scholarly journals Yapsins Are a Family of Aspartyl Proteases Required for Cell Wall Integrity in Saccharomyces cerevisiae

2005 ◽  
Vol 4 (8) ◽  
pp. 1364-1374 ◽  
Author(s):  
Damian J. Krysan ◽  
Elizabeth L. Ting ◽  
Claudia Abeijon ◽  
Lee Kroos ◽  
Robert S. Fuller
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Specific Activity ◽  
Wall Stress ◽  
Cell Wall Integrity ◽  
Dependent Manner ◽  
Glucan Synthesis ◽  
Aspartyl Proteases

ABSTRACT The yeast cell wall is a crucial extracellular organelle that protects the cell from lysis during environmental stress and morphogenesis. Here, we demonstrate that the yapsin family of five glycosylphosphatidylinositol-linked aspartyl proteases is required for cell wall integrity in Saccharomyces cerevisiae. Yapsin null mutants show hypersensitivity to cell wall perturbation, and both the yps1Δ2Δ mutant and the quintuple yapsin mutant (5ypsΔ) undergo osmoremedial cell lysis at 37°C. The cell walls of both 5ypsΔ and yps1Δ2Δ mutants have decreased amounts of 1,3- and 1,6-β-glucan. Although there is decreased incorporation of both 1,3- and 1,6-β-glucan in the 5ypsΔ mutant in vivo, in vitro specific activity of both 1,3- and 1,6-β-glucan synthesis is similar to wild type, indicating that the yapsins affect processes downstream of glucan synthesis and that the yapsins may be involved in the incorporation or retention of cell wall glucan. Presumably as a response to the significant alterations in cell wall composition, the cell wall integrity mitogen-activated kinase signaling cascade (PKC1-MPK pathway) is basally active in 5ypsΔ. YPS1 expression is induced during cell wall stress and remodeling in a PKC1-MPK1-dependent manner, indicating that Yps1p is a direct, and important, output of the cell wall integrity response. The Candida albicans (SAP9) and Candida glabrata (CgYPS1) homologues of YPS1 complement the phenotypes of the yps1Δ mutant. Taken together, these data indicate that the yapsins play an important role in glucan homeostasis in S. cerevisiae and that yapsin homologues may play a similar role in the pathogenic yeasts C. albicans and C. glabrata.

scholarly journals Differential Regulation of the Cell Wall Integrity Mitogen-Activated Protein Kinase Pathway in Budding Yeast by the Protein Tyrosine Phosphatases Ptp2 and Ptp3

1999 ◽  
Vol 19 (11) ◽  
pp. 7651-7660 ◽  
Author(s):  
Christopher P. Mattison ◽  
Scott S. Spencer ◽  
Kurt A. Kresge ◽  
Ji Lee ◽  
Irene M. Ota
Keyword(s):  
Cell Wall ◽  
Protein Tyrosine Phosphatases ◽  
Negative Regulator ◽  
Cell Wall Integrity ◽  
Dependent Manner ◽  
Tyrosine Phosphatases ◽  
Protein Tyrosine ◽  
Growth Defects ◽  
Mitogen Activated Protein

ABSTRACT Mitogen-activated protein kinases (MAPKs) are inactivated by dual-specificity and protein tyrosine phosphatases (PTPs) in yeasts. InSaccharomyces cerevisiae, two PTPs, Ptp2 and Ptp3, inactivate the MAPKs, Hog1 and Fus3, with different specificities. To further examine the functions and substrate specificities of Ptp2 and Ptp3, we tested whether they could inactivate a third MAPK, Mpk1, in the cell wall integrity pathway. In vivo and in vitro evidence indicates that both PTPs inactivate Mpk1, but Ptp2 is the more effective negative regulator. Multicopy expression of PTP2, but not PTP3, suppressed growth defects due to the MEK kinase mutation, BCK1-20, and the MEK mutation,MKK1-386, that hyperactivate this pathway. In addition, deletion of PTP2, but not PTP3, exacerbated growth defects due to MKK1-386. Other evidence supported a role for Ptp3 in this pathway. Expression of MKK1-386 was lethal in the ptp2Δ ptp3Δ strain but not in either single PTP deletion strain. In addition, the ptp2Δ ptp3Δ strain showed higher levels of heat stress-induced Mpk1-phosphotyrosine than the wild-type strain or strains lacking either PTP. The PTPs also showed differences in vitro. Ptp2 was more efficient than Ptp3 at binding and dephosphorylating Mpk1. Another factor that may contribute to the greater effectiveness of Ptp2 is its subcellular localization. Ptp2 is predominantly nuclear whereas Ptp3 is cytoplasmic, suggesting that active Mpk1 is present in the nucleus. Last, PTP2 but not PTP3 transcript increased in response to heat shock in a Mpk1-dependent manner, suggesting that Ptp2 acts in a negative feedback loop to inactivate Mpk1.

Abstract 3: Apolipoprotein A-I Inhibits Streptococcal Cell Wall-Induced Arthritis in the Rat in an ABCA1-dependent Manner

2013 ◽  
Vol 33 (suppl_1) ◽  
Author(s):  
Ben J Wu ◽  
Kwok L Ong ◽  
Sudichhya Shrestha ◽  
Kang Chen ◽  
Philip J Barter ◽  
...  
Keyword(s):  
Cell Wall ◽  
Inflammatory Cytokine ◽  
Density Lipoprotein ◽  
Joint Inflammation ◽  
Chronic Inflammatory Disease ◽  
Dependent Manner ◽  
Tlr2 Expression ◽  
Streptococcal Cell Wall

Introduction. Arthritis is a chronic inflammatory disease characterized by joint inflammation and destruction, reduced high-density lipoprotein (HDL) levels, and increased cardiovascular risk. Objective To determine if apolipoprotein (apo) A-I, the main HDL apolipoprotein, prevents joint inflammation in arthritis. Methods and Results In vivo: Arthritis was induced in female Lewis rats with a single 15 mg/kg intraperitoneal streptococcal cell wall peptidoglycan-polysaccharide (PG-PS) injection and quantified as a combined forepaw and hindpaw inflammation score. Arthritis progressed from an initial, acute phase of joint inflammation during the first 4 days post-PG-PS administration to remission by day 8, followed by chronic joint inflammation up to sacrifice at day 21. Two intravenous infusions of lipid-free apoA-I (8 mg/kg) 24 h pre- and 24 h post-PG-PS injection reduced the acute and chronic joint inflammation by 63±9% at day 3 and by 61±8% at day 21. Infusion of apoA-I at days 7, 9 and 11 post-PG-PS injection reduced the chronic response by 43±11% at day 21. ApoA-I infusions at 24 h prior to and at days 1, 7, 9, 11 post-PG-PS injection reduced joint inflammation by 61±5% at day 3 and by 90±5% at day 21 (p<0.05 for all vs saline infusion). These beneficial effects of apoA-I were accompanied by a reduced inflammatory white blood cell count, reduced pro-inflammatory cytokine levels in synovial fluid, and reduced macrophage accumulation, toll-like receptor 2 (TLR2) and inflammatory cytokine expression in synovial tissue. In vitro: Human monocyte-derived macrophages (HMDMs) were pre-incubated with lipid-free apoA-I, then stimulated with PG-PS (20 μg/mL). Pre-incubation with apoA-I inhibited PG-PS-induced TLR2 and MyD88, a TLR2 adapter protein, expression. Nuclear factor-κB activation and pro-inflammatory cytokine production were also attenuated. These anti-inflammatory effects of apoA-I were abolished in HMDMs transfected with ATP-binding cassette transporter 1 (ABCA1) siRNA. Conclusions These findings establish that apoA-I attenuates PG-PS induced arthritis in the rat. These effects may involve ABCA-1 and inhibition of TLR2 expression and activation.

scholarly journals Deletion of YLR358C Contributes to Reduce Cell Wall Integrity in Saccharomyces Cerevisiae

2022 ◽  
Author(s):  
Yu Zhang ◽  
Mengyan Li ◽  
Hanying Wang ◽  
Juqing Deng ◽  
Jianxing Liu ◽  
...  
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Cell Wall ◽  
Sodium Dodecyl ◽  
Wall Stress ◽  
Pathogenic Fungi ◽  
Cell Wall Integrity ◽  
Calcofluor White ◽  
Fungal Cell ◽  
Reading Frame ◽  
Stress Signals

Abstract The mechanism of fungal cell wall synthesis and assembly is still unclear. Saccharomyces cerevisiae (S. cerevisiae) and pathogenic fungi are conserved in cell wall construction and response to stress signals, and often respond to cell wall stress through activated cell wall integrity (CWI) pathways. Whether the YLR358C open reading frame regulates CWI remains unclear. This study found that the growth of S. cerevisiae with YLR358C knockout was significantly inhibited on the medium containing different concentrations of cell wall interfering agents Calcofluor White (CFW), Congo Red (CR) and sodium dodecyl sulfate (SDS). CFW staining showed that the cell wall chitin was down-regulated, and transmission electron microscopy also observed a decrease in cell wall thickness. Transcriptome sequencing and analysis showed that YLR358C gene may be involved in the regulation of CWI signaling pathway. It was found by qRT-PCR that WSC3, SWI4 and HSP12 were differentially expressed after YLR358C was knocked out. The above results suggest that YLR358C may regulate the integrity of the yeast cell walls and has some potential for application in fermentation.

scholarly journals Stimulation of cell proliferation in skeletal tissues of the rat by defined parathyroid hormone fragments

1991 ◽  
Vol 277 (3) ◽  
pp. 863-868 ◽  
Author(s):  
D Sömjen ◽  
K D Schlüter ◽  
E Wingender ◽  
H Mayer ◽  
A M Kaye
Keyword(s):  
Parathyroid Hormone ◽  
Bone Resorption ◽  
Thymidine Incorporation ◽  
Specific Activity ◽  
Fold Increase ◽  
Dependent Manner ◽  
Equimolar Concentration ◽  
Dose Dependent

We have found, in previous studies in vitro using skeletal derived cell cultures, that mid-region fragments of human parathyroid hormone (hPTH) stimulate [3H]thymidine incorporation into DNA and increase the specific activity of the brain-type isoenzyme of creatine kinase (CK). These changes occurred without an increase in cyclic AMP formation which is linked to bone resorption. In this study, we found that the mid-region fragment hPTH-(28-48) stimulated CK activity in diaphysis, epiphysis and kidney in a time- and dose-dependent manner, parallel to the effects of the whole molecule bovine (b)PTH-(1-84) and the fully active fragment hPTH-(1-34). The increase caused by hPTH-(28-48) at a dose of 1.25 micrograms/rat was not less than the 2-fold increase caused by a roughly equimolar concentration bPTH-(1-84). A significant increase was reached at 1 h after intraperitoneal injection in all cases. All three sequences of PTH caused an increase in [3H]thymidine incorporation into DNA in diaphysis and epiphysis, but not in kidney, 24 h after injection. A fragment further towards the C-terminal, hPTH-(34-47), was inactive compared with an equimolar concentration of the fragment hPTH-(25-39), which stimulated both CK activity and DNA synthesis. These results in vivo are in line with previous findings in vitro; they provide further support for the suggestion that mid-region fragments of the PTH molecule could be used to induce bone formation without incurring the deleterious effect of bone resorption.

scholarly journals O-Mannosyltransferase 1 in Aspergillus fumigatus (AfPmt1p) Is Crucial for Cell Wall Integrity and Conidium Morphology, Especially at an Elevated Temperature

2007 ◽  
Vol 6 (12) ◽  
pp. 2260-2268 ◽  
Author(s):  
Hui Zhou ◽  
Hongyan Hu ◽  
Lijuan Zhang ◽  
Ruoyu Li ◽  
Haomiao Ouyang ◽  
...  
Keyword(s):  
Cell Wall ◽  
Elevated Temperature ◽  
Aspergillus Fumigatus ◽  
Cell Wall Integrity ◽  
Secretory Proteins ◽  
Experimental Conditions ◽  
Human Fungal Pathogen ◽  
Conidium Formation

ABSTRACT Protein O-mannosyltransferases initiate O mannosylation of secretory proteins, which are of fundamental importance in eukaryotes. In this study, the PMT gene family of the human fungal pathogen Aspergillus fumigatus was identified and characterized. Unlike the case in Saccharomyces cerevisiae, where the PMT family is highly redundant, only one member of each PMT subfamily, namely, Afpmt1, Afpmt2, and Afpmt4, is present in A. fumigatus. Mutants with a deletion of Afpmt1 are viable. In vitro and in vivo activity assays confirmed that the protein encoded by Afpmt1 acts as an O-mannosyltransferase (AfPmt1p). Characterization of the ΔAfpmt1 mutant showed that a lack of AfPmt1p results in sensitivity to elevated temperature and defects in growth and cell wall integrity, thereby affecting cell morphology, conidium formation, and germination. In a mouse model, Afpmt1 was not required for the virulence of A. fumigatus under the experimental conditions used.

scholarly journals Opposite roles of trehalase activity in heat-shock recovery and heat-shock survival in Saccharomyces cerevisiae

1999 ◽  
Vol 343 (3) ◽  
pp. 621-626 ◽  
Author(s):  
Stefaan WERA ◽  
Ellen DE SCHRIJVER ◽  
Ilse GEYSKENS ◽  
Solomon NWAKA ◽  
Johan M. THEVELEIN
Keyword(s):  
Saccharomyces Cerevisiae ◽  
Heat Shock ◽  
Specific Activity ◽  
Point Mutations ◽  
Activity Levels ◽  
Protective Function ◽  
Trehalase Activity ◽  
Neutral Trehalase

A variety of results has been obtained consistent with activation of neutral trehalase in Saccharomyces cerevisiae through direct phosphorylation by cAMP-dependent protein kinase (PKA). A series of neutral trehalase mutant alleles, in which all evolutionarily conserved putative phosphorylation sites were changed into alanine, was tested for activation in vitro (by PKA) and in vivo (by glucose addition). None of the mutations alone affected the activation ratio, whereas all mutations combined resulted in an inactive enzyme. All mutant alleles were expressed to similar levels, as shown by Western blotting. Several of the point mutations significantly lowered the specific activity. Using this series of mutants with different activity levels we show an inverse relationship between trehalase activity and heat-shock survival during glucose-induced trehalose mobilization. This is consistent with a stress-protective function of trehalose. On the other hand, reduction of trehalase activity below a certain threshold level impaired recovery from a sublethal heat shock. This suggests that trehalose breakdown is required for efficient recovery from heat shock, and that the presence of trehalase protein alone is not sufficient for efficient heat-stress recovery.

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 TREM2 is a receptor for non-glycosylated mycolic acids of mycobacteria that limits anti-mycobacterial macrophage activation

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Ei’ichi Iizasa ◽  
Yasushi Chuma ◽  
Takayuki Uematsu ◽  
Mio Kubota ◽  
Hiroaki Kawaguchi ◽  
...  
Keyword(s):  
Cell Wall ◽  
Macrophage Activation ◽  
Mycolic Acid ◽  
Dependent Manner ◽  
Independent Manner ◽  
Lectin Receptors ◽  
Mycobacterial Cell ◽  
Mycobacterial Cell Wall

AbstractMycobacterial cell-wall glycolipids elicit an anti-mycobacterial immune response via FcRγ-associated C-type lectin receptors, including Mincle, and caspase-recruitment domain family member 9 (CARD9). Additionally, mycobacteria harbor immuno-evasive cell-wall lipids associated with virulence and latency; however, a mechanism of action is unclear. Here, we show that the DAP12-associated triggering receptor expressed on myeloid cells 2 (TREM2) recognizes mycobacterial cell-wall mycolic acid (MA)-containing lipids and suggest a mechanism by which mycobacteria control host immunity via TREM2. Macrophages respond to glycosylated MA-containing lipids in a Mincle/FcRγ/CARD9-dependent manner to produce inflammatory cytokines and recruit inducible nitric oxide synthase (iNOS)-positive mycobactericidal macrophages. Conversely, macrophages respond to non-glycosylated MAs in a TREM2/DAP12-dependent but CARD9-independent manner to recruit iNOS-negative mycobacterium-permissive macrophages. Furthermore, TREM2 deletion enhances Mincle-induced macrophage activation in vitro and inflammation in vivo and accelerates the elimination of mycobacterial infection, suggesting that TREM2-DAP12 signaling counteracts Mincle-FcRγ-CARD9-mediated anti-mycobacterial immunity. Mycobacteria, therefore, harness TREM2 for immune evasion.

Na(+)-K(+)-ATPase gene expression during in vitro development of rat fetal forebrain

1990 ◽  
Vol 258 (6) ◽  
pp. C1062-C1069 ◽  
Author(s):  
I. Corthesy-Theulaz ◽  
A. M. Merillat ◽  
P. Honegger ◽  
B. C. Rossier
Keyword(s):  
Cell Cultures ◽  
Specific Activity ◽  
Relative Rate ◽  
Adult Brain ◽  
Similar Proportion ◽  
Neuronal Cultures ◽  
Dependent Manner ◽  
Atpase Gene

The existence of at least three isoforms of Na(+)-K(+)-ATPase in adult brain tissues [alpha 1, kidney type; alpha 2 [or alpha(+)]; alpha 3] suggests that these genes might be regulated in a cell-specific and time-dependent manner during development. We have studied this question in serum-free aggregating cell cultures of mechanically dissociated rat fetal telencephalon. At the protein level, the relative rate of synthesis of the pool of alpha 1-, alpha 2-, and alpha 3-subunits increased approximately twofold over 15 days of culture, leading to a marked increase in the immunochemical pool of alpha-subunits as measured by a panspecific polyclonal antibody. Concomitantly, Na(+)-K(+)-ATPase enzyme-specific activity increased three- (lower forebrain) to sixfold (upper forebrain). The transcripts of all three alpha-isoforms and beta-subunit were detected in vitro in similar proportion to the level observed in vivo. alpha 3-mRNA (3.7 kb) was more abundant than alpha 1 (3.7 kb) or alpha 2 (5.3 and 3.4 kb). Cytosine arabinoside (0.4 microM) and cholera toxin (0.1 microM) were used to selectively eliminate glial cells or neurons, respectively. It was found that alpha 2-mRNA is predominantly transcribed in glial cell cultures, whereas alpha 3- and beta 1-mRNA (2.7, 2.3, and 1.8 kb) are predominant in neuronal cultures.

scholarly journals Mutations inmmpLand in the Cell Wall Stress Stimulon Contribute to Resistance to Oxadiazole Antibiotics in Methicillin-Resistant Staphylococcus aureus

2014 ◽  
Vol 58 (10) ◽  
pp. 5841-5847 ◽  
Author(s):  
Qiaobin Xiao ◽  
Sergei Vakulenko ◽  
Mayland Chang ◽  
Shahriar Mobashery
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Wall Stress ◽  
Methicillin Resistant ◽  
Content Type ◽  
Cell Wall Stress ◽  
A Cell ◽  
Putative Member

ABSTRACTStaphylococcus aureusis a leading cause of hospital- and community-acquired infections, which exhibit broad resistance to various antibiotics. We recently disclosed the discovery of the oxadiazole class of antibiotics, which hasin vitroandin vivoactivities against methicillin-resistantS. aureus(MRSA). We report herein that MmpL, a putative member of the resistance, nodulation, and cell division (RND) family of proteins, contributes to oxadiazole resistance in theS. aureusstrain COL. Through serial passages, we generated twoS. aureusCOL variants that showed diminished susceptibilities to an oxadiazole antibiotic. The MICs for the oxadiazole against one strain (designatedS. aureusCOLI) increased reproducibly 2-fold (to 4 μg/ml), while against the other strain (S. aureusCOLR), they increased >4-fold (to >8 μg/ml, the limit of solubility). The COLRstrain was derived from the COLIstrain. Whole-genome sequencing revealed 31 mutations inS. aureusCOLR, of which 29 were shared with COLI. Consistent with our previous finding that oxadiazole antibiotics inhibit cell wall biosynthesis, we found 13 mutations that occurred either in structural genes or in promoters of the genes of the cell wall stress stimulon. Two unique mutations inS. aureusCOLRwere substitutions in two genes that encode the putative thioredoxin (SACOL1794) and MmpL (SACOL2566). A role formmpLin resistance to oxadiazoles was discerned from gene deletion and complementation experiments. To our knowledge, this is the first report that a cell wall-acting antibiotic selects for mutations in the cell wall stress stimulon and the first to implicate MmpL in resistance to antibiotics inS. aureus.

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