Maintaining Cell Wall Integrity Research Articles

Transcriptome analysis provides insights into preservation mechanism of postharvest Muscat Hamburg grapes treated with SO2

Sulfur dioxide (SO2) is the most frequently used preservative for table grapes, yet the preservation mechanism remains unclear. To ascertain the specific genes and pathways involved in SO2 preservation, RNA-seq technology was employed to characterize the transcriptome profile of grapes during postharvest storage. A total of 22,288 genes were identified, of which 377 genes were differentially expressed (≥ 2-fold change) between SO2 and control groups, mainly enriched in secondary metabolism, plant-pathogen interactions, plant hormone signaling, etc. Numerous genes encoding pathogenesis-related (PR) proteins exhibited higher expression levels in SO2 group, while the activities of disease-resistant enzymes, such as β-1,3-glucanase (PR2) and chitinase (PR3) significantly increased by 28.9 % and 29.3 % (P < 0.05), indicating SO2-induced plant resistance. Exposure to SO2 also enhanced the expression of genes encoding essential enzymes of secondary metabolism, and significantly increased both the activities of critical enzymes for the biosynthesis of secondary metabolites, such as phenylalanine ammonia-lyase (PAL) and 4-coumarate-CoA ligase (4CL), and the contents of secondary metabolites such as total phenol, flavonoid, anthocyanin, and lignin, demonstrating an enhanced chemical and physical barriers in SO2-fumigated grapes. Meanwhile, the genes associated with ethylene signaling and cell wall degradation were down-regulated by SO2 preservative, and some ethylene-responsive elements were identified in the promoter regions of cell wall hydrolase genes, suggesting that SO2-inhibited ethylene signaling might contribute to maintaining cell wall integrity. Altogether, gene expression patterns and cellular physiological processes were altered in grapes exposed to SO2, which helped maintain fruit quality and prolong postharvest life by regulating the defense responses and fruit ageing.

COMPASS core subunits MpSet1 and MpSwd3 regulate Monascus pigments synthesis in Monascus purpureus.

In eukaryotes, methylation of histone H3 at lysine 4 (H3K4me) catalyzed by the complex of proteins associated with Set1 (COMPASS) is crucial for the transcriptional regulation of genes and the development of organisms. In Monascus, the functions of COMPASS in establishing H3K4me remain unclear. This study first identified the conserved COMPASS core subunits MpSet1 and MpSwd3 in Monascus purpureus and confirmed their roles in establishing H3K4me2/3. Loss of MpSet1 and MpSwd3 resulted in slower growth and development and inhibited the formation of cleistothecia, ascospores, and conidia. The loss of these core subunits also decreased the production of extracellular and intracellular Monascus pigments (MPs) by 94.2%, 93.5%, 82.7%, and 82.5%, respectively. In addition, RNA high-throughput sequencing and quantitative real-time polymerase chain reaction (qRT-PCR) showed that the loss of MpSet1 and MpSwd3 altered the expression of 2646 and 2659 genes, respectively, and repressed the transcription of MPs synthesis-related genes. In addition, the ΔMpset1 and ΔMpswd3 strains demonstrated increased sensitivity to cell wall stress with the downregulation of chitin synthase-coding genes. These results indicated that the COMPASS core subunits MpSet1 and MpSwd3 help establish H3K4me2/3 for growth and development, spore formation, and pigment synthesis in Monascus. These core subunits also assist in maintaining cell wall integrity.

A distinctive family of L,D-transpeptidases catalyzing L-Ala-mDAP crosslinks in Alpha- and Betaproteobacteria

The bacterial cell-wall peptidoglycan is made of glycan strands crosslinked by short peptide stems. Crosslinks are catalyzed by DD-transpeptidases (4,3-crosslinks) and LD-transpeptidases (3,3-crosslinks). However, recent research on non-model species has revealed novel crosslink types, suggesting the existence of uncharacterized enzymes. Here, we identify an LD-transpeptidase, LDTGo, that generates 1,3-crosslinks in the acetic-acid bacterium Gluconobacter oxydans. LDTGo-like proteins are found in Alpha- and Betaproteobacteria lacking LD3,3-transpeptidases. In contrast with the strict specificity of typical LD- and DD-transpeptidases, LDTGo can use non-terminal amino acid moieties for crosslinking. A high-resolution crystal structure of LDTGo reveals unique features when compared to LD3,3-transpeptidases, including a proline-rich region that appears to limit substrate access, and a cavity accommodating both glycan chain and peptide stem from donor muropeptides. Finally, we show that DD-crosslink turnover is involved in supplying the necessary substrate for LD1,3-transpeptidation. This phenomenon underscores the interplay between distinct crosslinking mechanisms in maintaining cell wall integrity in G. oxydans.

The plant cell wall-dynamic, strong, and adaptable-is a natural shapeshifter.

Mythology is replete with good and evil shapeshifters, who, by definition, display great adaptability and assume many different forms-with several even turning themselves into trees. Cell walls certainly fit this definition as they can undergo subtle or dramatic changes in structure, assume many shapes, and perform many functions. In this review, we cover the evolution of knowledge of the structures, biosynthesis, and functions of the 5 major cell wall polymer types that range from deceptively simple to fiendishly complex. Along the way, we recognize some of the colorful historical figures who shaped cell wall research over the past 100 years. The shapeshifter analogy emerges more clearly as we examine the evolving proposals for how cell walls are constructed to allow growth while remaining strong, the complex signaling involved in maintaining cell wall integrity and defense against disease, and the ways cell walls adapt as they progress from birth, through growth to maturation, and in the end, often function long after cell death. We predict the next century of progress will include deciphering cell type-specific wall polymers; regulation at all levels of polymer production, crosslinks, and architecture; and how walls respond to developmental and environmental signals to drive plant success in diverse environments.

WSC1 Regulates the Growth, Development, Patulin Production, and Pathogenicity of Penicillium expansum Infecting Pear Fruits.

In this study, the role of WSC1 in the infection of pear fruit by Penicillium expansum was investigated. The WSC1 gene was knocked out and complemented by Agrobacterium-mediated homologous recombination technology. Then, the changes in growth, development, and pathogenic processes of the knockout mutant and the complement mutant were analyzed. The results indicated that deletion of WSC1 slowed the growth rate, reduced the mycelial and spore yield, and reduced the ability to produce toxins and pathogenicity of P. expansum in pear fruits. At the same time, the deletion of WSC1 reduced the tolerance of P. expansum to cell wall stress factors, enhanced antioxidant capacity, decreased hypertonic sensitivity, decreased salt stress resistance, and was more sensitive to most metal ions. Our results confirmed that WSC1 plays an important role in maintaining cell wall integrity and responding to stress, toxin production, and the pathogenicity of P. expansum.

Proteins of the food-borne pathogen &lt;i&gt;Listeria monocytogenes&lt;/i&gt; strain F2365 relevant to lethal acidic stress and during rapid inactivation

&lt;abstract&gt; &lt;p&gt;&lt;italic&gt;Listeria monocytogenes&lt;/italic&gt;, which causes human listeriosis after consumption of contaminated food, can adapt and survive under a wide range of physiological and chemical stresses. In this study, the overall proteomic response of the &lt;italic&gt;L. monocytogenes&lt;/italic&gt; strain F2365—a strain with mutations limiting its ability to tolerate acidic conditions—to progressive non-thermal acidic inactivation was investigated. The challenge process was investigated in the early stationary growth phase where F2365 cultures were acidified (pH 3.0, HCl) at 5 min, 1 h, and 2 h, generating pH 4.8, pH 4.1, and pH 3.5, respectively, with protein abundance measured using iTRAQ. Approximately 73 proteins increased in abundance and 8 declined when acidic stress became non-growth-permissive (pH &amp;lt; 4.1) and inactivation accelerated to approximately 2 log units/h. The functional categories of responding proteins were broad but the proteins involved were specific in nature and did not include whole pathways. Many responses likely accentuate energy conservation and compensate vital metabolic processes. For example, further repression of FlaA, normally repressed under acidic stress, occurs accompanied by an increase in quinol oxidase subunit QoxA and glycerol kinase GlpK. Proteins maintaining cell wall integrity, such as Iap and CwlO, manifested the overall largest abundance increase trend. Virulence proteins were also induced, including InlA, InlC, Hyl, Mpl, PlcA, and PlcB, suggesting that acidification may have mimicked conditions inducing some host survival traits. The overall suite of proteins affected appears to be the "last ditch" responses to non-thermal inactivation above and beyond the standard protections afforded in the stationary-growth phase. The array of proteins found here may provide a deeper understanding of the physiological responses of this pathogen during non-thermal inactivation.&lt;/p&gt; &lt;/abstract&gt;

Toxic effects of nSiO2 and mPS on diatoms Nitzschia closterium f. minutissima

To investigate the toxic mechanism of SiO2 nanoparticles (nSiO2) and polystyrene microplastics (mPS) on microalgae Nitzschia closterium f. minutissima, growth inhibition tests were carried out. The growth and biological responses of the algae exposed to nSiO2 (0.5, 1, 2, 5, 10, 30 mg L−1) and mPS (1, 5, 10, 30 and 75 mg L−1) were explored in f/2 media for 96 h. Both micro-/nano-particles (MNPs) inhibited the growth of N. closterium f. minutissima in a concentration- and time-dependent manner. The toxic effect of mPS on N. closterium f. minutissima is higher than that of nSiO2, because silicon is essential for diatoms to maintain cell wall integrity, and the addition of appropriate amounts of nSiO2 can be absorbed and used as a nutrient to promote diatom growth and protect the integrity of the siliceous shell to some extent. Both MNPs induce the production of excess oxidation and activate the cellular antioxidant defense system, leading to increased SOD and CAT activity as a means to resist oxidative damage to the cell, and eliminating excess ROS and maintaining normal cell morphology and metabolism. SEM is consistent with the results of MDA, showing that mPS with high concentrations attach to the surface of algal cells to produce heterogeneous aggregates and disrupt the cell wall and cell membrane, causing the cells to expand and rupture. This study contributes to the understanding of the size effect of MNPs on the growth of marine diatom.

Unveiling the intricate mechanisms of plant defense

Plants may lack mobility, but they are not defenseless against the constant threats posed by pathogens and pests. Pattern Recognition Receptors (PRRs), which are located on the plasma membrane, enable plants to effectively recognize intruders. These receptors function by sensing elicitors or fragments of the cell wall that arise from damage. Recent studies underscore the significance of maintaining cell wall integrity in the coordination of defense mechanisms following the detection of parasitism. Pathogen invasion often triggers alterations in cell wall structure, which leads to the release of molecules like β-glucans and oligogalacturonides. These small molecules are then recognized by PRRs, which stimulate downstream signaling pathways that involve both receptor-like kinases and calcium-dependent signaling. Here, we present the latest insights into plant signaling that play a vital role in immunity: the maintenance of cell wall integrity; the intricate interplay between receptor-like kinases; and the involvement of calcium ions. The goal of the review is to provide readers with a deeper understanding of the intricate mechanisms underlying plant defense strategies.

ZmDRR206 functions in maintaining cell wall integrity during maize seedling growth and defense response to external stresses

Plants adaptively change their cell wall composition and structure during their growth, development, and interactions with environmental stresses. Dirigent proteins (DIRs) contribute to environmental adaptations by dynamically reorganizing the cell wall and/or by generating defense compounds. A maize DIR, ZmDRR206, was previously reported to play a dominant role in regulation of storage nutrient accumulation in endosperm during maize kernel development. Here we show that ZmDRR206 mediates maize seedling growth and disease resistance by coordinately regulating biosynthesis of cell wall components for cell-wall integrity (CWI) maintenance. Expression of ZmDRR206 was induced in maize seedlings upon pathogen infection. ZmDRR206 overexpression in maize resulted in reduced seedling growth and photosynthetic activity but increased disease resistance and drought tolerance, revealing a tradeoff between growth and defense. Consistently, ZmDRR206 overexpression reduced the contents of primary metabolites and down-regulated genes involved in photosynthesis, while increasing the contents of major cell wall components, defense phytohormones, and defense metabolites, and up-regulated genes involved in defense and cell-wall biosynthesis in seedlings. ZmDRR206-overexpressing seedlings were resistant to cell-wall stress imposed by isoxaben, and ZmDRR206 physically interacted with ZmCesA10, which is a cellulose synthase unit. Our findings suggest a mechanism by which ZmDRR206 coordinately regulates biosynthesis of cell-wall components for CWI maintenance during maize seedling growth, and might be exploited for breeding strong disease resistance in maize.

A genetic selection for Mycobacterium smegmatis mutants tolerant to killing by sodium citrate defines a combined role for cation homeostasis and osmotic stress in cell death.

Mycobacteria can colonize environments where the availability of metal ions is limited. Biological or inorganic chelators play an important role in limiting metal availability, and we developed a model to examine Mycobacterium smegmatis survival in the presence of the chelator sodium citrate. We observed that instead of restricting M. smegmatis growth, concentrated sodium citrate killed M. smegmatis. RNAseq analysis during sodium citrate treatment revealed transcriptional signatures of metal starvation and hyperosmotic stress. Notably, metal starvation and hyperosmotic stress, individually, do not kill M. smegmatis under these conditions. A forward genetic transposon selection was conducted to examine why sodium citrate was lethal, and several sodium-citrate-tolerant mutants were isolated. Based on the identity of three tolerant mutants, mgtE, treZ, and fadD6, we propose a dual stress model of killing by sodium citrate, where sodium citrate chelate metals from the cell envelope and then osmotic stress in combination with a weakened cell envelope causes cell lysis. This sodium citrate tolerance screen identified mutants in several other genes with no known function, with most conserved in the pathogen M. tuberculosis. Therefore, this model will serve as a basis to define their functions, potentially in maintaining cell wall integrity, cation homeostasis, or osmotolerance. IMPORTANCE Bacteria require mechanisms to adapt to environments with differing metal availability. When Mycobacterium smegmatis is treated with high concentrations of the metal chelator sodium citrate, the bacteria are killed. To define the mechanisms underlying killing by sodium citrate, we conducted a genetic selection and observed tolerance to killing in mutants of the mgtE magnesium transporter. Further characterization studies support a model where killing by sodium citrate is driven by a weakened cell wall and osmotic stress, that in combination cause cell lysis.

ZmDRR206 Regulates Nutrient Accumulation in Endosperm through Its Role in Cell Wall Biogenesis during Maize Kernel Development.

Dirigent proteins (DIRs) contribute to plant fitness by dynamically reorganizing the cell wall and/or by generating defense compounds during plant growth, development, and interactions with environmental stresses. ZmDRR206 is a maize DIR, it plays a role in maintaining cell wall integrity during seedling growth and defense response in maize, but its role in regulating maize kernel development is unclear. Association analysis of candidate genes indicated that the natural variations of ZmDRR206 were significantly associated with maize hundred-kernel weight (HKW). ZmDRR206 plays a dominant role in storage nutrient accumulation in endosperm during maize kernel development, ZmDRR206 overexpression resulted in small and shrunken maize kernel with significantly reduced starch content and significantly decreased HKW. Cytological characterization of the developing maize kernels revealed that ZmDRR206 overexpression induced dysfunctional basal endosperm transfer layer (BETL) cells, which were shorter with less wall ingrowth, and defense response was constitutively activated in developing maize kernel at 15 and 18 DAP by ZmDRR206 overexpression. The BETL-development-related genes and auxin signal-related genes were down-regulated, while cell wall biogenesis-related genes were up-regulated in developing BETL of the ZmDRR206-overexpressing kernel. Moreover, the developing ZmDRR206-overexpressing kernel had significantly reduced contents of the cell wall components such as cellulose and acid soluble lignin. These results suggest that ZmDRR206 may play a regulatory role in coordinating cell development, storage nutrient metabolism, and stress responses during maize kernel development through its role in cell wall biogenesis and defense response, and provides new insights into understanding the mechanisms of kernel development in maize.

The Pex3-mediated peroxisome biogenesis plays a critical role in metabolic biosynthesis, stress response, and pathogenicity in Alternaria alternata

Peroxisomes are microbodies involved in the metabolism of fatty acids and hydrogen peroxide (H2O2) in eukaryotes. In the current study, an AaPex3 gene encoding a peroxisome membrane protein was demonstrated to be required for peroxisome biogenesis and resistance to peroxides and superoxide-generating compounds. Deleting AaPex3 affected the expression of the genes encoding the NADPH oxidase (NoxA) and the Yap1 stress-responsive transcription regulator, both of which have been implicated in ROS resistance. The AaPex3-mediated peroxisome biogenesis negatively affected resistance to singlet oxygen-generating compounds, 2-chloro-5-hydroxypyridine (CHP), and 2,3,5-triiodobenzoic acid (TIBA), novel phenotypes associated with peroxisomes. Nile red staining revealed that ΔAaPex3 accumulated more lipid bodies than the wild type. ΔAaPex3 conidia had thinner cell walls than the wild type, suggesting the involvement of AaPex3 in maintaining cell wall integrity. Genetic evidence has also demonstrated that the AaPex3-mediated peroxisome biogenesis is required for conidiogenesis, conidia germination, siderophore biosynthesis, toxin production, and virulence. Biotin or lipids could restore ΔAaPex3 growth in axenic culture and on the surface of citrus leaves. In contrast, co-application of ΔAaPex3 with biotin and oleic acid on citrus leaves failed to induce necrotic lesions. Our results revealed the multifaceted functions of peroxisomes in the phytopathogenic fungus.

Evolutionary conservation of motifs within vanA and vanB of vancomycin-resistant enterococci.

Global Health is threatened by the rapid emergence of multidrug-resistant bacteria. Antibiotic resistomes rapidly evolve, yet conserved motifs elucidated in our study have the potential for future drug targets for precision medicine. This study aimed to identify conserved genetic sequences and their evolutionary pathways among vancomycin-resistant Enterococcus species such as Enterococcus faecium and Enterococcus faecalis. We retrieved a total of 26 complete amino acid and nucleotide sequences of resistance determinant genes against vancomycin (vanA and vanB), streptomycin (aac-aah), and penicillin (pbp5) from the publicly available genetic sequence database, GenBank. The sequences were comprised of bacteria classified under the genera of Enterococcus, Staphylococcus, Amycolatopsis, Ruminococcus, and Clostridium. Sequences were aligned with Clustal Omega Multiple Sequence Alignment program and Percent Identity Matrices were derived. Phylogenetic analyses to elucidate evolutionary relationships between sequences were conducted with the neighbor-end joining method through the Molecular Evolutionary Genetics Analysis (MEGAX) software, developed by the Institute of Molecular Evolutionary Genetics at Pennsylvania State University. Subsequent network analyses of the resistance gene, vanB, within E. faecium were derived from ScanProsite and InterPro. We observed the highest nucleotide sequence similarity of vanA regions within strains of E. faecium (100%) and E. faecalis (100%). Between Enterococcus genera, we continued to observe high sequence conservation for vanA and vanB, up to 99.9% similarity. Phylogenetic tree analyses suggest rapid acquisition of these determinants between strains within vanA and vanB, particularly between strains of Enterococcus genera, which may be indicative of horizontal gene transfer. Within E. faecium, Adenosine 5'-Triphosphate (ATP)-Grasp and D-ala-D-ala ligase (Ddl) were found as conserved domains of vanA and vanB. We additionally found that there is notable sequence conservation, up to 66.67%, between resistomes against vancomycin and streptomycin among E. faecium. Resistance genes against vancomycin have highly conserved sequences between strains of Enterococcus bacteria. These conserved sequences within vanA and vanB encode for ATP-Grasp and Ddl motifs, which have functional properties for maintaining cell wall integrity. High sequence conservation is also observed among resistance genes against penicillin and streptomycin, which can inform future drug targets for broader spectrum therapies.

In-situ visualizing selective lignin dissolution of tracheids wall in reaction wood

As a renewable biological macromolecule with aromatic structure, lignin can serve as matrix substance to maintain cell wall integrity and is regarded as the natural biomass recalcitrance. Substantial differences in the cell wall lignin topochemistry between opposite (Ow) and compression wood (Cw) trachieds in Pinus bungeana Zucc. were visualized during [Emim][OAc] pretreatment at room temperature. The ionic liqiuds treatment induced a more obvious wall swelling for highly lignified Cw tracheids than that of Ow, while dynamic Raman spectra analysis indicated the higher lignin and carbohydrates removal for Ow tracheids. Raman imaging further revealed that both lignin and carbohydrates were dissolved simultaneously within the middle lamella and secondary wall of Ow and pretreatment has little effects on Cw tracheids wall. Moreover, it was demonstrated that lignin composition was the key factor to affect the composition dissolution. In particular, lignin G-units were selectively removed from cell corner middle lamella (52.3 %) and secondary wall (62.0 %) of Ow tracheids. When cotton fiber, as a reference was treated under the same conditions, lattice conversion moving from cellulose I to II occurred. The findings confirmed the important role of lignin compostion in the dissolution behavior of carbohydrate dominant tracheids wall.

24-epibrassinolide enhances mango resistance to Colletotrichum gloeosporioides via activating multiple defense response

Brassinosteroids (BRs) are natural steroidal phytohormones that regulate many essential processes of plant growth, development and responses to stress. However, the role of BRs in the defense response against Colletotrichum gloeosporioides in mango fruit remains largely unknown. This study investigated the resistance against C. gloeosporioides and the possible mechanisms of exogenous 24-epibrassinolide (EBR) in mango fruit. Results showed 1 μmol L−1 EBR effectively reduced the disease incidence and disease severity on mango fruit after inoculating with C. gloeosporioides. However, EBR had no direct antifungal activity against C. gloeosporioides in vitro test. EBR significantly improved the activities of phenylalanine ammonia-lyase (PAL), peroxidase (POD), chitinase (CHT), cinnamate-4-hydroxylase (C4H), β-1,3-glucanase (GLU) and 4-hydroxycinnamate CoA ligase (4CL), as well as the accumulation of flavonoids, lignin and total phenolics in mango fruit. EBR treatment also increased hydrogen peroxide (H2O2) content in mango fruit at early storage stage after inoculation, while increased catalase (CAT), ascorbate peroxidase (APX) and superoxide dismutase (SOD) activities with low reactive oxygen species (ROS) level in later storage stage. Besides, EBR treatment markedly reduced the changes in pectin methylesterase (PME) and polygalacturonase (PG) activities, which slowed down the hydrolysis of pectin, thus maintaining cell wall integrity. The principal component analysis indicated that EBR treatment was strongly associated with the increase of defense enzyme activities and metabolite contents and the inhibition of pectin hydrolysis. These results indicated that exogenous EBR could be a promising strategy to increase resistance against C. gloeosporioides in mango fruit by improving the defense responses against pathogens.

The Tol-Pal System Plays an Important Role in Maintaining Cell Integrity During Elongation in Escherichia coli.

The Tol-Pal system is a transenvelope complex widely conserved among Gram-negative bacteria. It is recruited to the septal ring during cytokinesis, and its inactivation causes pleiotropic phenotypes mainly associated with the division process. From our genetic screen to identify factors required for delaying lysis upon treatment of beta lactams, we discovered that the tol-pal mutant shares similar defects with mutants of the major class A PBP system (PBP1b-LpoB) in terms of lysis prevention. Further phenotypic analyses revealed that the Tol-Pal system plays an important role in maintaining cell integrity not only during septation, but also during cell elongation. Simultaneous inactivation of the Tol-Pal system and the PBP1b-LpoB system leads to lysis during cell elongation as well as during division. Moreover, production of the Lpo activator-bypass PBP1b, but not wild-type PBP1b, partially suppressed the Tol-Pal defect in maintaining cell integrity upon treatment of mecillinam specific for the Rod system, suggesting that the Tol-Pal system is likely to be involved in the activation of aPBP by Lpo factors. Overall, our results indicate that the Tol-Pal system plays an important role in maintaining cell wall integrity during elongation in addition to its multifaceted roles during cytokinesis.

Sth1, the ATPase subunit of the RSC chromatin remodeler has important roles in stress response and DNA damage repair in the pathogenic fungi Candida albicans

Candida albicans, the most prevalent fungal pathogen, exists as a commensal in the human host. It is subjected to myriad physiological stress conditions in different host niches, which jeopardizes its fitness to survive and propagate as an established commensal. C. albicans has highly labile chromatin which gets remodeled in response to the stress conditions to facilitate the expression of several stress-responsive genes. Several epigenetic factors including histone variants, histone modifiers and chromatin remodelers that define the chromatin architecture play crucial roles in the regulation of the stress-responsive genes in this organism. Here we investigated the roles of the ATP-dependent chromatin remodeler RSC (Remodel the Structure of Chromatin) in several stress responses in C. albicans, by targeting the key ATPase component, Sth1, given its profound and similar roles exist in Saccharomyces cerevisiae. We have unraveled the crucial roles of the RSC complex (Sth1) in maintaining cell wall integrity and fighting against osmotic and oxidative stresses. We found that the mutant conditionally depleted of Sth1 was sensitive to the cell wall disrupting agents, and the mutant without exposure to any stressor accumulated higher chitin content in the cell wall as a defense mechanism to restore the cell wall integrity. Further, this was supported by the phosphorylation of MAPK1 protein Mkc1, which happens due to activation of the cell wall integrity pathway PKC1. We also observed the Sth1 mutant to be sensitive to oxidative and osmotic stresses in vitro, which are very important and imparted by the host defense mechanism. This suggests that the mutant could get attenuated and hence become less virulent than the wild-type when loss of function of Sth1 happens. We also found that Sth1 has a crucial role in maintaining genomic integrity as sth1 mutant cells accumulate extensive DNA damages and show the loss in cell viability. Overall this work suggests that Sth1 has an important role in fighting against some of the clinically relevant and physiologically important stresses. It also has a crucial role in fighting against stress to the genomic integrity and hence functions in DNA damage repair.

A Cell Wall Hydrolase MepH Is Negatively Regulated by Proteolysis Involving Prc and NlpI in Escherichia coli.

Cell wall assembly of Gram-negative bacteria requires DD-endopeptidase activity that cleaves peptidoglycan (PG) crosslinks in addition to PG synthetic activity, and the activity of DD-endopeptidases needs to be tightly regulated to maintain cell wall integrity during PG expansion. Among the major DD-endopeptidases functioning for PG assembly in Escherichia coli, MepS and MepM have been shown to be negatively controlled by the periplasmic protease Prc. In this study, we performed a genetic selection using the synthetic lethality between the mepS and mepM mutations in rich medium to uncover regulatory mechanisms controlling the activity of DD-endopeptidases other than MepS and MepM. This selection revealed mutations in prc and nlpI as suppressors. Gene deletion analyses revealed that MepH is required for suppression of the MepS— MepM— growth defect by the prc or nlpI mutation. We also discovered that MepH is directly degraded by Prc and that this degradation is further promoted by NlpI. Thus, our study showed that all three DD-endopeptidases which play major roles in PG assembly of E. coli under normal physiological conditions are controlled by a common periplasmic protease.

Glycosylphosphatidylinositol Anchor Biosynthesis Pathway-Related Protein GPI7 Is Required for the Vegetative Growth and Pathogenicity of Colletotrichum graminicola

Glycosylphosphatidylinositol (GPI) anchoring is a common post-translational modification in eukaryotic cells and has been demonstrated to have a wide range of biological functions, such as signal transduction, cellular adhesion, protein transport, immune response, and maintaining cell wall integrity. More than 25 proteins have been proven to participate in the GPI anchor synthesis pathway which occurs in the cytoplasmic and the luminal face of the ER membrane. However, the essential proteins of the GPI anchor synthesis pathway are still less characterized in maize pathogen Colletotrichum graminicola. In the present study, we analyzed the biological function of the GPI anchor synthesis pathway-related gene, CgGPI7, that encodes an ethanolamine phosphate transferase, which is localized in ER. The vegetative growth and conidia development of the ΔCgGPI7 mutant was significantly impaired in C. graminicola. and qRT-PCR results showed that the transcriptional level of CgGPI7 was specifically induced in the initial infection stage and that the pathogenicity of ΔCgGPI7 mutant was also significantly decreased compared with the wild type. Furthermore, the ΔCgGPI7 mutant displayed more sensitivity to cell wall stresses, suggesting that CgGPI7 may play a role in the cell wall integrity of C. graminicola. Cell wall synthesis-associated genes were also quantified in the ΔCgGPI7 mutant, and the results showed that chitin and β-1,3-glucans synthesis genes were significantly up-regulated in ΔCgGPI7 mutants. Our results suggested that CgGPI7 is required for vegetative growth and pathogenicity and might depend on the cell wall integrity of C. graminicola.

Physiological Response of the Resurrection Fern Subjected to Environmental Stress as Shown by Plant Cell Membrane Integrity

Resurrection fern has a unique ability to maintain cell wall integrity when the plant cell is desiccated. It uses proteins such as late embryogenesis proteins and heat shock proteins to maintain their cellular functions. The purpose of this experiment is to determine the effects of environmental stressors on the physiological response of the resurrection fern (Pleopeltis polypodioides). The physiological response of resurrection fern plants was subjected to various temperatures (-50°C, 0°C, 25°C, and 50°C) for 7 days. Results indicated that there was a significant difference between hydrated and desiccated ferns based on the temperature. Additionally, electrolyte leakage measurements confirmed cell damage following exposure to temperature extremes of -50°C and 50°C.