Arrestins are cytosolic proteins involved in the desensitization of G-protein-coupled receptors. We report the cloning of trout red blood cell arrestin which shows 76, 82 and 52% identity with bovine β-arrestin1, β-arrestin2 and retinal arrestin respectively. Antibodies were generated against the C-terminus of trout red blood cell arrestin. These antibodies detected arrestin in erythrocyte cytosol and were able to precipitate the native protein. The Na+/H+ antiporter of trout red blood cell is activated by β-adrenergic stimulation and is then desensitized whereas the transmembrane signalling pathway is not. To investigate the subcellular distribution of arrestin on β-adrenergic activation and desensitization of the antiporter, precipitation experiments were carried out on trout erythrocytes. A desensitization-dependent shift in cytosolic arrestin to the membranes could not be detected using the immunoprecipitation technique but we cannot exclude the possibility that a small number of cytosolic arrestins might be involved in the regulation of membrane proteins in trout erythrocyte. Recombinant trout arrestin was produced in a protease-deficient Escherichia coli strain and its functionality was tested in a reconstituted rhodopsin assay. The recombinant protein provides a suitable tool for investigating the target for arrestin in trout red blood cell, which still remains to be identified.

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Author notes

Present address: Department of Pharmacology, University of Würzburg, D-97078 Würzburg, Germany.

The sequence data for the three isoforms of trout red cell arrestin will appear in the EMBL/GenBank/DDBJ Nucleotide Sequence Databases under the accession numbers U48410, U48411 and U48412.