scholarly journals The vesicle transport protein Vac1p is required for virulence of Candida albicans

Microbiology ◽  
2006 ◽  
Vol 152 (10) ◽  
pp. 3111-3121 ◽  
Author(s):  
Kathrin Franke ◽  
Monika Nguyen ◽  
Albert Härtl ◽  
Hans-Martin Dahse ◽  
Georgia Vogl ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Mutant Strain ◽  
Protein Transport ◽  
Systemic Infection ◽  
Hyphal Growth ◽  
Transport Protein ◽  
Vesicle Transport ◽  
Antifungal Drugs ◽  
Null Mutant ◽  
Pathogenic Yeast

The putative vesicle transport protein Vac1p of the human pathogenic yeast Candida albicans plays an important role in virulence. To determine the cellular functions of Vac1p, a null mutant was generated by sequential disruption of both alleles. The vac1 null mutant strain showed defective endosomal vesicle transport, demonstrating a role of Vac1p in protein transport to the vacuole. Vac1p also contributes to resistance to metal ions, as the null mutant strain was hypersensitive to Cu2+, Zn2+ and Ni2+. In addition, the loss of Vac1p affected several virulence factors of C. albicans. In particular, the vac1 null mutant strain showed defective hyphal growth, even when hyphal formation was induced via different pathways. Furthermore, Vac1p affects chlamydospore formation, adherence to human vaginal epithelial cells, and the secretion of aspartyl proteinases (Saps). Avirulence in a mouse model of systemic infection of the vac1 null mutant strongly suggests that Vac1p of C. albicans is essential for pathogenicity. In summary, the Vac1p protein is required for several cellular pathways, in particular those that control virulence and pathogenicity. Consequently, Vac1p is a novel and interesting target for antifungal drugs.

Generation and functional in vivo characterization of a lipid kinase defective phosphatidylinositol 3-kinase Vps34p of Candida albicans

Microbiology ◽  
2005 ◽  
Vol 151 (1) ◽  
pp. 81-89 ◽  
Author(s):  
Juliane Günther ◽  
Monika Nguyen ◽  
Albert Härtl ◽  
Waldemar Künkel ◽  
Peter F. Zipfel ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Protein Transport ◽  
Kinase Activity ◽  
Hyphal Growth ◽  
Lipid Binding ◽  
Homologous Region ◽  
Null Mutant ◽  
Lipid Kinase ◽  
In Vivo Characterization

The phosphatidylinositol (PI) 3-kinase Vps34p of Candida albicans has lipid kinase and autophosphorylation activity and is involved in virulence and vesicular protein transport. In order to characterize the roles of lipid kinase activity, a chimeric Vps34 protein was created which lacks lipid kinase but retains autophosphorylation activity. To this end, six amino acids within the putative lipid-binding site of Vps34p were replaced by the homologous region of the PI 3-kinase-like C. albicans Tor protein. The resulting chimeric Vps34T protein was recombinantly expressed in Escherichia coli and shown to lack lipid kinase activity. The corresponding chimeric VPS34TOR gene was inserted into the genome of C. albicans, and this lipid-kinase-defective strain had a distinctive phenotype compared to those of the wild-type strain SC5314 and the vps34 null mutant. The lipid-kinase-defective strain was non-virulent, and showed altered hyphal growth, reduced adherence, as well as defective vacuole morphology and endosomal vesicle transport. These results demonstrate an important role for the lipid kinase activity of Vps34p in virulence and vesicular protein transport. On the other hand, the lipid-kinase-defective strain and the vps34 null mutant differ in their temperature- and osmotic-stress response. This indicates a possible role for activities different from the lipid kinase function of Vps34p.

The putative vacuolar ATPase subunit Vma7p of Candida albicans is involved in vacuole acidification, hyphal development and virulence

Microbiology ◽  
2005 ◽  
Vol 151 (5) ◽  
pp. 1645-1655 ◽  
Author(s):  
Sophia Poltermann ◽  
Monika Nguyen ◽  
Juliane Günther ◽  
Jürgen Wendland ◽  
Albert Härtl ◽  
...  
Keyword(s):  
Candida Albicans ◽  
Hyphal Growth ◽  
Null Mutant ◽  
Alkaline Ph ◽  
Phosphatidylinositol 3 Kinase ◽  
Pathogenic Yeast ◽  
Atpase Subunit ◽  
Mutant Strains ◽  
Toxic Ions ◽  
Null Mutants

The vacuolar H+-ATPase (V-ATPase) component Vma7p of the human-pathogenic yeast Candida albicans regulates hyphal growth induced by serum and Spider medium and is essential for virulence. In order to characterize the functions of the putative V-ATPase subunit Vma7p of C. albicans, null mutants were generated. The resulting mutants showed reduced vacuole acidification, which correlated with defective growth at alkaline pH. In addition, defects in degradation of intravacuolar putative endosomal structures were observed. vma7 null mutants were sensitive towards the presence of metal ions. It is concluded that the sequestration of toxic ions in the vacuole via a H+ gradient generated by the V-ATPase is affected. The vma7 null mutant strains were avirulent in a mouse model of systemic candidiasis. In addition, C. albicans vma7 null mutants and the null mutant strain of the Vma7p-interacting phosphatidylinositol 3-kinase Vps34p showed similar phenotypes. In summary, the V-ATPase subunit Vma7p is involved in vacuolar ion transport and this transport is required for hyphal growth and virulence of C. albicans.

scholarly journals Candida albicans sphingolipid C9-methyltransferase is involved in hyphal elongation

Microbiology ◽  
2010 ◽  
Vol 156 (4) ◽  
pp. 1234-1243 ◽  
Author(s):  
Takahiro Oura ◽  
Susumu Kajiwara
Keyword(s):  
Candida Albicans ◽  
Lipid Membrane ◽  
Hyphal Growth ◽  
Pathogenic Fungi ◽  
Wild Type ◽  
Membrane Structures ◽  
Pathogenic Yeast ◽  
Chain Base ◽  
Long Chain Base ◽  
Long Chain

C9-methylated glucosylceramide is a fungus-specific sphingolipid. This lipid is a major membrane component in the cell and is thought to play important roles in the growth and virulence of several fungal species. To investigate the importance of the methyl branch of the long-chain base in glucosylceramides in pathogenic fungi, we identified and characterized a sphingolipid C9-methyltransferase gene (MTS1, C9-MethylTransferase for Sphingolipid 1) in the pathogenic yeast Candida albicans. The mts1 disruptant lacked (E,E)-9-methylsphinga-4,8-dienine in its glucosylceramides and contained (E)-sphing-4-enine and (E,E)-sphinga-4,8-dienine. Reintroducing the MTS1 gene into the mts1 disruptant restored the synthesis of (E,E)-9-methylsphinga-4,8-dienine in the glucosylceramides. We also created a disruptant of the HSX11 gene, encoding glucosylceramide synthase, which catalyses the final step of glucosylceramide synthesis, in C. albicans and compared this mutant with the mts1 disruptant. The C. albicans mts1 and hsx11 disruptants both had a decreased hyphal growth rate compared to the wild-type strain. The hsx11 disruptant showed increased susceptibility to SDS and fluconazole, similar to a previously reported sld1 disruptant that contained only (E)-sphing-4-enine in its glucosylceramides, suggesting that these strains have defects in their cell membrane structures. In contrast, the mts1 disruptant grew similarly to wild-type in medium containing SDS or fluconazole. These results suggest that the C9-methyl group of a long-chain base in glucosylceramides plays an important role in the hyphal elongation of C. albicans independent of lipid membrane disruption.

scholarly journals Effects of Azole Antifungal Drugs on the Transition from Yeast Cells to Hyphae in Susceptible and Resistant Isolates of the Pathogenic Yeast Candida albicans

1999 ◽  
Vol 43 (4) ◽  
pp. 763-768 ◽  
Author(s):  
Kien C. Ha ◽  
Theodore C. White
Keyword(s):  
Drug Resistance ◽  
Candida Albicans ◽  
Molecular Mechanisms ◽  
Opportunistic Infections ◽  
Antifungal Drugs ◽  
Yeast Cells ◽  
Pathogenic Yeast ◽  
Oral Infections ◽  
Antifungal Drug Resistance ◽  
Hyphal Formation

ABSTRACT Oral infections caused by the yeast Candida albicansare some of the most frequent and earliest opportunistic infections in human immunodeficiency virus-infected patients. The widespread use of azole antifungal drugs has led to the development of drug resistance, creating a major problem in the treatment of yeast infections in AIDS patients and other immunocompromised individuals. Several molecular mechanisms that contribute to drug resistance have been identified. InC. albicans, the ability to morphologically switch from yeast cells (blastospores) to filamentous forms (hyphae) is an important virulence factor which contributes to the dissemination ofCandida in host tissues and which promotes infection and invasion. A positive correlation between the level of antifungal drug resistance and the ability to form hyphae in the presence of azole drugs has been identified. Under hypha-inducing conditions in the presence of an azole drug, resistant clinical isolates form hyphae, while susceptible yeast isolates do not. This correlation is observed in a random sample from a population of susceptible and resistant isolates and is independent of the mechanisms of resistance.35S-methionine incorporation suggests that growth inhibition is not sufficient to explain the inhibition of hyphal formation, but it may contribute to this inhibition.

scholarly journals PHR2 of Candida albicans encodes a functional homolog of the pH-regulated gene PHR1 with an inverted pattern of pH-dependent expression.

1997 ◽  
Vol 17 (10) ◽  
pp. 5960-5967 ◽  
Author(s):  
F A Mühlschlegel ◽  
W A Fonzi
Keyword(s):  
Candida Albicans ◽  
Mutant Strain ◽  
Low Ph ◽  
Predicted Amino Acid Sequence ◽  
Acidic Ph ◽  
Null Mutant ◽  
Alkaline Ph ◽  
Ph Values ◽  
Functional Homolog ◽  
Morphological Defects

Deletion of PHR1, a pH-regulated gene of Candida albicans, results in pH-conditional defects in growth, morphogenesis, and virulence evident at neutral to alkaline pH but absent at acidic pH. Consequently, we searched for a functional homolog of PHR1 active at low pH. This resulted in the isolation of a second pH-regulated gene, designated PHR2. The expression of PHR2 was inversely related to that of PHR1, being repressed at pH values above 6 and progressively induced at more acidic pH values. The predicted amino acid sequence of the PHR2 protein, Phr2p, was 54% identical to that of Phr1p. A PHR2 null mutant exhibited pH-conditional defects in growth and morphogenesis analogous to those of PHR1 mutants but manifest at acid rather than alkaline pH values. Engineered expression of PHR1 at acid pH in a PHR2 mutant strain and PHR2 at alkaline pH in a PHR1 mutant strain complemented the defects in the opposing mutant. Deletion of both PHR1 and PHR2 resulted in a strain with pH-independent, constitutive growth and morphological defects. These results indicate that PHR1 and PHR2 represent a novel pH-balanced system of functional homologs required for C. albicans to adapt to environments of diverse pH.

scholarly journals Disruption of the Candida albicans TPS2 Gene Encoding Trehalose-6-Phosphate Phosphatase Decreases Infectivity without Affecting Hypha Formation

2002 ◽  
Vol 70 (4) ◽  
pp. 1772-1782 ◽  
Author(s):  
Patrick Van Dijck ◽  
Larissa De Rop ◽  
Karolina Szlufcik ◽  
Elke Van Ael ◽  
Johan M. Thevelein
Keyword(s):  
Candida Albicans ◽  
High Throughput Screening ◽  
Deletion Mutant ◽  
Systemic Infection ◽  
Homozygous Deletion ◽  
Antifungal Drugs ◽  
Gene Encoding ◽  
Complete Inactivation ◽  
Hypha Formation ◽  
Stress Protectant

ABSTRACT Deletion of trehalose-6-phosphate phosphatase, encoded by TPS2, in Saccharomyces cerevisiae results in accumulation of trehalose-6-phosphate (Tre6P) instead of trehalose under stress conditions. Since trehalose is an important stress protectant and Tre6P accumulation is toxic, we have investigated whether Tre6P phosphatase could be a useful target for antifungals in Candida albicans. We have cloned the C. albicans TPS2 (CaTPS2) gene and constructed heterozygous and homozygous deletion strains. As in S. cerevisiae, complete inactivation of Tre6P phosphatase in C. albicans results in 50-fold hyperaccumulation of Tre6P, thermosensitivity, and rapid death of the cells after a few hours at 44°C. As opposed to inactivation of Tre6P synthase by deletion of CaTPS1, deletion of CaTPS2 does not affect hypha formation on a solid glucose-containing medium. In spite of this, virulence of the homozygous deletion mutant is strongly reduced in a mouse model of systemic infection. The pathogenicity of the heterozygous deletion mutant is similar to that of the wild-type strain. CaTPS2 is a new example of a gene not required for growth under standard conditions but required for pathogenicity in a host. Our results suggest that Tre6P phosphatase may serve as a potential target for antifungal drugs. Neither Tre6P phosphatase nor its substrate is present in mammals, and assay of the enzymes is simple and easily automated for high-throughput screening.

scholarly journals CRISPR-Cas9 mutagenesis and single gene reintegration suggests functional diversity within the Candida albicans TLO gene family

2021 ◽  
Vol 3 (12) ◽  
Author(s):  
Jessica Fletcher ◽  
Gary Moran ◽  
Derek Sullivan
Keyword(s):  
Growth Rate ◽  
Candida Albicans ◽  
Gene Family ◽  
Biofilm Formation ◽  
Single Gene ◽  
Hyphal Growth ◽  
Detectable Effect ◽  
Null Mutant ◽  
Phenotypic Analysis ◽  
Functional Differences

Candida albicans has between 10-15 Telomere-associated ORF family(TLO)genes, whereas its closest relative, Candida dubliniensis, has two. The Tlo proteins are components of the Mediator complex which plays an important role in transcriptional regulation. CRISPR-Cas9 mutagenesis was used to generate a TLOnull mutant of C. albicans. Phenotypic analysis of the mutant showed significantly reduced fitness, with major defects in growth rate, morphogenesis, stress resistance and virulence in a Galleria mellonellamodel. Clade representative TLOα1, TLOβ2 and TLOγ11constructs were reintroduced into the null mutant background to determine if members of the TLO gene family exhibit functional differences. The genes were reintroduced under the control of the TET1 and ENO1promoters. TLOα1and TLOβ2expression restored stress tolerance and growth rate, in some cases to the level of the WT. TLOβ2expression also showed a dramatic effect on morphology resulting in constitutive true hyphal growth. Moderate expression of TLOγ11 had no detectable effect on many of the phenotypes tested, however overexpression increased biofilm formation in Spider medium, and also conferred increased resistance to cell wall stressors. These data suggest that individual TLO genes have distinct functions and that the diversity within the TLO family may contribute to the relative success of C. albicans as a coloniser and pathogen of humans.

scholarly journals Functional Characterization of Myosin I Tail Regions in Candida albicans

2004 ◽  
Vol 3 (5) ◽  
pp. 1272-1286 ◽  
Author(s):  
Ursula Oberholzer ◽  
Tatiana L. Iouk ◽  
David Y. Thomas ◽  
Malcolm Whiteway
Keyword(s):  
Candida Albicans ◽  
Functional Characterization ◽  
Hyphal Growth ◽  
Tail Domain ◽  
Cortical Actin ◽  
Pathogenic Yeast ◽  
Rhodamine Phalloidin ◽  
Myosin I ◽  
Hyphal Formation ◽  
Hyphal Tip

ABSTRACT The molecular motor myosin I is required for hyphal growth in the pathogenic yeast Candida albicans. Specific myosin I functions were investigated by a deletion analysis of five neck and tail regions. Hyphal formation requires both the TH1 region and the IQ motifs. The TH2 region is important for optimal hyphal growth. All of the regions, except for the SH3 and acidic (A) regions that were examined individually, were required for the localization of myosin I at the hyphal tip. Similarly, all of the domains were required for the association of myosin I with pelletable actin-bound complexes. Moreover, the hyphal tip localization of cortical actin patches, identified by both rhodamine-phalloidin staining and Arp3-green fluorescent protein signals, was dependent on myosin I. Double deletion of the A and SH3 domains depolarized the distribution of the cortical actin patches without affecting the ability of the mutant to form hyphae, suggesting that myosin I has distinct functions in these processes. Among the six myosin I tail domain mutants, the ability to form hyphae was strictly correlated with endocytosis. We propose that the uptake of cell wall remodeling enzymes and excess plasma membrane is critical for hyphal formation.

scholarly journals The zinc cluster transcription factor Rha1 is a positive filamentation regulator in Candida albicans

Genetics ◽  
2021 ◽  
Author(s):  
Raha Parvizi Omran ◽  
Bernardo Ramírez-Zavala ◽  
Walters Aji Tebung ◽  
Shuangyan Yao ◽  
Jinrong Feng ◽  
...  
Keyword(s):  
Transcription Factor ◽  
Candida Albicans ◽  
Transcription Factors ◽  
Systemic Infection ◽  
Hyphal Growth ◽  
Growth Conditions ◽  
Hyphal Development ◽  
Zinc Cluster ◽  
Function Blocks ◽  
External Signals

Abstract Zinc cluster transcription factors are essential fungal regulators of gene expression. In the pathogen Candida albicans, the gene orf19.1604 encodes a zinc cluster transcription factor regulating filament development. Hyperactivation of orf19.1604, which we have named RHA1 for Regulator of Hyphal Activity, generates wrinkled colony morphology under non-hyphal growth conditions, triggers filament formation, invasiveness, and enhanced biofilm formation and causes reduced virulence in the mouse model of systemic infection. The strain expressing activated Rha1 shows up-regulation of genes required for filamentation and cell-wall-adhesion-related proteins. Increased expression is also seen for the hyphal-inducing transcription factors Brg1 and Ume6, while the hyphal repressor Nrg1 is downregulated. Inactivation of RHA1 reduces filamentation under a variety of filament-inducing conditions. In contrast to the partial effect of either single mutant, the double rha1 ume6 mutant strain is highly defective in both serum- and Spider-medium-stimulated hyphal development. While the loss of Brg1 function blocks serum-stimulated hyphal development, this block can be significantly bypassed by Rha1 hyperactivity, and the combination of Rha1 hyperactivity and serum addition can generate significant polarization even in brg1 ume6 double mutants. Thus, in response to external signals, Rha1 functions with other morphogenesis regulators including Brg1 and Ume6, to mediate filamentation.

scholarly journals Candida albicans hyphal expansion causes phagosomal membrane damage and luminal alkalinization

2018 ◽  
Author(s):  
Johannes Westman ◽  
Gary Moran ◽  
Selene Mogavero ◽  
Bernhard Hube ◽  
Sergio Grinstein
Keyword(s):  
Candida Albicans ◽  
Fungal Infections ◽  
Membrane Damage ◽  
Hyphal Growth ◽  
Antifungal Drugs ◽  
Apparent Paradox ◽  
Opportunistic Fungi ◽  
Life Threatening ◽  
Proton Leakage ◽  
Phagosomal Membrane

ABSTRACTMacrophages rely on phagosomal acidity to destroy engulfed microorganisms. To survive this hostile response, opportunistic fungi such as Candida albicans developed strategies to evade the acidic environment. C. albicans is polymorphic, able to convert from yeast to hyphae, and this transition is required to subvert the microbicidal activity of the phagosome. However, the phagosomal lumen, which is acidic and nutrient-deprived, inhibits yeast-to-hypha transition. To account for this apparent paradox, it was recently proposed that C. albicans produces ammonia that alkalinizes the phagosome, thus facilitating yeast-to-hypha transformation. We re-examined the mechanism underlying phagosomal alkalinization by applying dual-wavelength ratiometric pH measurements. The phagosomal membrane was found to be highly permeable to ammonia, which is therefore unlikely to account for the pH elevation. Instead, we find that yeast-to-hypha transition begins within acidic phagosomes, and that alkalinization is a consequence of proton leakage induced by excessive membrane distension caused by the expanding hypha.IMPORTANCEC. albicans is the most common nosocomial fungal infection, and over three million people acquire life-threatening invasive fungal infections every year. Even if antifungal drugs exist, almost half of these patients will die. Despite this, fungi remain underestimated as pathogens. Our study uses quantitative biophysical approaches to demonstrate that the yeast-to-hypha transition occurs within the nutrient deprived, acidic phagosome and that alkalinization is a consequence, as opposed to the cause of hyphal growth.

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