Desmin is a muscle-specific intermediate filament protein that has fundamental role in muscle structure and force transmission. Whereas human desmin protein is encoded by a single gene, two desmin paralogs (desma and desmb) exist in zebrafish. Desma and desmb show differential spatiotemporal expression during zebrafish embryonic and larval development, being similarly expressed in skeletal muscle until hatching, after which expression of desmb shifts to gut smooth muscle. We generated knockout (KO) mutant lines carrying loss-of-function mutations for each gene by using CRISPR/Cas9. Mutants are viable and fertile, and lack obvious skeletal muscle, heart or intestinal defects. In contrast to morphants, knockout of each gene did not cause any overt muscular phenotype, but did alter calcium flux in myofibres. These results point to a possible compensation mechanism in these mutant lines generated by targeting nonsense mutations to the first coding exon.

Current knowledge about the molecular properties of the crustacean ion channels is rather limited even if crustaceans have been widely used as a model in neuroscience. We cloned for the first time two different potassium channel genes from the freshwater crayfish Astacus leptodactylus (Eschscholtz, 1823), one of the genes functionally expressed in the Xenopus oocytes. The open-reading frames of the genes were 1,203 and 3,447 bp, respectively. The nucleic acid sequence of the genes and associated proteins were similar to those of a typical potassium channel. BLAST analyses indicated that one of the cloned genes had a substantial similarity to an inward-rectifier potassium channel whereas the other gene was similar to a high-conductance-KCa type potassium channel reported in related species. Transmembrane topology and three-dimensional structure of the coded proteins were calculated and functional regions of the channel proteins responsible for ion selectivity, voltage sensing, gating, and calcium binding were identified. One of the cloned channel genes has been expressed in the Xenopus oocytes. Analysis of the expressed potassium currents confirmed that the cloned gene was coding a typical Kir-type potassium channel with ATP sensitivity.

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Desmin is a muscle-specific intermediate filament protein that has fundamental role in muscle structure and force transmission. Whereas human desmin protein is encoded by a single gene, two desmin paralogs (desma and desmb) exist in zebrafish. Desma and desmb show differential spatiotemporal expression during zebrafish embryonic and larval development, being similarly expressed in skeletal muscle until hatching, after which expression of desmb shifts to gut smooth muscle. We generated knockout (KO) mutant lines carrying loss-of-function mutations for each gene by using CRISPR/Cas9. Desma;desmb double mutants are viable and fertile, and lack obvious skeletal muscle, heart or intestinal defects. In contrast to morphants, knockout of each gene did not cause any overt muscular phenotype, but did alter calcium flux in myofibres. These results point to a possible compensation mechanism in these mutant lines generated by targeting nonsense mutations to the first coding exon.