Abstract
Heterotrimeric G-proteins are key elements of signal transduction pathways, which participate in regulating multiple biological processes in fungi including growth, conidiation, antagonism, and mycoparasitism. Among G protein subunits, Gα3 showed diverse regulatory functions in different fungi. In this study, we cloned a Gα3 subunit coding gene thga3 from T. harzianum Th33 that can antagonize Rhizoctonia solani and some other plant pathogenic fungi. A thga3 deletion strain Δthga3 was generated using the double-crossover homologous recombination strategy, and Rthga3 was generated by transforming thga3-expressing vector into the protoplasts of Δthga3 by the PEG/CaCl2-mediated method. The biological characteristics of wild-type Th33, Δthga3 and Rthga3 were evaluated. Compared with wild-type Th33, Δthga3 showed 15%, 94%, and 23% decrease in hyphal growth, conidia yield, and chitinase activity, respectively, and Δthga3 showed lower antagonistic and mycoparasitism abilities, while there were no significant differences between wild-type Th33 and Rthga3. The hyphal surface hydrophobicity of Δthga3 significantly decreased compared with those of the wild-type Th33 and Rthga3. qRT-PCR analysis revealed that transcript abundance of the hydrophobin gene (tha_09745) of Δthga3 decreased by 80% compared with that of wild-type Th33 and Rthga3. The results showed that thga3 positively regulates the growth, conidiation, hydrophobicity, chitinase activities, and mycoparasitism of Th33 towards R. solani. We hence deduced that the expression level of Tha_09745 is correlated to the hyphal hydrophobicity of Th33 and therefore affects the other biological characteristics of Th33. The findings of this report provide a foundation for elucidating the G-protein signal regulatory mechanisms of fungi.
Highlights
Trichoderma spp. are widely used as biocontrol agents
Construction and validation of thga3 deleted and complemented mutants The Thga3 gene was cloned from wild-type Th33 genome, which contains six exons and five introns and encoding 355 amino acids
Gα3 subunits in T. viride, T. atroviride, and Rhodospirillum sp. negatively regulate conidiation (Schmoll et al 2009; Zeilinger et al 2005), and GanB (Gα3) in Aspergillus nidulans positively regulates conidial germination (Chang et al 2004)
Summary
Trichoderma spp. are widely used as biocontrol agents. Understanding their genetic regulation of biocontrol activities is beneficial to their genetic improvement and application as biofungicides. (Yu et al 2008; Guo et al 2016) These regulate the growth, conidiation, cellulase, and chitinase activities of fungi. GNA3 (Gα3) from T. reesei (do Nascimento et al 2009) and GNA3 (Gα3) in T. reesei (Schmoll et al 2009) are related to cellulase activity, while Tga (Gα3) in T. viride affects chitinase gene expression (Susanne et al 2005) and Tga (Gα3) in T. atroviride and GNA3 (Gα3) in T. viride negatively regulate conidiation. Light seems to play a role in regulating functions of Gα3 subunits in Trichoderma (Schmoll et al 2009; Susanne et al 2005). Studying the functions of Ga3 subunits of different Trichoderma strains may further clarify their functions and regulatory mechanisms
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