Department of Genetics, Cell Biology and Anatomy, University of Nebraska Medical Center, 985805 Nebraska Medical Center, Omaha, NE 68198, USA.
Biol Chem. 2010 Jan;391(1):9-19. doi: 10.1515/BC.2010.004.
Ecdysoneless (Ecd) is an evolutionarily conserved protein and its function is essential for embryonic development in Drosophila and cell growth in yeast. However, its function has remained unknown until recently. Studies in yeast suggested a potential role of Ecd in transcription; however, Ecd lacks a DNA-binding domain. Using a GAL4-luciferase reporter assay and a GAL4 DNA-binding domain fusion with Ecd or its mutants, we present evidence that human Ecd has a transactivation activity in its C-terminal region. Importantly, further analyses using point mutants showed that a single amino acid change at either Asp-484 or Leu-489 essentially completely abolishes the transactivation activity of Ecd. We further demonstrate that Ecd interacts with p300, a histone acetyltransferase, and coexpression of Ecd with p300 enhances the Ecd-mediated transactivation activity. Ecd localizes to both nucleus and cytoplasm and shuttles between the nucleus and cytoplasm; however, it exhibits strong nuclear export. Based on previous yeast studies and evidence provided here, we suggest that Ecd functions as a transcriptional regulator. Our results indicate an important function of human Ecd and provide a basis to explore the transcriptional partners of Ecd.
蜕皮激素缺失（Ecd）是一种进化上保守的蛋白质，其功能对于果蝇的胚胎发育和酵母的细胞生长是必不可少的。然而，直到最近，其功能仍然未知。酵母研究表明 Ecd 可能在转录中发挥作用；然而，Ecd 缺乏 DNA 结合域。我们使用 GAL4-荧光素酶报告基因测定和 GAL4 DNA 结合域融合 Ecd 或其突变体的方法，提供了证据表明人 Ecd 在其 C 末端区域具有转录激活活性。重要的是，使用点突变体的进一步分析表明，在 Asp-484 或 Leu-489 处的单个氨基酸变化基本上完全消除了 Ecd 的转录激活活性。我们进一步证明 Ecd 与组蛋白乙酰转移酶 p300 相互作用，并且 Ecd 与 p300 的共表达增强了 Ecd 介导的转录激活活性。Ecd 定位于细胞核和细胞质中，并在细胞核和细胞质之间穿梭；然而，它表现出强烈的核输出。基于先前的酵母研究和此处提供的证据，我们认为 Ecd 作为转录调节剂发挥作用。我们的结果表明了人 Ecd 的重要功能，并为探索 Ecd 的转录伙伴提供了基础。
