Abstract
Abstract: :
Purpose: Kinesin 2 is a microtubule plus end–directed motor, formed from two motor subunits, KIF3A plus KIF3B or KIF3C, and a non–motor subunit, KAP3, in a heterotrimeric complex. Kif3a CRE–lox knockout studies in mice have shown that kinesin–2 is required for the transport of opsin, and perhaps arrestin, along the photoreceptor cilium, and that complete removal of KIF3A results in very rapid photoreceptor cell death (within 1–2 weeks at ∼3 weeks of age). In the present study, we have determined whether the partial removal of KIF3A, as in Kif3a+/– mice, might be deleterious to photoreceptor cells, especially in the presence of compromised amounts of KIF3B or KIF3C. Methods: Kif3a+/–, Kif3b+/–, Kif3a+/–/Kif3b+/–, Kif3c–/–, and Kif3a+/–/Kif3c–/– mice were bred and aged with littermate WT controls for 12 months. Relative levels of the different kinesin–2 motor subunits in the retinas of these mice were determined by densitometry of western blots. Dorso–ventral sections of the retinas were examined by light microscopy, and the numbers of photoreceptor cell nuclei were counted at 500–µm intervals across each section. Results: In all mutant animals, there was a reduction in the expression level of the mutant gene(s). Moreover, in the single heterozygous Kif3a+/– and Kif3b+/– retinas, there was also a reduction in the amount of the other motor subunit from the non–mutant gene, Kif3a or Kif3b, respectively. In the double heterozygous, Kif3a+/–/Kif3b+/– retinas, the levels of KIF3A and KIF3B were depressed even further to 43% and 18% of WT levels, respectively. In none of these retinas, including the double heterozygous Kif3a+/–/Kif3b+/– retinas, and the Kif3c–/– and Kif3a+/–/Kif3c–/– retinas, was there any indication of photoreceptor cell death. Conclusions: Complete lack of KIF3C does not result in photoreceptor cell death. More than 57% removal of KIF3A is required for photoreceptor cell death, even in the presence of severely compromised levels of KIF3B or KIF3C. These results suggest that KIF3A is unlikely to be a human retinal disease gene, since homozygous mutant Kif3a is lethal. They also provide a cautionary note to RNAi studies where incomplete knockdown of motor protein levels may not be achieved.
Keywords: photoreceptors • protein structure/function • retina: distal (photoreceptors, horizontal cells, bipolar cells)