Heterotrimeric guanine nucleotide-binding regulatory proteins (G proteins), which consist of three subunits, a, b, and g, are involved in a variety of receptor-mediated signaling systems. Neurospora crassa contains two Ga subunit genes (gna-1+, and gna-2+). The predicted Gna2 protein sequence is 49% and 48% identical to the Gi-a homologue Gna-1 from N. crassa and the Gpa1 a subunit from Schizosaccharomyces pombe, respectively. In the upstream region of gna-2+, there are several potential promoter sequences, including a TATA and CAAT box, and several CTTTG (or CAAAG) consensus DNA sequences for a mating-type regulation in N. crassa. The 5Õ end of the cDNA is at position ø582 (position +1 is based on translation start site). Verification that this is the true mRNA start site is being analyzed using primer extension. The four introns in the gna-2+ coding region are in positions conserved in the Gi family of a subunits in mammals, but the gna-2+ gene does not belong to any of the known classes described in mammals based on amino acid identity. Deletion mutations have been made in the gna-2+ gene by replacing the promoter and coding region of gna-2+ with a selectable marker. Homokaryotic strains containing nuclei with only the gna-2 mutation can not live. The gna-2 RIP mutation was also created by crossing a strain with duplication of the gna-2+ gene to a wild type strain. However, the RIP mutation could only be recovered in the extra ectopic copy of gna-2+; the normal copy was always wild type. These preliminary results suggest that the gna-2+ gene is essential for vegetative growth in N. crassa.
Characterization of gna-2+, a G-Protein a Subunit Gene from Neurospora crassa