Retinitis pigmentosa is the most prevalent hereditary retinopathy of man, currently estimated to affect up to 1.5 million people. The recent attribution of choroideremia (an X-linked form of retinitis pigmentosa) to mutational inactivation of the Rab escort protein (REP-1) subunit of the Rab geranylgeranyl (GG) transferase suggests that post-translational isoprenylation of Rab proteins is critical to retinal long-term survival. Since Rab proteins regulate intracellular membrane transport, and prenylation is required for their function, a general deficiency in Rab geranylgeranylation would prove lethal. However, the sole functional consequence of the loss of the entire choroideremia gene is a progressive degeneration of the choroid and retina. The long-term goals of this project are to elucidate the mechanisms which regulate Rab protein isoprenylation and to understand how a general deficiency in Rab isoprenylation can result in a disease that is restricted in its clinical manifestations to the retina and choroid. As a means to realize these goals, we propose to characterize the functional domains and cellular distribution of the Rab escort proteins and identify novel retinal isoprenylated proteins.
The specific aims of this project are: 1) to use site-directed mutagenesis to generate altered forms of the known Rab escort proteins [REP-1 and choroideremia-like (CHML)]. These mutants will be analyzed in a battery of assays to test the hypothesis that each domain of REP is responsible for regulating a distinct aspect of Rab geranylgeranylation and cellular localization; 2) to examine the tissue and developmental expression patterns for both REP-l and CHML testing the hypothesis that cellular co-expression of CHML serves to protect the non retinal tissues of choroideremia patients, thereby preventing wide-spread tissue dysfunction; and 3) to isolate cDNAs encoding retinal isoprenylated proteins using two novel in vitro enzymatic screening systems. Identified proteins will be analyzed to determine their potential relevance to choroideremia. The hypothesis to be tested is that the restricted retinal pathogenesis of choroideremia results from the inability to prenylate a Rab GG transferase retinal-specific substrate, recognized only in the context of REP-1 and required for the long-term survival of retinal photoreceptors, pigment epithelium, or choroid. The proposed studies will provide fundamental new information on Rab protein geranylgeranylation and new insight into the functional role of prenylated proteins in retinal degenerative diseases, as well as possible avenues for diagnosis and treatment of both choroideremia and other forms of retinitis pigmentosa.
Showing the most recent 10 out of 13 publications
