Faculty Opinions recommendation of A metabolic checkpoint protein GlmR is important for diverting carbon into peptidoglycan biosynthesis in Bacillus subtilis.

ABSTRACTSelective chemical probes enable individual investigation of penicillin-binding proteins (PBPs) and provide critical information about their enzymatic activity with spatial and temporal resolution. To identify scaffolds for novel probes to study peptidoglycan biosynthesis in Bacillus subtilis, we evaluated the PBP inhibition profiles of 21 β-lactam antibiotics from different structural subclasses using a fluorescence-based assay. Most compounds readily labeled PBP1, PBP2a, PBP2b or PBP4. Almost all penicillin scaffolds were co-selective for all or combinations of PBP2a, 2b and 4. Cephalosporins, on the other hand, possessed the lowest IC50 values for PBP1 alone or along with PBP4 (ceftriaxone, cefoxitin), 2b (cefotaxime) or 2a, 2b and 4 (cephalothin). Overall, five selective inhibitors for PBP1 (aztreonam, faropenem, piperacillin, cefuroxime and cefsulodin), one selective inhibitor for PBP5 (6-aminopenicillanic acid) and various co-selective inhibitors for other PBPs in B. subtilis were discovered. Surprisingly, carbapenems strongly inhibited PBP3, formerly shown to have low affinity for β-lactams and speculated to be involved in resistance in B. subtilis. To investigate the specific roles of PBP3, we developed activity-based probes based on the meropenem core and utilized them to monitor the activity of PBP3 in living cells. We showed that PBP3 activity localizes as patches in single cells and concentrates as a ring at the septum and the division site during the cell growth cycle. Our activity-based approach enabled spatial resolution of the transpeptidation activity of individual PBPs in this model microorganism, which was not possible with previous chemical and biological approaches.

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A Checkpoint Protein That Scans the Chromosome for Damage at the Start of Sporulation in Bacillus subtilis

Cell ◽

10.1016/j.cell.2006.03.039 ◽

2006 ◽

Vol 125 (4) ◽

pp. 679-690 ◽

Cited By ~ 97

Author(s):

Michal Bejerano-Sagie ◽

Yaara Oppenheimer-Shaanan ◽

Idit Berlatzky ◽

Alex Rouvinski ◽

Mor Meyerovich ◽

...

Keyword(s):

Bacillus Subtilis ◽

Checkpoint Protein

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KinD Is a Checkpoint Protein Linking Spore Formation to Extracellular-Matrix Production in Bacillus subtilis Biofilms

mBio ◽

10.1128/mbio.00035-10 ◽

2010 ◽

Vol 1 (1) ◽

Cited By ~ 61

Author(s):

Claudio Aguilar ◽

Hera Vlamakis ◽

Alejandra Guzman ◽

Richard Losick ◽

Roberto Kolter

Keyword(s):

Extracellular Matrix ◽

Bacillus Subtilis ◽

Cell Types ◽

Deficient Mutant ◽

Content Type ◽

Checkpoint Protein ◽

Extracellular Matrix Production ◽

The Matrix ◽

Matrix Production ◽

Low Levels

ABSTRACTBacillus subtiliscells form multicellular biofilm communities in which spatiotemporal regulation of gene expression occurs, leading to differentiation of multiple coexisting cell types. These cell types include matrix-producing and sporulating cells. Extracellular matrix production and sporulation are linked in that a mutant unable to produce matrix is delayed for sporulation. Here, we show that the delay in sporulation is not due to a growth advantage of the matrix-deficient mutant under these conditions. Instead, we show that the link between matrix production and sporulation is through the Spo0A signaling pathway. Both processes are regulated by the phosphorylated form of the master transcriptional regulator Spo0A. When cells have low levels of phosphorylated Spo0A (Spo0A~P), matrix genes are expressed; however, at higher levels of Spo0A~P, sporulation commences. We have found that Spo0A~P levels are maintained at low levels in the matrix-deficient mutant, thereby delaying expression of sporulation-specific genes. This is due to the activity of one of the components of the Spo0A phosphotransfer network, KinD. A deletion ofkinDsuppresses the sporulation defect of matrix mutants, while its overproduction delays sporulation. Our data indicate that KinD displays a dual role as a phosphatase or a kinase and that its activity is linked to the presence of extracellular matrix in the biofilms. We propose a novel role for KinD in biofilms as a checkpoint protein that regulates the onset of sporulation by inhibiting the activity of Spo0A until matrix, or a component therein, is sensed.IMPORTANCEA question in the field of biofilm development has remained virtually unaddressed: how do the biofilm cells sense the completion of the synthesis of extracellular matrix? The presence of an extracellular matrix that holds the cells together is a defining feature of biofilms. How cells sense that matrix has been assembled and how this signal is transduced have not been investigated.Bacillus subtilisprovides an excellent system in which to address this question, as the molecular pathways involved in regulation of differentiation are well characterized. We provide the first evidence for a protein that senses the presence of matrix. We identify a membrane sensor histidine kinase, KinD, that alters its activity, depending on the presence or absence of the extracellular matrix. This activity feeds back to the master regulator Spo0A to regulate expression of genes involved in producing matrix and genes necessary for the progression into sporulation.

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Proteolysis of the replication checkpoint protein Sda is necessary for the efficient initiation of sporulation after transient replication stress in Bacillus subtilis

Molecular Microbiology ◽

10.1111/j.1365-2958.2006.05167.x ◽

2006 ◽

Vol 60 (6) ◽

pp. 1490-1508 ◽

Cited By ~ 35

Author(s):

Michael V. Ruvolo ◽

Kathleen E. Mach ◽

William F. Burkholder

Keyword(s):

Bacillus Subtilis ◽

Replication Stress ◽

Replication Checkpoint ◽

Checkpoint Protein

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A SpoIID Homolog Cleaves Glycan Strands at the Chlamydial Division Septum

mBio ◽

10.1128/mbio.01128-19 ◽

2019 ◽

Vol 10 (4) ◽

Cited By ~ 4

Author(s):

Nicolas Jacquier ◽

Akhilesh K. Yadav ◽

Trestan Pillonel ◽

Patrick H. Viollier ◽

Felipe Cava ◽

...

Keyword(s):

Bacillus Subtilis ◽

Intracellular Bacteria ◽

Human Pathogens ◽

Content Type ◽

Obligate Intracellular ◽

Peptidoglycan Biosynthesis ◽

Peptidoglycan Synthesis ◽

Lytic Transglycosylase ◽

Obligate Intracellular Bacteria

ABSTRACT Chlamydiales species are obligate intracellular bacteria lacking a classical peptidoglycan sacculus but relying on peptidoglycan synthesis for cytokinesis. While septal peptidoglycan biosynthesis seems to be regulated by MreB actin and its membrane anchor RodZ rather than FtsZ tubulin in Chlamydiales, the mechanism of peptidoglycan remodeling is poorly understood. An amidase conserved in Chlamydiales is able to cleave peptide stems in peptidoglycan, but it is not clear how peptidoglycan glycan strands are cleaved since no classical lytic transglycosylase is encoded in chlamydial genomes. However, a protein containing a SpoIID domain, known to possess transglycosylase activity in Bacillus subtilis, is conserved in Chlamydiales. We show here that the SpoIID homologue of the Chlamydia-related pathogen Waddlia chondrophila is a septal peptidoglycan-binding protein. Moreover, we demonstrate that SpoIID acts as a lytic transglycosylase on peptidoglycan and as a muramidase on denuded glycan strands in vitro. As SpoIID-like proteins are widespread in nonsporulating bacteria, SpoIID might commonly be a septal peptidoglycan remodeling protein in bacteria, including obligate intracellular pathogens, and thus might represent a promising drug target. IMPORTANCE Chlamydiales species are obligate intracellular bacteria and important human pathogens that have a minimal division machinery lacking the proteins that are essential for bacterial division in other species, such as FtsZ. Chlamydial division requires synthesis of peptidoglycan, which forms a ring at the division septum and is rapidly turned over. However, little is known of peptidoglycan degradation, because many peptidoglycan-degrading enzymes are not encoded by chlamydial genomes. Here we show that an homologue of SpoIID, a peptidoglycan-degrading enzyme involved in sporulation of bacteria such as Bacillus subtilis, is expressed in Chlamydiales, localizes at the division septum, and degrades peptidoglycan in vitro, indicating that SpoIID is not only involved in sporulation but also likely implicated in division of some bacteria.

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Linking the Peptidoglycan Synthesis Protein Complex with Asymmetric Cell Division during Bacillus subtilis Sporulation

International Journal of Molecular Sciences ◽

10.3390/ijms21124513 ◽

2020 ◽

Vol 21 (12) ◽

pp. 4513 ◽

Cited By ~ 1

Author(s):

Katarína Muchová ◽

Zuzana Chromiková ◽

Imrich Barák

Keyword(s):

Bacillus Subtilis ◽

Cell Division ◽

Protein Complex ◽

Asymmetric Cell Division ◽

Morphological Changes ◽

Direct Interaction ◽

Specific Protein ◽

Peptidoglycan Biosynthesis ◽

Peptidoglycan Synthesis ◽

Septum Formation

Peptidoglycan is generally considered one of the main determinants of cell shape in bacteria. In rod-shaped bacteria, cell elongation requires peptidoglycan synthesis to lengthen the cell wall. In addition, peptidoglycan is synthesized at the division septum during cell division. Sporulation of Bacillus subtilis begins with an asymmetric cell division. Formation of the sporulation septum requires almost the same set of proteins as the vegetative septum; however, these two septa are significantly different. In addition to their differences in localization, the sporulation septum is thinner and it contains SpoIIE, a crucial sporulation specific protein. Here we show that peptidoglycan biosynthesis is linked to the cell division machinery during sporulation septum formation. We detected a direct interaction between SpoIIE and GpsB and found that both proteins co-localize during the early stages of asymmetric septum formation. We propose that SpoIIE is part of a multi-protein complex which includes GpsB, other division proteins and peptidoglycan synthesis proteins, and could provide a link between the peptidoglycan synthesis machinery and the complex morphological changes required for forespore formation during B. subtilis sporulation.

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