Expression and Identification of Actin-binding Proteins in Light- and Dark-adapted Octopus Retinas

Abstract

Abstract: : Purpose: Rhabdomere microvilli dramatically reorganize in conditions of light and dark. In dark-adapted octopus retinas, new microvilli arise from a previously avillar membrane. We are using retinas of the octopus to identify actin-binding proteins that may be involved in this membrane remodeling event. Methods: Octopus retinas were light-/dark-adapted, dissected, separated into dorsal and ventral halves, and each sample homogenized. Protein levels in each extract were measured and samples of equal concentration subjected to SDS PAGE. The presence of actin-binding proteins in each sample was then determined using actin overlay blot assays. Results: Overall protein levels in dark-adapted eyes are higher than in retinas maintained in the light and subsequent actin overlays also showed that more actin-binding proteins are expressed in the dark than in the light. Light-/dark-adapted samples displayed nearly identical overlay patterns except for a band around 20kD that was present only in dark-adapted dorsal/ventral samples. Amino terminal sequencing of proteins identified in the actin overlays and a BLAST search using these sequences revealed that the band at 41 kD may be a novel actin-binding protein. The protein band identified at 20 kD, which is expressed only in the dark, is glutathione-S-transferase. Conclusions: A novel actin-binding protein was identified in octopus rhabdoms and may be involved in rhabdomere membrane remodeling in conditions of light and dark. Glutathione S- transferase, or S-crystallin, is a detoxification enzyme whose expression appears to be under light/dark control in the retina.

Keywords: cytoskeleton • photoreceptors • cell membrane/membrane specializations