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Dynamic expression of cell wall proteins of Candida albicans revealed by probes from cDNA clones
H M Alloush1, J L López-Ribot, W L Chaffin
1Department of Microbiology and Immunology, Texas Tech University Health Sciences Center, Lubbock 79430, USA.
Abstract:
Five cDNA clones were selected from the positive clones detected by screening a germ tube expression library constructed in lambda gt11 with rabbit antisera raised against cell wall extracts of Candida albicans. The selected clones were amplified and used to obtain affinity purified antibodies by eluting from the expressed proteins that had been previously transferred onto nitrocellulose discs. The antibodies obtained were used as probes in immunoblots of the cell wall extracts separated by denaturing polyacrylamide electrophoresis. A single protein band was detected for each clone. Detection of products of the cloned sequences varied according to the extraction procedure and/or cell morphology. These products included bands exhibiting apparent molecular weights of 40, 58, 68 and 70 kDa present in beta-mercaptoethanol (beta ME) extracts from both yeast and germ tubes, and a 30 kDa beta ME extracted protein specific for germ tubes. The expression of these products at the cell surface was confirmed by indirect immunofluorescence. Expression of the mRNAs of the different cDNA clones varied according to growth- and morphology-related factors and showed no direct correlation between expression and presence in the cell wall. These observations suggest that complex mechanisms are involved in the regulation and expression of cell surface components of C. albicans.
Insights
Researchers identified Candida albicans cell wall proteins using cDNA clones and antibodies. Protein expression varied with cell morphology and extraction, suggesting complex regulatory mechanisms for cell surface components.
Area of Science:
- Mycology
- Molecular Biology
- Immunology
Background:
- Candida albicans is an opportunistic fungal pathogen.
- Understanding its cell wall components is crucial for developing antifungal strategies.
- Gene expression and protein localization in C. albicans are complex.
Purpose of the Study:
- To identify and characterize genes encoding cell wall proteins in Candida albicans.
- To investigate the expression patterns of these proteins during different growth phases and morphologies.
- To elucidate the regulatory mechanisms governing cell surface component expression.
Main Methods:
- Screening of a lambda gt11 expression library using antibodies against C. albicans cell wall extracts.
- Amplification of cDNA clones and production of affinity-purified antibodies.
- Immunoblotting of cell wall extracts and indirect immunofluorescence for protein localization.
- Analysis of mRNA expression levels.
Main Results:
- Five cDNA clones encoding cell wall proteins were identified.
- Proteins of 40, 58, 68, and 70 kDa were detected in both yeast and germ tube extracts.
- A 30 kDa protein was specifically found in germ tube extracts.
- Protein expression varied with extraction methods and cell morphology.
- mRNA expression did not directly correlate with cell wall presence.
Conclusions:
- Complex regulatory mechanisms control the expression and localization of C. albicans cell surface components.
- Cell morphology and growth conditions significantly influence protein expression.
- Further research is needed to fully understand the regulation of cell wall biogenesis in C. albicans.