Membrane Stress Research Articles

Role of the anillin-like protein in growth of Cryptococcus neoformans at human host temperature

Cryptococcus neoformans, a basidiomycete yeast, causes lethal meningitis in immunocompromised individuals. The ability of C. neoformans to proliferate at 37°C is essential for virulence. We identified anillin-like protein, CnBud4, as essential for proliferation of C. neoformans at 37°C and for virulence in a heterologous host Galleria mellonella at 25°C. C. neoformans cells lacking CnBud4 were inviable at 25°C in the absence of active calcineurin and were hypersensitive to membrane stress and an anti-fungal agent fluconazole, phenotypes previously described for C. neoformans mutants lacking septins. CnBud4 localized to the mother-bud neck during cytokinesis in a septin-dependent manner. In the absence of CnBud4, septin complex failed to transition from a collar-like single ring to the double ring during cytokinesis. In an ascomycete yeast, Saccharomyces cerevisiae, the anillin-like homologue ScBud4 participates in the organization of the septin ring at the mother-bud neck and plays an important role in specifying location for new bud emergence, known as axial budding pattern. In contrast to their role in S. cerevisiae, neither septins nor CnBud4 were needed to direct the position of the new bud in C. neoformans, suggesting that this function is not conserved in basidiomycetous yeasts. Our data suggest that the requirement of CnBud4 for growth at 37°C and pathogenicity in C. neoformans is based on its conserved role in septin complex organization.

The effect of resting and compression time post-centrifugation on the characteristics of platelet rich fibrin (PRF) membranes.

To evaluate the effect of resting and compression time after centrifugation on the physical properties of platelet rich fibrin (PRF) membranes, and to provide optimal guidance regarding the clinical preparation of PRF. A total of 12 volunteers enrolled in this study divided into 2 groups equally. For each volunteer, 6 tubes of 10mL venous whole blood was drawn. To evaluate the influence of resting time after centrifugation, PRF clots were taken out 0, 1, 3, 5, 7, and 10min from tubes following centrifugation, and then the weight, size, maximum stress, and maximum strain of each group were measured. To evaluate the influence of compression time on the preparation of PRF membranes, the weight ratio of PRF membranes to PRF clots was calculated by compression for 10s, 30s, 60s, 90s, 120s, and 180s, respectively. Scanning electron microscopy was performed to observe the cross-linking of the fibers within membranes, and the maximum stress and strain of PRF membranes were tested followed by stress-strain curve analysis. The weight and volume of PRF clots and PRF membranes increased in size and weight reached the top at 3min, followed by a decrease after 7-min resting. The maximum strain of the PRF membranes after 10min decreased significantly compared to the 3-min and 5-min groups. The maximum stress was found at 3min followed by a statistical decrease when resting time went on. Scanning electron microscopy demonstrated that the internal fibrous structure of the PRF membranes was looser when the compression time was less than 60s when comparing the 90-s group. The maximum stress of PRF membranes was shown using a wait period of 3min post-centrifugation followed by compression for 120s. The findings from the present study demonstrate that the time post-centrifugation of PRF membranes showed a maximum weight, volume, and mechanical properties after resting for 3-5min in the tube post-centrifugation followed by a compression time of 120s. Although research to date has focused primarily on centrifugation protocols, this study revealed for the first time that the resting time post-centrifugation greatly affected the mechanical properties of PRF. This study demonstrated that the resting and compression time after centrifugation influences the mechanical strength of PRF membranes, which might explain differences in PRF characteristics prepared by different clinicians that may provide a standard guide for preparation of PRF membranes.

Tricalbin proteins regulate plasma membrane phospholipid homeostasis.

The evolutionarily conserved extended synaptotagmin (E-Syt) proteins are calcium-activated lipid transfer proteins that function at contacts between the ER and plasma membrane (ER-PM contacts). However, roles of the E-Syt family members in PM lipid organisation remain incomplete. Among the E-Syt family, the yeast tricalbin (Tcb) proteins are essential for PM integrity upon heat stress, but it is not known how they contribute to PM maintenance. Using quantitative lipidomics and microscopy, we find that the Tcb proteins regulate phosphatidylserine homeostasis at the PM. Moreover, upon heat-induced membrane stress, Tcb3 co-localises with the PM protein Sfk1 that is implicated in PM phospholipid asymmetry and integrity. The Tcb proteins also control the PM targeting of the known phosphatidylserine effector Pkc1 upon heat-induced stress. Phosphatidylserine has evolutionarily conserved roles in PM organisation, integrity, and repair. We propose that phospholipid regulation is an ancient essential function of E-Syt family members required for PM integrity.

Influence of geometric imperfections on the nonlinear forced vibration characteristics and stability of laminated angle-ply composite conical shells

The forced vibration characteristic of laminated composite truncated conical shell has been analyzed by including von Karman kinematic non-linearity. The geometric imperfection is included in the analysis by providing an initial doubly sinusoidal transverse displacement. The finite element analysis has been carried out using the displacement field of FSDT and the constrained strain terms are interpolated using field consistent modified shape functions in order to avoid membrane/transverse shear locking. The steady state periodic response has been obtained by solving the governing equations in time domain using modified shooting method and arc length/pseudo-arc length continuation techniques. The main aim of the paper is to explore the effects of initial imperfections, fiber-angle and curvature on the nonlinear periodic response characteristics. Further, the stability of the periodic response has also been investigated and the ensuing bifurcations resulting in loss/gain of stability has been discussed. The parameters affecting the softening/hardening nonlinear behavior of the conical shells have also been assessed. The nonlinear frequency response reveal greater amplitude during inward half cycle due to destabilizing nature of the nonlinear membrane stress resultants during inward motion at large amplitudes. Multiple stress reversals is observed during a periodic cycle which has large ramifications on the fatigue design.

The expression of virulence genes increases membrane permeability and sensitivity to envelope stress in Salmonella Typhimurium.

Virulence gene expression can represent a substantial fitness cost to pathogenic bacteria. In the model entero-pathogen Salmonella Typhimurium (S.Tm), such cost favors emergence of attenuated variants during infections that harbor mutations in transcriptional activators of virulence genes (e.g., hilD and hilC). Therefore, understanding the cost of virulence and how it relates to virulence regulation could allow the identification and modulation of ecological factors to drive the evolution of S.Tm toward attenuation. In this study, investigations of membrane status and stress resistance demonstrate that the wild-type (WT) expression level of virulence factors embedded in the envelope increases membrane permeability and sensitizes S.Tm to membrane stress. This is independent from a previously described growth defect associated with virulence gene expression in S.Tm. Pretreating the bacteria with sublethal stress inhibited virulence expression and increased stress resistance. This trade-off between virulence and stress resistance could explain the repression of virulence expression in response to harsh environments in S.Tm. Moreover, we show that virulence-associated stress sensitivity is a burden during infection in mice, contributing to the inherent instability of S.Tm virulence. As most bacterial pathogens critically rely on deploying virulence factors in their membrane, our findings could have a broad impact toward the development of antivirulence strategies.

Staphylococcus aureus Overcomes Anaerobe-Derived Short-Chain Fatty Acid Stress via FadX and the CodY Regulon.

Chronic rhinosinusitis (CRS) is characterized by immune dysfunction, mucus hypersecretion, and persistent infection of the paranasal sinuses. While Staphylococcus aureus is a primary CRS pathogen, recent sequence-based surveys have found increased relative abundances of anaerobic bacteria, suggesting that S. aureus may experience altered metabolic landscapes in CRS relative to healthy airways. To test this possibility, we characterized the growth kinetics and transcriptome of S. aureus in supernatants of the abundant CRS anaerobe Fusobacterium nucleatum. While growth was initially delayed, S. aureus ultimately grew to similar levels as in the control medium. The transcriptome was significantly affected by F. nucleatum metabolites, with the agr quorum sensing system notably repressed. Conversely, expression of fadX, encoding a putative propionate coenzyme A (CoA)-transferase, was significantly increased, leading to our hypothesis that short-chain fatty acids (SCFAs) produced by F. nucleatum could mediate S. aureus growth behavior and gene expression. Supplementation with propionate and butyrate, but not acetate, recapitulated delayed growth phenotypes observed in F. nucleatum supernatants. A fadX mutant was found to be more sensitive than wild type to propionate, suggesting a role for FadX in the S. aureus SCFA stress response. Interestingly, spontaneous resistance to butyrate, but not propionate, was observed frequently. Whole-genome sequencing and targeted mutagenesis identified codY mutants as resistant to butyrate inhibition. Together, these data show that S. aureus physiology is dependent on its cocolonizing microbiota and metabolites they exchange and indicate that propionate and butyrate may act on different targets in S. aureus to suppress its growth. IMPORTANCE Staphylococcus aureus is an important CRS pathogen, and yet it is found in the upper airways of 30% to 50% of people without complications. The presence of strict and facultative anaerobic bacteria in CRS sinuses has recently spurred research into bacterial interactions and how they influence S. aureus physiology and pathogenesis. We show here that propionate and butyrate produced by one such CRS anaerobe, namely, Fusobacterium nucleatum, alter the growth and gene expression of S. aureus. We show that fadX is important for S. aureus to resist propionate stress and that the CodY regulon mediates growth in inhibitory concentrations of butyrate. This work highlights the possible complexity of S. aureus-anaerobe interactions and implicates membrane stress as a possible mechanism influencing S. aureus behavior in CRS sinuses.

Balanites aegyptiaca (Heglig Dates) Reduces Oxidative Stress and Biophysical Alterations of Erythrocyte Membranes in Streptozotocin-Induced Diabetic Rats

Balanites aegyptiaca (Heglig Dates) Reduces Oxidative Stress and Biophysical Alterations of Erythrocyte Membranes in Streptozotocin-Induced Diabetic Rats

Tunable Antibacterial Activity of a Polypropylene Fabric Coated with Bristling Ti3C2Tx MXene Flakes Coupling the Nanoblade Effect with ROS Generation

Polypropylene (PP) is a thermoplastic polymer widely used as a medical textile in healthcare applications due to its low cost and superior performance. However, it does not show antibacterial properties leading to the possibility of pathogen transmission. Herein, we have developed an antibacterial medical fabric by facile self-assembly of delaminated two-dimensional (2D) Ti3C2Tx MXene flakes bristling on the surface of PP fibers. The increasing amount of MXene in the coating solution from 1 up to 32 mg/mL allowed for edge-on assembly of MXene flakes on the PP surface and tracking the evolution of the band gap for a restacked structure. Characterization of the PP/Ti3C2Tx nanocomposite has proven that it exhibited highly effective antibacterial, robust coating, and chemically/thermally stable properties. The in vitro microbiological studies against Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus have shown that PP/Ti3C2Tx reduced the bacterial viability up to 100%, as driven by synergistic membrane stress mediated by physical contact and light-induced reactive oxygen species (ROS) generation. Moreover, the use of l-ascorbic acid for MXene stabilization allowed for achieving excellent thermal stability of the PP/Ti3C2Tx nanocomposite upon accelerated thermal aging. Collectively, this work provides a facile surface engineering strategy for designing medical fabrics with outstanding functional performances. By demonstrating the exceptional performance of the stabilized MXene in a self-assembly nanocomposite structure, we are opening the door for MXenes to be applied in other biomedical fields.

Influence of Deposition Modes and Thermal Annealing on Residual Stresses in Magnetron-Sputtered YSZ Membranes.

Thin-film electrolyte made of 8-mol% yttria stabilized zirconia (8YSZ) for solid oxide fuel cells (SOFCs) was fabricated on anode substrates using reactive magnetron sputtering of Zr-Y targets in a mixture of Ar and O2 gases. The deposition of 4–6 µm thin-film electrolyte was in the transition or oxide modes differing by the oxygen concentration in the sputtering atmosphere. The half-cell bending of the anode-supported SOFCs was measured to determine the residual stresses in the electrolyte films after the deposition and thermal annealing in air. The dependences were studied between the deposition modes, residual stresses in the films, and the SOFC performance. At 800 °C, the maximum power density of SOFCs ranged between 0.58 and 1.2 W/cm2 depending on the electrolyte deposition mode. Scanning electron microscopy was carried out to investigate the surface morphology and structure of the YSZ electrolyte films after thermal annealing. Additionally, an X-ray diffraction analysis of the YSZ electrolyte films was conducted for the synchrotron radiation beam during thermal annealing at different temperatures up to 1300 °C. It was found that certain deposition modes provide the formation of the YSZ electrolyte films with acceptable residual stresses (<1 GPa) at room temperature, including films deposited on large area anodes (100 × 100 mm2).

ER-PM membrane contact site regulation by yeast ORPs and membrane stress pathways.

In yeast, at least seven proteins (Ice2p, Ist2p, Scs2/22p, Tcb1-Tcb3p) affect cortical endoplasmic reticulum (ER) tethering and contact with the plasma membrane (PM). In Δ-super-tether (Δ-s-tether) cells that lack these tethers, cortical ER-PM association is all but gone. Yeast OSBP homologue (Osh) proteins are also implicated in membrane contact site (MCS) assembly, perhaps as subunits for multicomponent tethers, though their function at MCSs involves intermembrane lipid transfer. Paradoxically, when analyzed by fluorescence and electron microscopy, the elimination of the OSH gene family does not reduce cortical ER-PM association but dramatically increases it. In response to the inactivation of all Osh proteins, the yeast E-Syt (extended-synaptotagmin) homologue Tcb3p is post-transcriptionally upregulated thereby generating additional Tcb3p-dependent ER-PM MCSs for recruiting more cortical ER to the PM. Although the elimination of OSH genes and the deletion of ER-PM tether genes have divergent effects on cortical ER-PM association, both elicit the Environmental Stress Response (ESR). Through comparisons of transcriptomic profiles of cells lacking OSH genes or ER-PM tethers, changes in ESR expression are partially manifested through the induction of the HOG (high-osmolarity glycerol) PM stress pathway or the ER-specific UPR (unfolded protein response) pathway, respectively. Defects in either UPR or HOG pathways also increase ER-PM MCSs, and expression of extra “artificial ER-PM membrane staples” rescues growth of UPR mutants challenged with lethal ER stress. Transcriptome analysis of OSH and Δ-s-tether mutants also revealed dysregulation of inositol-dependent phospholipid gene expression, and the combined lethality of osh4Δ and Δ-s-tether mutations is suppressed by overexpression of the phosphatidic acid biosynthetic gene, DGK1. These findings establish that the Tcb3p tether is induced by ER and PM stresses and ER-PM MCSs augment responses to membrane stresses, which are integrated through the broader ESR pathway.

Numerical investigation quantifies membrane stresses of tumor cells in blood vessels

Capillary size and appearance of red blood cells affect the tensions felt by blood vessel membranes.

Moderately large deflection of slightly curved layered beams with interlayer slip

This paper presents a beam theory for analyzing the static response of slightly curved three-layer beams with interlayer slip. Since the beams are supposed to be immovably supported, membrane stresses develop even at moderately large deflections and the response becomes geometrically nonlinear. The theory is based on a layerwise application of the Euler–Bernoulli theory and a linear elastic constitutive law for the interlaminar displacements. In three application examples, the accuracy of this theory is shown by comparing the results of this theory with the outcomes of a more complex finite element analysis assuming a plane stress state. These application examples demonstrate the effect of a small initial deflection on the nonlinear response of the considered layered structural members.

Numerical Simulation Study on Relationship between the Fracture Mechanisms and Residual Membrane Stresses of Metallic Material.

The distribution and dissipation energies in fracture mechanisms were a critical challenge to derive, especially for this ultra-thin sample. The membrane failure, which is the end of the fracture mechanisms, is a result of the cone wave reflections from the backend membrane boundaries. These reflections delay the failure processes due to the shock impacts. To compare these results with the experimental work, a numerical simulation was conducted for these processes. The cylinder-shaped rigid projectile was impacted using a frictionless Lagrange solver. The target was a cartridge brass circle plate clamped at its perimeter, and its zone was refined to a ten-times higher meshing density for better analysis. The erosion and cut-off controls involved a zero-gap interaction condition and an instantaneous geometric erosion strain of 200%. Due to the maximum projectile velocity of 382 m/s having the slowest perforation, the target thickness was found to be 5.5 mm. The fracture mechanism phenomena, such as tensile, compressive, through-thickness, and growth in-plane delamination, propagating delamination, and local punch shear waves were observed. After deducting tensile and flexural strengths from the last experiment, a total residual membrane stress of 650 MPa was found. This result indicated a relationship between the fracture mechanisms and residual membrane stresses of metallic material.

Large Deflections of Thin-Walled Plates under Transverse Loading-Investigation of the Generated In-Plane Stresses.

Thin-walled plates subjected to transverse loading undergoing large deflection have been the topic of a large number of studies. However, there is still a lack of information about the nature and the distribution membrane stresses generated under large deflections. The purpose of this paper is to calculate and display the distribution of the generated stresses and the respective deflections on the entire rectangular plate area. Finite element analysis results for thin-walled plates with aspect ratios of 1, 2 and 5, on movable and immovable edges simply supported and clamped boundary conditions are clearly visualized. The distribution of the normal and shear stresses enables a good understanding of the plate critical points locations. It was found that strong tensile and compressive membrane stresses exist at various points near the plate edges, creating potential failure hazards.

Identification of a novel cationic glycolipid in Streptococcus agalactiae that contributes to brain entry and meningitis.

Bacterial membrane lipids are critical for membrane bilayer formation, cell division, protein localization, stress responses, and pathogenesis. Despite their critical roles, membrane lipids have not been fully elucidated for many pathogens. Here, we report the discovery of a novel cationic glycolipid, lysyl-glucosyl-diacylglycerol (Lys-Glc-DAG), which is synthesized in high abundance by the bacterium Streptococcus agalactiae (Group B Streptococcus, GBS). To our knowledge, Lys-Glc-DAG is more positively charged than any other known lipids. Lys-Glc-DAG carries 2 positive net charges per molecule, distinct from the widely described lysylated phospholipid lysyl-phosphatidylglycerol (Lys-PG) that carries one positive net charge due to the presence of a negatively charged phosphate moiety. We use normal phase liquid chromatography (NPLC) coupled with electrospray ionization (ESI) high-resolution tandem mass spectrometry (HRMS/MS) and genetic approaches to determine that Lys-Glc-DAG is synthesized by the enzyme MprF in GBS, which covalently modifies the neutral glycolipid Glc-DAG with the cationic amino acid lysine. GBS is a leading cause of neonatal meningitis, which requires traversal of the endothelial blood–brain barrier (BBB). We demonstrate that GBS strains lacking mprF exhibit a significant decrease in the ability to invade BBB endothelial cells. Further, mice challenged with a GBSΔmprF mutant developed bacteremia comparably to wild-type (WT) infected mice yet had less recovered bacteria from brain tissue and a lower incidence of meningitis. Thus, our data suggest that Lys-Glc-DAG may contribute to bacterial uptake into host cells and disease progression. Importantly, our discovery provides a platform for further study of cationic lipids at the host–pathogen interface.

Glomerular Biomechanical Stress and Lipid Mediators during Cellular Changes Leading to Chronic Kidney Disease.

Hyperfiltration is an important underlying cause of glomerular dysfunction associated with several systemic and intrinsic glomerular conditions leading to chronic kidney disease (CKD). These include obesity, diabetes, hypertension, focal segmental glomerulosclerosis (FSGS), congenital abnormalities and reduced renal mass (low nephron number). Hyperfiltration-associated biomechanical forces directly impact the cell membrane, generating tensile and fluid flow shear stresses in multiple segments of the nephron. Ongoing research suggests these biomechanical forces as the initial mediators of hyperfiltration-induced deterioration of podocyte structure and function leading to their detachment and irreplaceable loss from the glomerular filtration barrier. Membrane lipid-derived polyunsaturated fatty acids (PUFA) and their metabolites are potent transducers of biomechanical stress from the cell surface to intracellular compartments. Omega-6 and ω-3 long-chain PUFA from membrane phospholipids generate many versatile and autacoid oxylipins that modulate pro-inflammatory as well as anti-inflammatory autocrine and paracrine signaling. We advance the idea that lipid signaling molecules, related enzymes, metabolites and receptors are not just mediators of cellular stress but also potential targets for developing novel interventions. With the growing emphasis on lifestyle changes for wellness, dietary fatty acids are potential adjunct-therapeutics to minimize/treat hyperfiltration-induced progressive glomerular damage and CKD.

Dynamic alteration of poroelastic attributes as determinant membrane nanorheology for endocytosis of organ specific targeted gold nanoparticles

BackgroundEfficacy of targeted drug delivery using nanoparticles relies on several factors including the uptake mechanisms such as phagocytosis, macropinocytosis, micropinocytosis and receptor mediated endocytosis. These mechanisms have been studied with respect to the alteration in signaling mechanisms, cellular morphology, and linear nanomechanical properties (NMPs). Commonly employed classical contact mechanics models to address cellular NMPs fail to address mesh like structure consisting of bilayer lipids and proteins of cell membrane. To overcome this technical challenge, we employed poroelastic model which accounts for the biphasic nature of cells including their porous behavior exhibiting both solid like (fluid storage) and liquid like (fluid dissipate) behavior.ResultsIn this study, we employed atomic force microscopy to monitor the influence of surface engineering of gold nanoparticles (GNPs) to the alteration of nonlinear NMPs such as drained Poisson’s ratio, effective shear stress, diffusion constant and pore dimensions of cell membranes during their uptake. Herein, we used pancreatic cancer (PDAC) cell lines including Panc1, AsPC-1 and endothelial cell (HUVECs) to understand the receptor-dependent and -independent endocytosis of two different GNPs derived using plectin-1 targeting peptide (PTP-GNP) and corresponding scrambled peptide (sPEP-GNP). Compared to untreated cells, in case of receptor dependent endocytosis of PTP-GNPs diffusion coefficient altered ~ 1264-fold and ~ 1530-fold and pore size altered ~ 320-fold and ~ 260-fold in Panc1 and AsPC-1 cells, respectively. Whereas for receptor independent mechanisms, we observed modest alteration in diffusion coefficient and pore size, in these cells compared to untreated cells. Effective shear stress corresponding to 7.38 ± 0.15 kPa and 20.49 ± 0.39 kPa in PTP-GNP treatment in Panc1 and AsPC-1, respectively was significantly more than that for sPEP-GNP. These results demonstrate that with temporal recruitment of plectin-1 during receptor mediated endocytosis affects the poroelastic attributes of the membrane.ConclusionThis study confirms that nonlinear NMPs of cell membrane are directly associated with the uptake mechanism of nanoparticles and can provide promising insights of the nature of endocytosis mechanism involved for organ specific drug delivery using nanoparticles. Hence, nanomechanical analysis of cell membrane using this noninvasive, label-free and live-cell analytical tool can therefore be instrumental to evaluate therapeutic benefit of nanoformulations.Graphical

Spontaneous local membrane curvature induced by transmembrane proteins

The (local) curvature of cellular membranes acts as a driving force for the targeting of membrane-associated proteins to specific membrane domains, as well as a sorting mechanism for transmembrane proteins, e.g., by accumulation in regions of matching spontaneous curvature. The latter measure was previously experimentally employed to study the curvature induced by the potassium channel KvAP and by aquaporin AQP0. However, the direction of the reported spontaneous curvature levels as well as the molecular driving forces governing the membrane curvature induced by these integral transmembrane proteins could not be addressed experimentally.Here, using both coarse-grained and atomistic molecular dynamics (MD) simulations, we report induced spontaneous curvature values for the homologous potassium channel Kv 1.2/2.1 Chimera (KvChim) and AQP0 embedded in unrestrained lipid bicelles that are in very good agreement with experiment. Importantly, the direction of curvature could be directly assessed from our simulations: KvChim induces a strong positive membrane curvature (≈0.036 nm−1) whereas AQP0 causes a comparably small negative curvature (≈−0.019 nm−1).Analyses of protein-lipid interactions within the bicelle revealed that the potassium channel shapes the surrounding membrane via structural determinants. Differences in shape of the protein-lipid interface of the voltage-gating domains between the extracellular and cytosolic membrane leaflets induce membrane stress and thereby promote a protein-proximal membrane curvature. In contrast, the water pore AQP0 displayed a high structural stability and an only faint effect on the surrounding membrane environment that is connected to its wedge-like shape.

Analytical prediction of tensile stress of photomask protective membranes based on measured frequency spectrum

The quality of photomask protective membranes is generally quantified in terms of its internal tensile stress. However, the tensile stress is not easily measured directly using experimental methods. Hence, the present research derives an theoretical model based on large deflection assumption for predicting the frequency spectrum of a pellicle. Using this model, the tensile stress of any membrane can be inversely derived by comparing the predicted frequency spectrum obtained using different tensile stress estimates with that determined experimentally using confocal microscopy and fast Fourier transform. The feasibility of the proposed approach is demonstrated by comparing the calculated frequencies of the main vibration modes of three commercial pellicles with the experimental results. The validated model is then used to investigate the effects of the membrane initial displacement, membrane width, and membrane thickness on the frequency response of a typical pellicle membrane.

Autophagy-related protein UvAtg7 contributes to mycelial growth, virulence, asexual reproduction and cell stress response in rice false smut fungus Ustilaginoidea virens

Autophagy is a conserved mechanism for nutrient and cytoplasmic components recycling in eukaryotic cell, in which E1-like enzyme Atg7 activates ubiquitin-like conjugation in the autophagy pathway. In plant pathogenic fungi Ustilaginoidea virens, UvAtg7, an ortholog of AAtg7 in baker’s yeast was identified and functionally investigated. UvAtg7 was confirmed to be essential for autophagy, because the disruption of UvATG7 gene in U. virens completely blocked the fusion of autophagosome-like into vacuoles and catalytic degradation of GFP-UvAtg8 under N-starving condition. The fluorescent signal indicated UvAtg7 protein was dispersed in cytoplasma, but spatially coordinated with core autophagy protein UvAtg8 on occasion. Interestingly, disruption of UvATG7 in U. virens caused slightly reduction in mycelial growth, but resulted in a considerable decrease in virulence, conidia production in YT broth and chlamydospore formation on rice false smut balls. Moreover, the UvATG7 deletion mutants exhibited increased sensitivity to cell wall integrity stress caused by congo red and calcofluor white, meanwhile the UvATG7 deletion mutants showed decreased sensitivity to osmotic stress, cell membrane stress and reactiveoxygen stress caused by sorbitol, sodium dodecyl sulfate and H2O2, respectively. All of these defects in UvATG7 deletion mutants could be partially or completely restored by gene complementation. In general, our study indicates that UvAtg7 is essential in autophagy pathway and contributes to mycelial growth, virulence, asexual reproduction and cell stress response in U. virens.