PALB2-PLK1在细胞周期检测点的调控作用

Genotoxic stress trigger a series of DNA damage signaling events that culminate into cell cycle arrest and DNA repair, dysregulation of which leads to genetic aberrations and tumorigenesis. Notably, exactly how cell proliferation is temporally coupled with DNA repair processes represents a fundamental yet outstanding question. In examining how the Fanconi anemia protein PALB2 elicits its pleotropic functions in DNA repair and cell cycle checkpoint regulation, we noticed that PALB2 becomes hyper-phosphorylated in response to ionising radiation (IR). Indeed, mass spectrometry analysis of PALB2 uncovered DNA damage-regulated phospho-residues that, when rendered phospho-mimetic, resulted in sustained checkpoint activation in cells challenged with a recoverable dose of IR. In support of its role in checkpoint control, we found that IR promoted a protein complex assembly composed of PALB2 and the mitotic driver PLK1. Together, these preliminary data revealed not only a molecular link between PALB2, a core component of the Homologous Recombination (HR)-based DNA repair machinery, with checkpoint components, but also highlights an unanticipated role of PALB2 phosphorylation as a regulatory means for checkpoint maintenance and recovery. We envision that PALB2, upon IR-induced phosphorylation, sequesters and inhibits the PLK1-dependent mitotic entrance and progression. To test this hypothesis, we will: (1) fully characterise the PALB2-PLK1 interaction. We will map the binding interfaceofthe PALB2-PLK1 complex,and will examine how IR enforces its functional interaction in checkpoint maintenance and control; (2) determine role of cell cycle progression in the formation the PALB2-PLK1 complex. We will study whether and how the PALB2-PLK1 module promotes genome stability and cell survival, and will investigate how PALB2 may couple DNA repair with checkpoint control; (3)explore utility of the PALB2-PLK1 module as therapeutic target. We will characterise PALB2 patient mutations and study how they may perturb PALB2 functions in the PLK1-dependent checkpoint regulation. The requirement of PALB2 in cell cycle checkpoint control may account for its tumour-suppressor functions in vivo,and is likely to exert substantial influence to cellular responses to current cancer therapeutics. Given that PLK1 is over-expressed in a wide range of human epithelial malignancies, our proposed work in characterizing the PALB2-PLK1 complex will advance the current understanding of PALB2-dependent cellular responses to genotoxic stress, and will aid in the development and rationalization of exploiting the PALB2-PLK1 module in the early detection and treatment of human cancers.

基因毒性压力触发DNA损伤信号引起细胞周期阻滞及DNA修复，其调控异常导致遗传变异和肿瘤生成。在检测范可尼贫血蛋白PALB2在DNA修复和细胞周期检测点调控复合功能时，我们发现电离辐射（IR）后PALB2被磷酸化。质谱分析显示存在受DNA损伤调控的PALB2磷酸化位点，当其突变为模拟磷酸化位点，细胞IR后会导致持续的检测点激活。随后我们发现IR能促进PALB2与有丝分裂驱动PLK1形成复合物。上述实验揭示DNA修复核心PALB2与检测点组分有分子连接，PALB2磷酸化对检测点的维持和恢复过程有调控。我们推测，电离辐射诱导的PALB2磷酸化，隔绝并抑制依赖于PLK1的有丝分裂的进入及维持。 我们将定义PALB2-PLK1的相互作用；确定PALB2-PLK1的形成在细胞周期的作用；探索其作为治疗靶点的应用。本工作将加深对PALB2功能的理解，并利于探索PALB2-PLK1对癌症早期检测和治疗。

PALB2是范可尼贫血症（Fanconi Anemia）的致病基因之一，又名FANCN。PALB2通过调控DNA修复和细胞周期检测点来保护基因组的完整性。然而我们现在还不确切知道如何将DNA损伤检测以及信号传递和PALB2功能联系起来。有趣的是，我们发现电离辐射（Ionizing Radiation, IR）后，PALB2被超磷酸化。电离辐射会特异性激发PALB2在丝氨酸157和376位点上磷酸化，并且这个磷酸化过程依赖于DNA损伤响应重要激酶ATM（Ataxia telangiectasia mutated）。这揭示了PALB2和ATM介导的DNA损伤响应之间存在着可能的分子机制。与之一致的是，PALB2磷酸化的调控失常会导致持续的DNA损伤响应的激活。PALB2磷酸化也需要乳腺卵巢易感基因BRCA1，说明了BRCA1-PALB2相互作用在分子响应基因毒素压力过程中起到重要作用。总而言之，我们对肿瘤抑制蛋白PALB2磷酸化分析揭示了新的基因组稳定和肿瘤抑制调控机制。