Genetic Screens Identify Additional Genes Implicated in Envelope Remodeling during the Engulfment Stage of Bacillus subtilis Sporulation

Chan, Helena; Taïb, Najwa; Gilmore, Michael C.; Mohamed, A. M.; Hanna, Kieran; Luhur, Johana; Nguyen, Hieu; Hafiz, Elham; Cava, Felipe; Gribaldo, Simonetta; Rudner, David Z.; Rodrigues, Christopher D. A.

doi:10.1128/mbio.01732-22

Cited by 7 publications

(9 citation statements)

References 62 publications

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“…Notably, these duplications contain murZ (Rued et al, 2017; Wamp et al, 2020), which encodes one of two homologs of the UDP‐N‐acetylglucosamine 1‐carboxyvinyltransferase that converts PEP and UDP‐GlcNAc to Pi and UDP‐N‐acetyl‐3‐O‐(1‐carboxyvinyl)‐alpha‐D‐glucosamine in the first committed step in the synthesis of the PG precursor Lipid II (Brown et al, 1995; Zhou et al, 2022). Like other low‐GC Gram‐positive bacteria, S. pneumoniae encodes two distinct homologs of this enzyme (Figure 1) (Blake et al, 2009; Chan et al, 2022; Du et al, 2000; Kedar et al, 2008; Kock et al, 2004; Mascari et al, 2022; Vesic & Kristich, 2012). The two homologs in S. pneumoniae strains were annotated as MurZ (MurA2) (Spd_0967) and MurA (MurA1) ( Spn )(Spd_1764) (Hoskins et al, 2001) (Figure 1).…”

Section: Introductionmentioning

confidence: 99%

Negative regulation of MurZ and MurA underlies the essentiality of GpsB‐ and StkP‐mediated protein phosphorylation in Streptococcus pneumoniaeD39

Tsui

Joseph

Zheng

et al. 2023

Molecular Microbiology

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GpsB links peptidoglycan synthases to other proteins that determine the shape of the respiratory pathogen Streptococcus pneumoniae (pneumococcus; Spn) and other low‐GC Gram‐positive bacteria. GpsB is also required for phosphorylation of proteins by the essential StkP(Spn) Ser/Thr protein kinase. Here we report three classes of frequently arising chromosomal duplications (≈21–176 genes) containing murZ (MurZ‐family homolog of MurA) or murA that suppress ΔgpsB or ΔstkP. These duplications arose from three different repeated sequences and demonstrate the facility of pneumococcus to modulate gene dosage of numerous genes. Overproduction of MurZ or MurA alone or overproduction of MurZ caused by ΔkhpAB mutations suppressed ΔgpsB or ΔstkP phenotypes to varying extents. ΔgpsB and ΔstkP were also suppressed by MurZ amino‐acid changes distant from the active site, including one in commonly studied laboratory strains, and by truncation or deletion of the homolog of IreB(ReoM). Unlike in other Gram‐positive bacteria, MurZ is predominant to MurA in pneumococcal cells. However, ΔgpsB and ΔstkP were not suppressed by ΔclpCP, which did not alter MurZ or MurA amounts. These results support a model in which regulation of MurZ and MurA activity, likely by IreB(Spn), is the only essential requirement for StkP‐mediated protein phosphorylation in exponentially growing D39 pneumococcal cells.

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Section: Introductionmentioning

confidence: 99%

Negative regulation of MurZ and MurA underlies the essentiality of GpsB‐ and StkP‐mediated protein phosphorylation in Streptococcus pneumoniaeD39

Tsui

Joseph

Zheng

et al. 2023

Molecular Microbiology

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“…These observations reinforce the interpretation that Dpp is a murein tripeptide transporter that functions in the uptake and recycling of cell wall peptides. Moreover, dpp regulation by CodY, and the persistence of the dpp transcript under nutrient-limiting conditions and during sporulation [55] are consistent with (i) the need to conserve resources in stationary phase and (ii) the extensive cell wall restructuring that accompanies asymmetric septation and mother cell engulfment of the forespore during spore formation [56].…”

Section: Discussionmentioning

confidence: 95%

Peptide transport in Bacillus subtilis – structure and specificity in the extracellular solute binding proteins OppA and DppE

et al. 2022

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Peptide transporters play important nutritional and cell signalling roles in Bacillus subtilis, which are pronounced during stationary phase adaptations and development. Three high-affinity ATP-binding cassette (ABC) family transporters are involved in peptide uptake – the oligopeptide permease (Opp), another peptide permease (App) and a less well-characterized dipeptide permease (Dpp). Here we report crystal structures of the extracellular substrate binding proteins, OppA and DppE, which serve the Opp and Dpp systems, respectively. The structure of OppA was determined in complex with endogenous peptides, modelled as Ser-Asn-Ser-Ser, and with the sporulation-promoting peptide Ser-Arg-Asn-Val-Thr, which bind with K d values of 0.4 and 2 µM, respectively, as measured by isothermal titration calorimetry. Differential scanning fluorescence experiments with a wider panel of ligands showed that OppA has highest affinity for tetra- and penta-peptides. The structure of DppE revealed the unexpected presence of a murein tripeptide (MTP) ligand, l-Ala-d-Glu-meso-DAP, in the peptide binding groove. The mode of MTP binding in DppE is different to that observed in the murein peptide binding protein, MppA, from Escherichia coli , suggesting independent evolution of these proteins from an OppA-like precursor. The presence of MTP in DppE points to a role for Dpp in the uptake and recycling of cell wall peptides, a conclusion that is supported by analysis of the genomic context of dpp, which revealed adjacent genes encoding enzymes involved in muropeptide catabolism in a gene organization that is widely conserved in Firmicutes .

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“…Remarkably, MurAB(Bsu) was discovered to be required for efficient spore engulfment during sporulation of B. subtilis (Chan et al, 2022). S. pneumoniae does not sporulate.…”

Section: Discussionmentioning

confidence: 99%

“…Likewise, MurZ( Spn ) and its MurZ-family homolog MurAB( Bsu ) play very different physiological roles. Remarkably, MurAB( Bsu ) was discovered to be required for efficient spore engulfment during sporulation of B. subtilis (Chan et al ., 2022). S. pneumoniae does not sporulate.…”

Section: Discussionmentioning

confidence: 99%

“…Like other low-GC Gram-positive bacteria, S. pneumoniae encodes two distinct homologs of this enzyme (Fig. 1) (Blake et al ., 2009, Chan et al ., 2022, Du et al ., 2000, Kedar et al ., 2008, Kock et al ., 2004, Mascari et al ., 2022, Vesic & Kristich, 2012). The two homologs in S. pneumoniae strains were annotated as MurZ (MurA2) (Spd_0967) and MurA (MurA1) ( Spn )(Spd_1764) (Hoskins et al ., 2001) (Fig.…”

Section: Introductionmentioning

confidence: 99%

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Chromosomal Duplications of MurZ (MurA2) or MurA (MurA1), Amino Acid Substitutions in MurZ (MurA2), and Absence of KhpAB Obviate the Requirement for Protein Phosphorylation inStreptococcus pneumoniaeD39

Tsui

Joseph

Zheng

et al. 2023

Preprint

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GpsB links peptidoglycan synthases to other proteins that determine the shape of the respiratory pathogenStreptococcus pneumoniae(pneumococcus;Spn) and other low-GC Gram-positive bacteria. GpsB is also required for phosphorylation of proteins by the essential StkP(Spn) Ser/Thr protein kinase. Here we report three classes of frequently arising chromosomal duplications (≈21-176 genes) containingmurZ(MurZ-family homolog of MurA) ormurAthat suppress ΔgpsBor ΔstkP. These duplications arose from three different repeated sequences and demonstrate the facility of pneumococcus to modulate gene dosage of numerous genes. Overproduction of MurZ or MurA alone or overexpression of MurZ caused by ΔkhpABmutations suppressed ΔgpsBor ΔstkPphenotypes to varying extents. ΔgpsBand ΔstkPwere also suppressed by MurZ amino-acid changes distant from the active site, including one in commonly studied laboratory strains, and by truncation or deletion of the homolog of IreB(ReoM). Unlike in other Gram-positive bacteria, MurZ is predominant to MurA in pneumococcal cells. However, ΔgpsBand ΔstkPwere not suppressed by ΔclpCP, which did not alter MurZ or MurA amounts. These results support a model in which regulation of MurZ and MurA activity, likely by IreB(Spn), is the only essential requirement for protein phosphorylation in exponentially growing D39 pneumococcal cells.

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Genetic Screens Identify Additional Genes Implicated in Envelope Remodeling during the Engulfment Stage of Bacillus subtilis Sporulation

Abstract: In bacteria, cell envelope remodeling is critical for cell growth and division. This is also the case during the development of bacteria into highly resistant endospores (spores), known as sporulation.

Cited by 7 publications

References 62 publications

Negative regulation of MurZ and MurA underlies the essentiality of GpsB‐ and StkP‐mediated protein phosphorylation in Streptococcus pneumoniaeD39

Negative regulation of MurZ and MurA underlies the essentiality of GpsB‐ and StkP‐mediated protein phosphorylation in Streptococcus pneumoniaeD39

Peptide transport in Bacillus subtilis – structure and specificity in the extracellular solute binding proteins OppA and DppE

Chromosomal Duplications of MurZ (MurA2) or MurA (MurA1), Amino Acid Substitutions in MurZ (MurA2), and Absence of KhpAB Obviate the Requirement for Protein Phosphorylation inStreptococcus pneumoniaeD39

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