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Cell Reports Jul 2023Bacterial cell-wall hydrolases must be tightly regulated during bacterial cell division to prevent aberrant cell lysis and to allow final separation of viable daughter...
Bacterial cell-wall hydrolases must be tightly regulated during bacterial cell division to prevent aberrant cell lysis and to allow final separation of viable daughter cells. In a multidisciplinary work, we disclose the molecular dialogue between the cell-wall hydrolase LytB, wall teichoic acids, and the eukaryotic-like protein kinase StkP in Streptococcus pneumoniae. After characterizing the peptidoglycan recognition mode by the catalytic domain of LytB, we further demonstrate that LytB possesses a modular organization allowing the specific binding to wall teichoic acids and to the protein kinase StkP. Structural and cellular studies notably reveal that the temporal and spatial localization of LytB is governed by the interaction between specific modules of LytB and the final PASTA domain of StkP. Our data collectively provide a comprehensive understanding of how LytB performs final separation of daughter cells and highlights the regulatory role of eukaryotic-like kinases on lytic machineries in the last step of cell division in streptococci.
Topics: Streptococcus pneumoniae; Protein Serine-Threonine Kinases; Teichoic Acids; Bacterial Proteins; Cell Division; Protein Kinases; Hydrolases; Cell Wall
PubMed: 37418323
DOI: 10.1016/j.celrep.2023.112756 -
Frontiers in Cellular and Infection... 2023The pilus is an extracellular structural part that can be detected in some () isolates (type I pili are found in approximately 30% of strains, while type II pili are... (Review)
Review
The pilus is an extracellular structural part that can be detected in some () isolates (type I pili are found in approximately 30% of strains, while type II pili are found in approximately 20%). It is anchored to the cell wall by LPXTG-like motifs on the peptidoglycan. Two kinds of pili have been discovered, namely, pilus-1 and pilus-2. The former is encoded by pilus islet 1 (PI-1) and is a polymer formed by the protein subunits RrgA, RrgB and RrgC. The latter is encoded by pilus islet 2 (PI-2) and is a polymer composed mainly of the structural protein PitB. Although pili are not necessary for the survival of , they serve as the structural basis and as virulence factors that mediate the adhesion of bacteria to host cells and play a direct role in promoting the adhesion, colonization and pathogenesis of . In addition, as candidate antigens for protein vaccines, pili have promising potential for use in vaccines with combined immunization strategies. Given the current understanding of the pili of regarding the genes, proteins, structure, biological function and epidemiological relationship with serotypes, combined with the immunoprotective efficacy of pilins as protein candidates for vaccines, we here systematically describe the research status and prospects of pili and provide new ideas for subsequent vaccine research and development.
Topics: Bacterial Proteins; Streptococcus pneumoniae; Fimbriae, Bacterial; Fimbriae Proteins; Vaccines; Polymers
PubMed: 37799336
DOI: 10.3389/fcimb.2023.1270848 -
MicrobiologyOpen Oct 2023Peptidoglycan for elongation in Escherichia coli is synthesized by the Rod complex, which includes RodZ. Although various mutant strains of the Rod complex have been...
Peptidoglycan for elongation in Escherichia coli is synthesized by the Rod complex, which includes RodZ. Although various mutant strains of the Rod complex have been isolated, the relationship between the activity of the Rod complex and the overall physical and chemical structures of the peptidoglycan have not been reported. We constructed a RodZ mutant, termed RMR, and analyzed the growth rate, morphology, and other characteristics of cells producing the Rod complexes containing RMR. The growth and morphology of RMR cells were abnormal, and we isolated suppressor mutants from RMR cells. Most of the suppressor mutations were found in components of the Rod complex, suggesting that these suppressor mutations increase the integrity and/or the activity of the Rod complex. We purified peptidoglycan from wild-type, RMR, and suppressor mutant cells and observed their structures in detail. We found that the peptidoglycan purified from RMR cells had many large holes and different compositions of muropeptides from those of WT cells. The Rod complex may be a determinant not only for the whole shape of peptidoglycan but also for its highly dense structure to support the mechanical strength of the cell wall.
Topics: Escherichia coli; Escherichia coli Proteins; Peptidoglycan; Cytoskeletal Proteins; Cell Wall
PubMed: 37877652
DOI: 10.1002/mbo3.1385 -
Proceedings of the National Academy of... Oct 2023Bacteria produce a structural layer of peptidoglycan (PG) that enforces cell shape, resists turgor pressure, and protects the cell. As bacteria grow and divide, the...
Bacteria produce a structural layer of peptidoglycan (PG) that enforces cell shape, resists turgor pressure, and protects the cell. As bacteria grow and divide, the existing layer of PG is remodeled and PG fragments are released. Enterics such as go to great lengths to internalize and reutilize PG fragments. is estimated to break down one-third of its cell wall, yet only loses ~0 to 5% of meso-diaminopimelic acid, a PG-specific amino acid, per generation. Two transporters were identified early on to possibly be the primary permease that facilitates PG fragment recycling, i) AmpG and ii) the Opp ATP binding cassette transporter in conjunction with a PG-specific periplasmic binding protein, MppA. The contribution of each transporter to PG recycling has been debated. Here, we have found that AmpG and MppA/Opp are differentially regulated by carbon source and growth phase. In addition, MppA/Opp is uniquely capable of high-affinity scavenging of muropeptides from growth media, demonstrating that AmpG and MppA/Opp allow for different strategies of recycling PG fragments. Altogether, this work clarifies environmental contexts under which utilizes distinct permeases for PG recycling and explores how scavenging by MppA/Opp could be beneficial in mixed communities.
Topics: Membrane Transport Proteins; Escherichia coli; Peptidoglycan; Bacterial Proteins; Bacteria; Cell Wall
PubMed: 37871219
DOI: 10.1073/pnas.2308940120 -
PeerJ 2023The rapid emergence of antibiotic-resistant bacteria directly contributes to a wave of untreatable infections. The lack of new drug development is an important driver of...
BACKGROUND
The rapid emergence of antibiotic-resistant bacteria directly contributes to a wave of untreatable infections. The lack of new drug development is an important driver of this crisis. Most antibiotics today are small molecules that block vital processes in bacteria. To optimize such effects, the three-dimensional structure of targeted bacterial proteins is imperative, although such a task is time-consuming and tedious, impeding the development of antibiotics. The development of RNA-based therapeutics has catalyzed a new platform of antibiotics-antisense oligonucleotides (ASOs). These molecules hybridize with their target mRNAs with high specificity, knocking down or interfering with protein translation. This study aims to develop a bioinformatics pipeline to identify potent ASO targets in essential bacterial genes.
METHODS
Three bacterial species (, , and ) were used to demonstrate the utility of the pipeline. Open reading frames of bacterial essential genes were downloaded from the Database of Essential Genes (DEG). After filtering for specificity and accessibility, ASO candidates were ranked based on their self-hybridization score, predicted melting temperature, and the position on the gene in an operon. Enrichment analysis was conducted on genes associated with putative potent ASOs.
RESULTS
A total of 45,628 ASOs were generated from 348 unique essential genes in . A total of 1,117 of them were considered putative. A total of 27,273 ASOs were generated from 191 unique essential genes in . A total of 847 of them were considered putative. A total of 175,606 ASOs were generated from 346 essential genes in . A total of 7,061 of them were considered putative. Critical biological processes associated with these genes include translation, regulation of cell shape, cell division, and peptidoglycan biosynthetic process. Putative ASO targets generated for each bacterial species are publicly available here: https://github.com/EricSHo/AOA. The results demonstrate that our bioinformatics pipeline is useful in identifying unique and accessible ASO targets in bacterial species that post major public health issues.
Topics: Oligonucleotides, Antisense; Anti-Bacterial Agents; Staphylococcus aureus; Oligonucleotides
PubMed: 38025700
DOI: 10.7717/peerj.16343 -
BioRxiv : the Preprint Server For... Aug 2023Daptomycin is a last-resort lipopeptide antibiotic that disrupts cell membrane (CM) and peptidoglycan homeostasis. Enterococcus faecalis has developed a sophisticated...
Daptomycin is a last-resort lipopeptide antibiotic that disrupts cell membrane (CM) and peptidoglycan homeostasis. Enterococcus faecalis has developed a sophisticated mechanism to avoid daptomycin killing by re-distributing CM anionic phospholipids away from the septum. The CM changes are orchestrated by a three-component regulatory system, designated LiaFSR, with a possible contribution of cardiolipin synthase (Cls). However, the mechanism by which LiaFSR controls the CM response and the role of Cls are unknown. Here, we show that cardiolipin synthase activity is essential for anionic phospholipid redistribution and daptomycin resistance since deletion of the two genes ( and ) encoding Cls abolished CM remodeling. We identified LiaY, a transmembrane protein regulated by LiaFSR, as an important mediator of CM remodeling required for re-distribution of anionic phospholipid microdomains via interactions with Cls1. Together, our insights provide a mechanistic framework on the enterococcal response to cell envelope antibiotics that could be exploited therapeutically.
PubMed: 37577577
DOI: 10.1101/2023.08.02.551704 -
Experimental Biology and Medicine... Oct 2023Bacterial cell wall formation is essential for cellular survival and morphogenesis. The peptidoglycan (PG), a heteropolymer that surrounds the bacterial membrane, is a... (Review)
Review
Bacterial cell wall formation is essential for cellular survival and morphogenesis. The peptidoglycan (PG), a heteropolymer that surrounds the bacterial membrane, is a key component of the cell wall, and its multistep biosynthetic process is an attractive antibacterial development target. Penicillin-binding proteins (PBPs) are responsible for cross-linking PG stem peptides, and their central role in bacterial cell wall synthesis has made them the target of successful antibiotics, including β-lactams, that have been used worldwide for decades. Following the discovery of penicillin, several other compounds with antibiotic activity have been discovered and, since then, have saved millions of lives. However, since pathogens inevitably become resistant to antibiotics, the search for new active compounds is continuous. The present review highlights the ongoing development of inhibitors acting mainly in the transpeptidase domain of PBPs with potential therapeutic applications for the development of new antibiotic agents. Both the critical aspects of the strategy, design, and structure-activity relationships (SAR) are discussed, covering the main published articles over the last 10 years. Some of the molecules described display activities against main bacterial pathogens and could open avenues toward the development of new, efficient antibacterial drugs.
Topics: Penicillin-Binding Proteins; Anti-Bacterial Agents; beta-Lactams; Penicillins; Bacteria; Bacterial Proteins
PubMed: 38030964
DOI: 10.1177/15353702231208407 -
BioRxiv : the Preprint Server For... Oct 2023Until recently only 11 distinct Sgls (single gene lysis proteins) have been experimentally identified. Of these, three have been shown to be specific inhibitors of...
Until recently only 11 distinct Sgls (single gene lysis proteins) have been experimentally identified. Of these, three have been shown to be specific inhibitors of different steps in the pathway that supplies Lipid II to the peptidoglycan (PG) biosynthesis machinery: Qβ A inhibits MurA, ϕX174 E inhibits MraY, and Lys from coliphage M inhibits MurJ. These Sgls have been called "protein antibiotics" because the lytic event is a septal catastrophe indistinguishable from that caused by cell wall antibiotics. Here we propose to designate these as members of type I Sgls, to distinguish them from another Sgl, the L protein of the paradigm ssRNA phage MS2. Although none of the other distinct Sgls have significant sequence similarity to L, alignments suggested the presence of four domains distinguished by hydrophobic and polar character. The simplest notion is that these other Sgls have the same autolytic mechanism and, based on this, constitute type II. Although the number of experimentally confirmed Sgls has not changed, recent environmental metagenomes and metatranscriptomes have revealed thousands of new ssRNA phage genomes, each of which presumably has at least one Sgl gene. Here we report on methods to distinguish type I and type II Sgls. Using phase-contrast microscopy, we show that both classes of Sgls cause the formation of blebs prior to lysis, but the location of the blebs differs significantly. In addition, we show that L and other type II Sgls do not inhibit net synthesis of PG, as measured by incorporation of [H]-diaminopimelic acid. Finally, we provide support for the unexpected finding by Adler and colleagues that the Sgl from Pseudomonas phage PP7 is a type I Sgl, as determined by the two methods. This shows that the sharing the putative 4-domain structure suggested for L is not a reliable discriminator for operational characterization of Sgls. Overall, this study establishes new ways to rapidly classify novel Sgls and thus may facilitate the identification of new cell envelope targets that will help generate new antibiotics.
PubMed: 37905155
DOI: 10.1101/2023.10.16.562596 -
Gut May 2024Pancreatic ductal adenocarcinoma (PDAC) has limited therapeutic options, particularly with immune checkpoint inhibitors. Highly chemoresistant 'stem-like' cells, known...
OBJECTIVE
Pancreatic ductal adenocarcinoma (PDAC) has limited therapeutic options, particularly with immune checkpoint inhibitors. Highly chemoresistant 'stem-like' cells, known as cancer stem cells (CSCs), are implicated in PDAC aggressiveness. Thus, comprehending how this subset of cells evades the immune system is crucial for advancing novel therapies.
DESIGN
We used the KPC mouse model () and primary tumour cell lines to investigate putative CSC populations. Transcriptomic analyses were conducted to pinpoint new genes involved in immune evasion. Overexpressing and knockout cell lines were established with lentiviral vectors. Subsequent coculture assays, mouse and zebrafish tumorigenesis studies, and database approaches were performed.
RESULTS
Using the KPC mouse model, we functionally confirmed a population of cells marked by EpCAM, Sca-1 and CD133 as authentic CSCs and investigated their transcriptional profile. Immune evasion signatures/genes, notably the gene peptidoglycan recognition protein 1 (PGLYRP1), were significantly overexpressed in these CSCs. Modulating PGLYRP1 impacted CSC immune evasion, affecting their resistance to macrophage-mediated and T-cell-mediated killing and their tumourigenesis in immunocompetent mice. Mechanistically, tumour necrosis factor alpha (TNFα)-regulated PGLYRP1 expression interferes with the immune tumour microenvironment (TME) landscape, promoting myeloid cell-derived immunosuppression and activated T-cell death. Importantly, these findings were not only replicated in human models, but clinically, secreted PGLYRP1 levels were significantly elevated in patients with PDAC.
CONCLUSIONS
This study establishes PGLYRP1 as a novel CSC-associated marker crucial for immune evasion, particularly against macrophage phagocytosis and T-cell killing, presenting it as a promising target for PDAC immunotherapy.
PubMed: 38754953
DOI: 10.1136/gutjnl-2023-330995 -
Frontiers in Immunology 2023Nucleotide-binding oligomerization domain-containing proteins, NOD1 and NOD2, are cytosolic receptors that recognize dipeptides and tripeptides derived from the... (Review)
Review
Nucleotide-binding oligomerization domain-containing proteins, NOD1 and NOD2, are cytosolic receptors that recognize dipeptides and tripeptides derived from the bacterial cell wall component peptidoglycan (PGN). During the past two decades, studies have revealed several roles for NODs beyond detecting PGN fragments, including activation of an innate immune anti-viral response, NOD-mediated autophagy, and ER stress induced inflammation. Recent studies have also clarified the dynamic regulation of NODs at cellular membranes to generate specific and balanced immune responses. This review will describe how NOD1 and NOD2 detect microbes and cellular stress and detail the molecular mechanisms that regulate activation and signaling while highlighting new evidence and the impact on inflammatory disease pathogenesis.
Topics: Humans; Nod Signaling Adaptor Proteins; Nod1 Signaling Adaptor Protein; Nod2 Signaling Adaptor Protein; Inflammation; Nucleotides
PubMed: 37869013
DOI: 10.3389/fimmu.2023.1242659