Originally published as Genetics Published Articles Ahead of Print on August 24, 2008.

Genetics, Vol. 180, 191-206, September 2008, Copyright © 2008
doi:10.1534/genetics.108.091603

Three {alpha}-Subunits of Heterotrimeric G Proteins and an Adenylyl Cyclase Have Distinct Roles in Fruiting Body Development in the Homothallic Fungus Sordaria macrospora

Jens Kamerewerd*,1, Malin Jansson*,1, Minou Nowrousian*, Stefanie Pöggeler{dagger} and Ulrich Kück*,2

* Lehrstuhl für Allgemeine und Molekulare Botanik, Ruhr-Universität Bochum, 44780 Bochum, Germany and {dagger} Institut für Mikrobiologie und Genetik, Abteilung Genetik Eukaryotischer Mikroorganismen, Georg-August-Universität, 37077 Göttingen, Germany

2 Corresponding author: Ruhr-Universität Bochum, Lehrstuhl für Allgemeine und Molekulare Botanik, ND7/131, Universitätsstraße 150, 44780 Bochum, Germany.
E-mail: ulrich.kueck{at}rub.de

Sordaria macrospora, a self-fertile filamentous ascomycete, carries genes encoding three different {alpha}-subunits of heterotrimeric G proteins (gsa, G protein Sordaria alpha subunit). We generated knockout strains for all three gsa genes ({Delta}gsa1, {Delta}gsa2, and {Delta}gsa3) as well as all combinations of double mutants. Phenotypic analysis of single and double mutants showed that the genes for G{alpha}-subunits have distinct roles in the sexual life cycle. While single mutants show some reduction of fertility, double mutants {Delta}gsa1{Delta}gsa2 and {Delta}gsa1{Delta}gsa3 are completely sterile. To test whether the pheromone receptors PRE1 and PRE2 mediate signaling via distinct G{alpha}-subunits, two recently generated {Delta}pre strains were crossed with all {Delta}gsa strains. Analyses of the corresponding double mutants revealed that compared to GSA2, GSA1 is a more predominant regulator of a signal transduction cascade downstream of the pheromone receptors and that GSA3 is involved in another signaling pathway that also contributes to fruiting body development and fertility. We further isolated the gene encoding adenylyl cyclase (AC) (sac1) for construction of a knockout strain. Analyses of the three {Delta}gsa{Delta}sac1 double mutants and one {Delta}gsa2{Delta}gsa3{Delta}sac1 triple mutant indicate that SAC1 acts downstream of GSA3, parallel to a GSA1–GSA2-mediated signaling pathway. In addition, the function of STE12 and PRO41, two presumptive signaling components, was investigated in diverse double mutants lacking those developmental genes in combination with the gsa genes. This analysis was further completed by expression studies of the ste12 and pro41 transcripts in wild-type and mutant strains. From the sum of all our data, we propose a model for how different G{alpha}-subunits interact with pheromone receptors, adenylyl cyclase, and STE12 and thus cooperatively regulate sexual development in S. macrospora.