Department of Cancer Biology and Genetics, College of Medicine, The Ohio State University, Columbus, OH 43210, USA.
Hum Mol Genet. 2018 Apr 1;27(7):1241-1251. doi: 10.1093/hmg/ddy038.
The recQ-like helicase BLM interacts directly with topoisomerase IIα to regulate chromosome breakage in human cells. We demonstrate that a phosphosite tri-serine cluster (S577/S579/S580) within the BLM topoisomerase IIα-interaction region is required for this function. Enzymatic activities of BLM and topoisomerase IIα are reciprocally stimulated in vitro by ten-fold for topoisomerase IIα decatenation/relaxation activity and three-fold for BLM unwinding of forked DNA duplex substrates. A BLM transgene encoding alanine substitutions of the tri-serine cluster in BLM-/- transfected cells increases micronuclei, DNA double strand breaks and anaphase ultra-fine bridges (UFBs), and decreases cellular co-localization of BLM with topoisomerase IIα. In vitro, these substitutions significantly reduce the topoisomerase IIα-mediated stimulation of BLM unwinding of forked DNA duplexes. Substitution of the tri-serine cluster with aspartic acids to mimic serine phosphorylation reverses these effects in vitro and in vivo. Our findings implicate the modification of this BLM tri-serine cluster in regulating chromosomal stability.
RecQ 样解旋酶 BLM 与拓扑异构酶 IIα 直接相互作用，以调节人类细胞中的染色体断裂。我们证明，BLM 与拓扑异构酶 IIα 相互作用区域内的磷酸丝氨酸三肽簇（S577/S579/S580）是该功能所必需的。体外实验中，BLM 和拓扑异构酶 IIα 的酶活性相互刺激十倍，以提高拓扑异构酶 IIα 的解连环/松弛活性，三倍以提高 BLM 分叉 DNA 双链底物的解旋活性。BLM 转染细胞中 BLM-/- 三丝氨酸簇的丙氨酸取代编码基因增加了微核、DNA 双链断裂和后期超细线（UFB），并减少了 BLM 与拓扑异构酶 IIα 的细胞共定位。在体外，这些取代显著降低了拓扑异构酶 IIα 介导的 BLM 对分叉 DNA 双链的解旋刺激。用天冬氨酸替代三丝氨酸簇模拟丝氨酸磷酸化，在体外和体内均可逆转这些效应。我们的研究结果表明，该 BLM 三丝氨酸簇的修饰可调节染色体稳定性。
