A G-Protein β Subunit Required for Sexual and Vegetative Development and Maintenance of Normal Gα Protein Levels in Neurospora crassa

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Abstract
The genome of the filamentous fungus Neurospora crassa contains a single gene encoding a heterotrimeric G-protein β subunit, gnb-1 . The predicted GNB-1 protein sequence is most identical to Gβ proteins from the filamentous fungi Cryphonectria parasitica and Aspergillus nidulans . N. crassa GNB-1 is also 65% identical to the human GNB-1 protein but only 38 and 45% identical to Gβ proteins from budding and fission yeasts. Previous studies in animal and fungal systems have elucidated phenotypes of Gβ null mutants, but little is known about the effects of Gβ loss on Gα levels. In this study, we analyzed a gnb-1 deletion mutant for cellular phenotypes and levels of the three Gα proteins. Δ gnb-1 strains are female-sterile, with production of aberrant fertilized reproductive structures. Δ gnb-1 strains conidiate more profusely and have altered mass on solid medium. Loss of gnb-1 leads to inappropriate conidiation and expression of a conidiation-specific gene during growth in submerged culture. Intracellular cyclic AMP levels are reduced by 60% in vegetative plate cultures of Δ gnb-1 mutants. Loss of gnb-1 leads to lower levels of the three Gα proteins under a variety of conditions. Analysis of transcript levels for the gna-1 and gna-2 Gα genes in submerged cultures indicates that regulation of Gα protein levels by gnb-1 is posttranscriptional. The results suggest that GNB-1 directly regulates apical extension rate and mass accumulation. In contrast, many other Δ gnb-1 phenotypes, including female sterility and defective conidiation, can be explained by altered levels of the three N. crassa Gα proteins.