control of an arabinose-inducible repressor and growing cells in the presence or absence of arabinose (ARA). Two strains were used: strain MA13748, which carries the native SPI-1 and strain MA14513, which carries a deleted version of SPI-1 with the Anhydrotetracycline (AHTc)-inducible Ptet promoter positioned at one edge of a H-NS patch. In the latter strain, the NusG-depletion treatment was carried out while concomitantly activating Ptet with AHTc.