ATP hydrolysis tunes specificity of a AAA+ protease

Mahmoud, Samar A.; Aldikacti, Berent

doi:10.1016/j.celrep.2022.111405

Cited by 7 publications

(10 citation statements)

References 69 publications

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“…To test this, we grew cells in either oxygen limiting or standard aerobic conditions, then measured growth kinetics when cells were shifted to standard aerobic conditions. Following standard aerobic growth overnight, Δ lon cells accumulate less cell mass upon entry to stationary phase (34), an effect that was not rescued by deletion of fixT (Fig 4A). Interestingly, while the loss of fixT was well tolerated in wild type cells, deletion of fixT led to a slower growth in Δ lon strains (Fig 4A).…”

Section: Resultsmentioning

confidence: 99%

“…As FixT acts as an inhibitor of FixL autophosphorylation (29), creating a negative feedback loop, we proposed that the elevated levels of FixT protein in Lon-deficient cells should diminish the FixLJ-dependent expression of genes. We analyzed RNA-seq data (34) to identify genes that showed significant changes in the Δ lon strain compared to the wild type and compared with previously published microarray data (17) to explore whether genes upregulated by FixL exhibited downregulation in Δ lon strains consistent with a surplus of FixT protein (Fig 3A, Table S2).…”

Section: Resultsmentioning

confidence: 99%

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Proteolytic control of FixT by the Lon protease impacts FixLJ signaling inCaulobacter crescentus

Yigit,

Chien

2024

Preprint

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Responding to changes in oxygen levels is critical for aerobic microbes. In Caulobacter crescentus, low oxygen is sensed by the FixL-FixJ two-component system which induces multiple genes, including heme biosynthesis, to accommodate microaerobic conditions. The FixLJ inhibitor FixT is also induced under low oxygen conditions and is degraded by the Lon protease, which together provides negative feedback proposed to adjust FixLJ signaling thresholds during changing conditions. Here, we address if the degradation of FixT by the Lon protease contributes to phenotypic defects associated with loss of Lon. We find that Δlon strains are deficient in FixLJ-dependent heme biosynthesis, consistent with elevated FixT levels as deletion of fixT suppresses this defect. Transcriptomics validate this result as there is diminished expression of many FixLJ-activated genes in Δlon. However, no physiological changes in response to microaerobic conditions occurred upon loss of Lon, suggesting that FixT dynamics are not a major contributor to fitness in oxygen limiting conditions. Similarly, stabilization of FixT in Δlon strains does not contribute to any known Lon-related fitness defect, such as cell morphology defects or stress sensitivity. In fact, cells lacking both FixT and Lon are compromised in viability during adaptation to long term aerobic growth. Our work highlights the complexity of protease-dependent regulation of transcription factors and explains the molecular basis of defective heme accumulation in Lon-deficient Caulobacter.

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Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Proteolytic control of FixT by the Lon protease impacts FixLJ signaling inCaulobacter crescentus

Yigit,

Chien

2024

Preprint

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“…We find that the peak induction occurs at a similar time point in the wild type (Figure 1A) and Δ lon strains (Figure 4A) but the fold-induction of RecA in the Δ lon strain is less than half that in the wild type strain. We reasoned that this was due to upregulation of DNA damage response genes in a Δ lon strain even in the absence of exogenous DNA damaging agents (24). Most of the detected known DNA damage-related proteins upregulated upon MMC treatment in the wild type strain are also upregulated in the Δ lon strain (See RecA, RuvB, and MmcA in Figure 4B).…”

Section: Resultsmentioning

confidence: 99%

“…While a similar set of proteins is upregulated upon DNA damage in the Δ lon and the wild type strains, fold-inductions of proteins during the DNA damage response were generally lower in the Δ lon strain (Figure 4). We propose this is due to the fact that many DNA damage genes are already upregulated in the Δ lon strain even in the absence of exogenous DNA damage treatment (24). These data may also explain the previous observation that the Δ lon strain has an elevated sensitivity to DNA damage (29).…”

Section: Discussionmentioning

confidence: 99%

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Proteomic survey of the DNA damage response inCaulobacter crescentus

Tashjian

Zeinert

Eyles³

2023

Preprint

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The bacterial DNA damage response is a critical, coordinated response to DNA replication stress. The canonical bacterial DNA damage response, first characterized in Escherichia coli, is controlled by the global transcriptional regulator LexA and the recombinase RecA. While genome-wide studies have described how the DNA damage response is regulated at a transcriptional level, relatively little is known about post-transcriptional regulation of this response. Here, we perform a proteome-wide survey of the DNA damage response in Caulobacter crescentus. We find that not all changes in protein abundance during the response to DNA damage are predicted by changes in transcription. We validate one of these post-transcriptionally regulated candidates to show its importance to survival of DNA damage. To investigate post-translational control of the DNA damage response, we perform a similar survey in cells lacking the Lon protease. This reveals that induction of the DNA damage response at the protein level is dampened in these strains, consistent with their reduced tolerance to DNA damage. Finally, proteome-wide stability measurements following damage nominate candidate Lon substrates that suggest post-translational regulation of the DNA damage response. The DNA damage response helps bacteria to respond to and potentially survive DNA damage. The mutagenesis that is induced as part of this response plays a role in bacterial evolution and is essential to the development and spread of antibiotic resistance. Understanding how bacteria coordinate their response to DNA damage could help us to combat this growing threat to human health. While the transcriptional regulation of the bacterial DNA damage response has been characterized, this study is the first to our knowledge to compare changes in RNA and protein levels to identify potential targets of post-transcriptional regulation in response to DNA damage.

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

Smith,

Panis,

Woldemeskel

et al. 2024

PNAS Nexus

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In response to nutrient deprivation, bacteria activate a conserved stress response pathway called the stringent response (SR). During SR activation in Caulobacter crescentus, SpoT synthesizes the secondary messengers (p)ppGpp, which affect transcription by binding RNA polymerase to downregulate anabolic genes. (p)ppGpp also impacts expression of anabolic genes by controlling the levels and activities of their transcriptional regulators. In Caulobacter, a major regulator of anabolic genes is the 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 downregulated post-translationally during starvation in a manner dependent on SpoT and the ClpXP protease. Artificial stabilization of CdnL during starvation causes misregulation of ribosomal and metabolic genes. Functionally, we demonstrate that the combined action of SR transcriptional regulators and CdnL clearance allows for rapid adaptation to nutrient repletion. Moreover, cells that are unable to clear CdnL during starvation are outcompeted by wild-type cells when subjected to nutrient fluctuations. We hypothesize that clearance of CdnL during the SR, in conjunction with direct binding of (p)ppGpp and DksA to RNAP, is critical for altering the transcriptome in order to permit cell survival during nutrient stress.

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ATP hydrolysis tunes specificity of a AAA+ protease

Cited by 7 publications

References 69 publications

Proteolytic control of FixT by the Lon protease impacts FixLJ signaling inCaulobacter crescentus

Proteolytic control of FixT by the Lon protease impacts FixLJ signaling inCaulobacter crescentus

Proteomic survey of the DNA damage response inCaulobacter crescentus

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

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