Cell Wall Protein Composition Research Articles

Changes in the Cell Wall Proteome of Leaves in Response to High Temperature Stress in Brachypodium distachyon.

High temperature stress leads to complex changes to plant functionality, which affects, i.a., the cell wall structure and the cell wall protein composition. In this study, the qualitative and quantitative changes in the cell wall proteome of Brachypodium distachyon leaves in response to high (40 °C) temperature stress were characterised. Using a proteomic analysis, 1533 non-redundant proteins were identified from which 338 cell wall proteins were distinguished. At a high temperature, we identified 46 differentially abundant proteins, and of these, 4 were over-accumulated and 42 were under-accumulated. The most significant changes were observed in the proteins acting on the cell wall polysaccharides, specifically, 2 over- and 12 under-accumulated proteins. Based on the qualitative analysis, one cell wall protein was identified that was uniquely present at 40 °C but was absent in the control and 24 proteins that were present in the control but were absent at 40 °C. Overall, the changes in the cell wall proteome at 40 °C suggest a lower protease activity, lignification and an expansion of the cell wall. These results offer a new insight into the changes in the cell wall proteome in response to high temperature.

COI1-dependent jasmonate signalling affects growth, metabolite production and cell wall protein composition in arabidopsis.

Background and AimsCultured cell suspensions have been the preferred model to study the apoplast as well as to monitor metabolic and cell cycle-related changes. Previous work showed that methyl jasmonate (MeJA) inhibits leaf growth in a CORONATINE INSENSITIVE 1 (COI1)-dependent manner, with COI1 being the jasmonate (JA) receptor. Here, the effect of COI1 overexpression on the growth of stably transformed arabidopsis cell cultures is described.MethodsTime-course experiments were carried out to analyse gene expression, and protein and metabolite levels.Key ResultsBoth MeJA treatment and the overexpression of COI1 modify growth, by altering cell proliferation and expansion. DNA content as well as transcript patterns of cell cycle and cell wall remodelling markers were altered. COI1 overexpression also increases the protein levels of OLIGOGALACTURONIDE OXIDASE 1, BETA-GLUCOSIDASE/ENDOGLUCANASES and POLYGALACTURONASE INHIBITING PROTEIN2, reinforcing the role of COI1 in mediating defence responses and highlighting a link between cell wall loosening and growth regulation. Moreover, changes in the levels of the primary metabolites alanine, serine and succinic acid of MeJA-treated Arabidopsis cell cultures were observed. In addition, COI1 overexpression positively affects the availability of metabolites such as β-alanine, threonic acid, putrescine, glucose and myo-inositol, thereby providing a connection between JA-inhibited growth and stress responses.ConclusionsThis study contributes to the understanding of the regulation of growth and the production of metabolic resources by JAs and COI1. This will have important implications in dissecting the complex relationships between hormonal and cell wall signalling in plants. The work also provides tools to uncover novel mechanisms co-ordinating cell division and post-mitotic cell expansion in the absence of organ developmental control.

Transcription Factor Efg1 Shows a Haploinsufficiency Phenotype in Modulating the Cell Wall Architecture and Immunogenicity of Candida albicans

The Candida albicans transcription factor Efg1 is known to be involved in many different cellular processes, including morphogenesis, general metabolism, and virulence. Here we show that besides its manifold roles, Efg1 also has a prominent effect on cell wall structure and composition, strongly affecting the structural glucan part. Deletion of only one allele of EFG1 already results in severe phenotypes for cell wall biogenesis, comparable to those with deletion of both alleles, indicative of a severe haploinsufficiency for EFG1. The observed defects in structural setup of the cell wall, together with previously reported alterations in expression of cell surface proteins, result in altered immunogenic properties of strains with compromised Efg1 function. This is shown by interaction studies with macrophages and primary dendritic cells. The structural changes in the cell wall carbohydrate meshwork presented here, together with the manifold changes in cell wall protein composition and metabolism reported in other studies, contribute to the altered immune response mounted by innate immune cells and to the altered virulence phenotypes observed for strains lacking EFG1.

Inactivation of the Ecs ABC Transporter of Staphylococcus aureus Attenuates Virulence by Altering Composition and Function of Bacterial Wall

BackgroundEcs is an ATP-binding cassette (ABC) transporter present in aerobic and facultative anaerobic Gram-positive Firmicutes. Inactivation of Bacillus subtilis Ecs causes pleiotropic changes in the bacterial phenotype including inhibition of intramembrane proteolysis. The molecule(s) transported by Ecs is (are) still unknown.Methodology/Principal FindingsIn this study we mutated the ecsAB operon in two Staphylococcus aureus strains, Newman and LS-1. Phenotypic and functional characterization of these Ecs deficient mutants revealed a defect in growth, increased autolysis and lysostaphin sensitivity, altered composition of cell wall proteins including the precursor form of staphylokinase and an altered bacterial surface texture. DNA microarray analysis indicated that the Ecs deficiency changed expression of the virulence factor regulator protein Rot accompanied by differential expression of membrane transport proteins, particularly ABC transporters and phosphate-specific transport systems, protein A, adhesins and capsular polysaccharide biosynthesis proteins. Virulence of the ecs mutants was studied in a mouse model of hematogenous S. aureus infection. Mice inoculated with the ecs mutant strains developed markedly milder infections than those inoculated with the wild-type strains and had consequently lower mortality, less weight loss, milder arthritis and decreased persistence of staphylococci in the kidneys. The ecs mutants had higher susceptibility to ribosomal antibiotics and plant alkaloids chelerythrine and sanguinarine.Conclusions/SignificanceOur results show that Ecs is essential for staphylococcal virulence and antimicrobial resistance probably since the transport function of Ecs is essential for the normal structure and function of the cell wall. Thus targeting Ecs may be a new approach in combating staphylococcal infection.

Thaumatin-like proteins and their possible role in protection against chilling injury in peach fruit

Peaches are highly perishable; they ripen and deteriorate quickly at ambient temperature, and cold storage is used to slow these processes. The cell wall protein composition of two peach cultivars, and total protein composition were examined at harvest and after cold storage (3 weeks, 5 °C) by two-dimensional polyacrylamide gel electrophoresis. The two peach cultivars used were ‘Oded’, a white-, melting-flesh, cling-stone, early season cultivar resistant to chilling injury, and ‘Hermoza’, a white-, melting-flesh, free-stone, mid-season cultivar susceptible to chilling injury. Following storage, peptides in the cell wall with molecular masses ranging from 18 kDa to 60 kDa were identified by amino acid sequence to be thaumatin-like protein 1 precursor and thaumatin-like protein 2 precursor. qRT-PCR analysis revealed that the thaumatin-like protein 1 precursor transcript accumulated significantly in both cultivars during storage. However, after 1 and 2 weeks of cold storage at 5 °C the thaumatin-like protein 1 precursor transcript levels were significantly higher in the chilling injury-resistant peach ‘Oded’ than the susceptible peach ‘Hermoza’. This early accumulation of the thaumatin-like protein 1 precursor transcript in the resistant peach suggests that thaumatin-like protein 1 precursor (and perhaps thaumatin-like protein 2 precursor) might be involved in protecting against chilling injury. Although thaumatin-like proteins accumulated to high levels in cell walls of chilling injury-sensitive ‘Hermoza’, the kinetics of transcript accumulation suggest that the early appearance of the transcript for this protein family might be involved in shielding the fruit from the dramatic cell wall-structure changes that accompany the onset of chilling injury in stone fruit, and that result in woolliness development.

Influence of carbohydrates on cell properties of Lactobacillus rhamnosus

AbstractLactobacilli represent normal commensals of the human body, particularly in the gut and vagina where they protect these environments from incoming pathogens via a variety of mechanisms. The influence of the carbohydrate source present in reconstituted MRS growth medium on the different cell properties of two Lactobacillus rhamnosus strains were examined. Two human vaginal isolates, BGHV719 and exopolysaccharide producer strain BGHV954 were analyzed. The results demonstrated that unlike in reconstituted MRS with glucose as a carbon source, the presence of fructose, mannose, or rhamnose, significantly reduced cell surface hydrophobicity of both strains. In addition, differences in cell wall protein composition of L. rhamnosus BGHV719 and alterations in colony mucoidity of L. rhamnosus BGHV954 were also demonstrated. Light and SEM microscopy revealed differences on the cellular level when BGHV719 was cultivated in the presence of different sugars. The results of this study point out the importance of complex relationships between growth medium composition and the different aspects of bacterial behavior, and call for more detailed analyses of versatile bacterial responses to the changes in the environment, including vaginal ecosystem. This is especially important since lactobacilli are amongst the most widely used of probiotics.

Cell wall proteome in the maize primary root elongation zone. II. Region-specific changes in water soluble and lightly ionically bound proteins under water deficit.

Previous work on the adaptation of maize (Zea mays) primary roots to water deficit showed that cell elongation is maintained preferentially toward the apex, and that this response involves modification of cell wall extension properties. To gain a comprehensive understanding of how cell wall protein (CWP) composition changes in association with the differential growth responses to water deficit in different regions of the elongation zone, a proteomics approach was used to examine water soluble and loosely ionically bound CWPs. The results revealed major and predominantly region-specific changes in protein profiles between well-watered and water-stressed roots. In total, 152 water deficit-responsive proteins were identified and categorized into five groups based on their potential function in the cell wall: reactive oxygen species (ROS) metabolism, defense and detoxification, hydrolases, carbohydrate metabolism, and other/unknown. The results indicate that stress-induced changes in CWPs involve multiple processes that are likely to regulate the response of cell elongation. In particular, the changes in protein abundance related to ROS metabolism predicted an increase in apoplastic ROS production in the apical region of the elongation zone of water-stressed roots. This was verified by quantification of hydrogen peroxide content in extracted apoplastic fluid and by in situ imaging of apoplastic ROS levels. This response could contribute directly to the enhancement of wall loosening in this region. This large-scale proteomic analysis provides novel insights into the complexity of mechanisms that regulate root growth under water deficit conditions and highlights the spatial differences in CWP composition in the root elongation zone.

Role of Protein O -Mannosyltransferase Pmt4 in the Morphogenesis and Virulence of Cryptococcus neoformans

Protein O mannosylation is initiated in the endoplasmic reticulum by protein O-mannosyltransferases (Pmt proteins) and plays an important role in the secretion, localization, and function of many proteins, as well as in cell wall integrity and morphogenesis in fungi. Three Pmt proteins, each belonging to one of the three respective Pmt subfamilies, are encoded in the genome of the human fungal pathogen Cryptococcus neoformans. Disruption of the C. neoformans PMT4 gene resulted in abnormal growth morphology and defective cell separation. Transmission electron microscopy revealed defective cell wall septum degradation during mother-daughter cell separation in the pmt4 mutant compared to wild-type cells. The pmt4 mutant also demonstrated sensitivity to elevated temperature, sodium dodecyl sulfate, and amphotericin B, suggesting cell wall defects. Further analysis of cell wall protein composition revealed a cell wall proteome defect in the pmt4 mutant, as well as a global decrease in protein mannosylation. Heterologous expression of C. neoformans PMT4 in a Saccharomyces cerevisiae pmt1pmt4 mutant strain functionally complemented the deficient Pmt activity. Furthermore, Pmt4 activity in C. neoformans was required for full virulence in two murine models of disseminated cryptococcal infection. Taken together, these results indicate a central role for Pmt4-mediated protein O mannosylation in growth, cell wall integrity, and virulence of C. neoformans.

Changes in the composition of cell wall proteins in barley roots during germination and growth in aluminium presence

Root growth inhibition and loss of cell viability in barley root cells were induced by Al during germination of barley caryopses on filter paper moistened with 2mM AlCl3. The inhibition of root growth as well as loss of cell viability started on the third day of germination. This time was also needed for induction of Al-induced changes in the composition of cell wall proteins. The accumulation of three salt-extractable polypeptides with relative molecular mass 14, 27, and 29 kDa started 72 h after the beginning of germination of barley caryopses on Al containing filter paper. However, the inhibition of developmentally regulated deposition of three (18, 23 and 28 kDa) salt-extractable CW polypeptides was observed at the same time. The pattern of detergent-extractable CW proteins was not influenced by Al regardless of the duration of Al treatment.

Sequential fractionation and two-dimensional gel analysis unravels the complexity of the dimorphic fungus Candida albicans cell wall proteome.

The cell wall proteins of Candida albicans play a key role in morphogenesis and pathogenesis and might be potential target sites for new specific antifungal drugs. However, these proteins are difficult to analyze because of their high heterogeneity, interconnections with wall polysaccharides (mannan, glucan, and chitin), low abundance, low solubility, and hydrophobic nature. Here we report a subproteomic approach for the study of the cell wall proteins (CWPs) from C. albicans yeast and hyphal forms. Most of the mannoproteins present in this compartment were extracted by cell wall fractionation according to the type of interactions that they establish with other structural components. CWPs were solubilized from isolated cell walls by hot SDS and dithiothreitol treatment followed by extraction either by mild alkali conditions or by enzymatic treatment with glucanases and chitinases. These highly enriched cell wall fractions were analyzed by two-dimensional PAGE, showing that a large number of proteins are involved in cell wall construction and that the wall remodeling that occurs during germ tube formation is related to changes in the composition of CWPs. We suggest that the CWP-chitin linkage is an important retention mechanism of CWPs in C. albicans mycelial forms. This article also highlights the usefulness of the combination of sequential fractionation and two-dimensional PAGE followed by Western blotting using specific antibodies against known CWPs in the characterization of incorporation mechanisms of such CWPs into the cell wall and of their interactions with other wall components. Mass spectrometry analyses have allowed the identification of several cell surface proteins classically associated with both the cell wall and other compartments. The physiological significance of the dual location of these moonlighting proteins is also discussed. This approach is therefore a powerful tool for obtaining a comprehensive and integrated view of the cell wall proteome.

A Comparative Study on the Porins of Rhodobacter capsulatus Strains 37b4 and B10 Grown Under Different Culture Conditions

Summary The cell wall protein composition and the isolated porins of Rhodobacter capsulatus strains 37b4 and B10 were comparatively studied. The porins from the two strains migrated differently on SDS-PAGE in the absence (oligomers; Mr 92 000 and 89 000, monomers; Mr 44 000 and 32 000, for strains 37b4 and B10, respectively) and in the presence of 8 M urea. N-terminal sequencing of the two purified porins demonstrated that the sequences were different in five positions out of 21 positions determined. With both strains, the cell walls showed no growth condition dependent differences in their protein patterns. In both cases, only the porin given above was expressed in photoorganoheterotrophically (organotrophic) and photolithoautotrophically (lithotrophic) grown cells. There was also no difference between cells grown with glucose instead of malic acid or even using cells grown under phosphate starvation. In lipid bilayer experiments the isolated porins from Rhodobacter capsulatus 37b4 and B10 had virtually the same single-channel properties. Furthermore, the porins obtained from cells grown under the given various different culture conditions revealed the same single-channel conductance. In all cases a slight cation-selectivity was observed.