scholarly journals Evolutionary Remodeling of the Cell Envelope in Bacteria of the Planctomycetes Phylum

2020 ◽  
Vol 12 (9) ◽  
pp. 1528-1548
Author(s):  
Mayank Mahajan ◽  
Christian Seeger ◽  
Benjamin Yee ◽  
Siv G E Andersson
Keyword(s):  
Cell Wall ◽  
Electron Tomography ◽  
Cell Envelope ◽  
Outer Membrane Proteins ◽  
Cell Wall Proteins ◽  
Secretion Systems ◽  
Peptidoglycan Biosynthesis ◽  
Multiple Copies ◽  
Cellular Phenotypes ◽  
Cell Envelopes

Abstract Bacteria of the Planctomycetes phylum have many unique cellular features, such as extensive membrane invaginations and the ability to import macromolecules. These features raise intriguing questions about the composition of their cell envelopes. In this study, we have used microscopy, phylogenomics, and proteomics to examine the composition and evolution of cell envelope proteins in Tuwongella immobilis and other members of the Planctomycetes. Cryo-electron tomography data indicated a distance of 45 nm between the inner and outer membranes in T. immobilis. Consistent with the wide periplasmic space, our bioinformatics studies showed that the periplasmic segments of outer-membrane proteins in type II secretion systems are extended in bacteria of the order Planctomycetales. Homologs of two highly abundant cysteine-rich cell wall proteins in T. immobilis were identified in all members of the Planctomycetales, whereas genes for peptidoglycan biosynthesis and cell elongation have been lost in many members of this bacterial group. The cell wall proteins contain multiple copies of the YTV motif, which is the only domain that is conserved and unique to the Planctomycetales. Earlier diverging taxa in the Planctomycetes phylum contain genes for peptidoglycan biosynthesis but no homologs to the YTV cell wall proteins. The major remodeling of the cell envelope in the ancestor of the Planctomycetales coincided with the emergence of budding and other unique cellular phenotypes. The results have implications for hypotheses about the process whereby complex cellular features evolve in bacteria.

scholarly journals Cryo-Electron Tomography Reveals the Comparative Three-Dimensional Architecture of Prochlorococcus, a Globally Important Marine Cyanobacterium

2007 ◽  
Vol 189 (12) ◽  
pp. 4485-4493 ◽  
Author(s):  
Claire S. Ting ◽  
Chyongere Hsieh ◽  
Sesh Sundararaman ◽  
Carmen Mannella ◽  
Michael Marko
Keyword(s):  
Cell Wall ◽  
Electron Tomography ◽  
Three Dimensional ◽  
Membrane System ◽  
Niche Differentiation ◽  
Dimensional Structure ◽  
Comparative Genomic ◽  
Photosynthetic Membrane ◽  
Peptidoglycan Biosynthesis ◽  
Cryo Electron Tomography

ABSTRACT In an age of comparative microbial genomics, knowledge of the near-native architecture of microorganisms is essential for achieving an integrative understanding of physiology and function. We characterized and compared the three-dimensional architecture of the ecologically important cyanobacterium Prochlorococcus in a near-native state using cryo-electron tomography and found that closely related strains have diverged substantially in cellular organization and structure. By visualizing native, hydrated structures within cells, we discovered that the MED4 strain, which possesses one of the smallest genomes (1.66 Mbp) of any known photosynthetic organism, has evolved a comparatively streamlined cellular architecture. This strain possesses a smaller cell volume, an attenuated cell wall, and less extensive intracytoplasmic (photosynthetic) membrane system compared to the more deeply branched MIT9313 strain. Comparative genomic analyses indicate that differences have evolved in key structural genes, including those encoding enzymes involved in cell wall peptidoglycan biosynthesis. Although both strains possess carboxysomes that are polygonal and cluster in the central cytoplasm, the carboxysomes of MED4 are smaller. A streamlined cellular structure could be advantageous to microorganisms thriving in the low-nutrient conditions characteristic of large regions of the open ocean and thus have consequences for ecological niche differentiation. Through cryo-electron tomography we visualized, for the first time, the three-dimensional structure of the extensive network of photosynthetic lamellae within Prochlorococcus and the potential pathways for intracellular and intermembrane movement of molecules. Comparative information on the near-native structure of microorganisms is an important and necessary component of exploring microbial diversity and understanding its consequences for function and ecology.

scholarly journals Structure of Vibrio phage XM1, a simple contractile DNA injection machine

2021 ◽  
Author(s):  
Zhiqing Wang ◽  
Andrei Fokine ◽  
Xinwu Guo ◽  
Wen Jiang ◽  
Michael G Rossmann ◽  
...  
Keyword(s):  
Conformational Changes ◽  
Pathogenic Bacteria ◽  
Virus Assembly ◽  
Cell Envelope ◽  
Neck Region ◽  
Secretion Systems ◽  
Atomic Structures ◽  
Dna Injection ◽  
Minimum Requirements ◽  
Cell Envelopes

Antibiotic resistance poses a growing risk to public health requiring new tools to combat pathogenic bacteria. Contractile injection systems, including bacteriophage tails, pyocins, and bacterial type VI secretion systems, can efficiently penetrate cell envelopes and become potential antibacterial agents. Bacteriophage XM1 is a dsDNA virus belonging to the Myoviridae family and infecting Vibrio bacteria. The XM1 virion, made of 18 different proteins, consists of an icosahedral head and a contractile tail, terminated with a baseplate. Here we report cryo-EM reconstructions of all components of the XM1 virion and describe atomic structures of 14 XM1 proteins. The XM1 baseplate is composed of a central hub surrounded by six wedge modules to which twelve spikes are attached. The XM1 tail contains a fewer number of smaller proteins compared with other reported phage baseplates, depicting the minimum requirements for building an effective cell-envelope-penetrating machine. We describe the tail sheath structure in the pre-infection post-infection states and its conformational changes during infection. In addition, we report, for the first time, the in situ structure of the phage neck region to near-atomic resolution. Based on these structures, we propose mechanisms of virus assembly and infection.

Ultrastructure of organohalide-respiring Dehalococcoidia revealed by cryo-electron tomography

2021 ◽  
Author(s):  
Danielle L. Sexton ◽  
Gao Chen ◽  
Fadime Kara Murdoch ◽  
Ameena Hashimi ◽  
Frank E. Löffler ◽  
...  
Keyword(s):  
Electron Tomography ◽  
Cell Envelope ◽  
Cell Structure ◽  
Lipid Vesicles ◽  
Cell Types ◽  
Reductive Dehalogenation ◽  
Outer Leaflet ◽  
New Findings ◽  
Cryo Electron Tomography ◽  
Cell Envelopes

Dehalococcoides mccartyi ( Dhc ) and Dehalogenimonas spp. ( Dhgm ) are members of the class Dehalococcoidia , phylum Chloroflexi, characterized by streamlined genomes and a strict requirement for organohalogens as electron acceptors. Here, we used cryo-electron tomography to reveal morphological and ultrastructural features of Dhc strain BAV1 and ‘ Candidatus Dehalogenimonas etheniformans’ strain GP cells at unprecedented resolution. Dhc cells were irregularly shaped discs (890 ± 110 nm long, 630 ± 110 nm wide and 130 ± 15 nm thick) with curved and straight sides that intersected at acute angles, whereas Dhgm cells appeared as slightly flattened cocci (760 ± 85 nm). The cell envelopes were composed of a cytoplasmic membrane (CM), a paracrystalline surface layer (S-layer) with hexagonal symmetry and ∼22 nm spacing between repeating units, and a layer of unknown composition separating the CM and the S-layer. Cell surface appendages were only detected in Dhc cells, whereas both cell types had bundled cytoskeletal filaments. Repetitive globular structures, ∼5 nm in diameter and ∼9 nm apart, were observed associated with the outer leaflet of the CM. We hypothesized that those represent organohalide respiration (OHR) complexes and estimated ∼30,000 copies per cell. In Dhgm cultures, extracellular lipid vesicles (20 - 110 nm in diameter) decorated with putative OHR complexes but lacking an S-layer were observed. The new findings expand our understanding of the unique cellular ultrastructure and biology of organohalide-respiring Dehalococcoidia . Importance: Dehalococcoidia respire organohalogen compounds and play relevant roles in bioremediation of groundwater, sediments and soils impacted with toxic chlorinated pollutants. Using advanced imaging tools, we have obtained 3-dimensional images at macromolecular resolution of whole Dehalococcoidia cells revealing their unique structural components. Our data detail the overall cellular shape, cell envelope architecture, cytoskeletal filaments, the likely localization of enzymatic complexes involved in reductive dehalogenation, and the structure of extracellular vesicles. The new findings expand our understanding of the cell structure-function relationship in Dehalococcoidia with implications for Dehalococcoidia biology and bioremediation.

scholarly journals Streptococcus thermophilus Cell Wall-Anchored Proteinase: Release, Purification, and Biochemical and Genetic Characterization

2000 ◽  
Vol 66 (11) ◽  
pp. 4772-4778 ◽  
Author(s):  
María Dolores Fernandez-Espla ◽  
Peggy Garault ◽  
Véronique Monnet ◽  
Françoise Rul
Keyword(s):  
Cell Wall ◽  
Lactic Acid ◽  
Lactic Acid Bacteria ◽  
Cell Envelope ◽  
Serine Proteinase ◽  
Streptococcus Thermophilus ◽  
Alumina Powder ◽  
A Cell ◽  
Cell Envelopes

ABSTRACT Streptococcus thermophilus CNRZ 385 expresses a cell envelope proteinase (PrtS), which is characterized in the present work, both at the biochemical and genetic levels. Since PrtS is resistant to most classical methods of extraction from the cell envelopes, we developed a three-step process based on loosening of the cell wall by cultivation of the cells in the presence of glycine (20 mM), mechanical disruption (with alumina powder), and enzymatic treatment (lysozyme). The pure enzyme is a serine proteinase highly activated by Ca2+ ions. Its activity was optimal at 37°C and pH 7.5 with acetyl-Ala-Ala-Pro-Phe-paranitroanilide as substrate. The study of the hydrolysis of the chromogenic and casein substrates indicated that PrtS presented an intermediate specificity between the most divergent types of cell envelope proteinases from lactococci, known as the PI and PIII types. This result was confirmed by the sequence determination of the regions involved in substrate specificity, which were a mix between those of PI and PIII types, and also had unique residues. Sequence analysis of the PrtS encoding gene revealed that PrtS is a member of the subtilase family. It is a multidomain protein which is maturated and tightly anchored to the cell wall via a mechanism involving an LPXTG motif. PrtS bears similarities to cell envelope proteinases from pyogenic streptococci (C5a peptidase and cell surface proteinase) and lactic acid bacteria (PrtP, PrtH, and PrtB). The highest homologies were found with streptococcal proteinases which lack, as PrtS, one domain (the B domain) present in cell envelope proteinases from all other lactic acid bacteria.

scholarly journals Native Cell Wall Organization Shown by Cryo-Electron Microscopy Confirms the Existence of a Periplasmic Space in Staphylococcus aureus

2006 ◽  
Vol 188 (3) ◽  
pp. 1011-1021 ◽  
Author(s):  
Valério R. F. Matias ◽  
Terry J. Beveridge
Keyword(s):  
Electron Microscopy ◽  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Plasma Membrane ◽  
Cell Envelope ◽  
High Density ◽  
Low Density ◽  
Periplasmic Space ◽  
Wall Zone ◽  
Cell Envelopes

ABSTRACT The current perception of the ultrastructure of gram-positive cell envelopes relies mainly on electron microscopy of thin sections and on sample preparation. Freezing of cells into a matrix of amorphous ice (i.e., vitrification) results in optimal specimen preservation and allows the observation of cell envelope boundary layers in their (frozen) hydrated state. In this report, cryo-transmission electron microscopy of frozen-hydrated sections of Staphylococcus aureus D2C was used to examine cell envelope organization. A bipartite wall was positioned above the plasma membrane and consisted of a 16-nm low-density inner wall zone (IWZ), followed by a 19-nm high-density outer wall zone (OWZ). Observation of plasmolyzed cells, which were used to artificially separate the membrane from the wall, showed membrane vesicles within the space associated with the IWZ in native cells and a large gap between the membrane and OWZ, suggesting that the IWZ was devoid of a cross-linked polymeric cell wall network. Isolated wall fragments possessed only one zone of high density, with a constant level of density throughout their thickness, as was previously seen with the OWZs of intact cells. These results strongly indicate that the IWZ represents a periplasmic space, composed mostly of soluble low-density constituents confined between the plasma membrane and OWZ, and that the OWZ represents the peptidoglycan-teichoic acid cell wall network with its associated proteins. Cell wall differentiation was also seen at the septum of dividing cells. Here, two high-density zones were sandwiched between three low-density zones. It appeared that the septum consisted of an extension of the IWZ and OWZ from the outside peripheral wall, plus a low-density middle zone that separated adjacent septal cross walls, which could contribute to cell separation during division.

scholarly journals In Situ Visualization of the pKM101-Encoded Type IV Secretion System Reveals a Highly Symmetric ATPase Energy Center

mBio ◽  
2021 ◽  
Author(s):  
Pratick Khara ◽  
Liqiang Song ◽  
Peter J. Christie ◽  
Bo Hu
Keyword(s):  
Electron Tomography ◽  
Cell Envelope ◽  
Type Iv Secretion ◽  
Secretion Systems ◽  
Type Iv ◽  
Cryo Electron Tomography ◽  
Plasmid Pkm101 ◽  
Energy Center ◽  
Bacterial Type

Bacterial type IV secretion systems (T4SSs) play central roles in antibiotic resistance spread and virulence. By cryo-electron tomography (CryoET), we solved the structure of the plasmid pKM101-encoded T4SS in the native context of the bacterial cell envelope.

scholarly journals Mutations in Sugar-Nucleotide Synthesis Genes Restore Holdfast Polysaccharide Anchoring toCaulobacter crescentusHoldfast Anchor Mutants

2017 ◽  
Vol 200 (3) ◽  
Author(s):  
Gail G. Hardy ◽  
Evelyn Toh ◽  
Cécile Berne ◽  
Yves V. Brun
Keyword(s):  
Biofilm Formation ◽  
Caulobacter Crescentus ◽  
Cell Envelope ◽  
Outer Membrane Proteins ◽  
Gene Mutations ◽  
Secretion Systems ◽  
Nucleotide Synthesis ◽  
Content Type ◽  
Polysaccharide Biosynthesis ◽  
Abiotic Surfaces

ABSTRACTAttachment is essential for microorganisms to establish interactions with both biotic and abiotic surfaces. Stable attachment ofCaulobacter crescentusto surfaces requires an adhesive polysaccharide holdfast, but the exact composition of the holdfast is unknown. The holdfast is anchored to the cell envelope by outer membrane proteins HfaA, HfaB, and HfaD.Holdfastanchor gene mutations result in holdfast shedding and reduced cell adherence. Translocation of HfaA and HfaD to the cell surface requires HfaB. The Wzx homolog HfsF is predicted to be a bacterial polysaccharide flippase. AnhfsFdeletion significantly reduced the amount of holdfast produced per cell and slightly reduced adherence. A ΔhfsFΔhfaDdouble mutant was completely deficient in adherence. A suppressor screen that restored adhesion in the ΔhfsFΔhfaDmutant identified mutations in three genes:wbqV,rfbB, andrmlA. Both WbqV and RfbB belong to a family of nucleoside-diphosphate epimerases, and RmlA has similarity to nucleotidyltransferases. The loss ofwbqVorrfbBin the ΔhfsFΔhfaDmutant reduced holdfast shedding but did not restore holdfast synthesis to parental levels. Loss ofwbqVorrfbBdid not restore adherence to a ΔhfsFmutant but did restore adherence and holdfast anchoring to a ΔhfaDmutant, confirming that suppression occurs through restoration of holdfast anchoring. The adherence and holdfast anchoring of a ΔhfaA ΔhfaDmutant could be restored bywbqVorrfbBmutation, but such mutations could not suppress these phenotypes in the ΔhfaBmutant. We hypothesize that HfaB plays an additional role in holdfast anchoring or helps to translocate an unknown factor that is important for holdfast anchoring.IMPORTANCEBiofilm formation results in increased resistance to both environmental stresses and antibiotics.Caulobacter crescentusrequires an adhesive holdfast for permanent attachment and biofilm formation, but the exact mechanism of polysaccharide anchoring to the cell and the holdfast composition are unknown. Here we identify novel polysaccharide genes that affect holdfast anchoring to the cell. We identify a new role for the holdfast anchor protein HfaB. This work increases our specific knowledge of the polysaccharide adhesin involved inCaulobacterattachment and the general knowledge regarding production and anchoring of polysaccharide adhesins by bacteria. This work also explores the interactions between different polysaccharide biosynthesis and secretion systems in bacteria.

scholarly journals The Integrity of the Cell Wall and Its Remodeling during Heterocyst Differentiation Are Regulated by Phylogenetically Conserved Small RNA Yfr1 in Nostoc sp. Strain PCC 7120

mBio ◽  
2020 ◽  
Vol 11 (1) ◽  
Author(s):  
Manuel Brenes-Álvarez ◽  
Agustín Vioque ◽  
Alicia M. Muro-Pastor
Keyword(s):  
Cell Wall ◽  
Outer Membrane ◽  
Small Rna ◽  
Cell Envelope ◽  
Outer Membrane Proteins ◽  
Site Directed Mutagenesis ◽  
Heterocyst Differentiation ◽  
Content Type ◽  
Bacterial Physiology ◽  
Pcc 7120

ABSTRACT Yfr1 is a strictly conserved small RNA in cyanobacteria. A bioinformatic prediction to identify possible interactions of Yfr1 with mRNAs was carried out by using the sequences of Yfr1 from several heterocyst-forming strains, including Nostoc sp. strain PCC 7120. The results of the prediction were enriched in genes encoding outer membrane proteins and enzymes related to peptidoglycan biosynthesis and turnover. Heterologous expression assays with Escherichia coli demonstrated direct interactions of Yfr1 with mRNAs of 11 of the candidate genes. The expression of 10 of them (alr2458, alr4550, murC, all4829, all2158, mraY, alr2269, alr0834, conR, patN) was repressed by interaction with Yfr1, whereas the expression of amiC2, encoding an amidase, was increased. The interactions between Yfr1 and the 11 mRNAs were confirmed by site-directed mutagenesis of Yfr1. Furthermore, a Nostoc strain with reduced levels of Yfr1 had larger amounts of mraY and murC mRNAs, supporting a role for Yfr1 in the regulation of those genes. Nostoc strains with either reduced or increased expression of Yfr1 showed anomalies in cell wall completion and were more sensitive to vancomycin than the wild-type strain. Furthermore, growth in the absence of combined nitrogen, which involves the differentiation of heterocysts, was compromised in the strain overexpressing Yfr1, and filaments were broken at the connections between vegetative cells and heterocysts. These results indicate that Yfr1 is an important regulator of cell wall homeostasis and correct cell wall remodeling during heterocyst differentiation. IMPORTANCE Bacterial small RNAs (sRNAs) are important players affecting the regulation of essentially every aspect of bacterial physiology. The cell wall is a highly dynamic structure that protects bacteria from their fluctuating environment. Cell envelope remodeling is particularly critical for bacteria that undergo differentiation processes, such as spore formation or differentiation of heterocysts. Heterocyst development involves the deposition of additional layers of glycolipids and polysaccharides outside the outer membrane. Here, we show that a cyanobacterial phylogenetically conserved small regulatory RNA, Yfr1, coordinates the expression of proteins involved in cell wall-related processes, including peptidoglycan metabolism and transport of different molecules, as well as expression of several proteins involved in heterocyst differentiation.

scholarly journals Association of a Myosin Immunoanalogue with Cell Envelopes ofAspergillus fumigatus Conidia and Its Participation in Swelling and Germination

1999 ◽  
Vol 67 (3) ◽  
pp. 1238-1244 ◽  
Author(s):  
Karine Esnault ◽  
Brahim el Moudni ◽  
Jean-Philippe Bouchara ◽  
Dominique Chabasse ◽  
Guy Tronchin
Keyword(s):  
Cell Wall ◽  
Immunoelectron Microscopy ◽  
Indirect Immunofluorescence ◽  
Actin Polymerization ◽  
Polyclonal Antibodies ◽  
Cell Envelope ◽  
Immunofluorescence Assay ◽  
Atpase Inhibitor ◽  
Butanedione Monoxime ◽  
Cell Envelopes

ABSTRACT A myosin immunoanalogue was identified in conidia ofAspergillus fumigatus by Western blotting, indirect immunofluorescence assay, and gold immunoelectron microscopy with two different antimyosin antibodies. The distribution pattern of this protein was followed during the early stages of germination. A single 180-kDa polypeptide, detected predominantly in a cell envelope extract, was found to cross-react with monoclonal and polyclonal antibodies raised against vertebrate muscle myosin. Immunoelectron microscopy permitted precise localization of this polypeptide, indicating that myosin analogue was mainly distributed along the plasma membrane of resting and swollen conidia. In germinating conidia, indirect immunofluorescence microscopy revealed myosin analogue at the periphery of germ tubes, whereas actin appeared as dispersed punctate structures in the cytoplasm that were more concentrated at the site of germ tube emergence. A myosin ATPase inhibitor, butanedione monoxime, greatly reduced swelling and blocked germination. In contrast, when conidia were treated with cytochalasin B, an inhibitor of actin polymerization, swelling was not affected and germination was only partially reduced. Butanedione monoxime-treated conidia showed accumulation of cytoplasmic vesicles and did not achieve cell wall reorganization, unlike swollen conidia. Collectively, these results suggest an essential role for this myosin analogue in the deposition of cell wall components during germination of A. fumigatus conidia and therefore in host tissue colonization.

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