Fusion junction sequence determination Creating an inducible ssrB allele The fusion junctions of 10 SsrB-regulated fusions unlinked to SPI-2 were determined .
The next column demonstrates the specificity of the regulation to the intracellular environment by comparing the expression ratio of the three most highly SsrB-regulated fusions in a wild-type versus ssrB : : cm background in DMEM .
Results from DNase-I-protection assays provide direct evidence that SsrB binds at ssrB , although the binding sites lie within the .
Expression of an episomal copy of ssrB resulted in expression of SPI-2 genes in each mutant background , suggesting that SsrB is epistatic to other regulators for SPI-2 transcription .
Thus , one explanation for the transcription we observe following overexpression of ssrB may be that both SlyA and SsrB counteract binding of small nucleoid-like proteins .
In addition , the SsrB regulator also directly binds and autoregulates the ssrB promoters to activate their expression .
Taken together , these lines of evidence suggest that the Fur repressor could interfere with binding of the SsrB activator to the ssrB promoter , leading to repression of ssrB transcription .
Even a recent report in which the proteomes of wild type , hilA null ( a transcriptional regulator of SPI-1 genes ) and ssrB null were analyzed by SILAC and compared with an existing CHIP dataset failed to identify csgD as an SsrB-regulated locus ( Brown et al. , 2014 ) , as sequence gazing alone does not help in identifying mechanisms of transcriptional regulation .
To confirm that SsrA-dependent phosphorylation was not required for SsrB-mediated regulation of bio-films , we compared them to ssrB null mutants .