Wag31, a homologue of the cell division protein DivIVA, regulates growth, morphology and polar cell wall synthesis in mycobacteria

SummaryStaphylococcus aureus needs to control the position and timing of cell division and cell wall synthesis to maintain its spherical shape. We identified two membrane proteins, named CozEa and CozEb, which together are important for proper cell division in S. aureus. CozEa and CozEb are homologs of the cell elongation regulator CozESpn of Streptococcus pneumoniae. While cozEa and cozEb were not essential individually, the ΔcozEaΔcozEb double mutant was lethal. To study the functions of cozEa and cozEb, we constructed a CRISPR interference (CRISPRi) system for S. aureus, allowing transcriptional knockdown of essential genes. CRISPRi knockdown of cozEa in the ΔcozEb strain (and vice versa) causes cell morphological defects and aberrant nucleoid staining, showing that cozEa and cozEb have overlapping functions and are important for normal cell division. We found that CozEa and CozEb interact with the cell division protein EzrA, and that EzrA-GFP mislocalizes in the absence of CozEa and CozEb. Furthermore, the CozE-EzrA interaction is conserved in S. pneumoniae, and cell division is mislocalized in cozESpn-depleted S. pneumoniae cells. Together, our results show that CozE proteins mediate control of cell division in S. aureus and S. pneumoniae, likely via interactions with key cell division proteins such as EzrA.

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Bacteriophage SP01 Gene Product 56 Inhibits Bacillus subtilis Cell Division by Interacting with FtsL and Disrupting Pbp2B and FtsW Recruitment

Journal of Bacteriology ◽

10.1128/jb.00463-20 ◽

2020 ◽

Vol 203 (2) ◽

pp. e00463-20

Author(s):

Amit Bhambhani ◽

Isabella Iadicicco ◽

Jules Lee ◽

Syed Ahmed ◽

Max Belfatto ◽

...

Keyword(s):

Cell Wall ◽

Bacillus Subtilis ◽

Cell Division ◽

Gene Product ◽

Fluorescent Protein ◽

Lytic Bacteriophage ◽

Cell Wall Synthesis ◽

Content Type ◽

Ftsz Ring ◽

Green Fluorescent

ABSTRACTPrevious work identified gene product 56 (gp56), encoded by the lytic bacteriophage SP01, as being responsible for inhibition of Bacillus subtilis cell division during its infection. Assembly of the essential tubulin-like protein FtsZ into a ring-shaped structure at the nascent site of cytokinesis determines the timing and position of division in most bacteria. This FtsZ ring serves as a scaffold for recruitment of other proteins into a mature division-competent structure permitting membrane constriction and septal cell wall synthesis. Here, we show that expression of the predicted 9.3-kDa gp56 of SP01 inhibits later stages of B. subtilis cell division without altering FtsZ ring assembly. Green fluorescent protein-tagged gp56 localizes to the membrane at the site of division. While its localization does not interfere with recruitment of early division proteins, gp56 interferes with the recruitment of late division proteins, including Pbp2b and FtsW. Imaging of cells with specific division components deleted or depleted and two-hybrid analyses suggest that gp56 localization and activity depend on its interaction with FtsL. Together, these data support a model in which gp56 interacts with a central part of the division machinery to disrupt late recruitment of the division proteins involved in septal cell wall synthesis.IMPORTANCE Studies over the past decades have identified bacteriophage-encoded factors that interfere with host cell shape or cytokinesis during viral infection. The phage factors causing cell filamentation that have been investigated to date all act by targeting FtsZ, the conserved prokaryotic tubulin homolog that composes the cytokinetic ring in most bacteria and some groups of archaea. However, the mechanisms of several phage factors that inhibit cytokinesis, including gp56 of bacteriophage SP01 of Bacillus subtilis, remain unexplored. Here, we show that, unlike other published examples of phage inhibition of cytokinesis, gp56 blocks B. subtilis cell division without targeting FtsZ. Rather, it utilizes the assembled FtsZ cytokinetic ring to localize to the division machinery and to block recruitment of proteins needed for septal cell wall synthesis.

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The Antituberculosis Drug Ethambutol Selectively Blocks Apical Growth in CMN Group Bacteria

mBio ◽

10.1128/mbio.02213-16 ◽

2017 ◽

Vol 8 (1) ◽

Cited By ~ 11

Author(s):

Karin Schubert ◽

Boris Sieger ◽

Fabian Meyer ◽

Giacomo Giacomelli ◽

Kati Böhm ◽

...

Keyword(s):

Cell Wall ◽

Cell Division ◽

Cell Growth ◽

Antibiotic Treatment ◽

Bacterial Infections ◽

Apical Cell ◽

Combined Treatment ◽

Mycolic Acid ◽

Cell Wall Synthesis ◽

Beta Lactam

ABSTRACT Members of the genus Mycobacterium are the most prevalent cause of infectious diseases. Mycobacteria have a complex cell envelope containing a peptidoglycan layer and an additional arabinogalactan polymer to which a mycolic acid bilayer is linked; this complex, multilayered cell wall composition (mAGP) is conserved among all CMN group bacteria. The arabinogalactan and mycolic acid synthesis pathways constitute effective drug targets for tuberculosis treatment. Ethambutol (EMB), a classical antituberculosis drug, inhibits the synthesis of the arabinose polymer. Although EMB acts bacteriostatically, its underlying molecular mechanism remains unclear. Here, we used Corynebacterium glutamicum and Mycobacterium phlei as model organisms to study the effects of EMB at the single-cell level. Our results demonstrate that EMB specifically blocks apical cell wall synthesis, but not cell division, explaining the bacteriostatic effect of EMB. Furthermore, the data suggest that members of the family Corynebacterineae have two dedicated machineries for cell elongation (elongasome) and cytokinesis (divisome). IMPORTANCE Antibiotic treatment of bacterial pathogens has contributed enormously to the increase in human health. Despite the apparent importance of antibiotic treatment of bacterial infections, surprisingly little is known about the molecular functions of antibiotic actions in the bacterial cell. Here, we analyzed the molecular effects of ethambutol, a first-line antibiotic against infections caused by members of the genus Mycobacterium. We find that this drug selectively blocks apical cell growth but still allows for effective cytokinesis. As a consequence, cells survive ethambutol treatment and adopt a pneumococcal cell growth mode with cell wall synthesis only at the site of cell division. However, combined treatment of ethambutol and beta-lactam antibiotics acts synergistically and effectively stops cell proliferation. IMPORTANCE Antibiotic treatment of bacterial pathogens has contributed enormously to the increase in human health. Despite the apparent importance of antibiotic treatment of bacterial infections, surprisingly little is known about the molecular functions of antibiotic actions in the bacterial cell. Here, we analyzed the molecular effects of ethambutol, a first-line antibiotic against infections caused by members of the genus Mycobacterium. We find that this drug selectively blocks apical cell growth but still allows for effective cytokinesis. As a consequence, cells survive ethambutol treatment and adopt a pneumococcal cell growth mode with cell wall synthesis only at the site of cell division. However, combined treatment of ethambutol and beta-lactam antibiotics acts synergistically and effectively stops cell proliferation.

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Faculty Opinions recommendation of FtsA acts through FtsW to promote cell wall synthesis during cell division in Escherichia coli.

Faculty Opinions – Post-Publication Peer Review of the Biomedical Literature ◽

10.3410/f.740724443.793588916 ◽

2021 ◽

Author(s):

David Weiss

Keyword(s):

Escherichia Coli ◽

Cell Wall ◽

Cell Division ◽

Cell Wall Synthesis

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The heat-shock response of Listeria monocytogenes comprises genes involved in heat shock, cell division, cell wall synthesis, and the SOS response

Microbiology ◽

10.1099/mic.0.2007/006361-0 ◽

2007 ◽

Vol 153 (10) ◽

pp. 3593-3607 ◽

Cited By ~ 81

Author(s):

Stijn van der Veen ◽

Torsten Hain ◽

Jeroen A. Wouters ◽

Hamid Hossain ◽

Willem M. de Vos ◽

...

Keyword(s):

Cell Wall ◽

Cell Division ◽

Listeria Monocytogenes ◽

Heat Shock ◽

Heat Shock Response ◽

Sos Response ◽

Cell Wall Synthesis ◽

Shock Cell ◽

Shock Response ◽

Division Cell

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DivIVA Is Required for Polar Growth in the MreB-Lacking Rod-Shaped Actinomycete Corynebacterium glutamicum

Journal of Bacteriology ◽

10.1128/jb.01934-07 ◽

2008 ◽

Vol 190 (9) ◽

pp. 3283-3292 ◽

Cited By ~ 94

Author(s):

Michal Letek ◽

Efrén Ordóñez ◽

José Vaquera ◽

William Margolin ◽

Klas Flärdh ◽

...

Keyword(s):

Cell Wall ◽

Bacillus Subtilis ◽

Cell Division ◽

Streptococcus Pneumoniae ◽

Corynebacterium Glutamicum ◽

Chromosome Segregation ◽

Polar Growth ◽

Cell Wall Synthesis ◽

Peptidoglycan Synthesis ◽

Polarized Cell Growth

ABSTRACT The actinomycete Corynebacterium glutamicum grows as rod-shaped cells by zonal peptidoglycan synthesis at the cell poles. In this bacterium, experimental depletion of the polar DivIVA protein (DivIVACg) resulted in the inhibition of polar growth; consequently, these cells exhibited a coccoid morphology. This result demonstrated that DivIVA is required for cell elongation and the acquisition of a rod shape. DivIVA from Streptomyces or Mycobacterium localized to the cell poles of DivIVACg-depleted C. glutamicum and restored polar peptidoglycan synthesis, in contrast to DivIVA proteins from Bacillus subtilis or Streptococcus pneumoniae, which localized at the septum of C. glutamicum. This confirmed that DivIVAs from actinomycetes are involved in polarized cell growth. DivIVACg localized at the septum after cell wall synthesis had started and the nucleoids had already segregated, suggesting that in C. glutamicum DivIVA is not involved in cell division or chromosome segregation.

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AUTORADIOGRAPHIC STUDIES OF BACTERIAL CELL WALL REPLICATION: I. CELL WALL GROWTH OF BACILLUS CEREUS IN THE PRESENCE OF CHLORAMPHENICOL

Canadian Journal of Microbiology ◽

10.1139/m67-046 ◽

1967 ◽

Vol 13 (4) ◽

pp. 341-350 ◽

Cited By ~ 7

Author(s):

K. L. Chung

Keyword(s):

Cell Wall ◽

Cell Division ◽

Bacillus Cereus ◽

First Generation ◽

Third Generation ◽

Cell Wall Synthesis ◽

Daughter Cells ◽

Entire Cell ◽

Wall Growth ◽

The Difference

The pattern of cell wall synthesis as measured by the incorporation of tritiated alanine into the cell wall of Bacillus cereus, and the number of synthesizing sites in the cell wall were studied by the direct and the reverse autoradiographic labelling methods.In the absence of chloramphenicol, the new cell wall was initiated at two or three segments, and later increased to four or five segments which continued to elongate but not to increase in number until the bacilli had made preparation for cell division. Shortly before the centripetal growth of the cell wall and constriction to separate daughter cells, two to three more new wall-segments were added to those already present. The second and third generation cells retained some old wall-segments from the first-generation mother, which remained as discrete clusters of grains, and could easily be distinguished from the new segments.In the presence of chloramphenicol, the new wall was initiated at 8 to 10 sites. Further incubation resulted in the uniform incorporation of labels at multiple sites along the entire cell length.The patterns of new wall replication as studied by the two methods were compared. To account for the difference in synthesizing sites when chloramphenicol is present, it is suggested that the cells have either used the maximum number of sites or have completely bypassed all the sites and allowed the tritiated alanine to diffuse into the wall to become incorporated.

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A conserved subcomplex within the bacterial cytokinetic ring activates cell wall synthesis by the FtsW-FtsI synthase

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.2004598117 ◽

2020 ◽

Vol 117 (38) ◽

pp. 23879-23885 ◽

Cited By ~ 1

Author(s):

Lindsey S. Marmont ◽

Thomas G. Bernhardt

Keyword(s):

Pseudomonas Aeruginosa ◽

Cell Wall ◽

Cell Division ◽

Polymerase Activity ◽

Bacterial Cell Division ◽

Cell Wall Synthesis ◽

Negative Regulators ◽

Major Function ◽

Osmotic Lysis ◽

Direct Activator

Cell division in bacteria is mediated by a multiprotein assembly called the divisome. A major function of this machinery is the synthesis of the peptidoglycan (PG) cell wall that caps the daughter poles and prevents osmotic lysis of the newborn cells. Recent studies have implicated a complex of FtsW and FtsI (FtsWI) as the essential PG synthase within the divisome; however, how PG polymerization by this synthase is regulated and coordinated with other activities within the machinery is not well understood. Previous results have implicated a conserved subcomplex of division proteins composed of FtsQ, FtsL, and FtsB (FtsQLB) in the regulation of FtsWI, but whether these proteins act directly as positive or negative regulators of the synthase has been unclear. To address this question, we purified a five-memberPseudomonas aeruginosadivision complex consisting of FtsQLB-FtsWI. The PG polymerase activity of this complex was found to be greatly stimulated relative to FtsWI alone. Purification of complexes lacking individual components indicated that FtsL and FtsB are sufficient for FtsW activation. Furthermore, support for this activity being important for the cellular function of FtsQLB was provided by the identification of two division-defective variants of FtsL that still form normal FtsQLB-FtsWI complexes but fail to activate PG synthesis. Thus, our results indicate that the conserved FtsQLB complex is a direct activator of PG polymerization by the FtsWI synthase and thereby define an essential regulatory step in the process of bacterial cell division.

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Aerosol survival of Serratia marcescens as a function of oxygen concentration, relative humidity, and time

Canadian Journal of Microbiology ◽

10.1139/m74-239 ◽

1974 ◽

Vol 20 (11) ◽

pp. 1529-1534 ◽

Cited By ~ 9

Author(s):

C. S. Cox ◽

S. J. Gagen ◽

Jean Baxter

Keyword(s):

Cell Wall ◽

Cell Division ◽

Relative Humidity ◽

Serratia Marcescens ◽

Oxygen Concentration ◽

Freeze Drying ◽

Cytoplasmic Membrane ◽

Cell Wall Synthesis ◽

Freeze Dried ◽

Kinetics Of

Previously the kinetics of loss of viability of freeze-dried Serratia marcescens 8UK were determined by Cox and Heckly as a function of oxygen concentration and time. Results are presented here when dehydration is brought about by aerosolization into atmospheres of low relative humidity (RH) rather than by freeze-drying. As for freeze-dried S. marcescens, oxygen was toxic and viable decay followed the same kinetics with respect to oxygen concentration and time. The influence of RH upon viable decay (which was not studied in the previous report) was that above 65% RH oxygen was not toxic but was progressively more toxic as the humidity was further reduced. Kinetic analyses of the results indicate that the site for the toxic action of oxygen lies in the interspace between the cytoplasmic membrane and the cell wall. Such a finding is consistent with other data which suggest that cell division and (or) cell wall synthesis in bacteria are inhibited by oxygen.

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