Elongation factor 3 (EF‐3) from <i>Candida albicans</i> shows both structural and functional similarity to EF‐3 from <i>Saccharomyces cerevisiae</i>

Colthurst, David R.; Schauder, Birgit; Hayes, Melvin B.; Tuite, Mick F.

doi:10.1111/j.1365-2958.1992.tb02168.x

Cited by 30 publications

(20 citation statements)

References 28 publications

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“…Complementation of two strains of haploid S. cerevisiae bearing a temperature-sensitive mutation in the YEF3 gene (strains ts22 and ts27) revealed the appearance of significantly slower-growing colonies at the nonpermissive temperature of 34°C. Similar complementation with wild-type YEF3 genes from S. cerevisiae and C. albicans gave normal-sized colonies, as reported previously (6,8). From these results, we conclude that CnEF3 complements the function of YEF3 poorly, although it does show that CnEF3 has functional capability.…”

Section: Resultssupporting

confidence: 67%

“…Since inhibition of protein synthesis by targeting EF3 might represent an effective drug strategy for the control of fungal diseases, identification and characterization of this factor among diverse fungi of medical importance are essential to allow design of pharmacological inhibitors having the broadest application. Thus far, EF3 has only been cloned and characterized from yeasts of the order Ascomycota as well as the fungus-like organism Pneumocystis carinii (6). Basidiomycete yeasts are distinct from ascomycete yeasts in both morphology (2) and 18S ribosomal DNA sequence criteria (21).…”

Section: Discussionmentioning

confidence: 99%

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Evolutionary Divergence of an Elongation Factor 3 from Cryptococcus neoformans

et al. 2001

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Elongation factor 3 (EF3) is considered a promising drug target for the control of fungal diseases because of its requirement for protein synthesis and survival of fungi and a lack of EF3 in the mammalian host. However, EF3 has been characterized only in ascomycete yeast. In order to understand the role of EF3 in a basidiomycete yeast, we cloned the gene encoding EF3 from Cryptococcus neoformans (CnEF3), an important fungal pathogen in immunocompromised patients, including those infected with human immunodeficiency virus. CnEF3 was found to encode a 1,055-amino-acid protein and has 44% identity with EF3 from Saccharomyces cerevisiae (YEF3). Expressed CnEF3 exhibited ATPase activity that was only modestly stimulated by ribosomes from S. cerevisiae. In contrast, CnEF3 showed tight binding to cryptococcal ribosomes, as shown by an inability to be removed under conditions which successfully remove Saccharomyces EF3 from ribosomes (0.5 M KCl or 2 M LiCl). CnEF3 also poorly complemented a YEF3 defect in a diploid null mutant and two temperature-sensitive mutants which have been shown previously to be complemented well by EF3 from other ascomycetes, such as Candida albicans. These data clearly identify the presence of a functioning EF3 in the basidiomycete yeast C. neoformans, which demonstrates an evolutionary divergence from EF3 of ascomycete yeast.Cryptococcus neoformans is an important fungal pathogen which causes a lethal meningoencephalitis in a significant number of persons with AIDS and afflicts an increasing number of immunocompromised patients on steroids, chemotherapy, or posttransplant immunosuppressives (18). Therapy of cryptococcosis is limited by toxicity of such agents as amphotericin B (1); newer agents such as the azole inhibitors are less toxic, but increasing reports of resistance may limit their eventual usefulness (3,15,17). Echinocandins and pneumocandins are important new antifungal agents which are inhibitors of 1,3-␤-glucan synthetases and show excellent activity against ascomycete pathogens such as Candida albicans and Aspergillus fumigatus, but alterations in this enzyme from basidiomycetes make this important class of agents ineffective against C. neoformans (27). The latter example shows the potential gap in antifungal coverage which may occur when inhibitors are chosen without consideration of possible evolutionary differences in drug targets within various fungal pathogens.Elongation factor 3 (EF3) has been shown to be a required translation cofactor in the ascomycete Saccharomyces cerevisiae (7). The factor is also present in a variety of pathogenic ascomycete fungi, including Candida albicans (8) and the pathogen Pneumocystis carinii (34), which has been shown to be closely related to ascomycete yeasts based on analysis of its rRNA gene as well as genes encoding dihydrofolate reductase, thymidylate synthetase, ␤-tubulin, and ATP (for a review, see reference 31). While an anti-EF3 antibody has been shown to react with basidiomycete yeasts (4), EF3 has not been characterized from this cl...

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Section: Resultssupporting

confidence: 67%

Section: Discussionmentioning

confidence: 99%

Evolutionary Divergence of an Elongation Factor 3 from Cryptococcus neoformans

et al. 2001

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“…mRNA levels were measured relative to the rRNAs by loading approximately equal amounts of total RNA in each lane of the Northern blots. In addition, the TEF3 mRNA (Colthurst et al, 1992) was probed as a positive (non-quantitative) control (Hube et al, 1994).…”

Section: Methodsmentioning

confidence: 99%

“…To analyse PLD1 gene disruption, a 430 bp long T7 (Stratagene) PLD1-2 PCR fragment of plasmid pHPLD1 or the random labelled hisG ::URA3 ::hisG cassette of pMB7 was used as probe. For Northern blots, the same fragment of subclone pHPLD1 and a 700 bp PCR fragment of TEF3 (Colthurst et al, 1992 ;Hube et al, 1994) were labelled with [α-$#P]dCTP (" 3000 Ci mmol − " ; 111 TBq mmol − ") (Amersham). mRNA levels were measured relative to the rRNAs by loading approximately equal amounts of total RNA in each lane of the Northern blots.…”

Section: Methodsmentioning

confidence: 99%

The role and relevance of phospholipase D1 during growth and dimorphism of Candida albicans

et al. 2001

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The phosphatidylcholine-specific phospholipase D1 (PLD1) in Saccharomyces cerevisiae is involved in vesicle transport and is essential for sporulation. The gene encoding the homologous phospholipase D1 from Candida albicans (PLD1) was used to study the role of PLD1 in this pathogenic fungus. In vitro and in vivo expression studies using Northern blots and reverse transcriptase-PCR showed low PLD1 mRNA levels in defined media supporting yeast growth and during experimental infection, while enhanced levels of PLD1 transcripts were detected during the yeast to hyphal transition. To study the relevance of PLD1 during yeast and hyphal growth, an essential part of the gene was deleted in both alleles of two isogenic strains. In vitro PLD1 activity assays showed that pld1 mutants produced no detectable levels of phosphatidic acid, the hydrolytic product of PLD1 activity, and strongly reduced levels of diacylglycerol, the product of lipid phosphate phosphohydrolase, suggesting no or a negligible background PLD1 activity in the pld1 mutants. The pld1 mutants showed no growth differences compared to the parental wild-type in liquid complex and minimal media, independent of the growth temperature. In addition, growth rates of pld1 mutants in media with protein as the sole source of nitrogen were similar to growth rates of the wild-type, indicating that secretion of proteinases was not reduced. Chlamydospore formation was normal in pld1 mutants. When germ tube formation was induced in liquid media, pld1 mutants showed similar rates of yeast to hyphal transition compared to the wild-type. However, no hyphae formation was observed on solid Spider medium, and cell growth on cornmeal/Tween 80 medium indicated aberrant morphogenesis. In addition, pld1 mutants growing on solid media had an attenuated ability to invade the agar. In a model of oral candidosis, pld1 mutants showed no attenuation of virulence. In contrast, the mutant was less virulent in two different mouse models. These data suggest that PLD1 is not essential for growth and oral infections. However, they also suggest that a prominent part of the phosphatidic acid and diacylglycerol pools is produced by PLD1 and that the level of these components is important for morphological transitions under certain conditions in C. albicans.

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“…GCN20 shows significant sequence similarities to genes that encode elongation factor 3 (EF-3). EF-3 genes are found in several fungi including C. albicans (Colthurst et al, 1992), Pneumocystis carinii (YpmaWong et al, 1992) and S. cerevisiae (Qin et al, 1990). The EF-3 proteins are attractive antifungal targets since they are not found in mammalian cells, but are essential for translation in fungal cells.…”

Section: Introductionmentioning

confidence: 99%

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

1998

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A 3.6 kb gene (ELF7) with homology to the ATP-binding cassette (ABC) gene family has been isolated from genomic libraries of Candida albicans. Members of this gene family include both membrane transport proteins which confer a drug-resistance phenotype, and proteins whose functions are associated with protein translation. ELF7 (Elongation Like Factor) showed greatest homology with a Sacchammyces cemwisiae ORF (YPu26w), whose function is unknown, and lower homology with fungal elongation factor 3 (EF-3) genes. In comparison, homology with a gene conferring a drug-resistant phenotype (CDR7) was low. To understand the function of ELF? in C. a/bkaf#S, geneknockout experiments were conducted using the hisG-URA3-hisG disruption cassette. Both single-copy (heterozygote) and double-disrupted strains in ELF7 were isolated. Phenotypically, the disrupted strains grew more slowly than wild-type and produced a mixture of large, irregular cells and apparently normal cells.

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Elongation factor 3 (EF‐3) from Candida albicans shows both structural and functional similarity to EF‐3 from Saccharomyces cerevisiae

Cited by 30 publications

References 28 publications

Evolutionary Divergence of an Elongation Factor 3 from Cryptococcus neoformans

Evolutionary Divergence of an Elongation Factor 3 from Cryptococcus neoformans

The role and relevance of phospholipase D1 during growth and dimorphism of Candida albicans

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

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