A Barrier to Diffusion of Opsins but not Peripheral Membrane Proteins at the Disk Rims of Cone Photoreceptor Sensory Cilia

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Localization and retention of transduction cascade proteins within ciliary signaling compartments are thought to be mediated, in part, by diffusion barriers at the transition zone. Mutations in cone opsins and other phototransduction cascade proteins often result in their mislocalization from the ciliary cone outer segment, and can lead to a spectrum of diseases, ranging from color blindness to progressive cone dystrophy, often followed by rod dystrophy, and eventually blindness.While the mechanisms for protein transport to and retention within rod outer segments have received considerable attention, little is known about these processes in cones. Retention of opsins in the rod outer segment is achieved by physical separation of the disc membranes from the plasma membrane. Cone discs, however, are contiguous with the plasma membrane, and therefore retention of opsin would require a diffusion barrier either at the cilium base, as has been demonstrated for primary cilia, or at the disc rim. Here we examine the diffusion of the transmembrane red cone opsin-GFP fusion protein and the peripheral membrane protein double geranylgeranyl-GFP expressed in cones of Xenopus laevis using multiphoton FRAP. We show that although both proteins diffuse laterally in the disc membranes with similar rates, diffusion of opsin-GFP between discs is significantly retarded compared with double geranylgeranyl-GFP.Our results are consistent with cone opsin retention in the outer segment being mediated by a selective barrier to free diffusion for trans-membrane proteins located at the end loops between adjacent discs. Peripheral membrane proteins, on the other hand, are not prevented from crossing this barrier.