Disruption of the Chaperonin Containing TCP-1 Function Affects Protein Networks Essential for Rod Outer Segment Morphogenesis and Survival

Abstract

Type II Chaperonin Containing TCP-1 (CCT, also known as TCP-1 Ring Complex, TRiC) is a multi-subunit molecular machine thought to assist in the folding of ∼10% of newly translated cytosolic proteins in eukaryotes. A number of proteins folded by CCT have been identified in yeast and cultured mammalian cells, however, the function of this chaperonin in vivo has never been addressed. Here we demonstrate that suppressing the CCT activity in mouse photoreceptors by transgenic expression of a dominant-negative mutant of the CCT cofactor, phosducin-like protein (PhLP), results in the malformation of the outer segment, a cellular compartment responsible for light detection, and triggers rapid retinal degeneration. Investigation of the underlying causes by quantitative proteomics identified distinct protein networks, encompassing ∼200 proteins, which were significantly affected by the chaperonin deficiency. Notably among those were several essential proteins crucially engaged in structural support and visual signaling of the outer segment such as peripherin 2, Rom1, rhodopsin, transducin, and PDE6. These data for the first time demonstrate that normal CCT function is ultimately required for the morphogenesis and survival of sensory neurons of the retina, and suggest the chaperonin CCT deficiency as a potential, yet unexplored, cause of neurodegenerative diseases. Type II Chaperonin Containing TCP-1 (CCT, also known as TCP-1 Ring Complex, TRiC) is a multi-subunit molecular machine thought to assist in the folding of ∼10% of newly translated cytosolic proteins in eukaryotes. A number of proteins folded by CCT have been identified in yeast and cultured mammalian cells, however, the function of this chaperonin in vivo has never been addressed. Here we demonstrate that suppressing the CCT activity in mouse photoreceptors by transgenic expression of a dominant-negative mutant of the CCT cofactor, phosducin-like protein (PhLP), results in the malformation of the outer segment, a cellular compartment responsible for light detection, and triggers rapid retinal degeneration. Investigation of the underlying causes by quantitative proteomics identified distinct protein networks, encompassing ∼200 proteins, which were significantly affected by the chaperonin deficiency. Notably among those were several essential proteins crucially engaged in structural support and visual signaling of the outer segment such as peripherin 2, Rom1, rhodopsin, transducin, and PDE6. These data for the first time demonstrate that normal CCT function is ultimately required for the morphogenesis and survival of sensory neurons of the retina, and suggest the chaperonin CCT deficiency as a potential, yet unexplored, cause of neurodegenerative diseases. Normal cellular function is hinged upon the ability of the endogenous machinery to properly process newly synthesized proteins, and this is often required for enabling their functional activity. Eukaryotic cells contain several protein-folding, or chaperone, systems assisting in this process. Despite the apparent redundancy, it is believed that each chaperone system plays an important and unique role in facilitating protein folding by acting on distinct sets of substrates at unique cellular locations and/or under certain conditions (1.Spiess C. Meyer A.S. Reissmann S. Frydman J. Mechanism of the eukaryotic chaperonin: protein folding in the chamber of secrets.Trends Cell Biol. 2004; 14: 598-604Abstract Full Text Full Text PDF PubMed Scopus (291) Google Scholar, 2.Clarke A.R. Cytosolic chaperonins: a question of promiscuity.Mol. Cell. 2006; 24: 165-167Abstract Full Text Full Text PDF PubMed Scopus (10) Google Scholar). One such chaperone system, unique to eukaryotic cells, is the Chaperonin Containing TCP-1 (CCT), 1The abbreviations used are:CCTchaperonin containing TCP-1TRiCTCP-1 ring complexTCP-1t-complex polypeptide 1PhLPphosducin-like proteinPDE6cyclic GMP phosphodiesterase 6MALDImatrix-assisted laser desorption ionizationM