The retinal degeneration slow or rds mutation in mice is characterized by complete non-development of photoreceptor outer segments, followed by slow degeneration of the photoreceptor cell bodies. Mutations in the human RDS gene are responsible for a significant fraction of dominantly inherited retinal dystrophies. Rds/peripherin, the protein product of rds, is an integral membrane glycoprotein that is located in the rims of outer segment discs. Rds/peripherin has been shown to interact non-covalently with the related protein, rom1. By the phenotype resulting from its absence, rds/peripherin is required for the formation of outer segments. Very little is known, however, about its specific function. This proposal combines in vivo genetic and biochemical approaches to study the role of rds/peripherin in outer segments.
The first aim i s to generate transgenic mice that express rds/rom1 chimeric proteins. Experiments in this aim should identify those domains responsible for the dramatic difference in phenotypes between rds-/- and rom1- /- null mutants. In a preliminary study, the applicant observed that rds/peripherin, but not rom1, associates with a cytoskeleton- enriched fraction, and he hypothesizes that this may be the key to understanding the role of rds/peripherin in the formation of outer segment discs.
The second aim of this proposal is to identify and clone the gene for the protein that couples rds/peripherin to the cytoskeleton. This interacting protein will be analyzed biochemically, and by generating mice with a knockout mutation in this gene.
The third aim i s to create transgenic mice with a pharmacologically-inducible rds null phenotype. These mice will be used to study the function of rd/peripherin by observing the effects of its arrested expression upon the ultrastructure of outer segments that had previously developed normally. Discs are continuously added to the bases of outer segments to keep up with their diurnal distal shedding, and this characteristic of photoreceptors will be exploited to define the earliest perturbations in disc structure caused by falling levels of rds/peripherin. The studies in this proposal should significantly deepen our understanding about the normal function of rds/peripherin. Further, valuable tools will have been created to study the biochemical etiologies of human retinal dystrophies caused by mutations in the RDS gene.
