Dataset Information

Characterization of agglutinin-like sequence genes from non-albicans Candida and phylogenetic analysis of the ALS family.

ABSTRACT: The ALS (agglutinin-like sequence) gene family of Candida albicans encodes cell-surface glycoproteins implicated in adhesion of the organism to host surfaces. Southern blot analysis with ALS-specific probes suggested the presence of ALS gene families in C. dubliniensis and C. tropicalis; three partial ALS genes were isolated from each organism. Northern blot analysis demonstrated that mechanisms governing expression of ALS genes in C. albicans and C. dubliniensis are different. Western blots with an anti-Als serum showed that cross-reactive proteins are linked by beta 1,6-glucan in the cell wall of each non-albicans Candida, suggesting similar cell wall architecture and conserved processing of Als proteins in these organisms. Although an ALS family is present in each organism, phylogenetic analysis of the C. albicans, C. dubliniensis, and C. tropicalis ALS genes indicated that, within each species, sequence diversification is extensive and unique ALS sequences have arisen. Phylogenetic analysis of the ALS and SAP (secreted aspartyl proteinase) families show that the ALS family is younger than the SAP family. ALS genes in C. albicans, C. dubliniensis, and C. tropicalis tend to be located on chromosomes that also encode genes from the SAP family, yet the two families have unexpectedly different evolutionary histories. Homologous recombination between the tandem repeat sequences present in ALS genes could explain the different histories for co-localized genes in a predominantly clonal organism like C. albicans.

SUBMITTER: Hoyer LL

PROVIDER: S-EPMC1461614 | biostudies-other | 2001 Apr

REPOSITORIES: biostudies-other

ACCESS DATA

Publications

Characterization of agglutinin-like sequence genes from non-albicans Candida and phylogenetic analysis of the ALS family.

Hoyer L L LL Fundyga R R Hecht J E JE Kapteyn J C JC Klis F M FM Arnold J J

Genetics 20010401 4

The ALS (agglutinin-like sequence) gene family of Candida albicans encodes cell-surface glycoproteins implicated in adhesion of the organism to host surfaces. Southern blot analysis with ALS-specific probes suggested the presence of ALS gene families in C. dubliniensis and C. tropicalis; three partial ALS genes were isolated from each organism. Northern blot analysis demonstrated that mechanisms governing expression of ALS genes in C. albicans and C. dubliniensis are different. Western blots wit ...[more]

PMID: 11290712

Similar Datasets

Project description:The agglutinin-like sequence (Als) proteins are best-characterized in Candida albicans and known for their role in adhesion of the fungal cell to host and abiotic surfaces. ALS sequences are often misassembled in whole-genome sequence data because each species has multiple ALS loci that contain similar sequences, most notably tandem copies of highly conserved repeated sequences. The Candida parapsilosis species complex includes Candida parapsilosis, Candida orthopsilosis, and Candida metapsilosis, three distinct but closely related species. Using publicly available genome resources, de novo genome assemblies, and laboratory experimentation including Sanger sequencing, five ALS genes were characterized in C. parapsilosis strain CDC317, three in C. orthopsilosis strain 90-125, and four in C. metapsilosis strain ATCC 96143. The newly characterized ALS genes shared similar features with the well-known C. albicans ALS family, but also displayed unique attributes such as novel short, imperfect repeat sequences that were found in other genes encoding fungal cell-wall proteins. Evidence of recombination between ALS sequences and other genes was most obvious in CmALS2265, which had the 5' end of an ALS gene and the repeated sequences and 3' end from the IFF/HYR family. Together, these results blur the boundaries between the fungal cell-wall families that were defined in C. albicans. TaqMan assays were used to quantify relative expression for each ALS gene. Some measurements were complicated by the assay location within the ALS gene. Considerable variation was noted in relative gene expression for isolates of the same species. Overall, however, there was a trend toward higher relative gene expression in saturated cultures rather than younger cultures. This work provides a complete description of the ALS genes in the C. parapsilosis species complex and a toolkit that promotes further investigations into the role of the Als proteins in host-fungal interactions.

Project description:The agglutinin-like sequence (ALS) gene family encodes cell-surface adhesins that interact with host and abiotic surfaces, promoting colonization by opportunistic fungal pathogens such as Candida tropicalis. Studies of Als protein contribution to C. tropicalis adhesion would benefit from an accurate catalog of ALS gene sequences as well as insight into relative gene expression levels. Even in the genomics era, this information has been elusive: genome assemblies are often broken within ALS genes because of their extensive regions of highly conserved, repeated DNA sequences and because there are many similar ALS genes at different chromosomal locations. Here, we describe the benefit of long-read DNA sequencing technology to facilitate characterization of C. tropicalis ALS loci. Thirteen ALS loci in C. tropicalis strain MYA-3404 were deduced from a genome assembly constructed from Illumina MiSeq and Oxford Nanopore MinION data. Although the MinION data were valuable, PCR amplification and Sanger sequencing of ALS loci were still required to complete and verify the gene sequences. Each predicted Als protein featured an N-terminal binding domain, a central domain of tandemly repeated sequences, and a C-terminal domain rich in Ser and Thr. The presence of a secretory signal peptide and consensus sequence for addition of a glycosylphosphatidylinositol (GPI) anchor was consistent with predicted protein localization to the cell surface. TaqMan assays were designed to recognize each ALS gene, as well as both alleles at the divergent CtrALS3882 locus. C. tropicalis cells grown in five different in vitro conditions showed differential expression of various ALS genes. To place the C. tropicalis data into a larger context, TaqMan assays were also designed and validated for analysis of ALS gene expression in Candida albicans and Candida dubliniensis. These comparisons identified the subset of highly expressed C. tropicalis ALS genes that were predicted to encode proteins with the most abundant cell-surface presence, prioritizing them for subsequent functional analysis. Data presented here provide a solid foundation for future experimentation to deduce ALS family contributions to C. tropicalis adhesion and pathogenesis.

Project description:The ALS (agglutinin-like sequence) gene family encodes eight large cell-surface glycoproteins. The work presented here focuses on Als2p and Als4p, and is part of a larger effort to deduce the function of each Als protein. Both ALS4 alleles were deleted from the Candida albicans genome and the phenotype of the mutant strain (als4Delta/als4Delta; named 2034) studied. Loss of Als4p slowed germ tube formation of cells grown in RPMI 1640 medium and resulted in decreased adhesion of C. albicans to vascular endothelial cells. Loss of Als4p did not affect adhesion to buccal epithelial cells, biofilm formation in a catheter model, or adhesion to or destruction of oral reconstituted human epithelium (RHE). Although deletion of one ALS2 allele was achieved readily, a strain lacking the second allele was not identified despite screening thousands of transformants. The remaining ALS2 allele was placed under control of the C. albicans MAL2 promoter to create an als2Delta/PMAL2-ALS2 strain (named 2342). Real-time RT-PCR analysis of strain 2342 grown in glucose-containing medium (non-inducing conditions) showed that although ALS2 transcript levels were greatly reduced compared to wild-type cells, some ALS2 transcript remained. The decreased ALS2 expression levels were sufficient to slow germ tube formation in RPMI 1640 and Lee medium, reduce adhesion to vascular endothelial cells and to RHE, decrease RHE destruction, and impair biofilm formation. Growth of strain 2342 in maltose-containing medium (inducing conditions) restored the wild-type phenotype in all assays. Real-time RT-PCR analysis demonstrated that in maltose-containing medium, strain 2342 overexpressed ALS2 compared to wild-type cells; however no overexpression phenotype was apparent. Microarray analysis revealed little transcriptional response to ALS4 deletion, but showed twofold up-regulation of orf19.4765 in the glucose-medium-grown als2Delta/PMAL2-ALS2 strain. orf19.4765 encodes a protein with features of a glycosylated cell wall protein with similarity to Saccharomyces cerevisiae Ccw12p, although initial analysis suggested functional differences between the two proteins. Real-time RT-PCR measurement of ALS2 and ALS4 transcript copy number showed a 2.8-fold increase in ALS2 expression in the als4Delta/als4Delta strain and a 3.2-fold increase in ALS4 expression in the als2Delta/PMAL2-ALS2 strain, suggesting the potential for compensatory function between these related proteins.

Dataset Information

Characterization of agglutinin-like sequence genes from non-albicans Candida and phylogenetic analysis of the ALS family.

Publications

Characterization of agglutinin-like sequence genes from non-albicans Candida and phylogenetic analysis of the ALS family.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets