Hahn S L, Wasylyk B, Criqui-Filipe P
Institut de Génétique et de Biologie Moléculaire et Cellulaire, CNRS-INSERM-ULP, Illkirch, France.
Oncogene. 1997 Sep 18;15(12):1489-95. doi: 10.1038/sj.onc.1201301.
Ets transcription factors have Ets DNA binding domains, that have a winged helix-turn-helix structure. ETS-1, the founding member of the family, is regulated by the Ras and Ca2+ signaling pathways and is implicated in various physiological processes leading to cell growth, differentiation and apoptosis. We have identified ETS-1 interacting factors with a yeast two-hybrid screen. The majority of the positive clones turned out to encode the human homologue of the yeast ubiquitin-conjugating enzymes UBC9 and Hus5. In two different yeast assays, ETS-1 interacted with huUBC9. In an in vitro GST 'pull-down' assay, ETS-1 and several other Ets family members complexed with huUBC9. Interestingly, in mammalian cells, coexpression of huUBC9 resulted in a substantial increase in the transcriptional activity of ETS-1. Coexpressed huUBC9 did not affect the ETS-1 protein level, and moreover, a point mutation at Cys93, an amino acid known to be essential for ubiquitination, did not abolish the stimulation of the ETS-1 transcriptional activity. Our results indicate that the modulation of ETS-1 activity by huUBC9 results from processes other than ubiquitination and ETS-1 stabilization.
Ets转录因子具有Ets DNA结合结构域，该结构域具有带翼的螺旋-转角-螺旋结构。ETS-1是该家族的创始成员，受Ras和Ca2+信号通路调控，并参与导致细胞生长、分化和凋亡的各种生理过程。我们通过酵母双杂交筛选鉴定了与ETS-1相互作用的因子。大多数阳性克隆结果显示编码酵母泛素结合酶UBC9和Hus5的人类同源物。在两种不同的酵母分析中，ETS-1与huUBC9相互作用。在体外GST“下拉”分析中，ETS-1和其他几个Ets家族成员与huUBC9形成复合物。有趣的是，在哺乳动物细胞中，huUBC9的共表达导致ETS-1的转录活性大幅增加。共表达的huUBC9不影响ETS-1蛋白水平，此外，已知对泛素化至关重要的Cys93位点的点突变并未消除对ETS-1转录活性的刺激。我们的结果表明，huUBC9对ETS-1活性的调节源于泛素化和ETS-1稳定化以外的过程。
