James Leonard, Eisenman Robert N
Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA.
Proc Natl Acad Sci U S A. 2002 Aug 6;99(16):10429-34. doi: 10.1073/pnas.162369299. Epub 2002 Jul 29.
The opposing transcriptional activities of the basic-helix-loop-helix-leucine zipper proteins Myc and Mad, taken together with information related to their expression patterns and biological effects, have led to a model of the Myc/Max/Mad network in which Myc and Mad proteins function as antagonists. This antagonism is presumed to operate at the level of genes targeted by these complexes, where Myc:Max activates and Mad:Max represses expression of the same set of genes. However, a detailed analysis of the DNA-binding preferences for Mad proteins has not been performed. Furthermore, the model does not address the findings that Myc:Max indirectly represses transcription of several regulatory genes. To examine these issues relating to DNA-binding specificity and biological responses, we have determined the DNA-binding preferences of Mad1 by using selection and amplification of randomized oligonucleotides and demonstrated that its intrinsic specificity is identical with that of c-Myc. We have also used a chimeric Myc protein, containing a substitution of the entire Mad basic-helix-loop-helix-leucine zipper motif, and shown that it can reproduce the growth-promoting activities of Myc, but not its apoptotic function. Our results suggest that Myc and Mad, although possessing identical in vitro DNA-binding specificities, do not have an identical set of target genes in vivo, and that apoptosis is one biological outcome in which the transcriptional effects of Myc are not directly antagonized by those of Mad.
碱性螺旋-环-螺旋-亮氨酸拉链蛋白Myc和Mad具有相反的转录活性，再结合与其表达模式和生物学效应相关的信息，形成了一个Myc/Max/Mad网络模型，其中Myc和Mad蛋白作为拮抗剂发挥作用。这种拮抗作用被认为在这些复合物靶向的基因水平上起作用，在该水平上，Myc:Max激活而Mad:Max抑制同一组基因的表达。然而，尚未对Mad蛋白的DNA结合偏好进行详细分析。此外，该模型未涉及Myc:Max间接抑制几个调控基因转录的研究结果。为了研究与DNA结合特异性和生物学反应相关的这些问题，我们通过随机寡核苷酸的选择和扩增确定了Mad1的DNA结合偏好，并证明其内在特异性与c-Myc相同。我们还使用了一种嵌合Myc蛋白，其包含整个Mad碱性螺旋-环-螺旋-亮氨酸拉链基序的替代，并表明它可以重现Myc的促生长活性，但不能重现其凋亡功能。我们的结果表明，Myc和Mad虽然在体外具有相同的DNA结合特异性，但在体内没有相同的靶基因集，并且凋亡是一种生物学结果，其中Myc的转录效应不会被Mad的转录效应直接拮抗。
