Pramanik Suravi, Chen Yingling, Bhakat Kishor K
Department of Genetics, Cell Biology and Anatomy, University of Nebraska Medical Center, Omaha, NE 68198, USA.
Fred & Pamela Buffett Cancer Center, University of Nebraska Medical Center, Omaha, NE 68198, USA.
Int J Mol Sci. 2024 Nov 27;25(23):12735. doi: 10.3390/ijms252312735.
Endogenous DNA damage occurs throughout the cell cycle, with cells responding differently at various stages. The base excision repair (BER) pathway predominantly repairs damaged bases in the genome. While extensively studied in interphase cells, it is unknown if BER operates in mitosis and how apurinic/apyrimidinic (AP) sites, intermediates in the BER pathway that inhibit transcriptional elongation, are processed for post-mitotic gene reactivation. In this study, using an alkaline comet assay, we demonstrate that BER is inefficient in mitosis and that AP endonuclease 1 (APE1), a key BER enzyme, is required for the repair of damage post-mitosis. We previously demonstrated that APE1 is acetylated (AcAPE1) in the chromatin. Using high-resolution microscopy, we show that AcAPE1 remains associated with specific regions in the condensed chromatin in each of the phases of mitosis. This association presumably occurs via the binding of APE1 to the G-quadruplex structure, a non-canonical DNA structure predominantly present in the transcribed gene regions. Additionally, using a nascent RNA detection strategy, we demonstrate that the knockdown of APE1 delayed the rapid post-mitotic transcriptional reactivation of genes. Our findings highlight the functional importance of APE1 in the mitotic chromosomes to facilitate faster repair of endogenous damage and rapid post-mitotic gene reactivation in daughter cells.
内源性DNA损伤在整个细胞周期中都会发生,细胞在不同阶段会有不同反应。碱基切除修复(BER)途径主要修复基因组中受损的碱基。虽然在间期细胞中已进行了广泛研究,但尚不清楚BER是否在有丝分裂中起作用,以及BER途径中抑制转录延伸的中间产物无嘌呤/无嘧啶(AP)位点是如何在有丝分裂后进行处理以实现基因重新激活的。在本研究中,我们使用碱性彗星试验证明,BER在有丝分裂中效率低下,并且关键的BER酶脱嘌呤/脱嘧啶核酸内切酶1(APE1)是有丝分裂后损伤修复所必需的。我们之前证明APE1在染色质中会发生乙酰化(AcAPE1)。使用高分辨率显微镜,我们发现AcAPE1在有丝分裂的每个阶段都与浓缩染色质中的特定区域保持关联。这种关联可能是通过APE1与G-四链体结构结合而发生的,G-四链体结构是一种主要存在于转录基因区域的非规范DNA结构。此外,使用新生RNA检测策略,我们证明敲低APE1会延迟基因在有丝分裂后的快速转录重新激活。我们的研究结果突出了APE1在有丝分裂染色体中的功能重要性,有助于更快地修复内源性损伤以及子细胞中有丝分裂后的快速基因重新激活。