scholarly journals Cardiolipin Alters Rhodobacter sphaeroides Cell Shape by Affecting Peptidoglycan Precursor Biosynthesis

mBio ◽  
2019 ◽  
Vol 10 (1) ◽  
Author(s):  
Ti-Yu Lin ◽  
William S. Gross ◽  
George K. Auer ◽  
Douglas B. Weibel
Keyword(s):  
Cell Wall ◽  
Rhodobacter Sphaeroides ◽  
Cell Morphology ◽  
Cell Shape ◽  
Molecular Mechanisms ◽  
Wild Type ◽  
Anionic Phospholipid ◽  
Content Type ◽  
Lipid Ii ◽  
Topological Features

ABSTRACT Cardiolipin (CL) is an anionic phospholipid that plays an important role in regulating protein biochemistry in bacteria and mitochondria. Deleting the CL synthase gene (Δcls) in Rhodobacter sphaeroides depletes CL and decreases cell length by 20%. Using a chemical biology approach, we found that a CL deficiency does not impair the function of the cell wall elongasome in R. sphaeroides; instead, biosynthesis of the peptidoglycan (PG) precursor lipid II is decreased. Treating R. sphaeroides cells with fosfomycin and d-cycloserine inhibits lipid II biosynthesis and creates phenotypes in cell shape, PG composition, and spatial PG assembly that are strikingly similar to those seen with R. sphaeroides Δcls cells, suggesting that CL deficiency alters the elongation of R. sphaeroides cells by reducing lipid II biosynthesis. We found that MurG—a glycosyltransferase that performs the last step of lipid II biosynthesis—interacts with anionic phospholipids in native (i.e., R. sphaeroides) and artificial membranes. Lipid II production decreases 25% in R. sphaeroides Δcls cells compared to wild-type cells, and overexpression of MurG in R. sphaeroides Δcls cells restores their rod shape, indicating that CL deficiency decreases MurG activity and alters cell shape. The R. sphaeroides Δcls mutant is more sensitive than the wild-type strain to antibiotics targeting PG synthesis, including fosfomycin, d-cycloserine, S-(3,4-dichlorobenzyl)isothiourea (A22), mecillinam, and ampicillin, suggesting that CL biosynthesis may be a potential target for combination chemotherapies that block the bacterial cell wall. IMPORTANCE The phospholipid composition of the cell membrane influences the spatial and temporal biochemistry of cells. We studied molecular mechanisms connecting membrane composition to cell morphology in the model bacterium Rhodobacter sphaeroides. The peptidoglycan (PG) layer of the cell wall is a dominant component of cell mechanical properties; consequently, it has been an important antibiotic target. We found that the anionic phospholipid cardiolipin (CL) plays a role in determination of the shape of R. sphaeroides cells by affecting PG precursor biosynthesis. Removing CL in R. sphaeroides alters cell morphology and increases its sensitivity to antibiotics targeting proteins synthesizing PG. These studies provide a connection to spatial biochemical control in mitochondria, which contain an inner membrane with topological features in common with R. sphaeroides.

scholarly journals Substrate Inhibition of VanA by d-Alanine Reduces Vancomycin Resistance in a VanX-Dependent Manner

2016 ◽  
Vol 60 (8) ◽  
pp. 4930-4939 ◽  
Author(s):  
Lizah T. van der Aart ◽  
Nicole Lemmens ◽  
Willem J. van Wamel ◽  
Gilles P. van Wezel
Keyword(s):  
Cell Wall ◽  
Streptomyces Coelicolor ◽  
Substrate Inhibition ◽  
Model Organism ◽  
Vancomycin Resistance ◽  
Dependent Manner ◽  
Wild Type ◽  
Content Type ◽  
Lipid Ii ◽  
Vancomycin Resistant

ABSTRACTThe increasing resistance of clinical pathogens against the glycopeptide antibiotic vancomycin, a last-resort drug against infections with Gram-positive pathogens, is a major problem in the nosocomial environment. Vancomycin inhibits peptidoglycan synthesis by binding to thed-Ala–d-Ala terminal dipeptide moiety of the cell wall precursor lipid II. Plasmid-transferable resistance is conferred by modification of the terminal dipeptide into the vancomycin-insensitive variantd-Ala–d-Lac, which is produced by VanA. Here we show that exogenousd-Ala competes withd-Lac as a substrate for VanA, increasing the ratio of wild-type to mutant dipeptide, an effect that was augmented by several orders of magnitude in the absence of thed-Ala–d-Ala peptidase VanX. Liquid chromatography-mass spectrometry (LC-MS) analysis showed that high concentrations ofd-Ala led to the production of a significant amount of wild-type cell wall precursors, whilevanX-null mutants produced primarily wild-type precursors. This enhanced the efficacy of vancomycin in the vancomycin-resistant model organismStreptomyces coelicolor, and the susceptibility of vancomycin-resistant clinical isolates ofEnterococcus faecium(VRE) increased by up to 100-fold. The enhanced vancomycin sensitivity ofS. coelicolorcells correlated directly to increased binding of the antibiotic to the cell wall. Our work offers new perspectives for the treatment of diseases associated with vancomycin-resistant pathogens and for the development of drugs that target vancomycin resistance.

scholarly journals VIBRIO CHOLERAE ADAPTS TO SESSILE AND MOTILE LIFESTYLES BY CYCLIC DI-GMP REGULATION OF CELL SHAPE

2020 ◽  
Author(s):  
Nicolas L. Fernandez ◽  
Nguyen T. Q. Nhu ◽  
Brian Y. Hsueh ◽  
Joshua L. Franklin ◽  
Yann S. Dufour ◽  
...  
Keyword(s):  
Cell Wall ◽  
Cell Morphology ◽  
Cell Shape ◽  
Guanosine Monophosphate ◽  
Flagellar Motility ◽  
Wild Type ◽  
Gene Encoding ◽  
Regulatory Pathways ◽  
Matrix Components ◽  
Biofilm Matrix

AbstractThe cell morphology of rod-shaped bacteria is determined by the rigid net of peptidoglycan forming the cell wall. While V. cholerae grows into a curved shape under most conditions, straight rods have been observed. However, the signals and regulatory pathways controlling cell shape transitions in V. cholerae and the benefits of switching between rod and curved shape have not been determined. We demonstrate that cell shape in V. cholerae is regulated by the bacterial second messenger cyclic dimeric guanosine monophosphate (c-di-GMP) by repressing expression of crvA, a gene encoding an intermediate filament-like protein necessary for curvature formation in V. cholerae. This regulation is mediated by the transcriptional cascade that also induces production of biofilm matrix components, indicating that cell shape is coregulated with V. cholerae’s induction of sessility. Wild-type V. cholerae cells adhering to a surface lose their characteristic curved shape to become as straight as cells lacking crvA while genetically engineering cells to maintain high curvature reduced microcolony formation and biofilm density. Conversely, straight V. cholerae mutants have reduced speed when swimming using flagellar motility in liquid. Our results demonstrate regulation of cell shape in bacteria is a mechanism to increase fitness in planktonic or biofilm lifestyles.

scholarly journals A Genome-WideHelicobacter pyloriMorphology Screen Uncovers a Membrane-Spanning Helical Cell Shape Complex

2019 ◽  
Vol 201 (14) ◽  
Author(s):  
Desirée C. Yang ◽  
Kris M. Blair ◽  
Jennifer A. Taylor ◽  
Timothy W. Petersen ◽  
Tate Sessler ◽  
...  
Keyword(s):  
Cell Wall ◽  
Helicobacter Pylori ◽  
Light Scattering ◽  
Protein Interactions ◽  
Cell Morphology ◽  
Cell Shape ◽  
Cross Linking ◽  
Protein Protein Interactions ◽  
Content Type ◽  
H Pylori

ABSTRACTEvident in its name, the gastric pathogenHelicobacter pylorihas a helical cell morphology which facilitates efficient colonization of the human stomach. An improved light-focusing strategy allowed us to robustly distinguish even subtle perturbations ofH. pyloricell morphology by deviations in light-scattering properties measured by flow cytometry. Profiling of an arrayed genome-wide deletion library identified 28 genes that influence different aspects of cell shape, including properties of the helix, cell length or width, cell filament formation, cell shape heterogeneity, and cell branching. Included in this mutant collection were two that failed to form any helical cells, a soluble lytic transglycosylase and a previously uncharacterized putative multipass inner membrane protein HPG27_0728, renamed Csd7. A combination of cell fractionation, mutational, and immunoprecipitation experiments show that Csd7 and Csd2 collaborate to stabilize the Csd1 peptidoglycan (PG) endopeptidase. Thus, bothcsd2andcsd7mutants show the same enhancement of PG tetra-pentapeptide cross-linking ascsd1mutants. Csd7 also links Csd1 with the bactofilin CcmA via protein-protein interactions. Although Csd1 is stable inccmAmutants, these mutants show altered PG tetra-pentapeptide cross-linking, suggesting that Csd7 may directly or indirectly activate as well as stabilize Csd1. These data begin to illuminate a highly orchestrated program to regulate PG modifications that promote helical shape, which includes nine nonessential nonredundant genes required for helical shape and 26 additional genes that further modifyH. pylori’s cell morphology.IMPORTANCEThe stomach ulcer and cancer-causing pathogenHelicobacter pylorihas a helical cell shape which facilitates stomach infection. Using light scattering to measure perturbations of cell morphology, we identified 28 genes that influence different aspects of cell shape. A mutant in a previously uncharacterized protein renamed Csd7 failed to form any helical cells. Biochemical analyses showed that Csd7 collaborates with other proteins to stabilize the cell wall-degrading enzyme Csd1. Csd7 also links Csd1 with a putative filament-forming protein via protein-protein interactions. These data suggest that helical cell shape arises from a highly orchestrated program to regulate cell wall modifications. Targeting of this helical cell shape-promoting program could offer new ways to block infectivity of this important human pathogen.

scholarly journals A Cardiolipin-Deficient Mutant of Rhodobacter sphaeroides Has an Altered Cell Shape and Is Impaired in Biofilm Formation

2015 ◽  
Vol 197 (21) ◽  
pp. 3446-3455 ◽  
Author(s):  
Ti-Yu Lin ◽  
Thiago M. A. Santos ◽  
Wayne S. Kontur ◽  
Timothy J. Donohue ◽  
Douglas B. Weibel
Keyword(s):  
Rhodobacter Sphaeroides ◽  
Biofilm Formation ◽  
Cell Shape ◽  
Bacterial Attachment ◽  
Growth Defect ◽  
Deficient Mutant ◽  
Membrane Composition ◽  
Anionic Phospholipid ◽  
Content Type ◽  
Altered Cell

ABSTRACTCell shape has been suggested to play an important role in the regulation of bacterial attachment to surfaces and the formation of communities associated with surfaces. We found that a cardiolipin synthase (Δcls) mutant of the rod-shaped bacteriumRhodobacter sphaeroides—in which synthesis of the anionic, highly curved phospholipid cardiolipin (CL) is reduced by 90%—produces ellipsoid-shaped cells that are impaired in biofilm formation. Reducing the concentration of CL did not cause significant defects inR. sphaeroidescell growth, swimming motility, lipopolysaccharide and exopolysaccharide production, surface adhesion protein expression, and membrane permeability. Complementation of the CL-deficient mutant by ectopically expressing CL synthase restored cells to their rod shape and increased biofilm formation. TreatingR. sphaeroidescells with a low concentration (10 μg/ml) of the small-molecule MreB inhibitorS-(3,4-dichlorobenzyl)isothiourea produced ellipsoid-shaped cells that had no obvious growth defect yet reducedR. sphaeroidesbiofilm formation. This study demonstrates that CL plays a role inR. sphaeroidescell shape determination, biofilm formation, and the ability of the bacterium to adapt to its environment.IMPORTANCEMembrane composition plays a fundamental role in the adaptation of many bacteria to environmental stress. In this study, we build a new connection between the anionic phospholipid cardiolipin (CL) and cellular adaptation inRhodobacter sphaeroides. We demonstrate that CL plays a role in the regulation ofR. sphaeroidesmorphology and is important for the ability of this bacterium to form biofilms. This study correlates CL concentration, cell shape, and biofilm formation and provides the first example of how membrane composition in bacteria alters cell morphology and influences adaptation. This study also provides insight into the potential of phospholipid biosynthesis as a target for new chemical strategies designed to alter or prevent biofilm formation.

scholarly journals Transcriptome and metabolome profiling provide insights into molecular mechanism of pseudostem elongation in banana

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Guiming Deng ◽  
Fangcheng Bi ◽  
Jing Liu ◽  
Weidi He ◽  
Chunyu Li ◽  
...  
Keyword(s):  
Cell Wall ◽  
Cell Elongation ◽  
Molecular Mechanisms ◽  
Specific Gene ◽  
Wild Type ◽  
Banana Plant ◽  
Cell Wall Modification ◽  
Dwarf Cultivar ◽  
Important Trait ◽  
Metabolome Profiling

AbstractBackgroundBanana plant height is an important trait for horticultural practices and semi-dwarf cultivars show better resistance to damages by wind and rain. However, the molecular mechanisms controlling the pseudostem height remain poorly understood. Herein, we studied the molecular changes in the pseudostem of a semi-dwarf banana mutant Aifen No. 1 (Musaspp. Pisang Awak sub-group ABB) as compared to its wild-type dwarf cultivar using a combined transcriptome and metabolome approach.ResultsA total of 127 differentially expressed genes and 48 differentially accumulated metabolites were detected between the mutant and its wild type. Metabolites belonging to amino acid and its derivatives, flavonoids, lignans, coumarins, organic acids, and phenolic acids were up-regulated in the mutant. The transcriptome analysis showed the differential regulation of genes related to the gibberellin pathway, auxin transport, cell elongation, and cell wall modification. Based on the regulation of gibberellin and associated pathway-related genes, we discussed the involvement of gibberellins in pseudostem elongation in the mutant banana. Genes and metabolites associated with cell wall were explored and their involvement in cell extension is discussed.ConclusionsThe results suggest that gibberellins and associated pathways are possibly developing the observed semi-dwarf pseudostem phenotype together with cell elongation and cell wall modification. The findings increase the understanding of the mechanisms underlying banana stem height and provide new clues for further dissection of specific gene functions.

scholarly journals Neisseria gonorrhoeaePBP3 and PBP4 Facilitate NOD1 Agonist Peptidoglycan Fragment Release and Survival in Stationary Phase

2018 ◽  
Vol 87 (2) ◽  
Author(s):  
Ryan E. Schaub ◽  
Krizia M. Perez-Medina ◽  
Kathleen T. Hackett ◽  
Daniel L. Garcia ◽  
Joseph P. Dillard
Keyword(s):  
Molecular Weight ◽  
Cell Wall ◽  
Inflammatory Response ◽  
Neisseria Gonorrhoeae ◽  
Wild Type ◽  
Cross Link ◽  
Content Type ◽  
Lytic Transglycosylases ◽  
Penicillin Binding Proteins ◽  
Proinflammatory Molecules

ABSTRACTNeisseria gonorrhoeaereleases peptidoglycan fragments during growth, and these molecules induce an inflammatory response in the human host. The proinflammatory molecules include peptidoglycan monomers, peptidoglycan dimers, and free peptides. These molecules can be released by the actions of lytic transglycosylases or an amidase. However, >40% of the gonococcal cell wall is cross-linked, where the peptide stem on one peptidoglycan strand is linked to the peptide stem on a neighboring strand, suggesting that endopeptidases may be required for the release of many peptidoglycan fragments. Therefore, we characterized mutants with individual or combined mutations in genes for the low-molecular-mass penicillin-binding proteins PBP3 and PBP4. Mutations in eitherdacB, encoding PBP3, orpbpG, encoding PBP4, did not significantly reduce the release of peptidoglycan monomers or free peptides. A mutation indacBcaused the appearance of a larger-sized peptidoglycan monomer, the pentapeptide monomer, and an increased release of peptidoglycan dimers, suggesting the involvement of this enzyme in both the removal of C-terminald-Ala residues from stem peptides and the cleavage of cross-linked peptidoglycan. Mutation of bothdacBandpbpGeliminated the release of tripeptide-containing peptidoglycan fragments concomitantly with the appearance of pentapeptide and dipeptide peptidoglycan fragments and higher-molecular-weight peptidoglycan dimers. In accord with the loss of tripeptide peptidoglycan fragments, the level of human NOD1 activation by thedacB pbpGmutants was significantly lower than that by the wild type. We conclude that PBP3 and PBP4 overlap in function for cross-link cleavage and that these endopeptidases act in the normal release of peptidoglycan fragments during growth.

scholarly journals The Ser/Thr Kinase PrkC Participates in Cell Wall Homeostasis and Antimicrobial Resistance inClostridium difficile

2019 ◽  
Vol 87 (8) ◽  
Author(s):  
Elodie Cuenot ◽  
Transito Garcia-Garcia ◽  
Thibaut Douche ◽  
Olivier Gorgette ◽  
Pascal Courtin ◽  
...  
Keyword(s):  
Cell Wall ◽  
Type Strain ◽  
Wild Type Strain ◽  
Cell Envelope ◽  
Wild Type ◽  
Hamster Model ◽  
Content Type ◽  
Physiological Processes ◽  
Signal Transduction Networks ◽  
Mean Size

ABSTRACTClostridium difficileis the leading cause of antibiotic-associated diarrhea in adults. During infection,C. difficilemust detect the host environment and induce an appropriate survival strategy. Signal transduction networks involving serine/threonine kinases (STKs) play key roles in adaptation, as they regulate numerous physiological processes. PrkC ofC. difficileis an STK with two PASTA domains. We showed that PrkC is membrane associated and is found at the septum. We observed that deletion ofprkCaffects cell morphology with an increase in mean size, cell length heterogeneity, and presence of abnormal septa. A ΔprkCmutant was able to sporulate and germinate but was less motile and formed more biofilm than the wild-type strain. Moreover, a ΔprkCmutant was more sensitive to antimicrobial compounds that target the cell envelope, such as the secondary bile salt deoxycholate, cephalosporins, cationic antimicrobial peptides, and lysozyme. This increased susceptibility was not associated with differences in peptidoglycan or polysaccharide II composition. However, the ΔprkCmutant had less peptidoglycan and released more polysaccharide II into the supernatant. A proteomic analysis showed that the majority ofC. difficileproteins associated with the cell wall were less abundant in the ΔprkCmutant than the wild-type strain. Finally, in a hamster model of infection, the ΔprkCmutant had a colonization delay that did not significantly affect overall virulence.

scholarly journals Vibrio choleraeadapts to sessile and motile lifestyles by cyclic di-GMP regulation of cell shape

2020 ◽  
Vol 117 (46) ◽  
pp. 29046-29054 ◽  
Author(s):  
Nicolas L. Fernandez ◽  
Brian Y. Hsueh ◽  
Nguyen T. Q. Nhu ◽  
Joshua L. Franklin ◽  
Yann S. Dufour ◽  
...  
Keyword(s):  
Cell Wall ◽  
Cell Shape ◽  
Guanosine Monophosphate ◽  
Human Pathogen ◽  
Flagellar Motility ◽  
Wild Type ◽  
Gene Encoding ◽  
Regulatory Pathways ◽  
Matrix Components ◽  
Curved Rod

The cell morphology of rod-shaped bacteria is determined by the rigid net of peptidoglycan forming the cell wall. Alterations to the rod shape, such as the curved rod, occur through manipulating the process of cell wall synthesis. The human pathogenVibrio choleraetypically exists as a curved rod, but straight rods have been observed under certain conditions. While this appears to be a regulated process, the regulatory pathways controlling cell shape transitions inV. choleraeand the benefits of switching between rod and curved shape have not been determined. We demonstrate that cell shape inV. choleraeis regulated by the bacterial second messenger cyclic dimeric guanosine monophosphate (c-di-GMP) by posttranscriptionally repressing expression ofcrvA, a gene encoding an intermediate filament-like protein necessary for curvature formation inV. cholerae.This regulation is mediated by the transcriptional cascade that also induces production of biofilm matrix components, indicating that cell shape is coregulated withV. cholerae’s induction of sessility. During microcolony formation, wild-typeV. choleraecells tended to exist as straight rods, while genetically engineering cells to maintain high curvature reduced microcolony formation and biofilm density. Conversely, straightV. choleraemutants have reduced swimming speed when using flagellar motility in liquid. Our results demonstrate regulation of cell shape in bacteria is a mechanism to increase fitness in planktonic and biofilm lifestyles.

scholarly journals AVibrio choleraeBolA-Like Protein Is Required for Proper Cell Shape and Cell Envelope Integrity

mBio ◽  
2019 ◽  
Vol 10 (4) ◽  
Author(s):  
Aurore Fleurie ◽  
Abdelrahim Zoued ◽  
Laura Alvarez ◽  
Kelly M. Hines ◽  
Felipe Cava ◽  
...  
Keyword(s):  
Vibrio Cholerae ◽  
Cell Morphology ◽  
Cell Shape ◽  
Cell Envelope ◽  
Intestinal Colonization ◽  
Iron Sulfur Cluster ◽  
Content Type ◽  
Sulfur Cluster ◽  
Iron Sulfur ◽  
Morphological Defects

ABSTRACTBolA family proteins are conserved in Gram-negative bacteria and many eukaryotes. While diverse cellular phenotypes have been linked to this protein family, the molecular pathways through which these proteins mediate their effects are not well described. Here, we investigated the roles of BolA family proteins inVibrio cholerae, the cholera pathogen. LikeEscherichia coli,V. choleraeencodes two BolA proteins, BolA and IbaG. However, in marked contrast toE. coli, wherebolAis linked to cell shape andibaGis not, inV. cholerae,bolAmutants lack morphological defects, whereasibaGproved critical for the generation and/or maintenance of the pathogen’s morphology. Notably, the bizarre-shaped, multipolar, elongated, and wide cells that predominated in exponential-phase ΔibaGV. choleraecultures were not observed in stationary-phase cultures. TheV. choleraeΔibaGmutant exhibited increased sensitivity to cell envelope stressors, including cell wall-acting antibiotics and bile, and was defective in intestinal colonization. ΔibaGV. choleraehad reduced peptidoglycan and lipid II and altered outer membrane lipids, likely contributing to the mutant’s morphological defects and sensitivity to envelope stressors. Transposon insertion sequencing analysis ofibaG’s genetic interactions suggested thatibaGis involved in several processes involved in the generation and homeostasis of the cell envelope. Furthermore, copurification studies revealed that IbaG interacts with proteins containing iron-sulfur clusters or involved in their assembly. Collectively, our findings suggest thatV. choleraeIbaG controls cell morphology and cell envelope integrity through its role in biogenesis or trafficking of iron-sulfur cluster proteins.IMPORTANCEBolA-like proteins are conserved across prokaryotes and eukaryotes. These proteins have been linked to a variety of phenotypes, but the pathways and mechanisms through which they act have not been extensively characterized. Here, we unraveled the role of the BolA-like protein IbaG in the cholera pathogenVibrio cholerae. The absence of IbaG was associated with dramatic changes in cell morphology, sensitivity to envelope stressors, and intestinal colonization defects. IbaG was found to be required for biogenesis of several components of theV. choleraecell envelope and to interact with numerous iron-sulfur cluster-containing proteins and factors involved in their assembly. Thus, our findings suggest that IbaG governsV. choleraecell shape and cell envelope homeostasis through its effects on iron-sulfur proteins and associated pathways. The diversity of processes involving iron-sulfur-containing proteins is likely a factor underlying the range of phenotypes associated with BolA family proteins.

scholarly journals Cross Talk between the Cell Wall Integrity and Cyclic AMP/Protein Kinase A Pathways in Cryptococcus neoformans

mBio ◽  
2014 ◽  
Vol 5 (4) ◽  
Author(s):  
Maureen J. Donlin ◽  
Rajendra Upadhya ◽  
Kimberly J. Gerik ◽  
Woei Lam ◽  
Laura G. VanArendonk ◽  
...  
Keyword(s):  
Cell Wall ◽  
Protein Kinase ◽  
Cyclic Amp ◽  
Protein Kinase A ◽  
Cryptococcus Neoformans ◽  
Signaling Pathway ◽  
Cell Wall Integrity ◽  
Wild Type ◽  
Content Type ◽  
Kinase A

ABSTRACTCryptococcus neoformans is a fungal pathogen of immunocompromised people that causes fatal meningitis. The fungal cell wall is essential to viability and pathogenesis ofC. neoformans, and biosynthesis and repair of the wall is primarily controlled by the cell wall integrity (CWI) signaling pathway. Previous work has shown that deletion of genes encoding the four major kinases in the CWI signaling pathway, namely,PKC1,BCK1,MKK2, andMPK1results in severe cell wall phenotypes, sensitivity to a variety of cell wall stressors, and for Mpk1, reduced virulence in a mouse model. Here, we examined the global transcriptional responses to gene deletions ofBCK1,MKK2, andMPK1compared to wild-type cells. We found that over 1,000 genes were differentially expressed in one or more of the deletion strains, with 115 genes differentially expressed in all three strains, many of which have been identified as genes regulated by the cyclic AMP (cAMP)/protein kinase A (PKA) pathway. Biochemical measurements of cAMP levels in the kinase deletion strains revealed significantly less cAMP in all of the deletion strains compared to the wild-type strain. The deletion strains also produced significantly smaller capsules than the wild-type KN99 strain did under capsule-inducing conditions, although the levels of capsule they shed were similar to those shed by the wild type. Finally, addition of exogenous cAMP led to reduced sensitivity to cell wall stress and restored surface capsule to levels near those of wild type. Thus, we have direct evidence of cross talk between the CWI and cAMP/PKA pathways that may have important implications for regulation of cell wall and capsule homeostasis.IMPORTANCECryptococcus neoformans is a fungal pathogen of immunocompromised people that causes fatal meningitis. The fungal cell wall is essential to viability and pathogenesis ofC. neoformans, and biosynthesis and repair of the wall are primarily controlled by the cell wall integrity (CWI) signaling pathway. In this study, we demonstrate that deletion of any of three core kinases in the CWI pathway impacts not only the cell wall but also the amount of surface capsule. Deletion of any of the kinases results in significantly reduced cellular cyclic AMP (cAMP) levels, and addition of exogenous cAMP rescues the capsule defect and some cell wall defects, supporting a direct role for the CWI pathway in regulation of capsule in conjunction with the cAMP/protein kinase A pathway.

Sign in / Sign up

Export Citation Format

Share Document