Exploring Heterotrimeric G Protein Diversity in Plants: Identification of a Novel Third γ Subunit and Other Aspects of Gβγ Signalling in Arabidopsis

David Chakravorty (2011). Exploring Heterotrimeric G Protein Diversity in Plants: Identification of a Novel Third γ Subunit and Other Aspects of Gβγ Signalling in Arabidopsis PhD Thesis, School of Biological Sciences, The University of Queensland.

       
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Author David Chakravorty
Thesis Title Exploring Heterotrimeric G Protein Diversity in Plants: Identification of a Novel Third γ Subunit and Other Aspects of Gβγ Signalling in Arabidopsis
School, Centre or Institute School of Biological Sciences
Institution The University of Queensland
Publication date 2011-08
Thesis type PhD Thesis
Supervisor Prof Jimmy Botella
Prof Robert Birch
Total pages 135
Total colour pages 5
Total black and white pages 130
Language eng
Subjects 06 Biological Sciences
Abstract/Summary Heterotrimeric G protein complexity in mammalian systems is well established, with multiple family members identified for each subunit (α, β and γ). However, only one α, one β and two γ subunits had been isolated in the model plant Arabidopsis. Despite the seeming lack of complexity, a vast array of functions and phenotypes has been attributed to heterotrimeric G proteins in plants. This apparent contradiction has raised many questions as to the function of plant heterotrimeric G proteins, and the possible existence of previously unidentified subunits. The major part of the work herein details the identification and characterization of a novel Arabidopsis γ subunit (AGG3). AGG3 accounts for many ‘orphan’ phenotypes previously identified in mutants of the α (GPA1) and β (AGB1) subunits, that were not explained by the two known γ subunits (AGG1 and AGG2). However, AGG3 is far from a canonical γ subunit. AGG3 has unique characteristics never before observed in plant or animal systems. These include, increased size (more than twice the size of a typical γ subunit) and the presence of a C-terminal Cys-rich domain, with the γ-like domain located on the N-terminus of the protein. Our searches have shown that AGG3 type γ subunits are ubiquitous across the plant kingdom, however, despite the abundance of plant homologues we have been unable to identify any animal AGG3 homologues, suggesting that AGG3 is a unique component of plant G protein signaling. Interestingly, as G protein researchers have recently widened the scope of their investigations, several other novel G protein components specific to plants have been identified by other groups. These include; a transmembrane regulator of G protein signaling (RGS) protein, extra large α subunits that may bind to AGB1 under specific conditions, and two extremely novel G protein coupled receptor-type G proteins (GTGs) that bind to GPA1 but are GTP-binding proteins in their own right. Two additional projects were undertaken to compare plant and mammalian G protein mechanisms. The first analysed the role of the C-terminal isoprenylation motif of AGG1. Isoprenylation has been shown to be critical to the function of mammalian γ subunits, however little was known about plant γ subunit isoprenylation past the fact that the motif is conserved. A combination of biochemical work by other groups, and phenotypic analysis of a non-isoprenylated AGG1 subunit presented here, has demonstrated that isoprenylation and correct membrane localisation of AGG1 is also critical to plant G protein function. Secondly, residues of human β subunits that have been proven to be crucial for Gβγ effector regulation were identified. The corresponding conserved residues of the sole Arabidopsis β subunit, AGB1, were individually mutated and into the null mutant line agb1-2. The mutated subunits were able to restore the majority of agb1-2 phenotypes back to wild-type, suggesting the selected residues are not as critical for effector regulation in plants. It was therefore speculated that AGB1 may use different mechanisms to signal to effectors, or may even regulate a different subset of effectors and pathways to its mammalian counterparts. The results presented here contribute to the growing belief that plant and animal G proteins have both shared and distinct evolutionary pathways. While some basic principles such as a requirement for isoprenylation of γ subunits appear to be conserved, there are increasing examples of divergent G protein mechanisms and components. Such unique aspects build upon the level of G protein complexity evident in plants, and therefore begin to explain the great diversity of G protein related phenotypes that have been uncovered in Arabidopsis.
Keyword heterotrimeric G proteins
signal transduction
Arabidopsis thaliana
AGG3
AGB1
Additional Notes Colour pages: 78, 79, 86, 101 and 108

 
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Created: Tue, 15 Nov 2011, 02:12:21 EST by Mr David Chakravorty on behalf of Library - Information Access Service