Miner J H, Wold B J
Division of Biology 156-29, California Institute of Technology, Pasadena 91125.
Mol Cell Biol. 1991 May;11(5):2842-51. doi: 10.1128/mcb.11.5.2842-2851.1991.
In vertebrate development, a prominent feature of several cell lineages is the coupling of cell cycle regulation with terminal differentiation. We have investigated the basis of this relationship in the skeletal muscle lineage by studying the effects of the proliferation-associated regulator, c-myc, on the differentiation of MyoD-initiated myoblasts. Transient cotransfection assays in NIH 3T3 cells using MyoD and c-myc expression vectors demonstrated c-myc suppression of MyoD-initiated differentiation. A stable cell system was also developed in which MyoD expression was constitutive, while myc levels could be elevated conditionally. Induction of this conditional c-myc suppressed myogenesis effectively, even in the presence of MyoD. c-myc suppression also prevented up-regulation of a relative of MyoD, myogenin, which is normally expressed at the onset of differentiation in all muscle cell lines examined and may be essential for differentiation. Additional experiments tested whether failure to differentiate in the presence of myc could be overcome by providing myogenin ectopically. Cotransfection of c-myc with myogenin, MyoD, or a mixture of myogenin and MyoD showed that neither myogenin alone nor myogenin plus MyoD together could bypass the c-myc block. The effects of c-myc were further dissected by showing that c-myc can inhibit differentiation independently of Id, a negative regulator of muscle differentiation. These results lead us to propose that c-myc and Id constitute independent negative regulators of muscle differentiation, while myogenin and any of the other three related myogenic factors (MyoD, Myf-5, and MRF4/herculin/Myf-6) act as positive regulators.
在脊椎动物发育过程中，几个细胞谱系的一个显著特征是细胞周期调控与终末分化的耦合。我们通过研究增殖相关调节因子c-myc对MyoD起始的成肌细胞分化的影响，来探究骨骼肌谱系中这种关系的基础。使用MyoD和c-myc表达载体在NIH 3T3细胞中进行的瞬时共转染实验表明，c-myc抑制MyoD起始的分化。还建立了一个稳定的细胞系统，其中MyoD表达是组成性的，而myc水平可以有条件地升高。即使存在MyoD，这种条件性c-myc的诱导也能有效地抑制肌生成。c-myc的抑制还阻止了MyoD的一个相关蛋白——生肌调节因子的上调，生肌调节因子通常在所有检测的肌肉细胞系分化开始时表达，可能对分化至关重要。额外的实验测试了在存在myc的情况下无法分化是否可以通过异位提供生肌调节因子来克服。c-myc与生肌调节因子、MyoD或生肌调节因子和MyoD的混合物共转染表明，单独的生肌调节因子或生肌调节因子加MyoD都不能绕过c-myc的阻断。通过表明c-myc可以独立于肌肉分化的负调节因子Id抑制分化，进一步剖析了c-myc的作用。这些结果使我们提出，c-myc和Id构成肌肉分化的独立负调节因子，而生肌调节因子和其他三个相关的生肌因子（MyoD、Myf-5和MRF4/赫库兰/Myf-6）中的任何一个都作为正调节因子。
