The functional role of RPGR protein complex in retinitis pigmentosa

Abstract

AbstractPurpose Retinitis pigmentosa (RP) is a progressive outer retinal dystrophy, affecting 1/3500 individuals in most populations. Mutations in the retinitis pigmentosa GTPase regulator (RPGR) gene are the most common single cause of RP, accounting for up to 20% of all cases. RPGR forms a protein complex by interacting with the RPGR‐interacting protein 1 (RPGRIP1) and/or RPGRIP1 like protein (RPGRIP1L). We aim to investigate the functional role of RPGR protein complex in the pathogenesis of RP.Methods Small interfering RNA (siRNA) was used to knock‐down RPGR, RPGRIP1 or RPGRIP1L in human retinal pigment epithelium (RPE) 1 cell line. The efficiency of siRNA knock‐down was assessed by qualitative RT‐PCR and western blotting. The knock‐down effects of RPGR, RPGRIP1, or RPGRIP1L were characterized by immnocytochemistry, enzyme linked immunosorbent assay (ELISA), and western blotting.Results Knock‐down of RPGR, RPGRIP1 or RPGRIP1L caused defects in ciliogenesis, remodelling of the actin cytoskeleton, and centrosome positioning. Since planar cell polarity (PCP) pathway regulates actin cytoskeleton rearrangement through activation of small RhoA GTPases. So, we performed ELISA and found that RhoA‐GTPase activity were upregulated in the absence of RPGR, RPGRIP1, or RPGRIP1L protein. We also found knock‐down of RPGR, RPGRIP1 or RPGRIP1L decreased the stability of DVl3 proteins, key components of polar cell polarity (PCP) pathway.Conclusion We provide compelling evidence that the RPGR protein complex is required for the stability of DVl3, and regulates RhoA for apical centrosome positioning required for cilia formation. We show a novel functional role of RPGR complex in cilia formation by regulating actin cytoskeleton through alteration of PCP pathway.