Effects of pretreatments with sulfhydryls and alkylating agents on spheroplast formation from Schwanniomyces species

1984 ◽  
Vol 30 (3) ◽  
pp. 368-374 ◽  
Author(s):  
T. M. Dowhanick ◽  
C. J. Panchal ◽  
G. G. Stewart
Keyword(s):  
Cell Wall ◽  
Stationary Phase ◽  
Growth Phase ◽  
Cell Walls ◽  
Alkylating Agents ◽  
Inhibitory Effects ◽  
Late Stationary Phase ◽  
Stages Of Growth ◽  
Degradative Enzymes ◽  
Sulfhydryl Compounds

Pretreatment of cells with β-mercaptoethanol, dithiothreitol, or cysteine increased the rate of spheroplast formation at all stages of growth in Schwanniomyces castellii and S. occidentalis, but had little effect on final yields of spheroplasts when compared with controls. Pretreatment with iodoacetate and cystine resulted in decreased rates of formation and lower yields of spheroplasts at all stages of growth for both species. The enhanced rates of spheroplast formation in the presence of the sulfhydryl compounds were more pronounced when the experimental cells were derived from early or late stationary phase cultures, whereas the inhibitory effects of cystine and iodoacetate were not associated with the growth phase. In agreement with extant data on other yeast genera, sulfhydryl compounds increased the susceptibility of the yeast cell wall to degradative enzymes, whilst alkylating agents decreased susceptibility of cell walls to these enzymes.

scholarly journals Requirement of Autolytic Activity for Bacteriocin-Induced Lysis

2000 ◽  
Vol 66 (8) ◽  
pp. 3174-3179 ◽  
Author(s):  
M. Carmen Martínez-Cuesta ◽  
Jan Kok ◽  
Elisabet Herranz ◽  
Carmen Peláez ◽  
Teresa Requena ◽  
...  
Keyword(s):  
Cell Wall ◽  
Sodium Dodecyl Sulfate ◽  
Growth Phase ◽  
Cell Walls ◽  
Cell Damage ◽  
Sodium Dodecyl ◽  
Exponential Growth Phase ◽  
Cell Wall Degradation ◽  
Intracellular Proteins ◽  
Autolytic Activity

ABSTRACT The bacteriocin produced by Lactococcus lactis IFPL105 is bactericidal against several Lactococcus andLactobacillus strains. Addition of the bacteriocin to exponential-growth-phase cells resulted in all cases in bacteriolysis. The bacteriolytic response of the strains was not related to differences in sensitivity to the bacteriocin and was strongly reduced in the presence of autolysin inhibitors (Co2+ and sodium dodecyl sulfate). When L. lactis MG1363 and its derivative deficient in the production of the major autolysin AcmA (MG1363acmAΔ1) were incubated with the bacteriocin, the latter did not lyse and no intracellular proteins were released into the medium. Incubation of cell wall fragments of L. lactisMG1363, or of L. lactis MG1363acmAΔ1 to which extracellular AcmA was added, in the presence or absence of the bacteriocin had no effect on the speed of cell wall degradation. This result indicates that the bacteriocin does not degrade cell walls, nor does it directly activate the autolysin AcmA. The autolysin was also responsible for the observed lysis of L. lactis MG1363 cells during incubation with nisin or the mixture of lactococcins A, B, and M. The results presented here show that lysis of L. lactis after addition of the bacteriocins is caused by the resulting cell damage, which promotes uncontrolled degradation of the cell walls by AcmA.

scholarly journals The Nature of Antibacterial Adaptive Immune Responses against Staphylococcus aureus Is Dependent on the Growth Phase and Extracellular Peptidoglycan

2019 ◽  
Vol 88 (1) ◽  
Author(s):  
Payal P. Balraadjsing ◽  
Lisbeth D. Lund ◽  
Yuri Souwer ◽  
Sebastian A. J. Zaat ◽  
Hanne Frøkiær ◽  
...  
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
T Cell ◽  
Stationary Phase ◽  
Growth Phase ◽  
Cell Response ◽  
Th17 Cell ◽  
Exponential Phase ◽  
Th1 Cell ◽  
Content Type

ABSTRACT Staphylococcus aureus has evolved different strategies to evade the immune response, which play an important role in its pathogenesis. The bacteria express and shed various cell wall components and toxins during different stages of growth that may affect the protective T cell responses to extracellular and intracellular S. aureus. However, if and how the dendritic cell (DC)-mediated T cell response against S. aureus changes during growth of the bacterium remain elusive. In this study, we show that exponential-phase (EP) S. aureus bacteria were endocytosed very efficiently by human DCs, and these DCs strongly promoted production of the T cell polarizing factor interleukin-12 (IL-12). In contrast, stationary-phase (SP) S. aureus bacteria were endocytosed less efficiently by DCs, and these DCs produced small amounts of IL-12. The high level of IL-12 production induced by EP S. aureus led to the development of a T helper 1 (Th1) cell response, which was inhibited after neutralization of IL-12. Furthermore, preincubation with the staphylococcal cell wall component peptidoglycan (PGN), characteristically shed during the exponential growth phase, modulated the DC response to EP S. aureus. PGN preincubation appeared to inhibit IL-12p35 expression, leading to downregulation of IL-12 and an increase of IL-23 production by DCs, enhancing Th17 cell development. Taken together, our data indicate that exponential-phase S. aureus bacteria induce a stronger IL-12-dependent Th1 cell response than stationary-phase S. aureus and that this Th1 cell response shifted toward a Th17 cell response in the presence of PGN.

scholarly journals Peptidoglycan Recycling in Gram-Positive Bacteria Is Crucial for Survival in Stationary Phase

mBio ◽  
2016 ◽  
Vol 7 (5) ◽  
Author(s):  
Marina Borisova ◽  
Rosmarie Gaupp ◽  
Amanda Duckworth ◽  
Alexander Schneider ◽  
Désirée Dalügge ◽  
...  
Keyword(s):  
Cell Wall ◽  
Stationary Phase ◽  
Exponential Growth ◽  
Model Organisms ◽  
Exponential Growth Phase ◽  
Rich Medium ◽  
Late Stationary Phase ◽  
Gram Positive ◽  
Term Survival ◽  
Gram Positive Bacteria

ABSTRACTPeptidoglycan recycling is a metabolic process by which Gram-negative bacteria reutilize up to half of their cell wall within one generation during vegetative growth. Whether peptidoglycan recycling also occurs in Gram-positive bacteria has so far remained unclear. We show here that three Gram-positive model organisms,Staphylococcus aureus,Bacillus subtilis, andStreptomyces coelicolor, all recycle the sugarN-acetylmuramic acid (MurNAc) of their peptidoglycan during growth in rich medium. They possess MurNAc-6-phosphate (MurNAc-6P) etherase (MurQ inE. coli) enzymes, which are responsible for the intracellular conversion of MurNAc-6P toN-acetylglucosamine-6-phosphate andd-lactate. By applying mass spectrometry, we observed accumulation of MurNAc-6P in MurNAc-6P etherase deletion mutants but not in either the isogenic parental strains or complemented strains, suggesting that MurQ orthologs are required for the recycling of cell wall-derived MurNAc in these bacteria. Quantification of MurNAc-6P in ΔmurQcells ofS. aureusandB. subtilisrevealed small amounts during exponential growth phase (0.19 nmol and 0.03 nmol, respectively, per ml of cells at an optical density at 600 nm [OD600] of 1) but large amounts during transition (0.56 nmol and 0.52 nmol) and stationary (0.53 nmol and 1.36 nmol) phases. The addition of MurNAc to ΔmurQcultures greatly increased the levels of intracellular MurNAc-6P in all growth phases. The ΔmurQmutants ofS. aureusandB. subtilisshowed no growth deficiency in rich medium compared to the growth of the respective parental strains, but intriguingly, they had a severe survival disadvantage in late stationary phase. Thus, although peptidoglycan recycling is apparently not essential for the growth of Gram-positive bacteria, it provides a benefit for long-term survival.IMPORTANCEThe peptidoglycan of the bacterial cell wall is turned over steadily during growth. As peptidoglycan fragments were found in large amounts in spent medium of exponentially growing Gram-positive bacteria, their ability to recycle these fragments has been questioned. We conclusively showed recycling of the peptidoglycan component MurNAc in different Gram-positive model organisms and revealed that a MurNAc-6P etherase (MurQ or MurQ ortholog) enzyme is required in this process. We further demonstrated that recycling occurs predominantly during the transition to stationary phase inS. aureusandB. subtilis, explaining why peptidoglycan fragments are found in the medium during exponential growth. We quantified the intracellular accumulation of recycling products in MurNAc-6P etherase gene mutants, revealing that about 5% and 10% of the MurNAc of the cell wall per generation is recycled inS. aureusandB. subtilis, respectively. Importantly, we showed that MurNAc recycling and salvaging does not sustain growth in these bacteria but is used to enhance survival during late stationary phase.

scholarly journals Global Role for ClpP-Containing Proteases in Stationary-Phase Adaptation of Escherichia coli

2003 ◽  
Vol 185 (1) ◽  
pp. 115-125 ◽  
Author(s):  
Dieter Weichart ◽  
Nadine Querfurth ◽  
Mathias Dreger ◽  
Regine Hengge-Aronis
Keyword(s):  
Escherichia Coli ◽  
Stationary Phase ◽  
Growth Phase ◽  
Proteome Analysis ◽  
Resting Cells ◽  
Wild Type ◽  
Late Stationary Phase ◽  
E Coli ◽  
In The Wild ◽  
Dps Protein

ABSTRACT To elucidate the involvement of proteolysis in the regulation of stationary-phase adaptation, the clpA, clpX, and clpP protease mutants of Escherichia coli were subjected to proteome analysis during growth and during carbon starvation. For most of the growth-phase-regulated proteins detected on our gels, the clpA, clpX, or clpP mutant failed to mount the growth-phase regulation found in the wild type. For example, in the clpP and clpA mutant cultures, the Dps protein, the WrbA protein, and the periplasmic lysine-arginine-ornithine binding protein ArgT did not display the induction typical for late-stationary-phase wild-type cells. On the other hand, in the protease mutants, a number of proteins accumulated to a higher degree than in the wild type, especially in late stationary phase. The proteins affected in this manner include the LeuA, TrxB, GdhA, GlnA, and MetK proteins and alkyl hydroperoxide reductase (AhpC). These proteins may be directly degraded by ClpAP or ClpXP, respectively, or their expression could be modulated by a protease-dependent mechanism. From our data we conclude that the levels of most major growth-phase-regulated proteins in E. coli are at some point controlled by the activity of at least one of the ClpP, ClpA, and ClpX proteins. Cultures of the strains lacking functional ClpP or ClpX also displayed a more rapid loss of viability during extended stationary phase than the wild type. Therefore, regulation by proteolysis seems to be more important, especially in resting cells, than previously suspected.

scholarly journals Reduced Lysis upon Growth of Lactococcus lactis on Galactose Is a Consequence of Decreased Binding of the Autolysin AcmA

2008 ◽  
Vol 74 (15) ◽  
pp. 4671-4679 ◽  
Author(s):  
Anton Steen ◽  
Girbe Buist ◽  
Naomi E. Kramer ◽  
Ruud Jalving ◽  
Germaine F. J. D. Benus ◽  
...  
Keyword(s):  
Cell Wall ◽  
Plasmodium Falciparum ◽  
Carbon Source ◽  
Fusion Protein ◽  
Stationary Phase ◽  
Lactococcus Lactis ◽  
Cell Walls ◽  
Trichloroacetic Acid ◽  
Binding Studies ◽  
Lipoteichoic Acids

ABSTRACT When Lactococcus lactis subsp. lactis IL1403 or L. lactis subsp. cremoris MG1363 is grown in a medium with galactose as the carbon source, the culture lyses to a lesser extent in stationary phase than when the bacteria are grown in a medium containing glucose. Expression of AcmA, the major autolysin of L. lactis, is not influenced by the carbon source. Binding studies with a fusion protein consisting of the MSA2 protein of Plasmodium falciparum and the C-terminal peptidoglycan-binding domain of AcmA revealed that cell walls of cells from both subspecies grown on galactose bind less AcmA than cell walls of cells grown on glucose. Cells grown on glucose or galactose and treated with trichloroacetic acid prior to AcmA binding bind similar amounts of AcmA. Analysis of the composition of the lipoteichoic acids (LTAs) of L. lactis IL1403 cells grown on glucose or galactose showed that the LTA composition is influenced by the carbon source: cells grown on galactose contain LTA with less galactose than cells grown on glucose. In conclusion, growth of L. lactis on galactose changes the LTA composition in the cell wall in such a way that less AcmA is able to bind to the peptidoglycan, resulting in a decrease in autolysis.

scholarly journals Sed1p Is a Major Cell Wall Protein ofSaccharomyces cerevisiae in the Stationary Phase and Is Involved in Lytic Enzyme Resistance

1998 ◽  
Vol 180 (13) ◽  
pp. 3381-3387 ◽  
Author(s):  
Hitoshi Shimoi ◽  
Hiroshi Kitagaki ◽  
Hisanobu Ohmori ◽  
Yuzuru Iimura ◽  
Kiyoshi Ito
Keyword(s):  
Cell Wall ◽  
Stationary Phase ◽  
Cell Walls ◽  
Secretory Pathway ◽  
Signal Sequence ◽  
Sodium Dodecyl ◽  
Cell Wall Protein ◽  
Lytic Enzyme ◽  
Cell Wall Proteins ◽  
Structural Cell

ABSTRACT A 260-kDa structural cell wall protein was purified from sodium dodecyl sulfate-treated cell walls of Saccharomyces cerevisiae by incubation with Rarobacter faecitabidusprotease I, which is a yeast-lytic enzyme. Amino acid sequence analysis revealed that this protein is the product of the SED1 gene.SED1 was formerly identified as a multicopy suppressor oferd2, which encodes a protein involved in retrieval of luminal endoplasmic reticulum proteins from the secretory pathway. Sed1p is very rich in threonine and serine and, like other structural cell wall proteins, contains a putative signal sequence for the addition of a glycosylphosphatidylinositol anchor. However, the fact that Sed1p, unlike other cell wall proteins, has six cysteines and seven putative N-glycosylation sites suggests that Sed1p belongs to a new family of cell wall proteins. Epitope-tagged Sed1p was detected in a β-1,3-glucanase extract of cell walls by immunoblot analysis, suggesting that Sed1p is a glucanase-extractable cell wall protein. The expression of Sed1p mRNA increased in the stationary phase and was accompanied by an increase in the Sed1p content of cell walls. Disruption of SED1 had no effect on exponentially growing cells but made stationary-phase cells sensitive to Zymolyase. These results indicate that Sed1p is a major structural cell wall protein in stationary-phase cells and is required for lytic enzyme resistance.

scholarly journals The effects of short and long shading, applied during different stages of growth, on the development, productivity and quality of forage maize (Zea mays L.).

1983 ◽  
Vol 31 (2) ◽  
pp. 101-124
Author(s):  
P.C. Struik
Keyword(s):  
Cell Wall ◽  
Cell Walls ◽  
Field Experiments ◽  
Zea Mays L ◽  
Vegetative Development ◽  
Forage Maize ◽  
Stages Of Growth ◽  
Yield And Quality ◽  
Stover Yield

In 2 field experiments, shading which differed in duration and date of initiation was applied to normal stands of forage maize cv. LG11. Short shading during vegetative development affected leaf area, plant ht., stem thickness and reproductive development but final effects on DM yield and quality were small. Short shading during silking drastically reduced ear size and final ear yield. Although the deleterious effect on ear yield was partly compensated for by higher stover yield, productivity was low after the shading tents were removed. Digestibility was also greatly reduced because production of total DM was affected more than production of partly indigestible cell walls. Short shading soon after silking curtailed cell-wall formation more than DM production and crop digestibility was not adversely affected. Reduction in DM production remained large especially in the ear because of grain abortion. Shading after grain set stimulated the depletion of short carbohydrates in the stover and slowed down the decrease in the cell wall content of the whole crop. Crops shaded for long periods yielded more than expected on the basis of the short treatments. Long shading treatments lasted until final sampling and the earlier a long treatment was initiated, the greater the reduction in yield. The same was true for whole-crop digestibility, except in the earliest shading treatment in which poor vegetative development accompanied poor ear development. (Abstract retrieved from CAB Abstracts by CABI’s permission)

scholarly journals Bactericidal/Permeability-Increasing Protein Inhibits Growth of a Strain of Acholeplasma laidlawii and L Forms of the Gram-Positive Bacteria Staphylococcus aureus andStreptococcus pyogenes

1999 ◽  
Vol 43 (9) ◽  
pp. 2314-2316 ◽  
Author(s):  
Arnold H. Horwitz ◽  
Robert E. Williams ◽  
Pei-Syan Liu ◽  
Rossana Nadell
Keyword(s):  
Staphylococcus Aureus ◽  
Cell Wall ◽  
Streptococcus Pyogenes ◽  
Cell Walls ◽  
Intact Cell ◽  
Inhibitory Effects ◽  
Gram Positive ◽  
Gram Positive Bacteria ◽  
Acholeplasma Laidlawii ◽  
Growth Inhibitory

ABSTRACT Bactericidal/permeability-increasing protein (BPI) inhibited growth of cell wall-deficient Acholeplasma laidlawii and L forms of certain strains of Staphylococcus aureus andStreptococcus pyogenes. However, the same strains ofS. aureus and S. pyogenes with intact cell walls were not susceptible to the growth-inhibitory effects of BPI.

scholarly journals Muropeptide Rescue in Bacillus subtilis Involves Sequential Hydrolysis by β-N-Acetylglucosaminidase and N-Acetylmuramyl-l-Alanine Amidase

2010 ◽  
Vol 192 (12) ◽  
pp. 3132-3143 ◽  
Author(s):  
Silke Litzinger ◽  
Amanda Duckworth ◽  
Katja Nitzsche ◽  
Christian Risinger ◽  
Valentin Wittmann ◽  
...  
Keyword(s):  
Cell Wall ◽  
Bacillus Subtilis ◽  
Stationary Phase ◽  
Particulate Material ◽  
Late Stationary Phase ◽  
Phosphotransferase System ◽  
E Coli ◽  
A Cell ◽  
Recycling Pathway ◽  
Hydrolysis Of

ABSTRACT We identified a pathway in Bacillus subtilis that is used for recovery of N-acetylglucosamine (GlcNAc)-N-acetylmuramic acid (MurNAc) peptides (muropeptides) derived from the peptidoglycan of the cell wall. This pathway is encoded by a cluster of six genes, the first three of which are orthologs of Escherichia coli genes involved in N-acetylmuramic acid dissimilation and encode a MurNAc-6-phosphate etherase (MurQ), a MurNAc-6-phosphate-specific transcriptional regulator (MurR), and a MurNAc-specific phosphotransferase system (MurP). Here we characterized two other genes of this cluster. The first gene was shown to encode a cell wall-associated β-N-acetylglucosaminidase (NagZ, formerly YbbD) that cleaves the terminal nonreducing N-acetylglucosamine of muropeptides and also accepts chromogenic or fluorogenic β-N-acetylglucosaminides. The second gene was shown to encode an amidase (AmiE, formerly YbbE) that hydrolyzes the N-acetylmuramyl-l-Ala bond of MurNAc peptides but not this bond of muropeptides. Hence, AmiE requires NagZ, and in conjunction these enzymes liberate MurNAc by sequential hydrolysis of muropeptides. NagZ expression was induced at late exponential phase, and it was 6-fold higher in stationary phase. NagZ is noncovalently associated with lysozyme-degradable particulate material and can be released from it with salt. A nagZ mutant accumulates muropeptides in the spent medium and displays a lytic phenotype in late stationary phase. The evidence for a muropeptide catabolic pathway presented here is the first evidence for cell wall recovery in a Gram-positive organism, and this pathway is distinct from the cell wall recycling pathway of E. coli and other Gram-negative bacteria.

Behavior of mitochondria during sporulation in yeast

1978 ◽  
Vol 36 (2) ◽  
pp. 406-407
Author(s):  
Barbara J. Stevens
Keyword(s):  
Cell Wall ◽  
Stationary Phase ◽  
Growth Phase ◽  
Serial Section ◽  
Logarithmic Phase ◽  
Serial Sections ◽  
Vegetative Cells ◽  
Thin Sections ◽  
Section Technique ◽  
Single Mitochondrion

The number and the form of mitochondria in vegetatively growing yeast, Saccharo-myces cerevisiae, have previously been determined by the use of serial thin sections of entire cells. It was found that the number of mitochondria is directly related to the growth phase: logarithmic phase cells contain few mitochondria, less than 10, while stationary phase cells have numerous organelles, from 30 to 50 individual mitochondria. A single mitochondrion was found in 2 log phase, glucose-repressed cells, but all other cells had more than one mitochondrion. In the present study, the behavior of the chondriome during meiosis and sporulation in yeast was followed by the serial section technique and was compared to that in vegetative cells.Diploid strains Z193, Z239-6B6B and s4l were obtained from R. and M. Esposito and S. Klapholz (Univ. of Chicago) and serial sections of strain 112 were kindly provided by D. Zickler (Univ. Paris-Sud). Cells were fixed in glutaraldehyde, treated with glusulase (Endo Lab.) or zymolyase (Kirin Brew. Co.) to remove the cell wall, and post-fixed in Dalton's chrome osmium.

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