Department of Human Genetics, Leiden University Medical Center, 2333 ZC Leiden, The Netherlands.
Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), 67404 Illkirch, France.
Genes Dev. 2019 Jun 1;33(11-12):684-704. doi: 10.1101/gad.321943.118. Epub 2019 May 2.
DNA double-strand breaks (DSBs) at RNA polymerase II (RNAPII) transcribed genes lead to inhibition of transcription. The DNA-dependent protein kinase (DNA-PK) complex plays a pivotal role in transcription inhibition at DSBs by stimulating proteasome-dependent eviction of RNAPII at these lesions. How DNA-PK triggers RNAPII eviction to inhibit transcription at DSBs remains unclear. Here we show that the HECT E3 ubiquitin ligase WWP2 associates with components of the DNA-PK and RNAPII complexes and is recruited to DSBs at RNAPII transcribed genes. In response to DSBs, WWP2 targets the RNAPII subunit RPB1 for K48-linked ubiquitylation, thereby driving DNA-PK- and proteasome-dependent eviction of RNAPII. The lack of WWP2 or expression of nonubiquitylatable RPB1 abrogates the binding of nonhomologous end joining (NHEJ) factors, including DNA-PK and XRCC4/DNA ligase IV, and impairs DSB repair. These findings suggest that WWP2 operates in a DNA-PK-dependent shutoff circuitry for RNAPII clearance that promotes DSB repair by protecting the NHEJ machinery from collision with the transcription machinery.
DNA 双链断裂 (DSB) 在 RNA 聚合酶 II (RNAPII) 转录基因处导致转录抑制。DNA 依赖性蛋白激酶 (DNA-PK) 复合物通过刺激蛋白酶体依赖性在这些损伤处驱逐 RNAPII，在 DSB 处的转录抑制中发挥关键作用。DNA-PK 如何触发 RNAPII 驱逐以抑制 DSB 处的转录尚不清楚。在这里，我们表明 HECT E3 泛素连接酶 WWP2 与 DNA-PK 和 RNAPII 复合物的成分相关联，并在 RNAPII 转录基因处被募集到 DSB。响应于 DSB，WWP2 将 RNAPII 亚基 RPB1 靶向 K48 连接的泛素化，从而驱动 DNA-PK 和蛋白酶体依赖性的 RNAPII 驱逐。缺乏 WWP2 或表达不可泛素化的 RPB1 会破坏非同源末端连接 (NHEJ) 因子的结合，包括 DNA-PK 和 XRCC4/DNA 连接酶 IV，并损害 DSB 修复。这些发现表明，WWP2 在 DNA-PK 依赖性的 RNAPII 清除关闭电路中起作用，通过防止 NHEJ 机制与转录机制碰撞来促进 DSB 修复。
