Alternative sigma factors play an important role in that they can respond with transcriptional activation to the presence of adverse conditions in their environment. The presence of multiple sigma factors is a ubiquitous feature in rhizobia. The genome of Sinorhizobium meliloti strain 1021 contains fourteen genes encoding for sigma factors, nine of which code for putative extracytoplasmic function sigma factors (RpoE) and two for heat shock sigma factors (RpoH). The purpose of this study was to gain detailed insight into the participation of sigma factors in the complex stress response system of S. meliloti 1021.

The RpoE sigma factors of S. meliloti share similarities in their predicted protein sequence and this could account for similar promoter selectivity and possible overlapping functions. Expression of sigma factors rpoE5 and rpoE1 was upregulated during oxidative stress. Induction of rpoE5 expression was also observed under cold shock and heat shock, as well as under pH stress. A deficiency in growth was observed for sigma factor mutants for rpoE2 and rpoE5 in heat shock and oxidative stress conditions. However, an extreme growth deficiency phenotype was only observed for the rpoH1 mutant at pH stress conditions.

At neutral pH, an upregulation of the rhizobactin biosynthesis operon was observed for the rpoH1 mutant. Analyses of the promoter regions of the genes involved in rhizobactin biosynthesis showed that there are possible binding sites for RpoH1 and RpoE2 in the upstream regions of rhbA, the first gene in the rhizobactin biosynthesis operon. Also, a reduction in rhbA expression was observed for the rpoH1 rpoE5 double mutant in comparison to the rpoH1 single mutant. This indicates that alternative complementarity between different sigma factors could play a role in the regulation of rhizobactin at neutral pH.

The involvement of RpoH1 in pH stress response was further analysed by time-course microarray analyes, which lead to the classification of three groups of genes, which were transcriptionally regulated in an RpoH1-independent, an RpoH1-dependent or in a complex manner. Genes regulated in an RpoH1-dependent manner are known to be involved in stress and heat shock response, like ibpA, grpE and groEL. The promoter consensus binding site for RpoH1 was identified in a number of the genes classified as being RpoH1-dependent upon pH stress, as well as for the rpoH1 gene itself. This study provided clear evidence that the sigma factor RpoH1 plays a major role in the pH stress response of S. meliloti.