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Maxim Sokolov, Satyabrata Sinha, Marycharmain Belcastro, Xueli Gao; Addressing the Role of PhLP in the Folding and Assembly of G-protein βγ dimer. Invest. Ophthalmol. Vis. Sci. 2013;54(15):1225.
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© ARVO (1962-2015); The Authors (2016-present)
To get better understanding of the roles of the cytosolic chaperonin containing T-complex protein 1 (CCT) and its co-factor phosducin-like protein (PhLP) in the folding and assembly of the heterotrimeric G-protein βγ subunits complex.
The interaction of PhLP and its short splice isoform (PhLPs) with CCT and G protein subunits was studied in both mouse photoreceptors and cell culture. The levels of mRNA and proteins were determined by quantitative real-time PCR and Western blotting, respectively. The assembly of epitope-tagged Gβ1 and Gγ1 subunits was monitored using a pull down assay.
We found that both PhLP and PhLPs form a complex with CCT, however, only PhLP facilitated the assembly of the Gβ1γ1 dimer. Interestingly, while doing so, PhLP interacted primarily with Gβ1 and not with the Gβ1γ1 complex. In contrast, PhLPs, which lacks an important Gβ-binding domain, showed a tendency to form a stable and apparently inactive tertiary complex with both CCT and the Gβ1 trapped inside. Such a mode of action was consistent with the pronounced cytotoxicity of PhLPs in rod photoreceptors expressing high levels of Gβ1. As a result, the levels of Gβ1 and, contingent on it, Gγ1 in rods became significantly reduced. Rather surprisingly, Gαt1 became down-regulated under this circumstance on the transcriptional level.
Our data suggest that PhLP assists G protein biosynthesis by shuttling newly folded Gβ1 away from the chaperonin, priming it for binding with Gγ1, which in turn, triggers PhLP’s own release. A splice isoform of PhLP, however, hinders this process via a complex mechanism.
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