G Protein activation without subunit dissociation depends on a Galpha(i)-specific
region
J Biol Chem, 280 (26):
24584-90 (July 2005)Frank, Monika Thumer, Leonore Lohse, Martin J Bunemann, Moritz United
States The Journal of biological chemistry J Biol Chem. 2005 Jul
1;280(26):24584-90. Epub 2005 May 2..Abstract
G proteins transmit a variety of extracellular signals into intracellular
responses. The Galpha and Gbetagamma subunits are both known to regulate
effectors. Interestingly, the Galpha subunit also determines subtype
specificity of Gbetagamma effector interactions. However, in light
of the common paradigm that Galpha and Gbetagamma subunits dissociate
during activation, a plausible mechanism of how this subtype specificity
is generated was lacking. Using a fluorescence resonance energy transfer
(FRET)-based assay developed to directly measure mammalian G protein
activation in intact cells, we demonstrate that fluorescent Galpha(i1,2,3),
Galpha(z), and Gbeta(1)gamma(2) subunits do not dissociate during
activation but rather undergo subunit rearrangement as indicated
by an activation-induced increase in FRET. In contrast, fluorescent
Galpha(o) subunits exhibited an activation-induced decrease in FRET,
reflecting subunit dissociation or, alternatively, a distinct subunit
rearrangement. The alpha(B/C)-region within the alpha-helical domain,
which is much more conserved within Galpha(i1,2,3) and Galpha(z)
as compared with that in Galpha(o), was found to be required for
exhibition of an activation-induced increase in FRET between fluorescent
Galpha and Gbetagamma subunits. However, the alpha(B/C)-region of
Galpha(il) alone was not sufficient to transfer the activation pattern
of Galpha(i) to the Galpha(o) subunit. Either residues in the first
91 amino acids or in the C-terminal remainder (amino acids 93-354)
of Galpha(il) together with the alpha(B/C)-helical region of Galpha(i1)
were needed to transform the Galpha(o)-activation pattern into a
Galpha(i1)-type of activation. The discovery of subtype-selective
mechanisms of G protein activation illustrates that G protein subfamilies
have specific mechanisms of activation that may provide a previously
unknown basis for G protein signaling specificity.