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Degradation of the non-palmitoylated invertebrate visual guanine-nucleotide binding protein, iGqα(C3,4A), by the ubiquitin-proteasomal pathway is regulated by its activation and translocation to the cytoplasm

Published online by Cambridge University Press:  19 July 2007

LYNLE GO
Affiliation:
Department of Pharmacology, University of Toronto, Ontario, Canada
JANE MITCHELL
Affiliation:
Department of Pharmacology, University of Toronto, Ontario, Canada

Abstract

Light-dependent translocation of invertebrate visual guanine-nucleotide binding protein, iGqα, from rhabdomeric membranes to the cytoplasm is one of many mechanisms that contribute to light adaptation in the invertebrate eye. We have previously cloned iGqα from a Loligo pealei photoreceptor cDNA library and shown that when expressed in HEK 293T cells it is palmitoylated. In this study we compared the activation, cytoplasmic translocation, and turnover of iGqα with that of a non-palmitoylated mutant, iGqα(C3,4A). In the HEK 293T cells, muscarinic M1 receptors coupled equally well to iGqα and iGqα(C3,4A) to activate phospholipase C. Activation of iGqα(C3,4A), but not iGqα, induced translocation of the α subunit from the membrane to cytosol with rapid degradation of the soluble protein resulting in a decreased half-life for iGqα(C3,4A) of 10 hours compared to 20 hours for iGqα. Degradation of iGqα(C3,4A) was inhibited by proteasomal inhibitors but not by inhibitors of lysosomal proteases or calpain. The presence of the proteasomal inhibitor led to the accumulation of polyubiquitinated species of either iGqα or iGqα(C3,4A). Our results suggest that palmitoylation of iGqα is required to maintain membrane association of the protein in its active conformation, and whereas membrane-bound and soluble iGqα can be polyubiquitinated, membrane association protects the protein from rapid degradation by the proteasomal pathway.

Type
Research Article
Copyright
2007 Cambridge University Press

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