Abstract

Purpose: Little is known about the functional domains of the protein CEP290, encoded by the gene most commonly associated with the devastating blinding disease Leber Congenital Amaurosis (LCA). This study aimed to identify different functional regions of the protein to better understand and define the regions necessary for protein function, with the hope of designing a truncated version of CEP290 that might recapitulate the full length protein’s function. Results: A 300 amino acid region of CEP290 was found to be necessary and sufficient for microtubule binding and stabilization. Using microtubule binding as a functional readout of CEP290 activity, regions of the protein were identified which cooperate to inhibit protein function. Conclusions: The ability of CEP290 to bind microtubules at the primary cilium has been postulated to be a critical role in the protein’s function. Our data show this hypothesis to be correct, and sharply define the region of the protein responsible for this activity. Furthermore, the identification and validation of novel inhibitory domains of CEP290 present the possibility that a truncated version of the protein lacking these inhibitory regions might retain the functionality of the full length protein. A mini-gene encoding this truncated protein could be used therapeutically for the treatment of LCA due to CEP290 mutations, and may be small enough to fit the limited cargo capacity of AAV gene therapy vectors.