Heterotrimeric G protein are involved in the regulation of multiple agronomic traits and stress tolerance in rice

Abstract

Background: The heterotrimeric G protein complex, consisting of Gα, Gβ, and Gγ subunits, are conserved signal transduction mechanism in eukaryotes. Recent molecular researches had demonstrated that G protein signaling participates in the regulation of yield related traits. However, the effects of G protein genes on yield components and stress tolerance are not well characterized.

Results: In this study, we generated heterotrimeric G protein mutants in rice using CRISPR/Cas9 (Clustered Regularly Interspaced Short Palindromic Repeats) gene-editing technology. The effects of heterotrimeric G proteins on the regulation of yield components and stress tolerance were investigated. The mutants of gs3 and dep1 generated preferable agronomic traits compared to the wild-type, whereas the mutants of rga1 showed an extreme dwarf phenotype, which led to a dramatic decrease in grain production. The mutants showed improved stress tolerance, especially under salinity treatment. We found four putative extra-large G proteins (PXLG)1-4 that also participate in the regulation of yield components and stress tolerance. A yeast two hybrid showed that the RGB1 might interact with PXLG2 but not with PXLG1, PXLG3 or PXLG4.

Conclusion: These findings will not only improve our understanding of the repertoire of heterotrimeric G proteins in rice but also contribute to the application of heterotrimeric G proteins in rice breeding.

Keywords: CRISPR/Cas9; Heterotrimeric G protein; Rice; Stress tolerance; Yield components.

Fig. 1

The schematic representation of the heterotrimeric G proteins and the sequences of the mutants generated by CRISPR/Cas9 gene editing. a The schematic representation of the heterotrimeric Gα and Gβ proteins in rice. The functional domains are shown in different colors. The black arrow indicates the position of the sgRNA. The number indicates the amino acid sequence. b The protein schematic representation of the heterotrimeric Gγ proteins in rice. The functional domains are shown in different colors. The black arrow indicates the position of the sgRNA. The number indicates the amino acid sequence. c The sequences of the heterotrimeric G protein mutants generated by CRISPR/Cas9 gene editing

Fig. 2

Genetic effects of the heterotrimeric G protein on plant architecture, panicle size, grain size, and yield components. a The whole plants of the heterotrimeric G protein mutants. Bar = 10 cm (b) The panicle of the heterotrimeric G protein mutants. Bar = 1 cm (c) The grains of the heterotrimeric G protein mutants. Bar = 1 cm (d–i) The plant heights, heading times, panicle numbers, grain numbers per panicle, 1000-grain weight, and setting rates of the heterotrimeric G protein mutants. * indicates significance at the 5% level

Fig. 3

The stress tolerance of the heterotrimeric G protein mutants. a The non-treatment control of WT and heterotrimeric G protein mutants. b–d Photos of the plants treated with drought, chilling, and salinity stress. e–g The survival rates after drought, chilling, and salinity treatment. Bar = 10 cm, * indicates significance at the 5% level

Fig. 4

The putative extra-large Gα protein in rice. a A phylogenetic analysis of the extra-large G protein in Arabidopsis and rice, in which identical and conserved residues are indicated by different colors. b The schematic representation of the putative extra-large Gα protein in rice. The black arrow indicates the position of the sgRNA. c The sequence of the putative extra-large Gα protein mutant generated by CRISPR/Cas9 gene editing

Fig. 5

Genetic effects of the putative extra-large Gα protein on plant architecture, panicle size, grain size, and yield components. a The whole plants of the putative extra-large Gα protein mutants. Bar = 10 cm. b The panicles of the putative extra-large Gα protein mutants. Bar = 1 cm. c The grains of the putative extra-large Gα protein mutants. Bar = 1 cm. d–i The plant heights, heading times, panicle numbers, grain numbers per panicle, 1000-grain weight, and setting rates of the putative extra-large Gα protein mutants. * indicates significance at the 5% level

Fig. 6

The stress tolerance of the putative extra-large Gα protein mutants. a The non-treatment control of the WT and putative extra-large Gα protein mutants. b–d Photos of plants treated with drought, chilling, and salinity stress. e–g The survival rates after drought, chilling, and salinity stress. Bar = 10 cm. * indicates significance at the 5% level

Fig. 7

Interaction of RGB1 with PXLGs. In the yeast two hybrid assay, RGB1 is used as a prey (AD), and the PXLGs are used as bait (BD) (a) The corresponding positions in the self-activation test and yeast two hybrid assay. b The result of self-activation test and yeast two hybrid assay. Two plasmids containing either an AD or BD construct were introduced into a yeast strain and transformants were grown on selective medium lacking Leu and Trp. pGBKT7-p53 + pGADT7-T was used as a positive control, and pGBKT7-p53 + pGADT7-LaminA was used as a negative control