Genetic and biochemical analysis of zebrafish with visual function defect

Zebrafish have been recognized as an excellent model organism for the study of vertebrate development and retinal function. Using N-ethyl-N-nitrosourea mutagenesis and a behavioral screening strategy based on the optokinetic response (OKR), several autosomal recessive zebrafish mutants with visual function defects have been isolated. Both the no optokinetic response a and f, noa and nof mutants have normal retinal morphology at 5 days postfertilization, electroretinograms indicative of defects in the outer retina, and reduced activation of transducin. The partial optokinetic response b (pob) mutant was identified as having no OKR in red light, a normal OKR in white light, and apoptosis-mediated red cone photoreceptor cell degeneration.;To gain a better understanding of these defects, genetic strategies were used to identify the mutated genes in noa, nof, and pob. Using the candidate gene approach, the mutated gene in not was identified as the cone transducin alpha subunit. For the pob mutant, positional cloning isolated the critical interval to a single PAC clone. A destabilizing mutation was uncovered in a 30 kDa protein of unknown function. Future experiments are warranted to characterize the function of this protein and to define its role in the survival of red cone photoreceptor cells.;Using a positional cloning strategy, the defective gene in the noa mutant was identified as dihydrolipoamide S-acetyltransferase (dlat), or the pyruvate dehydrogenase complex (PDHc) E2 subunit. Human mutations in PDHc cause a form of Leigh syndrome, a disease characterized by severe neurological dysfunction, mental and growth retardation, congenital lactic acidosis, and most often an early death. Development of effective therapies for this disease has been hampered by the absence of an appropriate animal model. In noa mutants, Dlat protein levels are reduced and lactate and pyruvate levels are elevated. To evaluate noa as a model for PDHc deficiency, I treated noa mutants with a ketogenic diet. This treatment successfully rescues blindness, promotes feeding behavior, reduces lethargy, increases survival, and reduces lactic acidosis in the noa mutants. The noa zebrafish mutant provides an excellent opportunity for improving current treatments and evaluating novel therapies for patients with PDHc deficiency and primary congenital lactic acidosis.