端粒维护是恶性细胞的标志，能让癌细胞无限分裂。在某些癌症中，这是通过替代端粒长度(ALT)途径实现的。尽管ATRX的缺失是ALT癌症的一个普遍特征，但单独缺失ATRX是不够的。因此，其他细胞事件必须是必要的 - 但是对这些次生事件的确切性质仍然是难以捉摸的。在这里，我们报告称DNA上蛋白质（如TOP1、TOP2A和PARP1）的限制导致缺乏ATRX的细胞诱导ALT。我们证明蛋白质限制的化疗药物，如依托泊苷、喜树碱和他拉祖帕，仅诱导ATRX-Null细胞中的ALT标志。此外，我们展示了用G4稳定化药物处理会导致被限制的TOP2A水平增加，从而导致ATRX-Null细胞的ALT诱导。该过程是MUS81-内切酶和断裂诱导的复制依赖型的，这表明蛋白质限制导致复制叉路的停滞，在缺乏ATRX的情况下，这些叉路被异常地处理。最后，我们表明ALT-阳性细胞携带着更高的全基因组被限制的蛋白质负荷，如TOP1，而TOP1的敲除会降低ALT活性。综上所述，这些发现说明蛋白质限制是ATRX缺失恶性肿瘤ALT生物学的一个基本驱动力。©作者(s)2023年。由牛津大学出版社代表核酸研究出版。

Telomere maintenance is a hallmark of malignant cells and allows cancers to divide indefinitely. In some cancers, this is achieved through the alternative lengthening of telomeres (ALT) pathway. Whilst loss of ATRX is a near universal feature of ALT-cancers, it is insufficient in isolation. As such, other cellular events must be necessary - but the exact nature of the secondary events has remained elusive. Here, we report that trapping of proteins (such as TOP1, TOP2A and PARP1) on DNA leads to ALT induction in cells lacking ATRX. We demonstrate that protein-trapping chemotherapeutic agents, such as etoposide, camptothecin and talazoparib, induce ALT markers specifically in ATRX-null cells. Further, we show that treatment with G4-stabilising drugs cause an increase in trapped TOP2A levels which leads to ALT induction in ATRX-null cells. This process is MUS81-endonuclease and break-induced replication dependent, suggesting that protein trapping leads to replication fork stalling, with these forks being aberrantly processed in the absence of ATRX. Finally, we show ALT-positive cells harbour a higher load of genome-wide trapped proteins, such as TOP1, and knockdown of TOP1 reduced ALT activity. Taken together, these findings suggest that protein trapping is a fundamental driving force behind ALT-biology in ATRX-deficient malignancies.© The Author(s) 2023. Published by Oxford University Press on behalf of Nucleic Acids Research.