Transcription dynamics prevents RNA-mediated genomic instability through SRPK2-dependent DDX23 phosphorylation

Genomic instability is frequently caused by nucleic acids structures formed during transcription termed R-loops. Despite their harmful potential, mechanisms that sense, signal and suppress these structures remain elusive. Here we report that oscillations in transcription dynamics are a major sensor of R-loops. We show that pausing of RNA polymerase II (RNA Pol II) initiates a signaling cascade whereby the serine/arginine protein kinase 2 (SRPK2) phosphorylates the DDX23 helicase culminating in the suppression of R-loops. We show that in the absence of either SRPK2 or DDX23, accumulation of R-loops leads to massive genomic instability revealed by high levels of DNA double-strand breaks (DSBs). Importantly, we found DDX23 mutations in several cancers and detected homozygous deletions of the entire DDX23 locus in 10 (17%) adenoid cystic carcinoma samples. Our results unravel molecular details of a novel link between transcription dynamics and RNA-mediated genomic instability that may play important roles in cancer development. Overall design: To map the genomic occupancy of splicing factors SRPK1 and SRPK2, ChIP-seq was performed in MCF10A cells.

基因组不稳定性（Genomic instability）常由转录过程中形成的被称为R环（R-loops）的核酸结构所引发。尽管这类结构具有潜在危害，但其感知、信号传导与抑制机制仍未阐明。本研究证实，转录动态的振荡是R环的主要感知因子。我们发现，RNA聚合酶II（RNA polymerase II, RNA Pol II）的暂停会启动一条信号级联反应，其中丝氨酸/精氨酸蛋白激酶2（serine/arginine protein kinase 2, SRPK2）会对DDX23解旋酶进行磷酸化，最终实现R环的抑制。实验表明，当SRPK2或DDX23缺失时，R环的积累会引发大规模基因组不稳定性，这可通过高水平的DNA双链断裂（DNA double-strand breaks, DSBs）得以体现。值得注意的是，我们在多种癌症中发现了DDX23突变，并在10例（17%）腺样囊性癌样本中检测到了整个DDX23基因座的纯合缺失。本研究揭示了转录动态与RNA介导的基因组不稳定性之间全新关联的分子细节，这一关联可能在癌症发生发展中发挥重要作用。整体实验设计：为绘制剪接因子SRPK1与SRPK2的基因组结合位点，我们在MCF10A细胞中开展了染色质免疫共沉淀测序（chromatin immunoprecipitation sequencing, ChIP-seq）实验。