Intraflagellar Transport (IFT) Proteins Necessary for Photoreceptor Outer Segment Formation

Purpose: The photoreceptor outer segment holds hundreds of tightly stacked membranous discs that contain the proteins necessary for phototransduction. The photoreceptor outer segments regularly, and rapidly, turn over by shedding membrane from the tips; thus efficient protein transport mechanisms through the connecting cilium are essential to provide photoreceptors with a sufficient supply of protein to replenish those molecules lost during outer segment turnover. Defects that compromise outer segment formation or protein transport to the outer segment result in photoreceptor degeneration. Recently, components of the intraflagellar transport (IFT) complex have been implicated in sensory cilia formation and protein transport and the genes encoding proteins represent likely candidates for retinitis pigmentosa. Zebrafish mutants in 4 IFT proteins (IFT172, IFT88, IFT81, IFT57) have been identified. The purpose of this study was to investigate the effects of these 4 zebrafish mutations on photoreceptor structure, protein transport, and outer segment formation. Methods: Photoreceptor morphology in IFT mutants was examined under light- and transmission electron microscopy at 60 hours post fertilization (hpf), 72 hpf, and 96 hpf. Immunohistochemistry was used to determine the localization of several photoreceptor proteins, including rod and cone opsins. Mutant phenotypes were verified by antisense morpholino injections. Results: In IFT172, IFT88, and IFT57 mutants, light microscopy revealed a slight increase in cell death and holes in the outer nuclear layer as well as absence of photoreceptor outer segments. Surprisingly, IFT81 mutants did not exhibit any observable retina phenotype at 96 hpf. Cone photoreceptors were disorganized and opsin mislocalization was observed in rods. No outer segment staining was observed in any of the IFT mutants. Conclusions: Mutations in zebrafish IFT genes result in photoreceptor outer segment defects. Defects in opsin localization were observed in three mutants. Interestingly, IFT81 mutants did not exhibit a retina phenotype, despite being a component of the same IFT complex. This suggests that some, but not all, IFT proteins participate in outer segment formation and opsin transport. Given the importance of opsin transport and outer segment formation for the survival of photoreceptors, the IFT genes represent likely candidates for retinal degenerations.