A novel cytoskeletal antigen, RET52, has been identified in the mouse retina. This 52 kD polypeptide is antigenically related to dematin (band 4.9), an actin-bundling phosphoprotein component of the erythrocyte membrane skeleton. Like dematin, RET52 is also a substrate for cAMP-dependent protein kinase. Within the retina, RET52 is primarily concentrated in two regions-the rod inner segment and the outer synaptic layers-although the developmental expression of RET52 differs in these areas. RET52 is present at birth in the inner segment, but appears about the time of initial synapse formation (postnatal day 4-6) in the outer plexiform layer. No differences in RET52 expression have been detected in early-stage mouse retinas with the retinal degeneration (rd) phenotype. RET52 localization, developmental expression, homologies to dematin, and in vitro phosphorylation pattern suggest a possible role for cytoskeleton-associated proteins in the initiation or control of disk membrane assembly and/or synapse formation in the rod photoreceptor.