Institute of Biotechnology, College of Bioscience and Biotechnology, National Cheng Kung University, Tainan, 701, Taiwan.
Division of Applied Bioscience, Graduate School of Agriculture, Kyoto University, Kyoto, 606-8502, Japan.
Sci Rep. 2017 Sep 12;7(1):11435. doi: 10.1038/s41598-017-09966-9.
Myostatin (MSTN) suppresses skeletal muscle development and growth in mammals, but its role in fish is less well understood. Here we used CRISPR/Cas9 to mutate the MSTN gene in medaka (Oryzias latipes) and evaluate subsequent growth performance. We produced mutant F0 fish that carried different frameshifts in the OlMSTN coding sequence and confirmed the heritability of the mutant genotypes to the F1 generation. Two F1 fish with the same heterozygous frame-shifted genomic mutations (a 22 bp insertion in one allele; a 32 bp insertion in the other) were then crossbred to produce subsequent generations (F2F5). Body length and weight of the MSTN F4 medaka were significantly higher than in the wild type fish, and muscle fiber density in the inner and outer compartments of the epaxial muscles was decreased, suggesting that MSTN null mutation induces muscle hypertrophy. From 34 weeks post hatching (wph), the expression of three major myogenic related factors (MRFs), MyoD, Myf5 and Myogenin, was also significantly upregulated. Some medaka had a spinal deformity, and we also observed a trade-off between growth and immunity in MSTN F4 medaka. Reproduction was unimpaired in the fast-growth phenotypes.
肌肉生长抑制素 (MSTN) 抑制哺乳动物的骨骼肌发育和生长，但它在鱼类中的作用知之甚少。在这里，我们使用 CRISPR/Cas9 对斑马鱼（Oryzias latipes）中的 MSTN 基因进行突变，并评估随后的生长性能。我们产生了携带 OlMSTN 编码序列中不同移码突变的 F0 突变鱼，并将突变基因型的遗传性传递到 F1 代。然后，将两条具有相同杂合移码基因组突变的 F1 鱼（一个等位基因中有 22 bp 的插入；另一个等位基因中有 32 bp 的插入）杂交，以产生后续世代（F2F5）。MSTN F4 斑马鱼的体长和体重明显高于野生型鱼，并且背肌内外区的肌纤维密度降低，表明 MSTN 缺失突变诱导肌肉肥大。从孵化后 34 周（wph）开始，三种主要的肌生成相关因子（MRFs），MyoD、Myf5 和 Myogenin 的表达也显著上调。一些斑马鱼出现脊柱畸形，我们还观察到 MSTN F4 斑马鱼在生长和免疫之间存在权衡。快速生长表型的繁殖不受影响。
