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Regulation of the transcription factor CdnL promotes adaptation to nutrient stress in Caulobacter

ABSTRACT: In response to nutrient deprivation, bacteria activate a conserved stress response pathway called the stringent response (SR). In Caulobacter crescentus, SpoT synthesizes the secondary messengers (p)ppGpp, which affect transcriptional reprogramming by binding to RNA polymerase to downregulate anabolic gene transcription. (p)ppGpp can also impact the expression of anabolic genes by controlling the levels and activities of their transcriptional regulators. In Caulobacter, a major regulator of anabolic genes is the conserved transcription factor CdnL. If and how CdnL is controlled during the SR and why that might be functionally important is unclear. Here, we show that CdnL is regulated post-translationally in a manner dependent on SpoT and the ClpXP protease. We find that stabilization of CdnL causes misregulation of ribosomal and metabolic genes during starvation. Functionally, we demonstrate that the combined action of SR transcriptional regulators and CdnL clearance allows for rapid adaptation to nutrient repletion. We also find that cells that are unable to clear CdnL during starvation are outcompeted by wild-type cells when subjected to fluctuations in nutrients. We hypothesize that post-transcriptional clearance of CdnL during the SR, in conjunction with direct binding of (p)ppGpp and DksA to RNAP, are critical for altering the transcriptome in order to permit cell survival during nutrient stress.

ORGANISM(S): Caulobacter vibrioides

PROVIDER: GSE249185 | GEO | 2024/05/10

REPOSITORIES: GEO

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Project description:Transcription profiling of wild type, relA-, and relA-spoT-, crp-, dksA-, rpoS-, lrp- mutant strains of E. coli starved for isoleucine Bacteria comprehensively reorganize their global gene expression when faced with nutrient exhaustion. In Escherichia coli and other free-living bacteria, the alarmone ppGpp facilitates this massive response by directly or indirectly coordinating the down-regulation of genes of the translation apparatus, and the induction of biosynthetic genes and the general stress response. Such a large reorientation likely requires the cooperative activities of many different genetic regulators, yet the structure of the transcription network below the level of ppGpp remains poorly defined. Using isoleucine starvation as an experimental model system for amino acid starvation, we identified genes that required ppGpp, Lrp, and RpoS for their induction. Surprisingly, despite the fact that the overwhelming majority of genes controlled by Lrp and RpoS required ppGpp for their activation, we found that these two regulons were not induced simultaneously. The data reported here suggest that metabolic genes, such as those of the Lrp regulon, require only a low basal level of ppGpp for their efficient induction. In contrast, the RpoS-dependent general stress response is not robustly induced until relatively high levels of ppGpp accumulate. Here we describe a data-driven conceptual model that explains how bacterial cells allocate transcriptional resources between metabolic and stress survival processes by discretely tuning regulatory activities to a central indicator of cellular physiology. The regulatory structure that emerges is consistent with a rheostatic model of the stringent response that allows cells to efficiently adapt to a wide range of nutritional environments. Keywords: genetic modification design; stress response; isoleucine starvation Two experiments were run. First experiment: WT and several mutant strains were starved for isoleucine (exhaustion of 60 uM ile). 40 minutes after starvation, RNA was extracted. All samples were compared to RNA from rapidly growing WT cells in identical medium replete with isoleucine (400 uM). 16 samples were hybridized: duplicates of 8 strains/conditions. Wildtype in log phase (OD = 0.4) with replete ile was control for ile starved samples. Second experiment: WT strain was starved for isoleucine (exhaustion of 60 uM ile). RNA was extracted at 12 timepoints as the cells entered stationary phase. All samples were compared to RNA from rapidly growing WT cells in identical medium replete with isoleucine (400 uM). 14 samples were hybridized: duplicates of the control condition and single timepoints during starvation. Wildtype in log phase (OD = 0.4) with replete ile was control for ile starved samples.

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Regulation of the transcription factor CdnL promotes adaptation to nutrient stress in Caulobacter

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