Cell Surface最新文献

Cellulose microfibril patterning influences many of the mechanical attributes of plant cell walls. We developed a simple, fluorescence microscopy-based method to detect the orientation of newly-synthesized cellulose microfibrils in epidermal peels of onion and Arabidopsis. It is based on Alexa Fluor 488-tagged carbohydrate binding module 3a (CBM3a) from Clostridium thermocellum which displayed a nearly 4-fold greater binding to cell walls at pH 5.5 compared with pH 8. Binding to isolated cellulose did not display this pH dependence. At pH 7.5 fibrillar patterns at the surface of the epidermal peels were visible, corresponding to the directionality of surface cellulose microfibrils, as verified by atomic force microscopy. The fibrillar pattern was not visible as the labeling intensity increased at lower pH. The pH of greatest cell wall labeling corresponds to the isoelectric point of CBM3a, suggesting that electrostatic forces limit CBM3a penetration into the wall. Consistent with this, digestion of the wall with pectate lyase to remove homogalacturonan increased labeling intensity. We conclude that electrostatic interactions strongly influence labeling of cell walls with CBM3 and potentially other proteins, holding implications for any work that relies on penetration of protein probes such as CBMs, antibodies, or enzymes into charged polymeric substrates.

In plants, cell adhesion relies on balancing the integrity of the pectin-rich middle lamella with wall loosening during tissue expansion. Mutation of QUASIMODO2 (QUA2), a pectin methyltransferase, causes defective hypocotyl elongation and cell adhesion in Arabidopsis thaliana hypocotyls. However, the molecular function of QUA2 in cell adhesion is obscured by complex genetic and environmental interactions. To dissect the role of QUA2 in cell adhesion, we investigated a qua2 loss-of-function mutant and a suppressor mutant with restored cell adhesion, qua2 esmeralda1, using a combination of imaging and biochemical techniques. We found that qua2 hypocotyls have reductions in middle lamellae integrity, pectin methyl-esterase (PME) activity, pectin content and molecular mass, and immunodetected Ca²⁺-crosslinking at cell corners, but increased methyl-esterification and polygalacturonase (PG) activity, with qua2 esmd1 having wild type-like or intermediate phenotypes. Our findings suggest that excessive pectin degradation prevents pectin accumulation and the formation of a sufficiently Ca²⁺-crosslinked network to maintain cell adhesion in qua2 mutants. We propose that PME and PG activities balance tissue-level expansion and cell separation. Together, these data provide insight into the cause of cell adhesion defects in qua2 mutants and highlight the importance of harmonizing pectin modification and degradation during plant growth and development.

The immunogenicity of Candida albicans cells is influenced by changes in the exposure of microbe-associated molecular patterns (MAMPs) on the fungal cell surface. Previously, the degree of exposure on the C. albicans cell surface of the immunoinflammatory MAMP β-(1,3)-glucan was shown to correlate inversely with colonisation levels in the gastrointestinal (GI) tract. This is important because life-threatening systemic candidiasis in critically ill patients often arises from translocation of C. albicans strains present in the patient’s GI tract. Therefore, using a murine model, we have examined the impact of gut-related factors upon β-glucan exposure and colonisation levels in the GI tract.

The degree of β-glucan exposure was examined by imaging flow cytometry of C. albicans cells taken directly from GI compartments, and compared with colonisation levels. Fungal β-glucan exposure was lower in the cecum than the small intestine, and fungal burdens were correspondingly higher in the cecum. This inverse correlation did not hold for the large intestine.

The gut fermentation acid, lactate, triggers β-glucan masking in vitro, leading to attenuated anti-Candida immune responses. Additional fermentation acids are present in the GI tract, including acetate, propionate, and butyrate. We show that these acids also influence β-glucan exposure on C. albicans cells in vitro and, like lactate, they influence β-glucan exposure via Gpr1/Gpa2-mediated signalling. Significantly, C. albicans gpr1Δ gpa2Δ cells displayed elevated β-glucan exposure in the large intestine and a corresponding decrease in fungal burden, consistent with the idea that Gpr1/Gpa2-mediated β-glucan masking influences colonisation of this GI compartment. Finally, extracts from the murine gut and culture supernatants from the mannan grazing gut anaerobe Bacteroides thetaiotaomicron promote β-glucan exposure at the C. albicans cell surface. Therefore, the local microbiota influences β-glucan exposure levels directly (via mannan grazing) and indirectly (via fermentation acids), whilst β-glucan masking appears to promote C. albicans colonisation of the murine large intestine.

Gram-positive and Gram-negative bacterial pathogens are commonly found in Urinary Tract Infection (UTI), particularly infected in females like pregnant women, elder people, sexually active, or individuals prone to other risk factors for UTI. In this article, we review the expression of virulence surface proteins and their interaction with host cells for the most frequently isolated uropathogens: Escherichia coli, Enterococcus faecalis, Proteus mirabilis, Klebsiella pneumoniae, and Staphylococcus saprophyticus. In addition to the host cell interaction, surface protein regulation was also discussed in this article. The surface protein regulation serves as a key tool in differentiating the pathogen isotypes. Furthermore, it might provide insights on novel diagnostic methods to detect uropathogen that are otherwise easily overlooked due to limited culture-based assays. In essence, this review shall provide an in-depth understanding on secretion of virulence factors of various uropathogens and their role in host-pathogen interaction, this knowledge might be useful in the development of therapeutics against uropathogens.

Mycobacterium tuberculosis causes the disease tuberculosis and affects a third of the world’s population. The recent COVID-19 pandemic exacerbated the situation with a projected 27% increase in tuberculosis related deaths. M. tuberculosis has an elaborate cell wall consisting of peptidoglycan, arabinogalactan and mycolic acids which shield the bacilli from the toxic bactericidal milieu within phagocytes. Amongst, the numerous glycosyltransferase enzymes involved in mycobacterial cell wall biosynthesis, arabinofuranosyltransferase C (aftC) is responsible for the branching of the arabinan domain in both arabinogalactan and lipoarabinomannan. Using Clustered Regularly Interspaced Short Palindromic Repeats interference (CRISPRi) we have generated aftC knockdowns in Mycobacterium bovis BCG and demonstrated the generation of a truncated, immunogenic lipoarabinomannan within its cell envelope. The aftC depleted BCG mutants were unable to form characteristic mycobacterial pellicular biofilms and elicit a potent immunostimulatory phenotype compared to wild type M. bovis BCG in a THP1 cell line. This study paves the way to further explore novel BCG mutants as promising vaccine boosters in preventing pulmonary tuberculosis.

Proteus mirabilis harbours a variety of O antigens, permitting evasion of the host immune response. LPS decoration with phosphocholine increases cell surface hydrophobicity and decreases electrokinetic potential, which may interfere with antibody interaction and bacterial surface recognition. The decoration does not influence adherence to solid surfaces.

GH16 chitin transferases, GH17 β-1,3-glucan transferases, and GH72 β-1,3-glucan/lichenin transferases are important fungal cell wall crosslinking enzymes. The Neurospora crassa genome encodes three genes from the GH17 gene family and five members in the GH16 subfamily 18 and 19 fungal chitin transferases. We created deletion mutants lacking all three GH17 genes and determined that they had wild type morphology and are more sensitive to cell wall perturbation reagents than the wild type. We also created deletion mutants lacking all five GH16 subfamily 18 and 19 genes and found that they had wild type morphology and are more sensitive to cell wall perturbation reagents than the wild type. We conclude that GH16 and GH17 enzymes play roles in cell wall biogenesis. In N. crassa, GH72 enzymes have been reported to be lichenin transferases, while in other fungi they have been shown to be the β-1,3-glucan transferases. Neurospora triple GH72 deletions give rise to a tight colonial morphology, sensitivity to cell wall perturbation reagents, and release of cell wall proteins into the medium. To ask if GH72 and GH17 enzymes might be redundant in N. crassa, we created sextuple mutants lacking the three GH72 genes and the three GH17 genes and found that they were indistinguishable from the GH72 triple mutant. We also found that a recombinant GH72 enzyme is able to form a lichenin-enzyme intermediate demonstrating that GH72 enzymes are lichenin transferases. The N. crassa GH72 enzymes are lichenin transferases and are not redundant with the GH17 β-1,3-glucan transferases.

Peptidoglycan (PG) is an essential component of the cell envelope in most bacteria, responsible for maintaining the shape of the cell and protecting the cell from environmental stresses. The growth of the PG layer during cell elongation and division is facilitated by the coordinated activities of PG synthases and hydrolases. PG synthases are regulated from inside the cell by components of the elongasome and divisome complexes driven by the cytoskeletal proteins MreB and FtsZ. In Escherichia coli the PG synthases PBP1A and PBP1B require the activation by outer membrane (OM)-anchored lipoproteins LpoA and LpoB, respectively. These have an elongated structure and are capable to span the periplasm to reach their cognate, cytoplasmic membrane (CM)-anchored PG synthase through the PG layer. Presumably, the Lpo proteins activate the PBPs at sites where the PG mesh is stretched or defective, resulting in coupling of PG synthase activation with cell growth or PG repair. Here we investigated the importance of OM-anchoring on the function of Lpo proteins in regulating PG synthesis in response to environmental stresses. We investigated the effects of an artificially CM-tethered LpoB on cell morphology and PG synthesis. Our results indicate that mis-localization of LpoB affects the growth and morphology of cells in high osmolarity growth medium, and PG synthesis rate upon an osmotic upshift.

Cell walls are at the front line of interactions between walled-organisms and their environment. They support cell expansion, ensure cell integrity and, for multicellular organisms such as plants, they provide cell adherence, support cell shape morphogenesis and mediate cell–cell communication. Wall-sensing, detecting perturbations in the wall and signaling the cell to respond accordingly, is crucial for growth and survival. In recent years, plant signaling research has suggested that a large family of receptor-like kinases (RLKs) could function as wall sensors partly because their extracellular domains show homology with malectin, a diglucose binding protein from the endoplasmic reticulum of animal cells. Studies of several malectin/malectin-like (M/ML) domain-containing RLKs (M/MLD-RLKs) from the model plant Arabidopsis thaliana have revealed an impressive array of biological roles, controlling growth, reproduction and stress responses, processes that in various ways rely on or affect the cell wall. Malectin homologous sequences are widespread across biological kingdoms, but plants have uniquely evolved a highly expanded family of proteins with ML domains embedded within various protein contexts. Here, we present an overview on proteins with malectin homologous sequences in different kingdoms, discuss the chromosomal organization of Arabidopsis M/MLD-RLKs and the phylogenetic relationship between these proteins from several model and crop species. We also discuss briefly the molecular networks that enable the diverse biological roles served by M/MLD-RLKs studied thus far.