Identification of Novel Murine- and Human-Specific RPGRIP1 Splice Variants with Distinct Expression Profiles and Subcellular Localization

Leber congenital amaurosis (LCA, Mendelian Inheritance in Man [MIM] 204000) is the most severe and the leading cause of blindness in children.1 Affected children exhibit strongly reduced or extinguished electroretinograms (ERGs), typically by 12 months of life. Several genes with diverse retinal function such as the aryl hydrocarbon receptor-interacting protein-like 1 (AILP1),2 Crumbs homolog 1 (CRB1),3,4 cone-rod homeobox (CRX),5 guanylate cyclase 2D (GUCYD2),6 RPE65,7 retinal dehydrogenase 12 (RDH12),8 and RPGRIP1,9,10 have been implicated in LCA and account for approximately 50% of cases. All mutations in RPGRIP1 lead only to LCA, whereas genetic lesions in the remaining genes also lead to other clinically heterogeneous retinal dystrophies with early postnatal and adult onsets.1 RPGRIP1 was originally found to be an interacting substrate of retinitis pigmentosa GTPase regulator (RPGR),11–14 thus implicating RPGRIP1 in the molecular pathogenesis of X-linked retinitis pigmentosa type 3 (XlRP3). All missense mutations in RPGR are clustered in its RCC1-homologous domain,15 and some of these have been shown to uncouple the interaction of RPGR with RPGRIP1.11 Human mutations in RPGR lead to retinitis pigmentosa (RP)16–19 and several other retinal degenerative diseases such as cone–rod,20 cone,21 and recessive atrophic macular degeneration.22 In addition, two distinct mutations (845-846delTG and G173R) in exon 8 of RPGR segregate with systemic disorders associated with hearing loss, sinusitis, and chronic recurrent respiratory tract and ear infections.23–26 Immunocytochemical analysis of human retina, bronchi, sinuses, and cochlea localized RPGR to the outer segments of photoreceptors and to nonocular tissues, such as the epithelial cells lining the lumen of the sinuses and bronchi cavities and the nonciliated cochlear tissues, stria vascularis, suprastrial cells, and spiral limbus.24 This finding is consistent with the manifestations of the ocular and systemic diseases described.23–26 Likewise, we have found that RPGR and RPGRIP1 isoforms localize to the outer segment of human and bovine photoreceptors,11,27 whereas in mouse photoreceptors, they localize to the connecting cilium.27,28 However, RPGRIP1 was also strongly expressed in a subset of inner retinal neurons, the amacrine cells.27,28 Hence, the differential expression of RPGR and RPGRIP1 among retinal neurons may provide a rationale for the distinct phenotypes caused by genetic lesions in RPGR and RPGRIP1 in the human.28 RPGRIP1 is subjected to significant alternative splicing in the human and bovine,11,13 and products thereof have been shown to be susceptible to various degrees of limited proteolysis, depending on the subcellular localization of RPGRIP1.28 This led to the proposal that the repertoire of RPGRIP1 products generated may mediate distinct functions and subcellular processes with pathologic outcomes still to be determined.28 To investigate further the implications of the heterogeneity of RPGRIP1 isoforms among and within species and the function of these in subcellular processes, we report the identification of novel murine- and human-specific RPGRIP1 isoforms with distinct expression profiles and subcellular properties.