Lingbeck Jody M, Trausch-Azar Julie S, Ciechanover Aaron, Schwartz Alan L
Edward Mallinckrodt Department of Pediatrics and Molecular Biology and Pharmacology, Washington University School of Medicine and St. Louis Children's Hospital, St. Louis, Missouri 63110, USA.
FASEB J. 2008 Jun;22(6):1694-701. doi: 10.1096/fj.07-095000. Epub 2008 Jan 15.
MyoD, a skeletal muscle transcription factor, is rapidly degraded by the ubiquitin-proteasome system. MyoD interacts with ubiquitously expressed E2A or inhibitor of DNA binding (Id) proteins to activate or inhibit transcription, respectively. Furthermore, MyoD has been shown to modulate the ubiquitin-mediated degradation of Id1 and E2A proteins, E12 and E47. The molecular mechanisms governing these events are not clear but are hypothesized to occur via heterodimer formation. Fluorescence resonance energy transfer (FRET) is a technique for evaluation of protein-protein interactions in vivo. Using acceptor photobleaching FRET and chimeric proteins composed of MyoD, Id1, E12, E47, E12(NLS), or MyoD(NLS) and either cyan fluorescent protein or yellow fluorescent protein, we show that each of the wild-type proteins is capable of homodimerization. In addition, heterodimers form between Id1 and E2A proteins, as well as between MyoD and E2A proteins. The Id1:E2A interaction is stronger than the MyoD:E2A interaction, which is consistent with the notion that inhibition of MyoD action occurs by the sequestration of E2A proteins by Id. The stronger interaction of Id1 with E2A may also explain the decrease in the rate of ubiquitin-proteasome degradation of Id1 that is significantly greater than that of MyoD when E2A proteins are abundant. Thus, these studies extend our understanding of the molecular mechanisms of MyoD action.
肌分化因子（MyoD）是一种骨骼肌转录因子，可被泛素 - 蛋白酶体系统快速降解。MyoD分别与普遍表达的E2A或DNA结合抑制剂（Id）蛋白相互作用，以激活或抑制转录。此外，已有研究表明MyoD可调节Id1、E2A蛋白、E12和E47的泛素介导降解。调控这些事件的分子机制尚不清楚，但据推测是通过异源二聚体形成发生的。荧光共振能量转移（FRET）是一种用于评估体内蛋白质 - 蛋白质相互作用的技术。使用受体光漂白FRET以及由MyoD、Id1、E12、E47、E12（核定位信号）或MyoD（核定位信号）与青色荧光蛋白或黄色荧光蛋白组成的嵌合蛋白，我们发现每种野生型蛋白都能够形成同二聚体。此外，Id1与E2A蛋白之间以及MyoD与E2A蛋白之间会形成异二聚体。Id1：E2A的相互作用强于MyoD：E2A的相互作用，这与Id通过隔离E2A蛋白来抑制MyoD作用的观点一致。当E2A蛋白丰富时，Id1与E2A更强的相互作用也可能解释了Id1泛素 - 蛋白酶体降解速率的降低，其降低幅度明显大于MyoD。因此，这些研究扩展了我们对MyoD作用分子机制的理解。
