An RpoHI-Dependent Response Promotes Outgrowth after Extended Stationary Phase in the Alphaproteobacterium Rhodobacter sphaeroides

Abstract

Under unfavorable growth conditions, bacteria enter stationary phase and can maintain cell viability over prolonged periods with no increase in cell number. To obtain insights into the regulatory mechanisms that allow bacteria to resume growth when conditions become favorable again (outgrowth), we performed global transcriptome analyses at different stages of growth for the alphaproteobacterium Rhodobacter sphaeroides. The majority of genes were not differentially expressed across growth phases. After a short stationary phase (about 20 h after growth starts to slow down), only 7% of the genes showed altered expression (fold change of Ͼ1.6 or less than Ϫ1.6, corresponding to a log 2 fold change of Ͼ0.65 or less than Ϫ0.65, respectively) compared to expression at exponential phase. Outgrowth induced a distinct response in gene expression which was strongly influenced by the length of the preceding stationary phase. After a long stationary phase (about 64 h after growth starts to slow down), a much larger number of genes (15.1%) was induced in outgrowth than after a short stationary phase (1.7%). Many of those genes are known members of the RpoHI/RpoHII regulons and have established functions in stress responses. A main effect of RpoHI on the transcriptome in outgrowth after a long stationary phase was confirmed. Growth experiments with mutant strains further support an important function in outgrowth after prolonged stationary phase for the RpoHI and RpoHII sigma factors.IMPORTANCE In natural environments, the growth of bacteria is limited mostly by lack of nutrients or other unfavorable conditions. It is important for bacterial populations to efficiently resume growth after being in stationary phase, which may last for long periods. Most previous studies on growth-phase-dependent gene expression did not address outgrowth after stationary phase. This study on growth-phase-dependent gene regulation in a model alphaproteobacterium reveals, for the first time, that the length of the stationary phase strongly impacts the transcriptome during outgrowth. The alternative sigma factors RpoHI and RpoHII, which are important regulators of stress responses in alphaproteobacteria, play a major role during outgrowth following prolonged stationary phase. These findings provide the first insight into the regulatory mechanisms enabling efficient outgrowth.

Adaptation of the Alphaproteobacterium Rhodobacter sphaeroides to stationary phase

“…The subset of genes with decreased transcripts in the transition phase by the presence of oxygen provide further support for the reliability of our data since this set of genes include not only genes responsible for photosynthesis (Fig. S2) but also cco genes (cbb3 type cytochrome oxidase), feoA and feoB (Fe 2+ transport), nuo genes (NADH dehydrogenase), and genes for some other metabolic enzymes that are known to strongly depend on the redox conditions (Zeilstra‐Ryalls and Kaplan, ; Arai et al ., ; Remes et al ., ).…”

“…Previous work has highlighted the importance of the RpoHI sigma factor during standard growth and following the transfer from the stationary phase to fresh medium (Remes et al ., ). Here, our transcriptome data revealed the expression pattern of a gene coding for another sigma factor, RSP_3095.…”

“…For example, although the RpoH/RpoE sigma factors did not appear among our candidates with changed expression in the transition phase (see Fig. for the promoter activity of rpoH1 ), deletions of these sigma factor genes cause poor growth under standard growth conditions (Nuss et al ., ; Nuss et al ., ; Remes et al ., ). Finally, our analysis did not consider all genes with increased transcript levels in the transition phase.…”

“…sphaeroides rpoZ‐relA locus shares a similar genetic topology. Other promoters included in our analysis are the promoter of rpoHI , coding for an alternative sigma factor (Remes et al ., ), the promoter of takP , a TRAP transporter for the uptake of carbon (Adnan et al ., ) and the promoter of the 16S rRNA gene, previously used in R . sphaeroides for the strong constitutive overexpression of target genes (Mank et al ., ).…”

“…How do the bacteria achieve this? The first step is generally thought to be a remodelling of their transcriptomes through diverse transcriptional regulators such as alternative sigma factors (Navarro Llorens et al, 2010), which are capable of rapidly inducing changes in gene expression within minutes of detecting environmental change (Rolfe et al, 2012;Remes et al, 2017).…”