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The MFF phenotype of nrg2a mutants is rescued upon concomitant loss of Lgl2 function. (A) At 52 hpf, morpholino (MO)-mediated knockdown of lgl2 significantly ameliorated the nrg2a mutant phenotype (12%, n = 639, p < 0.0001) relative to uninjected embryos of an nrg2a -/+ intercross (25%, n = 943). Percentages represent the mean of means (MOM); error bars represent the standard deviations (SD). (B) Percentages of genotyped nrg2a-/- mutants with a strong, medium, weak, or wild-type MFF phenotype, classified by morphological criteria at 52 hpf. While uninjected nrg2a mutants (n = 20) all display a strong phenotype, lgl2 MO-injected mutants (n = 26) show medium, weak or no MFF defects. (C) PCR products obtained via nrg2a genotyping of representative nrg2a -/+, +/+ and-/- embryos at 52 hpf (see Materials and Methods). (D-H) Tail fins of representative live embryos at 52 hpf, as used for quantitative classification in panel B: wild-type (D), uninjected nrg2a-/- mutant with strong MFF phenotype (E), and lgl2 MO-injected nrg2a -/- mutant embryos with medium (F), weak (G) or wild-type (H) phenotype. (I, J) Tail fins of genotyped uninjected (I) and lgl2 MO-injected (J) nrg2a -/- mutant embryo at 48 hpf. Col II immunostaining reveals a normalized organization of collagenous actinotrichia within the dermal space of the Nrg2a/Lgl2-double-deficient embryo (J; compare with Fig 5C for wild-type condition). (K, L) Transverse sections through the dorsal MFF of a genotyped uninjected (K) and an lgl2 MO-injected (L) nrg2a -/- mutant embryo at 52 hpf; CellMask (red) and DAPI (blue) staining reveals a rescue of the dermal space (indicated by arrowheads) from a serpentine-like organization (K) to a straight organization (L) in the Nrg2a/Lgl2-double-deficient embryo (L; compare with Fig 7D for wild-type condition).
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