The conserved transcriptional regulator CdnL is required for metabolic homeostasis and morphogenesis in Caulobacter

Woldemeskel, Selamawit Abi; Daitch, Allison K.; Álvarez, Laura; Panis, Gaël; Zeinert, Rilee; González, Diego; Smith, Erika; Collier, Justine; Cava, Felipe; Viollier, Patrick H.; Goley, Erin D.

doi:10.1371/journal.pgen.1008591

Cited by 22 publications

(46 citation statements)

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“…Depletion of CarD from M. smegmatis and M. tuberculosis in vivo ( 12 , 18 ), or deletion of the C. crescentus homolog ( 19 ), suggested that CarD affects gene expression broadly, although the approaches did not distinguish direct from indirect effects. CarD colocalized with M. smegmatis RNAP in ChIP-seq experiments at all promoters recognized by the primary σ-factor in vivo ( 14 , 32 ).…”

Section: Resultsmentioning

confidence: 99%

“…However, the previous studies did not correlate the absence of −7T with activation by CarD. Deletion of C. crescentus CarD (CdnL) was reported to affect transcription from many promoters in vivo, but it was proposed that the number of promoters regulated directly by CarD was small and that most effects of CarD on transcription were likely to be indirect ( 19 ). In contrast, our data suggest that α-proteobacterial CarD is likely to affect transcription initiation from many promoters directly.…”

Section: Discussionmentioning

confidence: 98%

“…CarD family members are found in the Actinomycetes, Firmicutes , Deinococcus/Thermus , Spirochaetes , δ-proteobacteria, and most classes of α-proteobacteria, but are not found in β- and γ-proteobacteria ( 13 , 14 , 17 ). Although carD is essential in some species, and its depletion or deletion affects expression of many genes in M. tuberculosis and C. crescentus ( 12 , 17 – 19 ), a direct, widespread role for CarD in transcription from non-rRNA promoters has not been demonstrated previously with purified components in vitro.…”

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confidence: 99%

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A majority of Rhodobacter sphaeroides promoters lack a crucial RNA polymerase recognition feature, enabling coordinated transcription activation

Henry

Ross

Myers

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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Using an in vitro transcription system with purified RNA polymerase (RNAP) to investigate rRNA synthesis in the photoheterotrophic α-proteobacteriumRhodobacter sphaeroides, we identified a surprising feature of promoters recognized by the major holoenzyme. Transcription fromR. sphaeroidesrRNA promoters was unexpectedly weak, correlating with absence of −7T, the very highly conserved thymine found at the last position in −10 elements of promoters in most bacterial species. Thymine substitutions for adenine at position −7 in the three rRNA promoters strongly increased intrinsic promoter activity, indicating thatR. sphaeroidesRNAP can utilize −7T when present. rRNA promoters were activated by purifiedR. sphaeroidesCarD, a transcription factor found in many bacterial species but not in β- and γ-proteobacteria. Overall, CarD increased the activity of 15 of 16 nativeR. sphaeroidespromoters tested in vitro that lacked −7T, whereas it had no effect on three of the four native promoters that contained −7T. Genome-wide bioinformatic analysis of promoters fromR. sphaeroidesand two other α-proteobacterial species indicated that 30 to 43% contained −7T, whereas 90 to 99% of promoters from non–α-proteobacteria contained −7T. Thus, promoters lacking −7T appear to be widespread in α-proteobacteria and may have evolved away from consensus to enable their coordinated regulation by transcription factors like CarD. We observed a strong reduction inR. sphaeroidesCarD levels when cells enter stationary phase, suggesting that reduced activation by CarD may contribute to inhibition of rRNA transcription when cells enter stationary phase, the stage of growth when bacterial ribosome synthesis declines.

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Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 98%

mentioning

confidence: 99%

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A majority of Rhodobacter sphaeroides promoters lack a crucial RNA polymerase recognition feature, enabling coordinated transcription activation

Henry

Ross

Myers

et al. 2020

Proc. Natl. Acad. Sci. U.S.A.

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“…C. crescentus NA1000 cells were grown at 30 °C in peptone yeast extract (PYE) media. Antibiotics concentrations used in liquid (solid) media for C. crescentus were as follows: gentamycin 1 (5) µg mL −1 , kanamycin 5 (25) µg mL −1 , spectinomycin 25 (100) µg mL −1 , streptomycin (5 µg mL −1 ). For experiments with inducible expression of genes, inducer concentrations used were as follows: xylose – 0.3% (w/v), vanillate – 0.5 mM, glucose – 0.2% (w/v).…”

Section: Methodsmentioning

confidence: 99%

“…Cells were then outlined with meshes using phase images and fluorescence signal was analyzed. FWHM calculations of midcell mNG signal from Oufti output were performed using a custom MATLAB script (25) which fit the normalized signal output from Oufti into an eighth term Fourier series model and determined the width of the fluorescence curve at 50% of maximum intensity. Z-rings were defined using the maxima detection function in MicrobeJ and verified manually.…”

Section: Methodsmentioning

confidence: 99%

Determinants of FtsZ C-terminal linker-dependent regulation of cell wall metabolism in Caulobacter crescentus

Sundararajan

Barrows

Goley

2019

Preprint

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19Bacterial cell division requires assembly of a multi-protein machinery or "divisome" that 20 remodels the cell envelope to cause constriction. The cytoskeletal protein FtsZ forms a ring-like 21 scaffold for the divisome at the incipient division site. FtsZ has three major regions -a 22 conserved, polymerizing GTPase domain; a C-terminal conserved (CTC) peptide required for 23 binding membrane-anchoring proteins; and a C-terminal linker (CTL) of poor length and 24 sequence conservation. We previously demonstrated that, in Caulobacter crescentus, the CTL 25 regulates FtsZ polymerization in vitro and cell wall metabolism in vivo. To understand the 26 mechanism of CTL-dependent regulation of cell wall metabolism, here we investigated the 27 impact of the CTL on Z-ring structure in cells and employed genetics to identify molecular 28 determinants of the dominant lethal effects of ∆CTL. Deleting the CTL specifically resulted in 29 formation of dense, asymmetric, non-ring FtsZ assemblies in vivo. Moreover, we observed that 30 production of an FtsZ variant with the GTPase domain of Escherichia coli FtsZ fused to the CTC 31 of C. crescentus FtsZ phenocopied the effects of C. crescentus ∆CTL, suggesting the CTC 32 mediates signaling to cell wall metabolism. Finally, whereas overproduction of ZapA, FzlC, or 33 FtsEX had slight protective effects against ∆CTL, depletion of FtsA partially suppressed the 34 effects of ∆CTL. From these results, we propose that the cell wall misregulation downstream of 35 ∆CTL results from its aberrant assembly properties and is propagated through the interaction 36 between the CTC of FtsZ and FtsA. Our study provides mechanistic insights into CTL-dependent 37 regulation of cell wall enzymes downstream of FtsZ. 38 39Importance: 40 Bacterial cell division is essential and requires the recruitment and regulation of a complex 41 network of proteins needed to initiate and guide constriction and cytokinesis. FtsZ serves as a 42 master regulator for this process, and its function is highly dependent on both its self-assembly 43 into a canonical "Z-ring" and interaction with protein binding partners, which results in the 44 activation of enzymes that remodel the cell wall to drive constriction. Using mutants of FtsZ and 45 its binding partners, we have established the role of its C-terminal linker domain in regulating Z-46 ring organization, as well as the requirement for its C-terminal conserved peptide and interaction 47 with the membrane-anchoring protein FtsA for regulating cell wall remodeling for constriction. 48Introduction: 50 Bacterial cell division requires spatially-and temporally-coordinated remodeling of the cell 51 envelope to cause constriction. To this end, a multi-protein machinery called the divisome is 52 assembled at the incipient site of division. The first and most conserved protein of the divisome, 53 FtsZ, is a tubulin homolog that polymerizes into a discontinuous ring-like scaffold or "Z-ring" 54 for the recruitment of other members of the divisome (1). Over tw...

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EstG is a novel esterase required for cell envelope integrity in Caulobacter

Daitch

Orsburn²,

Chen³

et al. 2023

Current Biology

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The conserved transcriptional regulator CdnL is required for metabolic homeostasis and morphogenesis in Caulobacter

Cited by 22 publications

References 55 publications

A majority of Rhodobacter sphaeroides promoters lack a crucial RNA polymerase recognition feature, enabling coordinated transcription activation

A majority of Rhodobacter sphaeroides promoters lack a crucial RNA polymerase recognition feature, enabling coordinated transcription activation

Determinants of FtsZ C-terminal linker-dependent regulation of cell wall metabolism in Caulobacter crescentus

EstG is a novel esterase required for cell envelope integrity in Caulobacter

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