title: Zebrafish foxP2 zinc finger nuclease mutant has normal axon pathfinding.
abstract: foxP2, a forkhead-domain transcription factor, is critical for speech and language development in humans, but its role in the establishment of CNS connectivity is unclear. While in vitro studies have identified axon guidance molecules as targets of foxP2 regulation, and cell culture assays suggest a role for foxP2 in neurite outgrowth, in vivo studies have been lacking regarding a role for foxP2 in axon pathfinding. We used a modified zinc finger nuclease methodology to generate mutations in the zebrafish foxP2 gene. Using PCR-based high resolution melt curve analysis (HRMA) of G0 founder animals, we screened and identified three mutants carrying nonsense mutations in the 2(nd) coding exon: a 17 base-pair (bp) deletion, an 8bp deletion, and a 4bp insertion. Sequence analysis of cDNA confirmed that these were frameshift mutations with predicted early protein truncations. Homozygous mutant fish were viable and fertile, with unchanged body morphology, and no apparent differences in CNS apoptosis, proliferation, or patterning at embryonic stages. There was a reduction in expression of the known foxP2 target gene cntnap2 that was rescued by injection of wild-type foxP2 transcript. When we examined axon pathfinding using a pan-axonal marker or transgenic lines, including a foxP2-neuron-specific enhancer, we did not observe any axon guidance errors. Our findings suggest that foxP2 is not necessary for axon pathfinding during development.
authors.affiliation: Division of Pediatric Neurology, Department of Pediatrics, University of Utah School of Medicine, Salt Lake City, Utah, United States of America.
authors.forename: Lingyan
authors.initials: L
authors.lastname: Xing
authors.forename: Kazuyuki
authors.initials: K
authors.lastname: Hoshijima
authors.forename: David J
authors.initials: DJ
authors.lastname: Grunwald
authors.forename: Esther
authors.initials: E
authors.lastname: Fujimoto
authors.forename: Tyler S
authors.initials: TS
authors.lastname: Quist
authors.forename: Jacob
authors.initials: J
authors.lastname: Sneddon
authors.forename: Chi-Bin
authors.initials: CB
authors.lastname: Chien
authors.forename: Tamara J
authors.initials: TJ
authors.lastname: Stevenson
authors.forename: Joshua L
authors.initials: JL
authors.lastname: Bonkowsky
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D001369:Axons / Q000378:metabolism*; D001921:Brain / Q000378:metabolism*; D051858:Forkhead Transcription Factors / Q000235:genetics* / Q000378:metabolism; D009154:Mutation; D016501:Neurites / Q000378:metabolism; D015027:Zebrafish / Q000235:genetics* / Q000378:metabolism; D029961:Zebrafish Proteins / Q000235:genetics* / Q000378:metabolism; D016335:Zinc Fingers / Q000235:genetics
chemical_list: D051858:Forkhead Transcription Factors; C506553:FoxP2 protein, zebrafish; D029961:Zebrafish Proteins