Yanas Amber, Shweta Him, Owens Michael C, Liu Kathy Fange, Goldman Yale E
bioRxiv. 2023 Nov 29:2023.11.29.569239. doi: 10.1101/2023.11.29.569239.
DEAD-box helicases, which are crucial for many aspects of RNA metabolism, often contain intrinsically disordered regions (IDRs), whose functions remain unclear. Using multiparameter confocal microscopy, we reveal that sex chromosome-encoded homologous RNA helicases, DDX3X and DDX3Y, form nano-sized RNA-protein clusters (RPCs) that foster their catalytic activities and in cells. The IDRs are critical for the formation of these RPCs. A thorough analysis of the catalytic cycle of DDX3X and DDX3Y by ensemble biochemistry and single molecule photon bursts in the confocal microscope showed that RNA release is a major step that differentiates the unwinding activities of DDX3X and DDX3Y. Our findings provide new insights that the nano-sized helicase RPCs may be the normal state of these helicases under non-stressed conditions that promote their RNA unwinding and act as nucleation points for liquid-liquid phase separation under stress. This mechanism may apply broadly among other members of the DEAD-box helicase family.
DEAD盒解旋酶对RNA代谢的许多方面至关重要，通常包含内在无序区域（IDR），其功能尚不清楚。通过多参数共聚焦显微镜，我们发现性染色体编码的同源RNA解旋酶DDX3X和DDX3Y形成纳米级的RNA-蛋白质簇（RPC），促进其在细胞中的催化活性。IDR对这些RPC的形成至关重要。通过共聚焦显微镜中的整体生物化学和单分子光子爆发对DDX3X和DDX3Y的催化循环进行的全面分析表明，RNA释放是区分DDX3X和DDX3Y解旋活性的主要步骤。我们的研究结果提供了新的见解，即纳米级解旋酶RPC可能是这些解旋酶在非应激条件下的正常状态，促进其RNA解旋，并在应激下作为液-液相分离的成核点。这种机制可能广泛适用于DEAD盒解旋酶家族的其他成员。
