Department of Biomedical Science, College of Osteopathic Medicine, New York Institute of Technology, Old Westbury, NY 11568.
Department of Cell Biology, Albert Einstein College of Medicine, New York, NY 10461.
Proc Natl Acad Sci U S A. 2017 Jul 18;114(29):E5940-E5949. doi: 10.1073/pnas.1708065114. Epub 2017 Jul 3.
In the mammalian genome, certain genomic loci/regions pose greater challenges to the DNA replication machinery (i.e., the replisome) than others. Such known genomic loci/regions include centromeres, common fragile sites, subtelomeres, and telomeres. However, the detailed mechanism of how mammalian cells cope with the replication stress at these loci/regions is largely unknown. Here we show that depletion of FANCM, or of one of its obligatory binding partners, FAAP24, MHF1, and MHF2, induces replication stress primarily at the telomeres of cells that use the alternative lengthening of telomeres (ALT) pathway as their telomere maintenance mechanism. Using the telomere-specific single-molecule analysis of replicated DNA technique, we found that depletion of FANCM dramatically reduces the replication efficiency at ALT telomeres. We further show that FANCM, BRCA1, and BLM are actively recruited to the ALT telomeres that are experiencing replication stress and that the recruitment of BRCA1 and BLM to these damaged telomeres is interdependent and is regulated by both ATR and Chk1. Mechanistically, we demonstrated that, in FANCM-depleted ALT cells, BRCA1 and BLM help to resolve the telomeric replication stress by stimulating DNA end resection and homologous recombination (HR). Consistent with their roles in resolving the replication stress induced by FANCM deficiency, simultaneous depletion of BLM and FANCM, or of BRCA1 and FANCM, leads to increased micronuclei formation and synthetic lethality in ALT cells. We propose that these synthetic lethal interactions can be explored for targeting the ALT cancers.
在哺乳动物基因组中，某些基因组位点/区域比其他区域对 DNA 复制机制（即复制体）构成更大的挑战。已知的基因组位点/区域包括着丝粒、常见的脆弱位点、端粒和端粒。然而，哺乳动物细胞如何应对这些位点/区域的复制应激的详细机制在很大程度上尚不清楚。在这里，我们发现 FANCM 或其必需结合伙伴之一 FAAP24、MHF1 和 MHF2 的耗竭主要诱导使用端粒延长替代（ALT）途径作为其端粒维持机制的细胞中的端粒处的复制应激。使用复制 DNA的端粒特异性单分子分析技术，我们发现 FANCM 的耗竭显着降低了 ALT 端粒处的复制效率。我们进一步表明，FANCM、BRCA1 和 BLM 被主动募集到经历复制应激的 ALT 端粒，并且 BRCA1 和 BLM 到这些受损端粒的募集是相互依赖的，并且受到 ATR 和 Chk1 的调节。从机制上讲，我们证明，在 FANCM 耗竭的 ALT 细胞中，BRCA1 和 BLM 通过刺激 DNA 末端切除和同源重组（HR）有助于解决端粒复制应激。与它们在解决由 FANCM 缺乏引起的复制应激中的作用一致，BLM 和 FANCM 的同时耗竭，或 BRCA1 和 FANCM 的同时耗竭，导致 ALT 细胞中微核形成增加和合成致死。我们提出可以探索这些合成致死相互作用来靶向 ALT 癌症。
