Supakar Anushka, Davis Richoo B, Biswas Subhadip, Yang Sean, Potoyan Davit A, Banerjee Priya R
Department of Biological Sciences, The State University of New York at Buffalo, Buffalo, NY 14260, USA.
Department of Physics, The State University of New York at Buffalo, Buffalo, NY 14260, USA.
J Mol Biol. 2025 Aug 21:169398. doi: 10.1016/j.jmb.2025.169398.
Many transcription factors (TFs) contain intrinsically disordered regions (IDRs) and are thought to form biomolecular condensates in the nucleus. These proteins can be conceptualized as block co-polymers, with the IDRs driving both homotypic and heterotypic protein-protein interactions and the DNA-binding domain (DBD) mediating heterotypic interactions with chromatin. While in vitro studies have predominantly reported micron-scale, spherical condensates in the absence of chromatin, TF condensates in live cells exhibit strikingly different behavior-adopting diverse, nanoscale, often aspherical morphologies and displaying sub-diffusive dynamics. Here, we show that this distinct phase behavior can arise from TF-chromatin interactions, using engineered fusion proteins with tunable IDR-DBD architectures. Specifically, we fused the prion-like domain (PLD) of the SS18 subunit from the mammalian SWI/SNF complex-a domain known to drive homotypic phase separation-to the DBD of the pioneer factor FOXA1. While SS18 on overexpression forms large, spherical condensates in cells, its fusion with FOXA1 leads to condensates that re-localize to chromatin, adopt aspherical morphologies, and exhibit chromatin-wetting behavior. Disruption of DBD-chromatin binding shifts condensate morphology toward a mixed or spherical state, implicating chromatin affinity as a key regulator of condensate coarsening and spatial organization. Coarse-grained simulations recapitulate these observations, revealing a finely balanced interplay between PLD-PLD and DBD-DNA interactions that collectively determine condensate dynamics and structure. Together, our findings demonstrate that chromatin binding is a critical modulator of transcriptional condensate behavior in vivo.
许多转录因子（TFs）含有内在无序区域（IDRs），并被认为在细胞核中形成生物分子凝聚物。这些蛋白质可以被概念化为嵌段共聚物，其中IDRs驱动同型和异型蛋白质-蛋白质相互作用，而DNA结合结构域（DBD）介导与染色质的异型相互作用。虽然体外研究主要报道了在没有染色质的情况下微米级的球形凝聚物，但活细胞中的TF凝聚物表现出截然不同的行为——呈现出多样的、纳米级的、通常为非球形的形态，并表现出亚扩散动力学。在这里，我们使用具有可调节IDR-DBD结构的工程融合蛋白表明，这种独特的相行为可能源于TF-染色质相互作用。具体来说，我们将哺乳动物SWI/SNF复合物中SS18亚基的朊病毒样结构域（PLD）——一个已知能驱动同型相分离的结构域——与先驱因子FOXA1的DBD融合。虽然SS18过表达时在细胞中形成大的球形凝聚物，但其与FOXA1的融合导致凝聚物重新定位于染色质，呈现非球形形态，并表现出染色质湿润行为。DBD-染色质结合的破坏使凝聚物形态向混合或球形状态转变，这表明染色质亲和力是凝聚物粗化和空间组织的关键调节因子。粗粒度模拟重现了这些观察结果，揭示了PLD-PLD和DBD-DNA相互作用之间精细平衡的相互作用，这些相互作用共同决定了凝聚物的动力学和结构。总之，我们的研究结果表明，染色质结合是体内转录凝聚物行为的关键调节因子。
