G protein signaling depends on the ability of the individual subunits of the G protein heterotrimer to assemble into a functional complex. Formation of the G protein βγ (Gβγ) dimer is particularly challenging because it is an obligate dimer in which the individual subunits are unstable on their own. Recent studies have revealed an intricate chaperone system that brings the Gβ and Gγ subunits together. This system includes the cytosolic chaperonin containing TCP-1 (CCT) and a co-chaperone phosducin-like protein 1 (PhLP1). Two key intermediates in the Gβγ assembly process, the Gβ-CCT and the PhLP1-Gβ-CCT complexes, were isolated and their structures determined by cryo-electron microscopy, chemical cross-linking coupled with mass spectrometry, and unnatural amino acid cross-linking. These structures show that Gβ interacts with CCT in a near-native state through interactions of the Gγ-binding region of Gβ with the CCTγ subunit. PhLP1 binding stabilizes the Gβ β-propeller, disrupting interactions with CCT and releasing a PhLP1-Gβ dimer for assembly with Gγ. We also investigated the role of CCT and PhLP1 in folding and assembling mTOR complexes, which regulate cell growth through phosphorylation. We found that the β-propeller protein mLST8 and one of its binding partners called raptor, which is a large protein in which one domain forms a β-propeller, both bind to CCT. PhLP1 forms a ternary complex with mLST8 and CCT and may play a co-chaperone role. Depletion of PhLP1 or CCT reduces assembly of mTOR complexes in the cell. Collectively, this report reveals diversity in the contributions of CCT to the formation of protein complexes in signaling pathways and presents a molecular mechanism of Gβ folding by CCT and PhLP1.
| Identifer | oai:union.ndltd.org:BGMYU2/oai:scholarsarchive.byu.edu:etd-6550 |
| Date | 01 December 2014 |
| Creators | Plimpton, Rebecca L |
| Publisher | BYU ScholarsArchive |
| Source Sets | Brigham Young University |
| Detected Language | English |
| Type | text |
| Format | application/pdf |
| Source | Theses and Dissertations |
| Rights | http://lib.byu.edu/about/copyright/ |
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