Tafazzin基因缺失对斑马鱼生长及肌纤维收缩的影响

Effects of the loss of tafazzin on growth and muscle fiber contraction in zebrafish

摘要:

鱼类骨骼肌中富含大量线粒体，尤其在红肌中线粒体含量很高。线粒体对维持鱼类骨骼肌的收缩具有重要作用。Tafazzin（taz）基因作为合成线粒体特异性磷脂心磷脂的重要基因，对线粒体ATP的产生有关键作用。通过对斑马鱼的肌肉组织进行taz基因切片原位杂交，结果显示：taz基因在斑马鱼骨骼肌中大量表达，且特异性表达在肌原纤维的I带中。通过CRISPR/Cas9技术在构建斑马鱼taz缺失突变体taz^-/-过程中发现taz缺失后，斑马鱼生长发育缓慢，突变体的体长、体质量、体宽、体高均显著小于野生型斑马鱼（P<0.05）。受精后48 h胚胎的肌动蛋白荧光染色结果显示，taz突变体肌纤维排列变疏松，肌动蛋白长度显著变长（P<0.05）。整体骨骼染色结果显示，taz突变后肌间骨的生长受到抑制。以上结果表明，在斑马鱼中，taz基因影响斑马鱼生长、骨骼肌的收缩及肌间骨的发育。

Abstract:

Fish skeletal muscle is rich in mitochondria， especially in red muscle fibers. Mitochondria play an important role in maintaining the contraction of fish skeletal muscle. Tafazzin （taz） gene， as an important gene for the synthesis of mitochondrial-specific phospholipid cardiolipin， plays a key role in the production of mitochondrial ATP. Through in situ hybridization of zebrafish muscle tissue sections， the results showed the taz gene was expressed in zebrafish skeletal muscle， and specifically expressed in the I zone of myofibrils. The taz^-/- mutant was further constructed in zebrafish by CRISPR/Cas9 technology， and it was found that after taz deletion， the body length， weight， body width， and height of taz^-/- were significantly smaller than wild-type zebrafish （P<0.05）. It shows that mutation of taz gene lead to growth and developmental delay in zebrafish. The fluorescence staining of 48 hpf embryos showed that the muscle fibers from the taz^-/- mutant was looser than WT. Statistical analysis of F-actin length showed that the H-H length of taz^-/- was（1.12±0.03）μm， while the wild-type zebrafish H-H length was（0.59±0.01）μm， indicating that the length of actin was significantly longer （P<0.05）. Whole Alizarin stain of zebrafish showed the length of intermuscular bones became smaller in taz^-/- than WT. The above results indicate that taz affects the growth， actin contraction and ossification of intermuscular bones.