The photoreceptor-specific 33 kDa phosphoprotein of mammalian retina: Generation of monospecific antibodies and localization by immunocytochemistry

The distribution in mouse retina of a 33000 Da phosphoprotein (33 kDa) that complexes with the beta/gamma subunits of transducin (T βγ) and undergoes light-induced dephosphorylation was determined by immunocytochemistry. An antiserum containing antibodies for the 33 kDa protein and β-transducin of mouse and bovine retinas was generated against the purified 33 kDa-T βγ complex from bovine retina. The antiserum reacts with β-transducin derived from either 33 kDa-T βγ complex or transducin complex (T αβγ), but not with the α- or γ-transducin. It also reacts with both the phosphorylated and unphosphorylated form of the 33 kDa-T βγ complex. Antibodies, monospecific for the 33 kDa and β-transducin subunits respectively, were purified from the antiserum by immunoadsorption and used in immunocytochemical analysis of the respective antigens. The 33 kDa protein was found to be associated exclusively with the photoreceptor cells of the retinas, with the most intense staining in the inner and outer segments' layers and lighter staining in the synaptic terminal layers. β-Transducin also is found in the photoreceptors, but some T β immunoreactivity exists within the inner plexiform layer. The specific localization of the 33 kDa protein together with its light-modulated phosphorylation suggest that the 33 kDa-T βγ complex is involved in light-regulated activities of the rod photoreceptor cells.