Berroyer Alexandra, Bacolla Albino, Tainer John A, Kim Nayun
Department of Microbiology and Molecular Genetics, University of Texas Health Science Center at Houston, Houston, TX 77030, USA.
MD Anderson Cancer Center UT Health Graduate School of Biomedical Sciences, Houston, TX, 77030, USA.
Microb Cell. 2022 Jan 31;9(3):52-68. doi: 10.15698/mic2022.03.771. eCollection 2022 Mar 7.
Topoisomerase 1 (Top1) removes transcription-associated helical stress to suppress G4-formation and its induced recombination at genomic loci containing guanine-run containing sequences. Interestingly, Top1 binds tightly to G4 structures, and its inhibition or depletion can cause elevated instability at these genomic loci. Top1 is targeted by the widely used anti-cancer chemotherapeutic camptothecin (CPT) and its derivatives, which stabilize Top1 covalently attached on a DNA nick and prevent the re-ligation step. Here we investigated how CPT-resistance conferring Top1 mutants, which emerge in cancer patients and cells treated with CPT, affect G4-induced genomic instability in . We found that Top1 mutants form stable complexes with G4 DNA and that expression of Top1 cleavage-defective mutants but not a DNA-binding-defective mutant lead to significantly elevated instability at a G4-forming genomic locus. Elevated recombination rates were partly suppressed by their proteolytic removal by SPRTN homolog Wss1 SUMO-dependent metalloprotease . Furthermore, interaction between G4-DNA binding protein Nsr1, a homolog to clinically-relevant human nucleolin, and Top1 mutants lead to a synergistic increase in G4-associated recombination. These results in the yeast system are strengthened by our cancer genome data analyses showing that functionally detrimental mutations in Top1 correlate with an enrichment of mutations at G4 motifs. Our collective experimental and computational findings point to cooperative binding of Top1 cleavage-defective mutants and Nsr1 as promoting DNA replication blockage and exacerbating genomic instability at G4-motifs, thus complicating patient treatment.
拓扑异构酶1（Top1）消除转录相关的螺旋应力，以抑制G4形成及其在含有鸟嘌呤重复序列的基因组位点诱导的重组。有趣的是，Top1与G4结构紧密结合，其抑制或缺失会导致这些基因组位点的不稳定性增加。Top1是广泛使用的抗癌化疗药物喜树碱（CPT）及其衍生物的作用靶点，这些药物可稳定共价连接在DNA切口上的Top1，并阻止重新连接步骤。在这里，我们研究了在癌症患者和接受CPT治疗的细胞中出现的赋予CPT抗性的Top1突变体如何影响G4诱导的基因组不稳定性。我们发现Top1突变体与G4 DNA形成稳定复合物，并且Top1切割缺陷突变体而非DNA结合缺陷突变体的表达导致在形成G4的基因组位点的不稳定性显著增加。通过SPRTN同源物Wss1 SUMO依赖性金属蛋白酶对其进行蛋白水解去除，部分抑制了重组率的升高。此外，G4-DNA结合蛋白Nsr1（临床相关人类核仁素的同源物）与Top1突变体之间的相互作用导致G4相关重组的协同增加。我们对癌症基因组数据分析加强了酵母系统中的这些结果，表明Top1中的功能有害突变与G4基序处的突变富集相关。我们的实验和计算结果共同表明，Top1切割缺陷突变体和Nsr1的协同结合促进了DNA复制阻滞并加剧了G4基序处的基因组不稳定性，从而使患者治疗复杂化。
