CLONING OF A CRYPTOCOCCUS-NEOFORMANS GENE, GPA1, ENCODING A G-PROTEINALPHA-SUBUNIT HOMOLOG

We have isolated a gene, GPA1, from Cryptococcus neoformans by the PCR technique. DNA sequencing of the GPA1 clone suggested that it encodes a protein homologous to the G-protein ar-subunit family. Comparison o f the deduced amino acid sequence of the GPA1-encoded protein revealed that it is about 45% identical to several mammalian Gi alpha subunits and 48% identical to the Co protein Gpa2 from Saccharomyces cerevisia e. Co proteins are known to be involved in mating of other yeasts, suc h as S. cerevisiae and Schizosaccharomyces pombe. Southern analysis de monstrated that GPA1 is present in a single copy within the Cryptococc us genome. Isolation of the cDNA for GPA1 confirmed that the gene cont ains six introns within the coding region. The GPA1 transcript was ide ntified by Northern (RNA) analysis as a 1.6-kb RNA present in exponent ially growing cells of both the alpha and a mating types. Moreover, th e abundance of this transcript increased in cells shifted to starvatio n medium. Coincubation of alpha and a cells on starvation medium is re quired for mating of cryptococcal cells. Thus, our results are consist ent with the involvement of C. neoformans GPA1 in mating.