THESIS
2009
xv, 122 p. : ill. ; 30 cm
Abstract
Heterotrimeric G proteins regulate multiple effectors of which some are mediated via the Gβγ dimers. There is evidence to suggest that the functions of Gβγ dimers are not shared by all possible permutations of Gβγ complexes. The aim of this research is to define the formation of distinct Gβγ dimers, their functional differences in activating downstream effectors such as phospholipase Cβ (PLCβ) isoforms, the molecular determinant of Gβγ formation and the dissociation of different G protein family members upon activation. Co-immunoprecipitation assays using COS7 cells transiently expressing 48 different combinations of Gβ
(1-4) and Gγ
1-5, 7-13) subunits showed that Gβ
1 and Gβ
4 could form dimers with all known Gγ subunits, whereas several dimers could not be observed for Gβ
2 and Gβ
3. All Gβ...[
Read more ]
Heterotrimeric G proteins regulate multiple effectors of which some are mediated via the Gβγ dimers. There is evidence to suggest that the functions of Gβγ dimers are not shared by all possible permutations of Gβγ complexes. The aim of this research is to define the formation of distinct Gβγ dimers, their functional differences in activating downstream effectors such as phospholipase Cβ (PLCβ) isoforms, the molecular determinant of Gβγ formation and the dissociation of different G protein family members upon activation. Co-immunoprecipitation assays using COS7 cells transiently expressing 48 different combinations of Gβ
(1-4) and Gγ
1-5, 7-13) subunits showed that Gβ
1 and Gβ
4 could form dimers with all known Gγ subunits, whereas several dimers could not be observed for Gβ
2 and Gβ
3. All Gβ
1γ and Gβ
2γ dimers significantly stimulated PLCβ isoforms (PLCβ
2 ≥ PLCβ
3 > PLCβ
1), but Gβ
3γ and Gβ
4γ dimers were poor activators of PLCβ
1 and exhibited preference for PLCβ
3 and PLCβ
2, respectively. Mutation of certain residues in Gβ
3 enhanced PLCβ
2-activating functions when co-expressed with different Gγ subunits, while the corresponding stimulation on PLCβ
3 was not altered. All these results show that the exact composition of a Gβγ dimer can determine its selectivity for activating PLCβ isoforms, and certain residues in Gβ
3 may account for the preferential stimulation of PLCβ
3 by Gβ
3γ dimers. Apart from the N-terminal α helix and blades 5 – 7 of Gβ subunits, the region of blades 1 – 4 also play a minor but significant role in Gβγ dimer formation. Furthermore, different from the classical model of G protein activation upon receptor-mediated activation, some activated Gα subunits, including Gα
q, Gα
11, Gα
14, Gα
16, Gα
z, Gα
t1, Gα
t2, Gα
s, Gα
olf, Gα
12 and Gα
13, may remain associate with Gβγ dimer upon G protein activation, indicating that a new model of G protein activation mechanism is required for replacement of the classical model in which G protein activation is always coupled with subunit dissociation.
Post a Comment