title: Site-directed mutagenesis of the Arabidopsis heterotrimeric G protein β subunit suggests divergent mechanisms of effector activation between plant and animal G proteins.
abstract: Heterotrimeric G proteins are integral components of signal transduction in humans and other mammals and have been therefore extensively studied. However, while they are known to mediate many processes, much less is currently known about the effector pathways and molecular mechanisms used by these proteins to regulate effectors in plants. We designed a complementation strategy to study G protein signaling in Arabidopsis thaliana, particularly the mechanism of action of AGB1, the sole identified β subunit. We used biochemical and effector regulation data from human G protein studies to identify four potentially important residues for site-directed mutagenesis (T65, M111, D250 and W361 of AGB1). Each residue was individually mutated and the resulting mutated protein introduced in the agb1-2 mutant background under the control of the native AGB1 promoter. Interestingly, even though these mutations have been shown to have profound effects on effector signaling in humans, all the mutated subunits were able to restore thirteen of the fifteen Gβ-deficient phenotypes characterized in this study. Only one mutated protein, T65A was unable to complement the hypersensitivity to mannitol during germination observed in agb1 mutants; while only D250A failed to restore lateral root numbers in the agb1 mutant to wild-type levels. Our results suggest that the mechanisms used in mammalian G protein signaling are not well conserved in plant G protein signaling, and that either the effectors used by plant G proteins, or the mechanisms used to activate them, are at least partially divergent from the well-studied mammalian G proteins.
authors: Plant Genetic Engineering Laboratory, School of Agriculture and Food Sciences, University of Queensland, Brisbane, QLD 4072, Australia.
authors: David
authors: D
authors: Chakravorty
authors: Yuri
authors: Y
authors: Trusov
authors: José Ramón
authors: JR
authors: Botella
mesh_terms: D000818:Animals; D017360:Arabidopsis / Q000235:genetics / Q000378:metabolism* / Q000382:microbiology; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism*; D005670:Fusarium / Q000502:physiology; D044387:GTP-Binding Protein beta Subunits / Q000235:genetics / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D030821:Plants, Genetically Modified / Q000235:genetics / Q000378:metabolism / Q000382:microbiology; D015398:Signal Transduction / Q000235:genetics / Q000502:physiology
chemical_list: C089590:AGB1 protein, Arabidopsis; D029681:Arabidopsis Proteins; D044387:GTP-Binding Protein beta Subunits; D019204:GTP-Binding Proteins