Cloning and characterization of PRB1, a Candida albicans gene encoding a putative novel endoprotease B and factors affecting its expression

• Published in: Research in microbiology 2002-11, Vol.153 (9), p.611-620
• Main Authors: Orozco, Iranzu, Ortiz, Lucila, Elorza, Maria-Victoria, Ruiz-Herrera, José, Sentandreu, Rafael
• Format: Article
• Language: English
• Subjects: Amino Acid Sequence; Base Sequence; Biological and medical sciences; Blotting, Southern; Candida albicans; Candida albicans - enzymology; Candida albicans - genetics; Chromosomes, Fungal - genetics; Cloning, Molecular; Differential display; DNA, Fungal - chemistry; DNA, Fungal - genetics; Electrophoresis, Gel, Pulsed-Field; Factors controlling expression of Ca PRB1; Fundamental and applied biological sciences. Psychology; Fungal Proteins - biosynthesis; Fungal Proteins - chemistry; Fungal Proteins - genetics; Gene Expression Profiling; Gene Expression Regulation, Fungal - genetics; Gene Expression Regulation, Fungal - physiology; Gene Library; Generation of PRB1 by PCR; Genes. Genome; Humans; Molecular and cellular biology; Molecular genetics; Molecular Sequence Data; Mutagenesis; Overexpressing prb1 strains; PRB1 null mutants, construction; Sequence Alignment; Sequence Analysis, DNA; Serine Endopeptidases - biosynthesis; Serine Endopeptidases - chemistry; Serine Endopeptidases - genetics
• ISSN: 0923-2508; 1769-7123
• DOI: 10.1016/S0923-2508(02)01373-6

Several cDNA fragments corresponding to transcripts differentially expressed under conditions that favor mycelial growth of Candida albicans were identified by the “differential display” technique. One of these was cloned and used as a probe to rescue the full gene from a genomic library of the fungus. The sequence identified a single, uninterrupted open reading frame of 1395 nucleotides encoding a putative protein of 465 residues and a theoretical molecular weight of 50.3 kDa, present in the genome as a single copy located at chromosome 2 in different strains. The gene product showed high homology with subtilisin-like proteases, mainly PRB1, the vacuolar B protease from Saccharomyces cerevisiae, and for this reason it was designated as a putative CaPRB1. Expression of the gene was not directly related to fungal morphogenesis, but to the initial response to inducers: Heat shock and the presence of N-acetyl glucosamine. It was also subject to nitrogen, but not to carbon catabolite repression, although glucose inhibited the GlcNAc stimulatory effect. The gene was, in our hands, unable to complement PRB1 mutation in S. cerevisiae. C. albicans prb null mutants did not show any distinct alteration in the phenotype. CaPRB1 is the first gene coding for a putative vacuolar serine protease cloned from C. albicans.