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Molecular cloning of the CRM1 gene from Candida albicans
M Raymond1, D Dignard, A M Alarco
1Institut de recherches cliniques de Montréal, Montréal, Québec, Canada. raymonm@ircm.qc.ca
Abstract:
In a screen for Candida albicans genes capable of supressing a ste20Delta mutation in Saccharomyces cerevisiae, a homologue of the exportin-encoding gene CRM1 was isolated. The CaCRM1 gene codes for a protein of 1079 amino acids with a predicted molecular weight of 124 029 and isoelectric point of 5.04. Crm1p from C. albicans displays significant amino acid sequence homology with Crm1p from Saccharomyces cerevisiae (65% identity, 74% similarity), Schizosaccharomyces pombe (55% identity, 66% similarity), Caenorhabditis elegans (45% identity, 57% similarity), and Homo sapiens (48% identity, 59% similarity). Interestingly, CaCRM1 encodes a threonine rather than a cysteine at position 533 in the conserved central region, suggesting that CaCrm1p is leptomycin B-insensitive, like S. cerevisiae Crm1p. CaCRM1 on a high copy vector can complement a thermosensitive allele of CRM1 (xpo1-1) in S. cerevisiae, showing that CaCrm1p and S. cerevisiae Crm1p are functionally conserved. Southern blot analysis suggests that CaCRM1 is present at a single locus within the C. albicans genome. The nucleotide sequence of the CaCRM1 gene has been deposited at GenBank under Accession No. AF178855.
Insights
A novel Candida albicans gene, CaCRM1, was identified for its ability to suppress a mutation in yeast. This exportin homologue shows functional conservation with its Saccharomyces cerevisiae counterpart.
Area of Science:
- Molecular Biology
- Mycology
- Genetics
Background:
- The exportin-encoding gene CRM1 plays a crucial role in nuclear transport.
- Understanding CRM1 homologs in pathogenic fungi like Candida albicans is important for their biology.
Purpose of the Study:
- To identify and characterize the Candida albicans homolog of the CRM1 gene.
- To investigate the functional conservation and properties of the CaCRM1 protein.
Main Methods:
- Gene screening in Saccharomyces cerevisiae to identify suppressor genes.
- Sequence homology analysis with known CRM1 proteins.
- Functional complementation assays using a thermosensitive yeast CRM1 allele.
- Southern blot analysis for gene locus determination.
Main Results:
- The Candida albicans CRM1 (CaCRM1) gene was isolated and characterized.
- CaCrm1p shares significant sequence homology with Crm1p from various organisms, including yeast and humans.
- CaCrm1p demonstrated functional conservation by complementing a thermosensitive S. cerevisiae CRM1 mutant.
- CaCRM1 appears to be a single-copy gene in the C. albicans genome.
- The protein sequence suggests CaCrm1p is likely insensitive to leptomycin B.
Conclusions:
- CaCRM1 is a functional homolog of CRM1, conserved across species.
- The identified CaCRM1 gene provides a new tool for studying nuclear export in Candida albicans.
- Functional and sequence similarities suggest conserved roles for CRM1 in eukaryotic organisms.