Over-expression of a truncated Arabidopsis thaliana heterotrimeric G protein γ subunit results in a phenotype similar to α and β subunit knockouts

Heterotrimeric G proteins (G-proteins) are a diverse class of signal transducing proteins which have been implicated in a variety of important roles in plants. When G-proteins are activated, they dissociate into two functional subunits (a and the beta gamma, dinner) that effectively relay the signal to a multitude of effectors. In animal systems, the beta gamma dimer is anchored to the plasma membrane by a prenyl group present in the gamma subunit and membrane localization has proven vital for heterotrimer function. A semi-dominant negative strategy was designed aiming to disrupt heterotrimer function in Arabidopsis thaliana (ecotype Columbia) plants by over-expressing a truncated y subunit lacking the isoprenylation motif (gamma*). Northern analysis shows that the levels of expression of the mutant -y subunit in several transgenic lines (35S-gamma*) are orders of magnitude higher than that of the native subunits. In-depth characterization of the 35S-gamma* lines has been carried out, specifically focusing on a number of developmental characteristics and responses to several stimuli previously shown to be affected in alpha- and [beta-deficient mutants. In all cases, the transgenic lines expressing the mutant gamma subunit behave in the same way as the alpha- and/or the beta-deficient mutants, albeit with reduced severity of the phenotype. Our data indicates that signaling from both functional subunits, a and the beta/gamma dimer, is disrupted in the transgenic plants. Even though physical association of the subunits has been previously reported, our research provides evidence of the functional association of alpha and beta with the gamma subunits in Arabidopsis, while also suggesting that plasma membrane localization may be critical for function of plant heterotrimeric G proteins. (C) 2007 Elsevier B.V. All rights reserved.