Loss-of-function mutations in SCN1A cause Dravet syndrome (DS), a catastrophic childhood epilepsy in which patients experience comorbid behavioral conditions, including movement disorders, sleep abnormalities, anxiety, and intellectual disability. To study the functional consequences of voltage-gated sodium channel mutations, we use zebrafish with a loss-of-function mutation in scn1lab, a zebrafish homolog of human SCN1A. Homozygous scn1lab^s552/s552 mutants exhibit early-life seizures, metabolic deficits, and early death. Here, we developed in vivo assays using scn1lab^s552 mutants between 3 and 6 d postfertilization (dpf). To evaluate sleep disturbances, we monitored larvae for 24 h with locomotion tracking software. Locomotor activity during dark (night phase) was significantly higher in mutants than in controls. Among anticonvulsant drugs, clemizole and diazepam, but not trazodone or valproic acid, decreased distance moved at night for scn1lab^s552 mutant larvae. To monitor exploratory behavior in an open field, we tracked larvae in a novel arena. Mutant larvae exhibited impaired exploratory behavior, with increased time spent near the edge of the arena and decreased mobility, suggesting greater anxiety. Both clemizole and diazepam, but not trazodone or valproic acid, decreased distance moved and increased time spent in the center of the arena. Counting inhibitory neurons in vivo revealed no differences between scn1lab^s552 mutants and siblings. Taken together, our results demonstrate conserved features of sleep, anxiety, and movement disorders in scn1lab mutant zebrafish, and provide evidence that a zebrafish model allows effective tests of treatments for behavioral comorbidities associated with DS.

Precise developmental control of jaw length is critical for survival, but underlying molecular mechanisms remain poorly understood. The jaw skeleton arises from neural crest mesenchyme (NCM), and we previously demonstrated that these progenitor cells express more bone-resorbing enzymes including Matrix metalloproteinase 13 (Mmp13) when they generate shorter jaws in quail embryos versus longer jaws in duck. Moreover, if we inhibit bone resorption or Mmp13, we can increase jaw length. In the current study, we uncover mechanisms establishing species-specific levels of Mmp13 and bone resorption. Quail show greater activation of and sensitivity to transforming growth factor beta (TGFβ) signaling than duck; where intracellular mediators like SMADs and targets like Runt-related transcription factor 2 (Runx2), which bind Mmp13, become elevated. Inhibiting TGFβ signaling decreases bone resorption, and overexpressing Mmp13 in NCM shortens the duck lower jaw. To elucidate the basis for this differential regulation, we examine the Mmp13 promoter. We discover a SMAD-binding element and single nucleotide polymorphisms (SNPs) near a RUNX2-binding element that distinguish quail from duck. Altering the SMAD site and switching the SNPs abolish TGFβ sensitivity in the quail Mmp13 promoter but make the duck promoter responsive. Thus, differential regulation of TGFβ signaling and Mmp13 promoter structure underlie avian jaw development and evolution.

Finding effective therapeutic targets to treat NRAS-mutated melanoma remains a challenge. Long non-coding RNAs (lncRNAs) recently emerged as essential regulators of tumorigenesis. Using a discovery approach combining experimental models and unbiased computational analysis complemented by validation in patient biospecimens, we identified a nuclear-enriched lncRNA (AC004540.4) that is upregulated in NRAS/MAPK-dependent melanoma, and that we named T-RECS. Considering potential innovative treatment strategies, we designed antisense oligonucleotides (ASOs) to target T-RECS. T-RECS ASOs reduced the growth of melanoma cells and induced apoptotic cell death, while having minimal impact on normal primary melanocytes. Mechanistically, treatment with T-RECS ASOs downregulated the activity of pro-survival kinases and reduced the protein stability of hnRNPA2/B1, a pro-oncogenic regulator of MAPK signaling. Using patient- and cell line- derived tumor xenograft mouse models, we demonstrated that systemic treatment with T-RECS ASOs significantly suppressed the growth of melanoma tumors, with no noticeable toxicity. ASO-mediated T-RECS inhibition represents a promising RNA-targeting approach to improve the outcome of MAPK pathway-activated melanoma.