Department of Biochemistry & Biophysics, University of Kalyani, Kalyani 741235, India.
Department of Biochemistry & Biophysics, University of Kalyani, Kalyani 741235, India.
Life Sci. 2021 Jul 15;277:119556. doi: 10.1016/j.lfs.2021.119556. Epub 2021 May 1.
Poly(ADP-ribose) polymerase1 (PARP1) interacts and poly(ADP-ribosyl)ates telomere repeat binding factor 2 (TRF2), which acts as a platform to recruit a large number of proteins at the telomere. Since the discovery of TRF2-SLX4 interaction, SLX4 is becoming the key player in telomere length (TL) maintenance and repair by telomere sister chromatid exchange (T-SCE). Defective TL maintenance pathway results in a spectrum of diseases called telomeropathies like dyskeratosis congenita, aplastic anemia, fanconi anemia, cancer. We aimed to study the role of SLX4 and PARP1 on each other's telomere localization, T-SCE, and TL maintenance in human telomerase-negative osteosarcoma U2OS cells to understand some of the molecular mechanisms of telomere homeostasis.
We checked the role of SLX4 and PARP1 on each other's telomere localization by telomere immunofluorescence. We have cloned full-length wild-type and catalytically inactive mutant PARP1 to understand the role of poly(ADP-ribosyl)ation reaction by PARP1 in telomere length homeostasis. TL of U2OS cells was measured by Q-FISH. T-SCE was measured by Telomere-FISH.
We observed that SLX4 has no role in the telomere localization of PARP1. However, reduced localization of SLX4 at undamaged and damaged telomere upon PARP1 depletion was reversed by overexpression of exogenous wild-type PARP1 but not by overexpression of catalytically inactive mutant PARP1. PARP1 depletion synergized SLX4 depletion-mediated reduction of T-SCE. Furthermore, SLX4 depletion elongated TL, and combined insufficiency of SLX4 with PARP1 further elongated TL.
So, PARP1 controls SLX4 recruitment at telomere by poly(ADP-ribosyl)ation reaction, thereby regulating SLX4-mediated T-SCE and TL homeostasis.
聚（ADP-核糖）聚合酶 1（PARP1）与端粒重复结合因子 2（TRF2）相互作用并聚（ADP-核糖基）化，TRF2 作为平台在端粒处募集大量蛋白质。自从发现 TRF2-SLX4 相互作用以来，SLX4 通过端粒姐妹染色单体交换（T-SCE）成为端粒长度（TL）维持和修复的关键因素。TL 维持途径的缺陷导致一系列疾病，称为端粒体病，如先天性角化不良、再生障碍性贫血、范可尼贫血、癌症。我们旨在研究 SLX4 和 PARP1 对彼此端粒定位、T-SCE 和人端粒酶阴性骨肉瘤 U2OS 细胞中 TL 维持的作用，以了解端粒动态平衡的一些分子机制。
我们通过端粒免疫荧光检查 SLX4 和 PARP1 对彼此端粒定位的作用。我们已经克隆了全长野生型和催化失活突变型 PARP1，以了解 PARP1 的聚（ADP-核糖基）化反应在端粒长度动态平衡中的作用。通过 Q-FISH 测量 U2OS 细胞的 TL。通过端粒-FISH 测量 T-SCE。
我们观察到 SLX4 对 PARP1 的端粒定位没有作用。然而，PARP1 耗尽后，未受损和受损端粒上 SLX4 的定位减少，通过过表达外源性野生型 PARP1 而不是过表达催化失活突变型 PARP1得到逆转。PARP1 耗竭与 SLX4 耗竭介导的 T-SCE 减少协同作用。此外，SLX4 耗竭使 TL 伸长，并且 SLX4 与 PARP1 的联合不足进一步使 TL 伸长。
因此，PARP1 通过聚（ADP-核糖基）化反应控制 SLX4 在端粒上的募集，从而调节 SLX4 介导的 T-SCE 和 TL 动态平衡。
