The Heterotrimeric G-Protein Subunits GNG-1 and GNB-1 Form a Gβγ Dimer Required for Normal Female Fertility, Asexual Development, and Gα Protein Levels in Neurospora crassa

Abstract

We have identified a gene encoding a heterotrimeric G protein γ subunit, gng-1 , from the filamentous fungus Neurospora crassa. gng-1 possesses a gene structure similar to that of mammalian Gγ genes, consisting of three exons and two introns, with introns present in both the open reading frame and 5′-untranslated region. The GNG-1 amino acid sequence displays high identity to predicted Gγ subunits from other filamentous fungi, including Giberella zeae , Cryphonectria parasitica , Trichoderma harzianum , and Magnaporthe grisea . Deletion of gng-1 leads to developmental defects similar to those previously characterized for Δ gnb-1 (Gβ) mutants. Δ gng-1 , Δ gnb-1 , and Δ gng-1 Δ gnb-1 strains conidiate inappropriately in submerged cultures and are female sterile, producing aberrant female reproductive structures. Similar to previous results obtained with Δ gnb-1 mutants, loss of gng-1 negatively influences levels of Gα proteins (GNA-1, GNA-2, and GNA-3) in plasma membrane fractions isolated from various tissues of N. crassa and leads to a significant reduction in the amount of intracellular cyclic AMP. In addition, we show that GNB-1 is essential for maintenance of normal steady-state levels of GNG-1, suggesting a functional interaction between GNB-1 and GNG-1. Direct evidence for a physical association between GNB-1 and GNG-1 in vivo was provided by coimmunoprecipitation.