Importance of RpoD- and Non-RpoD-Dependent Expression of Horizontally Acquired Genes in Cupriavidus metallidurans

Abstract

The genome of the metal-resistant, hydrogen-oxidizing bacterium Cupriavidus metallidurans contains a large number of horizontally acquired plasmids and genomic islands that were integrated into its chromosome or chromid. For the C. metallidurans CH34 wild-type strain growing under nonchallenging conditions, 5,763 transcriptional starting sequences (TSSs) were determined. Using a custom-built motif discovery software based on hidden Markov models, patterns upstream of the TSSs were identified. The pattern TTGACA, -35.6 ± 1.6 bp upstream of the TSSs, in combination with a TATAAT sequence 15.8 ± 1.4 bp upstream occurred frequently, especially upstream of the TSSs for 48 housekeeping genes, and these were assigned to promoters used by RNA polymerase containing the main housekeeping sigma factor RpoD. From patterns upstream of the housekeeping genes, a score for RpoD-dependent promoters in C. metallidurans was derived and applied to all 5,763 TSSs. Among these, 2,572 TSSs could be associated with RpoD with high probability, 373 with low probability, and 2,818 with no probability. In a detailed analysis of horizontally acquired genes involved in metal resistance and not involved in this process, the TSSs responsible for the expression of these genes under nonchallenging conditions were assigned to RpoD- or non-RpoD-dependent promoters. RpoD-dependent promoters occurred frequently in horizontally acquired metal resistance and other determinants, which should allow their initial expression in a new host. However, other sigma factors and sense/antisense effects also contribute-maybe to mold in subsequent adaptation steps the assimilated gene into the regulatory network of the cell. IMPORTANCE In their natural environment, bacteria are constantly acquiring genes by horizontal gene transfer. To be of any benefit, these genes should be expressed. We show here that the main housekeeping sigma factor RpoD plays an important role in the expression of horizontally acquired genes in the metal-resistant hydrogen-oxidizing bacterium C. metallidurans. By conservation of the RpoD recognition consensus sequence, a newly arriving gene has a high probability to be expressed in the new host cell. In addition to integrons and genes travelling together with that for their sigma factor, conservation of the RpoD consensus sequence may be an important contributor to the overall evolutionary success of horizontal gene transfer in bacteria. Using C. metallidurans as an example, this publication sheds some light on the fate and function of horizontally acquired genes in bacteria.

Keywords: Cupriavidus metallidurans; RpoD; metal resistance.

FIG 1

Overview of the motifs discovered in the −10 and −35 regions. For each motif, a sequence logo and a histogram of the positions of motif matches are shown. In the −10 region, 5 motifs were discovered (from the top to the bottom, components 0,1 to 0,5), of which one (component 0,1 at the top) covered the majority of TSSs and showed strict positioning. Only promoters of TSSs containing this motif were considered when searching for a second, upstream motif. Of the 5 motifs discovered, only two components (C = 1,1 and C = 2,1) were clearly positioned and showed similarities to known −35 consensus sequences.

FIG 2

Map of the cobalt-zinc-cadmium resistance determinant czc on plasmid pMOL30. The map shows the first part of the czc determinants in the indicated regions with NPKM values on one DNA strand (red) or the other direction of transcription (blue). Above are the Rmet locus and gene names, the mean NPKM, and response values (24). TSSs (flags) are indicated with the corresponding TSS score as white with a score of <50, shades of red for strong (>1,000) or orange for medium (50 to 1000) transcription initiation from RpoD promoters (medium or strong RpoD score), and shades of blue if not associated or only weakly associated with the RpoD model. Transcripts (48) and operons (25) are also indicated in the transcript map. TSSs with scores of <50 (white arrows) are shown without TSS scores.