Brian Morrow

Transcription of the gene for a pepsinogen, PEP1, is regulated by white-opaque switching in Candida albicans.

Cells of Candida albicans WO-1 spontaneously switch between a white and opaque CFU, and this phase transition involves a dramatic change in cellular phenotype. By using a differential hybridization screen, an opaque-specific cDNA, Op1a, which represents the transcript of a gene regulated by switching, has been isolated. The gene for Op1a is transcribed by opaque but not by white cells. The nucleotide sequence of the Op1a cDNA reveals over 99% base homology with an acid protease gene of C. albicans, and the predicted amino acid sequence demonstrates that the product of this gene is a member of the family of pepsinogens, which possess a hydrophobic leader sequence for secretion and two catalytic aspartate domains. Southern blots of both genomic DNA digested with 14 different endonucleases and electrophoretically separated chromosomes were probed with the Op1a cDNA. No polymorphisms were detected in either case between white and opaque cells, suggesting that no genomic reorganization occurs in the proximity of the gene during the white-opaque transition. Although transcription of Op1a correlates with the high levels of extracellular protease activity in opaque cell cultures and the absence of activity in white cell cultures, stimulation of extracellular protease activity by addition of serum albumin is not accompanied by Op1a transcription in cultures of WO-1 white cells or cultures of two additional clinical isolates of C. albicans, suggesting that expression of one or more other protease genes is stimulated in these cases. The results demonstrate that transcription of the Op1a gene is under the rigid control of switching in strain WO-1.

High-frequency phenotypic switching in Candida albicans

Most strains of Candida albicans are capable of switching spontaneously and at high frequencies between a number of phenotypes distinguished by colony morphology. Unlike switching in many other microbial pathogens, switching in C. albicans is pleiotropic, affecting several morphological and physiological parameters. Recently, the first phase-specific genes were identified and shown to be regulated at the level of gene transcription.

The frequency of integrative transformation at phase-specific genes of Candida albicans correlates with their transcriptional state.

The phase transition between the white and opaque phenotypes in the switching system of Candida albicans strain WO-1 is accompanied by the differential expression of the white-specific gene WH11 and the opaque-specific gene PEP1. The frequency of integrative transformation at the white-specific gene locus WH11 is between 4.5 and 7.0 times more frequent in white than in opaque spheroplasts, and the frequency of disruptive transformation at the opaque-specific gene locus PEP1 is 30.5 times more frequent in opaque spheroplasts than in white spheroplasts. In contrast, the frequencies of integrative transformation at the constitutively expressed loci ADE2 and EF1α2 are similar in the white and opaque phases. Therefore, the frequency of integration of linear plasmid DNA containing sequences of phase-specific genes correlates with the transcriptional state of the targeted locus.

Candida Albicans Acid Proteinase: Characterization and Role in Candidiasis

Candida albicans and related species are medically important yeast-like dimorphic fungi that are responsible, in part, for the rising incidence of serious, life-threatening opportunistic infections seen in immunocompromised and debilitated patients. In addition to minor localized infections of cutaneous, oropharyngeal and vaginal epithelium, Candida spp. produce transient or persistent fungemia that leads to systemic infections of virtually any organ (esp. liver, kidney and lung). Hematologic malignancy and transplant patients with prolonged neutropenia and antibiotic resistant fevers are particularly at risk. C. albicans and C. tropicalis are medically the most important, although other species do cause disease, but less frequently.

Developmental and molecular biology of switching in Candida albicans

Candida albicans and related species switch frequently and reversibly between a number of general phenotypes usually discriminated by colony morphology and in some cases by cellular morphology. Switching has been demonstrated to affect a number of physiologic and architectural characteristics of single cells including most of the putative virulence factors of C.albicans. In the past few years, we have cloned several genes regulated by switching in the white-opaque transition of C.albicans strain WO-1. Two of the genes, PEP1 and Op4, are transcribed only in the opaque phase, and one of the genes, Wh11, is transcribed only in the white phase. These coordinately regulated genes are unlinked in the genome and do not undergo sequence reorganization in switching. With the identification of a cis-acting regulatory sequence in the five-prime flanking sequence of Wh11, we now believe that phase-specific genes are regulated by transacting factors and that these factors may be coded for or under the direct regulation of a single master regulatory gene at which site the basic switch event occurs.

Partial nucleotide sequence of a single ribosomal RNA coding region and secondary structure of the large subunit 25 s rRNA of Candida albicans

A rDNA cistron of Candida albicans strain WO-1 was cloned and the ITS1, ITS2, 5.8 s rDNA and 25 s rDNA coding regions sequenced in their entirety. These sequences were compared to those of three related yeast species (Saccharomyces cerevisiae, Saccharomyces carlsbergensis, and Thermomyces lanuginosus), and the 5.8 s rDNA was compared to seven additional 5.8 s rDNAs from organisms ranging in complexity from D. discoideum to H. sapiens. The C. albicans ITS regions are shorter than those of most other eukaryotes. The 25 s and 5.8 s rDNA sequences were folded into a secondary structure model based on comparative methods. In a comparison of regional similarities between the large subunit rDNAs of C. albicans, the three related yeasts and other eukaryotes, it is demonstrated that the additional sequences not present in the E. coli 23 s rDNA are more variable than the regions present in both prokaryotes and eukaryotes.