scholarly journals The Antifungal Protein AFP from Aspergillus giganteus Inhibits Chitin Synthesis in Sensitive Fungi

2007 ◽  
Vol 73 (7) ◽  
pp. 2128-2134 ◽  
Author(s):  
Silke Hagen ◽  
Florentine Marx ◽  
Arthur F. Ram ◽  
Vera Meyer
Keyword(s):  
Cell Wall ◽  
Wall Stress ◽  
Chitin Synthase ◽  
Antifungal Protein ◽  
Cell Integrity ◽  
Class Iii ◽  
Chitin Synthesis ◽  
Aspergillus Giganteus ◽  
Cell Wall Stress ◽  
Sytox Green

ABSTRACT The antifungal protein AFP from Aspergillus giganteus is highly effective in restricting the growth of major human- and plant-pathogenic filamentous fungi. However, a fundamental prerequisite for the use of AFP as an antifungal drug is a complete understanding of its mode of action. In this study, we performed several analyses focusing on the assumption that the chitin biosynthesis of sensitive fungi is targeted by AFP. Here we show that the N-terminal domain of AFP (amino acids 1 to 33) is sufficient for efficient binding of AFP to chitin but is not adequate for inhibition of the growth of sensitive fungi. AFP susceptibility tests and SYTOX Green uptake experiments with class III and class V chitin synthase mutants of Fusarium oxysporum and Aspergillus oryzae showed that deletions made the fungi less sensitive to AFP and its membrane permeabilization effect. In situ chitin synthase activity assays revealed that chitin synthesis is specifically inhibited by AFP in sensitive fungi, indicating that AFP causes cell wall stress and disturbs cell integrity. Further evidence that there was AFP-induced cell wall stress was obtained by using an Aspergillus niger reporter strain in which the cell wall integrity pathway was strongly induced by AFP.

scholarly journals Saccharomyces cerevisiae Mid2p Is a Potential Cell Wall Stress Sensor and Upstream Activator of thePKC1-MPK1 Cell Integrity Pathway

1999 ◽  
Vol 181 (11) ◽  
pp. 3330-3340 ◽  
Author(s):  
Troy Ketela ◽  
Robin Green ◽  
Howard Bussey
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Wall Stress ◽  
Structural Features ◽  
Mitogen Activated Protein Kinase ◽  
Cell Integrity ◽  
Chitin Synthesis ◽  
Calcofluor White ◽  
Reading Frame ◽  
Cell Wall Stress

ABSTRACT The MID2 gene of Saccharomyces cerevisiaeencodes a protein with structural features indicative of a plasma membrane-associated cell wall sensor. MID2 was isolated as a multicopy activator of the Skn7p transcription factor. Deletion ofMID2 causes resistance to calcofluor white, diminished production of stress-induced cell wall chitin under a variety of conditions, and changes in growth rate and viability in a number of different cell wall biosynthesis mutants. Overexpression ofMID2 causes hyperaccumulation of chitin and increased sensitivity to calcofluor white. α-Factor hypersensitivity ofmid2Δ mutants can be suppressed by overexpression of upstream elements of the cell integrity pathway, includingPKC1, RHO1, WSC1, andWSC2. Mid2p and Wsc1p appear to have overlapping roles in maintaining cell integrity since mid2Δ wsc1Δ mutants are inviable on medium that does not contain osmotic support. A role for MID2 in the cell integrity pathway is further supported by the finding that MID2 is required for induction of Mpk1p tyrosine phosphorylation during exposure to α-factor, calcofluor white, or high temperature. Our data are consistent with a role for Mid2p in sensing cell wall stress and in activation of a response that includes both increased chitin synthesis and the Mpk1p mitogen-activated protein kinase cell integrity pathway. In addition, we have identified an open reading frame, MTL1, which encodes a protein with both structural and functional similarity to Mid2p.

scholarly journals Chitin Synthesis in Saccharomyces cerevisiae in Response to Supplementation of Growth Medium with Glucosamine and Cell Wall Stress

2003 ◽  
Vol 2 (5) ◽  
pp. 886-900 ◽  
Author(s):  
Dorota A. Bulik ◽  
Mariusz Olczak ◽  
Hector A. Lucero ◽  
Barbara C. Osmond ◽  
Phillips W. Robbins ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Plasma Membrane ◽  
Growth Medium ◽  
Specific Activity ◽  
Neck Region ◽  
Wall Stress ◽  
Chitin Synthesis ◽  
Fourfold Increase ◽  
Cell Wall Stress

ABSTRACT In Saccharomyces cerevisiae most chitin is synthesized by Chs3p, which deposits chitin in the lateral cell wall and in the bud-neck region during cell division. We have recently found that addition of glucosamine (GlcN) to the growth medium leads to a three- to fourfold increase in cell wall chitin levels. We compared this result to the increases in cellular chitin levels associated with cell wall stress and with treatment of yeast with mating pheromone. Since all three phenomena lead to increases in precursors of chitin, we hypothesized that chitin synthesis is at least in part directly regulated by the size of this pool. This hypothesis was strengthened by our finding that addition of GlcN to the growth medium causes a rapid increase in chitin synthesis without any pronounced change in the expression of more than 6,000 genes monitored with Affymetrix gene expression chips. In other studies we found that the specific activity of Chs3p is higher in the total membrane fractions from cells grown in GlcN and from mutants with weakened cell walls. Sucrose gradient analysis shows that Chs3p is present in an inactive form in what may be Golgi compartments but as an active enzyme in other intracellular membrane-bound vesicles, as well as in the plasma membrane. We conclude that Chs3p-dependent chitin synthesis in S. cerevisiae is regulated both by the levels of intermediates of the UDP-GlcNAc biosynthetic pathway and by an increase in the activity of the enzyme in the plasma membrane.

scholarly journals Cbk1 kinase and Bck2 control MAP kinase activation and inactivation during heat shock

2011 ◽  
Vol 22 (24) ◽  
pp. 4892-4907 ◽  
Author(s):  
Venkata K. Kuravi ◽  
Cornelia Kurischko ◽  
Manasi Puri ◽  
Francis C. Luca
Keyword(s):  
Gene Expression ◽  
Cell Wall ◽  
Heat Shock ◽  
Wall Stress ◽  
Mitogen Activated Protein Kinase ◽  
Stress Signaling ◽  
Cell Integrity ◽  
Loss Of Function ◽  
Kinase Activation ◽  
Cell Wall Stress

Saccharomyces cerevisiae Cbk1 kinase is a LATS/NDR tumor suppressor orthologue and component of the Regulation of Ace2 and Morphogenesis signaling network. Cbk1 was previously implicated in regulating polarized morphogenesis, gene expression, and cell integrity. Here we establish that Cbk1 is critical for heat shock and cell wall stress signaling via Bck2, a protein associated with the Pkc1-Mpk1 cell integrity pathway. We demonstrate that cbk1 and bck2 loss-of-function mutations prevent Mpk1 kinase activation and Mpk1-dependent gene expression but do not disrupt Mpk1 Thr-190/Tyr-192 phosphorylation. Bck2 overexpression partially restores Mpk1-dependent Rlm1 transcription factor activity in cbk1 mutants, suggesting that Bck2 functions downstream of Cbk1. We demonstrate that Bck2 precisely colocalizes with the mitogen-activated protein kinase (MAPK) phosphatase Sdp1. During heat shock, Bck2 and Sdp1 transiently redistribute from nuclei and the cytosol to mitochondria and other cytoplasmic puncta before returning to their pre-stressed localization patterns. Significantly, Cbk1 inhibition delays the return of Bck2 and Sdp1 to their pre-stressed localization patterns and delays Mpk1 Thr-190/Tyr-192 dephosphorylation upon heat shock adaptation. We conclude that Cbk1 and Bck2 are required for Mpk1 activation during heat shock and cell wall stress and for Mpk1 dephosphorylation during heat shock adaptation. These data provide the first evidence that Cbk1 kinase regulates MAPK-dependent stress signaling and provide mechanistic insight into Sdp1 phosphatase regulation.

scholarly journals A novel checkpoint pathway controls actomyosin ring constriction trigger in fission yeast

eLife ◽  
2020 ◽  
Vol 9 ◽  
Author(s):  
Tomás Edreira ◽  
Rubén Celador ◽  
Elvira Manjón ◽  
Yolanda Sánchez
Keyword(s):  
Cell Wall ◽  
Fission Yeast ◽  
Wall Stress ◽  
Cell Integrity ◽  
Wild Type ◽  
Cell Wall Stress ◽  
Checkpoint Pathway ◽  
In The Wild ◽  
Septation Initiation Network ◽  
Ring Constriction

In fission yeast, the septation initiation network (SIN) ensures temporal coordination between actomyosin ring (CAR) constriction with membrane ingression and septum synthesis. However, questions remain about CAR regulation under stress conditions. We show that Rgf1p (Rho1p GEF), participates in a delay of cytokinesis under cell wall stress (blankophor, BP). BP did not interfere with CAR assembly or the rate of CAR constriction, but did delay the onset of constriction in the wild type cells but not in the rgf1Δ cells. This delay was also abolished in the absence of Pmk1p, the MAPK of the cell integrity pathway (CIP), leading to premature abscission and a multi-septated phenotype. Moreover, cytokinesis delay correlates with maintained SIN signaling and depends on the SIN to be achieved. Thus, we propose that the CIP participates in a checkpoint, capable of triggering a CAR constriction delay through the SIN pathway to ensure that cytokinesis terminates successfully.

scholarly journals The chitin synthase genes chsA and chsC are not required for cell wall stress responses in the human pathogen Aspergillus fumigatus

2011 ◽  
Vol 411 (3) ◽  
pp. 549-554 ◽  
Author(s):  
Luise E. Rogg ◽  
Jarrod R. Fortwendel ◽  
Praveen Rao Juvvadi ◽  
Amanda Lilley ◽  
William J. Steinbach
Keyword(s):  
Cell Wall ◽  
Aspergillus Fumigatus ◽  
Stress Responses ◽  
Wall Stress ◽  
Chitin Synthase ◽  
Human Pathogen ◽  
Cell Wall Stress

scholarly journals Boric Acid Disturbs Cell Wall Synthesis inSaccharomyces cerevisiae

2010 ◽  
Vol 2010 ◽  
pp. 1-10 ◽  
Author(s):  
Martin Schmidt ◽  
Jaron Z. Schaumberg ◽  
Courtney M. Steen ◽  
Michael P. Boyer
Keyword(s):  
Antimicrobial Activity ◽  
Cell Wall ◽  
Alternative Medicine ◽  
Boric Acid ◽  
Wall Stress ◽  
Cell Wall Synthesis ◽  
Chitin Synthesis ◽  
Cell Wall Stress ◽  
Bud Neck ◽  
Irregular Cell

Boric acid (BA) has broad antimicrobial activity that makes it a popular treatment for yeast vaginitis in complementary and alternative medicine. In the model yeastS. cerevisiae, BA disturbs the cytoskeleton at the bud neck and impairs the assembly of the septation apparatus. BA treatment causes cells to form irregular septa and leads to the synthesis of irregular cell wall protuberances that extend far into the cytoplasm. The thick, chitin-rich septa that are formed during BA exposure prevent separation of cells after abscission and cause the formation of cell chains and clumps. As a response to the BA insult, cells signal cell wall stress through the Slt2p pathway and increase chitin synthesis, presumably to repair cell wall damage.

Potent Chitin Synthase Inhibitors from Plants

2020 ◽  
Vol 16 (1) ◽  
pp. 58-63
Author(s):  
Amrutha Vijayakumar ◽  
Ajith Madhavan ◽  
Chinchu Bose ◽  
Pandurangan Nanjan ◽  
Sindhu S. Kokkal ◽  
...  
Keyword(s):  
Cell Wall ◽  
Candida Albicans ◽  
Caenorhabditis Elegans ◽  
Aspergillus Niger ◽  
Small Molecules ◽  
Tip Growth ◽  
Chitin Synthase ◽  
Chitin Synthesis ◽  
Hyphal Tip Growth ◽  
Hyphal Tip

Background: Chitin is the main component of fungal, protozoan and helminth cell wall. They help to maintain the structural and functional characteristics of these organisms. The chitin wall is dynamic and is repaired, rearranged and synthesized as the cells develop. Active synthesis can be noticed during cytokinesis, laying of primary septum, maintenance of lateral cell wall integrity and hyphal tip growth. Chitin synthesis involves coordinated action of two enzymes namely, chitin synthase (that lays new cell wall) and chitinase (that removes the older ones). Since chitin synthase is conserved in different eukaryotic microorganisms that can be a ‘soft target’ for inhibition with small molecules. When chitin synthase is inhibited, it leads to the loss of viability of cells owing to the self- disruption of the cell wall by existing chitinase. Methods: In the described study, small molecules from plant sources were screened for their ability to interfere with hyphal tip growth, by employing Hyphal Tip Burst assay (HTB). Aspergillus niger was used as the model organism. The specific role of these small molecules in interfering with chitin synthesis was established with an in-vitro method. The enzyme required was isolated from Aspergillus niger and its activity was deduced through a novel method involving non-radioactively labelled substrate. The activity of the potential lead molecules were also checked against Candida albicans and Caenorhabditis elegans. The latter was adopted as a surrogate for the pathogenic helminths as it shares similarity with regard to cell wall structure and biochemistry. Moreover, it is widely studied and the methodologies are well established. Results: Out of the 11 compounds and extracts screened, 8 were found to be prospective. They were also found to be effective against Candida albicans and Caenorhabditis elegans. Conclusion: Purified Methyl Ethyl Ketone (MEK) Fraction1 (F1) of Coconut (Cocos nucifera) Shell Extract (COSE) was found to be more effective against Candida albicans with an IC50 value of 3.04 μg/mL and on L4 stage of Caenorhabditis elegans with an IC50 of 77.8 μg/mL.

In vitro Cell Wall Stress Assay for Fusarium oxysporum

BIO-PROTOCOL ◽  
2016 ◽  
Vol 6 (17) ◽  
Author(s):  
Elena Pérez-Nadales ◽  
Antonio Di Pietro
Keyword(s):  
Cell Wall ◽  
Fusarium Oxysporum ◽  
Wall Stress ◽  
Cell Wall Stress

Identification of cell-wall stress as a hexose-dependent and osmosensitive regulator of plant responses

2009 ◽  
Vol 57 (6) ◽  
pp. 1015-1026 ◽  
Author(s):  
Thorsten Hamann ◽  
Mark Bennett ◽  
John Mansfield ◽  
Christopher Somerville
Keyword(s):  
Cell Wall ◽  
Wall Stress ◽  
Plant Responses ◽  
Cell Wall Stress
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