Department of Microbiology and Molecular Genetics, McGovern Medical School, University of Texas Health Science Center at Houston, TX 77030, USA.
Nucleic Acids Res. 2019 Sep 5;47(15):7973-7988. doi: 10.1093/nar/gkz476.
DNA damage-induced cell cycle checkpoints serve as surveillance mechanisms to maintain genomic stability, and are regulated by ATM/ATR-mediated signaling pathways that are conserved from yeast to humans. Trypanosoma brucei, an early divergent microbial eukaryote, lacks key components of the conventional DNA damage-induced G2/M cell cycle checkpoint and the spindle assembly checkpoint, and nothing is known about how T. brucei controls its cell cycle checkpoints. Here we discover a kinetochore-based, DNA damage-induced metaphase checkpoint in T. brucei. MMS-induced DNA damage triggers a metaphase arrest by modulating the abundance of the outer kinetochore protein KKIP5 in an Aurora B kinase- and kinetochore-dependent, but ATM/ATR-independent manner. Overexpression of KKIP5 arrests cells at metaphase through stabilizing the mitotic cyclin CYC6 and the cohesin subunit SCC1, mimicking DNA damage-induced metaphase arrest, whereas depletion of KKIP5 alleviates the DNA damage-induced metaphase arrest and causes chromosome mis-segregation and aneuploidy. These findings suggest that trypanosomes employ a novel DNA damage-induced metaphase checkpoint to maintain genomic integrity.
DNA 损伤诱导的细胞周期检查点作为维持基因组稳定性的监测机制，受 ATM/ATR 介导的信号通路调控，该信号通路在从酵母到人等生物中都保守。布氏锥虫是一种早期分化的微生物真核生物，缺乏传统的 DNA 损伤诱导的 G2/M 细胞周期检查点和纺锤体组装检查点的关键成分，目前尚不清楚布氏锥虫如何控制其细胞周期检查点。在这里，我们在布氏锥虫中发现了一种基于着丝粒的 DNA 损伤诱导的中期检查点。MMS 诱导的 DNA 损伤通过调节着丝粒蛋白 KKIP5 的丰度来触发中期阻滞，这种调节方式依赖于 Aurora B 激酶和着丝粒，但不依赖于 ATM/ATR。KKIP5 的过表达通过稳定有丝分裂周期蛋白 CYC6 和黏合蛋白亚基 SCC1 来将细胞阻滞在中期，模拟 DNA 损伤诱导的中期阻滞，而 KKIP5 的耗竭则缓解了 DNA 损伤诱导的中期阻滞，并导致染色体错误分离和非整倍体。这些发现表明，锥虫采用了一种新的 DNA 损伤诱导的中期检查点来维持基因组的完整性。
