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Joseph Fogerty, Brian D Perkins; Genetic ablation of miR-129-3p in zebrafish does not affect cone outer segment development. Invest. Ophthalmol. Vis. Sci. 2016;57(12):1773.
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© ARVO (1962-2015); The Authors (2016-present)
MicroRNAs (miRs) function as important regulators of photoreceptor development and physiology. The purpose of this study was to determine if photoreceptor sensory cilium development requires miR-129-3p, a key regulator of ciliogenesis.
Both morpholinos and TALENs were used to inactivate miR-129-3p in zebrafish. Zebrafish contain two mir-129 paralogs, mir-129-1 and mir-129-2. We designed TALENs against each locus and selected mutations that are predicted to inactivate the seed sequence of each miR-129-3p paralog without significantly altering the pre-miRNA hairpin structure. We used PNA to examine cone outer segment length in 5dpf larvae of both single and double mir-129 mutants.
At 5 days post fertilization (dpf), zebrafish larvae injected with morpholinos targeting miR-129-3p exhibited phenotypes consistent with malfunctioning cilia, including shortened photoreceptor outer segments, kidney cysts, ectopic otoliths, and severe ventral body curvature. Mir-129-1-3p-/-; mir-129-2-3p-/- fish, which are predicted to lack all functional miR-129-3p, were viable and were produced in the expected Mendelian ratio. At 5 dpf, these mutant animals lacked the severe body curvature and kidney cysts that were readily apparent in morphant fish, and PNA staining showed normal cone outer segment lengths.
Contrary to our hypothesis and in stark contrast to miR-129-3p morphant zebrafish, miR-129-3p mutants have no overt cilia phenotypes at 5 dpf. This suggests that the morpholino may non-selectively target additional RNAs that can compensate for loss of miR-129-3p in mutant fish.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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