Dual RpoH sigma factors and transcriptional plasticity in a symbiotic bacterium

Abstract

Sinorhizobium meliloti can live as a soil saprophyte and can engage in a nitrogen-fixing symbiosis with plant roots. To succeed in such diverse environments, the bacteria must continually adjust gene expression. Transcriptional plasticity in eubacteria is often mediated by alternative sigma (σ) factors interacting with core RNA polymerase. The S. meliloti genome encodes 14 of these alternative σ factors, including two putative RpoH ("heat shock") σ factors. We used custom Affymetrix symbiosis chips to characterize the global transcriptional response of S. meliloti rpoH1, rpoH2, and rpoH1 rpoH2 mutants during heat shock and stationary-phase growth. Under these conditions, expression of over 300 genes is dependent on rpoH1 and rpoH2. We mapped transcript start sites of 69 rpoH-dependent genes using 5' RACE (5' rapid amplification of cDNA ends), which allowed us to determine putative RpoH1-dependent, RpoH2-dependent, and dual-promoter (RpoH1- and RpoH2-dependent) consensus sequences that were each used to search the genome for other potential direct targets of RpoH. The inferred S. meliloti RpoH promoter consensus sequences share features of Escherichia coli RpoH promoters but lack extended -10 motifs.

Fig 1

Intersection of rpoH1-dependent genes with heat shock- and acid shock-induced genes. Genes induced by heat shock were identified in this study, and genes induced by acid shock were determined by de Lucena et al. (10). Seventeen genes whose expression was significantly decreased in the rpoH1 rpoH2 double mutant during heat shock and whose expression was also decreased in the rpoH1 mutant but with an SLR of more than −0.96 are also shown in this diagram. Genes whose expression was rpoH1 dependent during acid shock but that are not themselves induced by acid shock in the wild type were not identified by de Lucena et al.; therefore, this diagram shows only 85 genes that were rpoH1 dependent in heat shock, but not themselves induced by heat or acid shock. Not shown on the diagram are 12 genes whose expression increased with both heat and acid shock treatments but that were rpoH1 dependent in only one of these treatments: 4 of these genes were rpoH1 dependent during heat shock, and 8 were rpoH1 dependent during acid shock.

Fig 2

S. meliloti rpoH-dependent promoter consensus motifs. The sequence logos for three gene sets are shown. Gene set 1 shows RpoH1-specific promoter consensus motifs (20 genes). Gene set 2 shows RpoH2-specific promoter consensus motifs (11 genes). Gene set 3 shows dual-promoter (RpoH1 and RpoH2) consensus motifs (14 genes). Colored sequence logos for gene sets 1, 2, and 3 of Table 1 were generated using MEME as described in Materials and Methods. The total information content of each logo is given in bits; the height of each nucleotide in the logo represents the positional probability of that nucleotide multiplied by the information content of the logo. −35 consensus motifs are displayed on the left side of the figure, and −10 consensus motifs are displayed on the right side of the figure. Nonconserved spacer regions of 15 to 16 nt between the motifs are not shown.