Synthesis and Assembly of G Protein βγ Dimers: Comparison of In Vitro and In Vivo Studies

Abstract

The heterotrimeric GTP-binding proteins (G proteins) are the canonical cellular machinery used with the approximately 700 G protein-coupled receptors (GPCRs) in the human genome to transduce extracellular signals across the plasma membrane. The synthesis of the constituent G protein subunits, and their assembly into Gβγ dimers and G protein heterotrimers, determines the signaling repertoire for G-protein/GPCR signaling in cells. These synthesis/assembly -processes are intimately related to two other overlapping events in the intricate pathway leading to formation of G protein signaling complexes, posttranslational modification and intracellular trafficking of G proteins. The assembly of the Gβγ dimer is a complex process involving multiple accessory proteins and organelles. The mechanisms involved are becoming increasingly appreciated, but are still incompletely understood. In vitro and in vivo (cellular) studies provide different perspectives of these processes, and a comparison of them can provide insight into both our current level of understanding and directions to be taken in future investigations.

Comprehensive analysis of heterotrimeric G-protein complex diversity and their interactions with GPCRs in solution

“…With the exception of the β 5 -subunit, all combinations of the other four β-subunits (β 1 -β 4 ) and 12 γ-subunits (γ 1 -γ 5 and γ 7 -γ 13 ) were found to interact with the two different α-subunits. This finding was unexpected because other investigators had indicated that certain βγ-dimers (e.g., β 2 γ 1 , β 2 γ 11 , β 2 γ 13 , β 3 γ 1 , and β 3 γ 11 ) do not exist (6,(26)(27)(28)(29)(30)(31)(32)(33)(34).…”

“…Interestingly, those combinations-especially for β 3 and β 5 -that cannot be generated by in vitro translation could still be found in cell-based assays like the yeast two-hybrid screen (29), coimmunoprecipitation (31), and bimolecular fluorescence complementation (47), as summarized in detail in ref. 6. Similarly, the β 2 γ 13 -dimer, which could not be identified when mixing subunits that had been separately translated in vitro (32), could be identified when both subunits were cotranslated in vitro (37).…”

“…S5D). The finding that β 5 exhibits a lower conservation throughout those residues may explain why β 5 only poorly forms βγ-dimers, which are furthermore often unstable (6,7).…”

“…The heterotrimeric G-protein complex is assembled from a pool of 16 α-subunits, 5 β-subunits, and 12 γ-subunits (3)(4)(5). Extensive analyses of the βγ-dimer formation potential had indicated unrestricted dimer formation for β 1 and β 4 (i.e., dimers with all γ-subunits are found), restricted dimer formation of β 2 and β 3 (e.g., no dimers with γ 1 or γ 11 ), and no or only weak dimer formation for β 5 (6,7). Although a comprehensive analysis of Gαβγ complex formation is missing, it is likely that most of the Gαβγ combinations are capable of forming a functional complex (8)(9)(10).…”

Proteasome overload is a common stress factor in multiple forms of inherited retinal degeneration

“…1D). Newly synthetized G-protein β-subunit polypeptides assume their final shape inside the cytosolic chaperonin containing T-complex protein 1 (CCT), whereas Gγ subunits (aided by the phosducin-like protein, PhLP) (13) play a critical role in stabilizing and releasing Gβ from CCT to yield mature Gβγ complexes (14)(15)(16)(17). Therefore, the lack of Gγ 1 in rods may result in Gβ 1 retention inside CCT, ultimately reducing the CCT availability to process the dozens of other cytosolic proteins normally folded inside this chaperonin (18).…”