title: Identification of a redox-sensitive cysteine in GCP60 that regulates its interaction with golgin-160.
abstract: Golgin-160 is ubiquitously expressed in vertebrates. It localizes to the cytoplasmic side of the Golgi and has a large C-terminal coiled-coil domain. The noncoiled-coil N-terminal head domain contains Golgi targeting information, a cryptic nuclear localization signal, and three caspase cleavage sites. Caspase cleavage of the golgin-160 head domain generates different fragments that can translocate to the nucleus by exposing the nuclear localization signal. We have previously shown that GCP60, a Golgi resident protein, interacts weakly with the golgin-160 head domain but has a strong interaction with one of the caspase-generated golgin-160 fragments (residues 140-311). This preferential interaction increases the Golgi retention of the golgin-160 fragment in cells overexpressing GCP60. Here we studied the interaction of golgin-160-(140-311) with GCP60 and identified a single cysteine residue in GCP60 (Cys-463) that is critical for the interaction of the two proteins. Mutation of the cysteine blocked the interaction in vitro and disrupted the ability to retain the golgin-160 fragment at the Golgi in cells. We also found that Cys-463 is redox-sensitive; in its reduced form, interaction with golgin-160 was diminished or abolished, whereas oxidation of the Cys-463 by hydrogen peroxide restored the interaction. In addition, incubation with a nitric oxide donor promoted this interaction in vitro. These findings suggest that nuclear translocation of golgin-160-(140-311) is a highly coordinated event regulated not only by cleavage of the golgin-160 head but also by the oxidation state of GCP60.
authors: Department of Cell Biology, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA.
authors: Juan I
authors: JI
authors: Sbodio
authors: Carolyn E
authors: CE
authors: Machamer
mesh_terms: D021581:Active Transport, Cell Nucleus; D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D001324:Autoantigens / Q000378:metabolism*; D003593:Cytoplasm / Q000378:metabolism; D004926:Escherichia coli / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect / Q000379:methods; D006056:Golgi Apparatus / Q000378:metabolism; D000077614:Golgi Matrix Proteins; D006367:HeLa Cells; D006801:Humans; D006861:Hydrogen Peroxide / Q000494:pharmacology; D007460:Iodoacetamide / Q000737:chemistry; D008565:Membrane Proteins / Q000378:metabolism*; D008954:Models, Biological; D009584:Nitrogen / Q000737:chemistry; D010084:Oxidation-Reduction; D010100:Oxygen / Q000737:chemistry / Q000378:metabolism
chemical_list: C449459:ACBD3 protein, human; D048868:Adaptor Proteins, Signal Transducing; D001324:Autoantigens; C108388:GOLGA3 protein, human; D000077614:Golgi Matrix Proteins; D008565:Membrane Proteins; D006861:Hydrogen Peroxide; D009584:Nitrogen; D010100:Oxygen; D007460:Iodoacetamide