BRD4-NUT, a driver fusion mutant in rare and highly aggressive NUT carcinoma, acts in aberrant transcription of anti-differentiation genes by recruiting histone acetyltransferase (HAT) p300 and promoting p300-driven histone hyperacetylation and nuclear condensation in chromatin. However, the molecular basis of how BRD4-NUT recruits and activates p300 remains elusive. Here, we report that BRD4-NUT contains two transactivation domains (TADs) in NUT that bind to the TAZ2 domain in p300. Our NMR structures reveal that NUT TADs adopt amphipathic helices when bound to the four-helical bundle TAZ2 domain. The NUT protein forms liquid-like droplets in-vitro that are enhanced by TAZ2 binding in 1:2 stoichiometry. The TAD/TAZ2 bipartite binding in BRD4-NUT/p300 triggers allosteric activation of p300 and acetylation-driven liquid-like condensation on chromatin that comprise histone H3 lysine 27 and 18 acetylation and transcription proteins BRD4L/S, CDK9, MED1, and RNA polymerase II. The BRD4-NUT/p300 chromatin condensation is key for activating transcription of pro-proliferation genes such as ALX1, resulting ALX1/Snail signaling and epithelial-to-mesenchymal transition. Our study provides a previously underappreciated structural mechanism illuminating BRD4-NUT’s bipartite p300 recruitment and activation in NUT carcinoma that nucleates a feed-forward loop for propagating histone hyperacetylation and chromatin condensation to sustain aberrant anti-differentiation gene transcription and perpetual tumor cell growth.

BRD4-NUT 是罕见且高度侵袭性 NUT 癌中的驱动融合突变体，通过招募组蛋白乙酰转移酶 (HAT) p300 并促进 p300 驱动的组蛋白过度乙酰化和染色质中的核浓缩，在抗分化基因的异常转录中发挥作用。然而，BRD4-NUT 如何招募和激活 p300 的分子基础仍然难以捉摸。在这里，我们报告 BRD4-NUT 在 NUT 中包含两个反式激活域 (TAD)，它们与 p300 中的 TAZ2 域结合。我们的 NMR 结构表明，NUT TAD 在与四螺旋束 TAZ2 结构域结合时采用两亲性螺旋。 NUT 蛋白在体外形成液体样液滴，TAZ2 以 1:2 化学计量比结合可增强这种液滴的结合能力。 BRD4-NUT/p300 中的 TAD/TAZ2 二分结合触发 p300 的变构激活和乙酰化驱动的染色质上的液体状缩合，该染色质包含组蛋白 H3 赖氨酸 27 和 18 乙酰化和转录蛋白 BRD4L/S、CDK9、MED1 和 RNA 聚合酶二. BRD4-NUT/p300 染色质浓缩是激活ALX1等促增殖基因转录的关键，从而产生 ALX1/Snail 信号传导和上皮间质转化。我们的研究提供了一种先前未被充分认识的结构机制，阐明了 BRD4-NUT 在 NUT 癌中的二分 p300 募集和激活，该机制形成了一个前馈环，用于传播组蛋白高度乙酰化和染色质浓缩，以维持异常的抗分化基因转录和永久肿瘤细胞生长。