Regulator of G Protein Signaling Proteins Control Growth, Development and Cellulase Production in Neurospora crassa

Abstract

Heterotrimeric (αβγ) G protein signaling pathways are critical environmental sensing systems found in eukaryotic cells. Exchange of GDP for GTP on the Gα subunit leads to its activation. In contrast, GTP hydrolysis on the Gα is accelerated by Regulator of G protein Signaling (RGS) proteins, resulting in a return to the GDP-bound, inactive state. Here, we analyzed growth, development and extracellular cellulase production in strains with knockout mutations in the seven identified RGS genes (rgs-1 to rgs-7) in the filamentous fungus, Neurospora crassa. We compared phenotypes to those of strains with either knockout mutations or expressing predicted constitutively activated, GTPase-deficient alleles for each of the three Gα subunit genes (gna-1Q204L, gna-2Q205L or gna-3Q208L). Our data revealed that six RGS mutants have taller aerial hyphae than wild type and all seven mutants exhibit reduced asexual sporulation, phenotypes shared with strains expressing the gna-1Q204L or gna-3Q208L allele. In contrast, Δrgs-1 and Δrgs-3 were the only RGS mutants with a slower growth rate phenotype, a defect in common with gna-1Q204L strains. With respect to female sexual development, Δrgs-1 possessed defects most similar to gna-3Q208L strains, while those of Δrgs-2 mutants resembled strains expressing the gna-1Q204L allele. Finally, we observed that four of the seven RGS mutants had significantly different extracellular cellulase levels relative to wild type. Of interest, the Δrgs-2 mutant had no detectable activity, similar to the gna-3Q208L strain. In contrast, the Δrgs-1 and Δrgs-4 mutants and gna-1Q204L and gna-2Q205L strains exhibited significantly higher cellulase activity than wild type. With the exception of sexual development, our results demonstrate the greatest number of genetic interactions between rgs-1 and gna-1 and rgs-2 and gna-3 in N. crassa.

Mechanistic insights into the role of Ca2+-stimulated AMPK in the secretion of cellulases during carbon stress

“…Another recent study highlighted the role of GNA3 as well as regulator of G-protein signaling, RGS2 in extracellular cellulase production in N. crassa 29 . Furthermore, studies on Aspergillus nidulans and N. crassa identified the activation of the HOG1 MAPK pathway during cellulase production 30,31 .…”

“…Additionally, G protein receptors relay cellulase inducing signal by activating Gα subunits, GNA1/3, and elevate cAMP levels to activate Protein kinase A (PKA) pathway in T. reesei 27,28 . Another recent study highlighted the role of GNA3 as well as regulator of G-protein signaling, RGS2 in extracellular cellulase production in N. crassa 29 . Furthermore, studies on Aspergillus nidulans and N. crassa identified the activation of the HOG1 MAPK pathway during cellulase production 30,31 .…”

Phosducin-like protein PoPlp1 impacts cellulase and amylase expression and development in Penicillium oxalicum via the G protein–cAMP signaling pathway

“…The efficient synthesis of cellulase and several developmental processes (e.g., mycelial growth and conidiation) are co-regulated synergistically by multiple transcription factors in P. oxalicum (Li et al, 2015(Li et al, , 2020Guo et al, 2021;Xu et al, 2022). The regulation of these processes in filamentous fungi is usually triggered by environmental signals (i.e., soluble carbon sources and light), which act on target genes through an intracellular signaling pathway, i.e., the G protein-cAMP signaling pathway (Hu et al, 2013;Cabrera et al, 2022;Schmoll and Hinterdobler, 2022).…”

“…The G protein-cAMP signaling pathway is involved in multiple biological processes, such as development, morphogenesis, induced enzyme synthesis, and secondary metabolism (Lafon et al, 2006;Hinterdobler et al, 2020;Cabrera et al, 2022). Heterotrimeric G proteins (consisting of α, β, and γ subunits), G-protein-coupled receptors (GPCRs), regulators of G protein signaling (RGS), the secondary messenger cAMP, and cAMP-dependent protein kinase (PKA) all play pivotal roles in the G protein-cAMP signaling pathway.…”