Multiple functions of mfa-1, a putative pheromone precursor gene of Neurospora crassa.
A putative pheromone precursor gene of Neurospora crassa, mfa-1 (which encodes mating factor a-1), was identified as the most abundant clone in starved mycelial and perithecial cDNA libraries. Northern analysis demonstrated high mfa-1 expression in all mating type a tissues and suggested low expression levels in mat A tissues. The mfa-1 gene was expressed as an approximately 1.2-kb transcript predicted to encode a 24-residue peptide, followed by a long 3' untranslated region (3' UTR). The predicted MFA1 sequence showed 100% sequence identity to PPG2 of Sordaria macrospora and structural similarity (a carboxy-terminal CAAX motif) to many hydrophobic fungal pheromone precursors. Mutants with a disrupted open reading frame (ORF) in which the critical cysteine residue had been changed to a nonprenylatable residue, tyrosine (YAAX mutants), were isolated, as were mfa-1 mutants with intact ORFs but multiple mutations in the 3' noncoding region (CAAX mutants). The 3' UTR is required for the full range of mfa-1 gene activity. Both classes of mutants showed delayed and reduced vegetative growth (which was suppressed by supplementation with a minute amount [30 micro M] of ornithine, citrulline, or arginine), as well as aberrant sexual development. When crossed as female parents to wild-type males, the CAAX and YAAX mutants showed greatly reduced ascospore production. No ascospores were produced in homozygous mfa-1 crosses. As males, YAAX mat a mutants were unable to attract wild-type mat A trichogynes (female-specific hyphae) or to initiate sexual development, while CAAX mat a mutants were able to mate and produce sexual progeny despite their inability to attract mat A trichogynes. In the mat A background, both CAAX and YAAX mutants showed normal male fertility but defective vegetative growth and aberrant female sexual development. Thus, the mfa-1 gene appears to have multiple roles in N. crassa development: (i) it encodes a hydrophobic pheromone with a putative farnesylated and carboxymethylated C-terminal cysteine residue, required by mat a to attract trichogynes of mat A; (ii) it is involved in female sexual development and ascospore production in both mating types; and (iii) it functions in vegetative growth of both mating types.
Kim
Hyojeong
H
Department of Biology, The University of New Mexico, Albuquerque, New Mexico 87131, USA.
Metzenberg
Robert L
RL
Nelson
Mary Anne
MA
D020413:3' Untranslated Regions; D001483:Base Sequence; D015139:Blotting, Southern; D002455:Cell Division; D002874:Chromosome Mapping; D003433:Crosses, Genetic; D003545:Cysteine / Q000737:chemistry; D018076:DNA, Complementary / Q000378:metabolism; D005656:Fungal Proteins / Q000737:chemistry* / Q000378:metabolism; D015723:Gene Library; D005800:Genes, Fungal; D049770:Genes, Mating Type, Fungal; D005816:Genetic Complementation Test; D008745:Methylation; D008969:Molecular Sequence Data; D009154:Mutation; D009492:Neurospora crassa / Q000235:genetics* / Q000502:physiology*; D009690:Nucleic Acid Conformation; D016366:Open Reading Frames; D010675:Pheromones / Q000737:chemistry* / Q000378:metabolism; D010957:Plasmids / Q000378:metabolism; D017354:Point Mutation; D012150:Polymorphism, Restriction Fragment Length; D017368:Protein Prenylation; D017434:Protein Structure, Tertiary; D017422:Sequence Analysis, DNA; D012689:Sequence Homology, Nucleic Acid
D020413:3' Untranslated Regions; D018076:DNA, Complementary; D005656:Fungal Proteins; D010675:Pheromones; D003545:Cysteine