Disruption studies of a Candida albicans gene, ELF1: a member of the ATP-binding cassette family

Sturtevant, Joy; Cihlar, R. L.; Calderone, Richard

doi:10.1099/00221287-144-8-2311

Cited by 18 publications

(16 citation statements)

References 32 publications

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“…In some instances, a rapid protocol was used to isolate genomic DNA (Burke et al, 2000). Southern analysis was performed as previously described (Sturtevant et al, 1998). Blots were probed with the C. albicans BMH1 ORF isolated from plasmid pJP1.…”

Section: Dna Manipulationsmentioning

confidence: 99%

“…These clones were manually sequenced and one clone that showed remarkable homology to the 14-3-3 proteins (Muhlschlegel et al, 1998;Sturtevant, 2000) was used to screen an EcoRI C. albicans SC5314 Lambda Zap II library (Sturtevant et al, 1998). A 5 kb EcoRI genomic fragment in pBluescriptSK + phagemid was excised from a tertiary positive plaque from the Lambda Zap II SC5314 library.…”

Section: Isolation Of C Albicans Bmh1mentioning

confidence: 99%

“…A 5 kb EcoRI fragment was identified. Subsequently, an EcoRI-digested SC5314 genomic DNA library constructed in lZap II (Sturtevant et al, 1998) was screened and a 5 kb clone isolated. The entire ORF and 5k and 3k flanking regions were sequenced.…”

Section: Isolation Of C Albicans Bmh1mentioning

confidence: 99%

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The Candida albicans 14‐3‐3 gene, BMH1, is essential for growth

Cognetti

Davis

Sturtevant

2001

Yeast

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The 14-3-3 proteins are a family of conserved small acidic proteins that have been implicated in playing major roles in a wide variety of signalling cascades. In Saccharomyces cerevisiae, the 14-3-3 genes (BMH1 and BMH2) are essential for normal pseudohyphal induction and normal bud cell development. The Bmh proteins function in the cAMPdependent RAS/MAPK and rapamycin-sensitive signalling cascades. Deletion of only one BMH gene demonstrates no phenotypic differences under normal growth conditions. Strains deleted of both BMH1 and BMH2 are either non-viable or demonstrate sensitivity to environmental stresses. In Schizosaccharomyces pombe, the BMH homologues (RAD24 and RAD25) are essential for cell cycle control after DNA damage and deletion of both genes renders the cell inviable. The 14-3-3 gene in Candida albicans (BMH1) was identified using a novel adherence assay and differential display RT-PCR. Unlike other yeasts, C. albicans has only one 14-3-3 gene (BMH1). It was not possible to construct double knockouts by routine methods. These results suggested that the C. albicans BMH1 gene is essential. The essentiality of C. albicans BMH1 was confirmed by a PCR disruption technique. The C. albicans bmh1D/BMH1 heterozygotes exhibit growth and morphogenetic defects. Therefore, the BMH1 gene in C. albicans (Accession No. AF038154) is an excellent candidate to improve our understanding of the coordinate regulation of cell cycle and morphogenesis.

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Section: Dna Manipulationsmentioning

confidence: 99%

Section: Isolation Of C Albicans Bmh1mentioning

confidence: 99%

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The Candida albicans 14‐3‐3 gene, BMH1, is essential for growth

Cognetti

Davis

Sturtevant

2001

Yeast

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“…Genomic DNA from C. albicans was isolated using glass beads (9). Southern blot analysis was performed as previously described (43). Probes were radiolabeled on purified plasmid inserts with [␣-32 P]dCTP by using Ready-To-Go DNA Labeling Beads (Amersham Pharmacia Biotech).…”

Section: Methodsmentioning

confidence: 99%

Candida albicans VPS11 Is Required for Vacuole Biogenesis and Germ Tube Formation

2003

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The Candida albicans vacuole has previously been observed to undergo rapid expansion during the emergence of a germ tube from a yeast cell, to occupy the majority of the parent yeast cell. Furthermore, the yeast-to-hypha switch has been implicated in the virulence of this organism. The class C vps (vacuolar protein sorting) mutants of Saccharomyces cerevisiae are defective in multiple protein delivery pathways to the vacuole and prevacuole compartment. In this study C. albicans homologues of the S. cerevisiae class C VPS genes have been identified. Deletion of a C. albicans VPS11 homologue resulted in a number of phenotypes that closely resemble those of the class C vps mutants of S. cerevisiae, including the absence of a vacuolar compartment. The C. albicans vps11⌬ mutant also had much-reduced secreted lipase and aspartyl protease activities. Furthermore, vps11⌬ strains were defective in yeast-hypha morphogenesis. Upon serum induction of filamentous growth, mutants showed delayed emergence of germ tubes, had a reduced apical extension rate compared to those of control strains, and were unable to form mature hyphae. These results suggest that Vps11p-mediated trafficking steps are necessary to support the rapid emergence and extension of the germ tube from the parent yeast cell.

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“…Genomic DNA from C. albicans was isolated using glass beads (Burke et al, 2000). Southern analysis was performed as previously described (Sturtevant et al, 1998). DNA sequence of selected clones was determined by MWG Biotech.…”

Section: Methodsmentioning

confidence: 99%

Mutant alleles of the essential 14-3-3 gene in Candida albicans distinguish between growth and filamentation

et al. 2004

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The opportunistic fungal pathogen Candida albicans has the ability to exploit diverse host environments and can either reside commensally or cause disease. In order to adapt to its new environment it must respond to new physical conditions, nutrient sources, and the host immune response. This requires the co-regulation of multiple signalling networks. The 14-3-3 family of proteins is highly conserved in all eukaryotic species. These proteins regulate signalling pathways involved in cell survival, the cell cycle, and differentiation, and effect their functions via interactions with phosphorylated serines/threonines. In C. albicans there is only one 14-3-3 protein, Bmh1p, and it is required for vegetative growth and optimal filamentation. In order to dissect separate functions of Bmh1p in C. albicans, site-directed nucleotide substitutions were made in the C. albicans BMH1 gene based on studies in other species. Putative temperature-sensitive, ligand-binding and dimerization mutants were constructed. In addition two mutant strains identified through random mutagenesis were analysed. All five mutant strains demonstrated varying defects in growth and filamentation. This paper begins to segregate functions of Bmh1p that are required for optimal growth and the different filamentation pathways. These mutant strains will allow the identification of 14-3-3 target interactions and correlate the individual functions of Bmh1p to cellular processes involved in pathogenesis.

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Disruption studies of a Candida albicans gene, ELF1: a member of the ATP-binding cassette family

Cited by 18 publications

References 32 publications

The Candida albicans 14‐3‐3 gene, BMH1, is essential for growth

The Candida albicans 14‐3‐3 gene, BMH1, is essential for growth

Candida albicans VPS11 Is Required for Vacuole Biogenesis and Germ Tube Formation

Mutant alleles of the essential 14-3-3 gene in Candida albicans distinguish between growth and filamentation

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