Abstract：
ADP-ribosylation (ADPr) signaling plays a crucial role in DNA damage response. Inhibitors against the main enzyme catalyzing ADPr after DNA damage, poly(ADP-ribose) polymerase 1 (PARP1), are used to treat patients with breast cancer harboring BRCA1/2 mutations. However, resistance to PARP inhibitors (PARPi) is a major obstacle in treating patients. To understand the role of ADPr in PARPi sensitivity, we use liquid chromatography-tandem mass spectrometry (LC-MS/MS) to analyze ADPr in six breast cancer cell lines exhibiting different PARPi sensitivities. We identify 1,632 sites on 777 proteins across all cell lines, primarily on serine residues, with site-specific overlap of targeted residues across DNA-damage-related proteins across all cell lines, demonstrating high conservation of serine ADPr-signaling networks upon DNA damage. Furthermore, we observe site-specific differences in ADPr intensities in PARPi-sensitive BRCA mutants and unique ADPr sites in PARPi-resistant BRCA-mutant HCC1937 cells, which have low poly(ADP-ribose) glycohydrolase (PARG) levels and longer ADPr chains on PARP1.
摘要：
ADP核糖基化（ADPr）信号在DNA损伤反应中起着至关重要的作用。针对DNA损伤后催化ADPr的主要酶的抑制剂，聚(ADP-核糖)聚合酶1(PARP1)，用于治疗携带BRCA1/2突变的乳腺癌患者。然而,对PARP抑制剂（PARPi）的耐药性是治疗患者的主要障碍。为了了解ADPr在PARPi敏感性中的作用，我们使用液相色谱-串联质谱（LC-MS/MS）分析了六种表现出不同PARPi敏感性的乳腺癌细胞系中的ADPr。我们在所有细胞系的777个蛋白质上鉴定了1,632个位点，主要是在丝氨酸残基上，所有细胞系中DNA损伤相关蛋白的靶向残基存在位点特异性重叠，证明DNA损伤后丝氨酸ADPr信号网络的高度保守性。此外，我们观察到PARPi敏感BRCA突变体中ADPr强度的位点特异性差异和PARPi抗性BRCA突变体HCC1937细胞中独特的ADPr位点，其具有低的聚(ADP-核糖)糖水解酶(PARG)水平和PARP1上更长的ADPr链。
