Characterization of the MFalpha pheromone of the human fungal pathogen cryptococcus neoformans.
Cryptococcus neoformans is an important human pathogenic fungus with a defined sexual cycle and well-developed molecular and genetic approaches. C. neoformans is predominantly haploid and has two mating types, MATa and MATalpha. Mating is known to be regulated by nutritional limitation and thought also to be regulated by pheromones. Previously, a portion of the MATalpha locus was cloned, and a presumptive pheromone gene, MFalpha1, was identified by its ability to induce conjugation tube-like filaments when introduced by transformation into MATa cells. Here, the ability of the MFalpha1 gene to induce these morphological changes in MATa cells was used as a phenotypic assay to perform a structure-function analysis of the gene. We show that the MFalpha1 open reading frame is required for the morphological response of MATa cells. We also find that the cysteine residue of the C-terminal CAAX motif is required for activity of the MFalpha1 pheromone. In addition, we use a reporter system to measure the expression levels of the MFalpha1 pheromone gene and find that two signals, nutrient starvation and the presence of factors secreted by mating partner cells, impinge on this promoter and regulate MFalpha1 expression. We identify a second pheromone gene, MFalpha2, and show phenotypically that this gene is also expressed. Finally, we have synthesized the MFalpha1 pheromone and show that only the predicted mature modified form of the alpha-factor peptide triggers morphological responses in MATa cells.
Davidson
R C
RC
Departments of Genetics, Pharmacology and Cancer Biology, Biochemistry, Microbiology and Medicine, and the Howard Hughes Medical Institute, 322 Carl Building, Box 3546, Duke University Medical Center, Durham, NC 27710, USA.
Moore
T D
TD
Odom
A R
AR
Heitman
J
J
D003455:Cryptococcus neoformans / Q000235:genetics* / Q000502:physiology; D003470:Culture Media; D004271:DNA, Fungal; D006801:Humans; D000072235:Mating Factor; D016366:Open Reading Frames; D010455:Peptides / Q000737:chemistry / Q000235:genetics*; D010675:Pheromones / Q000737:chemistry / Q000235:genetics*; D016133:Polymerase Chain Reaction; D013329:Structure-Activity Relationship
D003470:Culture Media; D004271:DNA, Fungal; D010455:Peptides; D010675:Pheromones; D000072235:Mating Factor