Department of Molecular Epigenetics, Helmholtz Center Munich and Center for Integrated Protein Science Munich (CIPSM), Marchioninistrasse 25, 81377, Munich, Germany.
Institute for Informatics, Ludwig-Maximilians-Universität München, Amalienstr. 17, Munich, 80333, Germany.
Sci Rep. 2017 May 10;7(1):1684. doi: 10.1038/s41598-017-01943-6.
The bromodomain protein Brd4 is an epigenetic reader and plays a critical role in the development and maintenance of leukemia. Brd4 binds to acetylated histone tails and activates transcription by recruiting the positive elongation factor P-TEFb. Small molecule inhibitor JQ1 competitively binds the bromodomains of Brd4 and displaces the protein from acetylated histones. However, it remains unclear whether genes targeted by JQ1 are mainly regulated by Brd4 or by other bromodomain proteins such as Brd2 and Brd3. Here, we describe anti-proliferative dominant-negative Brd4 mutants that compete with the function of distinct Brd4 domains. We used these Brd4 mutants to compare the Brd4-specific transcriptome with the transcriptome of JQ1-treated cells. We found that most JQ1-regulated genes are also regulated by dominant-negative Brd4 mutants, including the mutant that competes with the P-TEFb recruitment function of Brd4. Importantly, JQ1 and dominant-negative Brd4 mutants regulated the same set of target genes of c-Myc, a key regulator of the JQ1 response in leukemia cells. Our results suggest that Brd4 mediates most of the anti-cancer effects of JQ1 and that the major function of Brd4 in this process is the recruitment of P-TEFb. In summary, our studies define the molecular targets of JQ1 in more detail.
溴结构域蛋白 Brd4 是一种表观遗传读码器，在白血病的发生和维持中起着关键作用。Brd4 与乙酰化组蛋白尾部结合，并通过募集正延伸因子 P-TEFb 来激活转录。小分子抑制剂 JQ1 竞争性地结合 Brd4 的溴结构域，并将蛋白从乙酰化组蛋白上置换出来。然而，目前尚不清楚 JQ1 靶向的基因主要是由 Brd4 还是其他溴结构域蛋白（如 Brd2 和 Brd3）调控。在这里，我们描述了抗增殖的显性负 Brd4 突变体，它们与 Brd4 不同结构域的功能竞争。我们使用这些 Brd4 突变体来比较 JQ1 处理细胞的 Brd4 特异性转录组与转录组。我们发现，大多数 JQ1 调控的基因也受显性负 Brd4 突变体的调控，包括与 Brd4 的 P-TEFb 募集功能竞争的突变体。重要的是，JQ1 和显性负 Brd4 突变体调节了 c-Myc 的相同一组靶基因，c-Myc 是白血病细胞中 JQ1 反应的关键调节因子。我们的研究结果表明，Brd4 介导了 JQ1 的大部分抗癌作用，而 Brd4 在这一过程中的主要功能是募集 P-TEFb。总之，我们的研究更详细地定义了 JQ1 的分子靶点。
