Tom20-mediated mitochondrial protein import in muscle cells during differentiation
Tom20-mediated mitochondrial protein import in muscle cells during differentiation
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DOI：
10.1152/ajpcell.2000.279.5.c1393
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发表时间：
2000-11-01
影响因子：
5.5
通讯作者：
Hood, DA
中科院分区：
文献类型：
--
作者：
Grey, JY;Connor, MK;Hood, DA
线粒体生物合成伴随着蛋白质输入机制成分表达的增加，以及运往基质的蛋白质输入的增加。我们通过改变外膜受体 Tom20 的表达并测量苹果酸脱氢酶 (MDH) 在分化 C2C12 肌细胞中的输入变化来评估其作用。用Tom20转染的细胞的水平比对照细胞高两倍。对细胞进行标记，然后进行 MDH 免疫沉淀，结果显示 MDH 导入量有同等增加。由于甲状腺激素治疗，输入率和 Tom20 水平之间的这种相似性也很明显。使用反义寡脱氧核苷酸，我们将 Tom20 表达抑制了 40%，导致 MDH 导入减少 40-60%。体外测定还表明，输入基质对 Tom20 抑制比输入外膜更敏感。这些数据表明 Tom20 诱导的变化与基质蛋白的输入之间存在密切关系，表明 Tom20 参与确定输入动力学。然而，这种关系在正常分化过程中被分离，因为 Tom20 的表达保持相对恒定，而输入的 MDH 增加了 12 倍。因此，Tom20 在确定细胞器生物发生过程中的输入方面很重要，但其他机制（例如线粒体内蛋白质降解或核转录）也可能在正常肌肉分化过程中建立最终线粒体表型中发挥作用。
Mitochondrial biogenesis is accompanied by an increased expression of components of the protein import machinery, as well as increased import of proteins destined for the matrix. We evaluated the role of the outer membrane receptor Tom20 by varying its expression and measuring changes in the import of malate dehydrogenase (MDH) in differentiating C2C12 muscle cells. Cells transfected with Tom20 had levels that were twofold higher than in control cells. Labeling of cells followed by immunoprecipitation of MDH revealed equivalent increases in MDH import. This parallelism between import rate and Tom20 levels was also evident as a result of thyroid hormone treatment. Using antisense oligodeoxynucleotides, we inhibited Tom20 expression by 40%, resulting in 40-60% reductions in MDH import. In vitro assays also revealed that import into the matrix was more sensitive to Tom20 inhibition than import into the outer membrane. These data indicate a close relationship between induced changes in Tom20 and the import of a matrix protein, suggesting that Tom20 is involved in determining the kinetics of import. However, this relationship was dissociated during normal differentiation, since the expression of Tom20 remained relatively constant, whereas imported MDH increased 12-fold. Thus Tom20 is important in determining import during organelle biogenesis, but other mechanisms (e.g., intramitochondrial protein degradation or nuclear transcription) likely also play a role in establishing the final mitochondrial phenotype during normal muscle differentiation.