诱导粒细胞分化的研究有助于揭示细胞成熟的分子机制。核蛋白质组代表了调节分子的丰富来源,包括转录因子(TFs)。重要的是在分化过程的早期阶段了解分子扰动。通过使用等量异位标记应用蛋白质组学定量分析,我们发现214、319、376、426和391蛋白质的含量在3、6、9、12和72小时发生了变化,分别,与全反式维甲酸(ATRA)处理下HL-60细胞核部分的0h相比。从1860年鉴定的核蛋白,231个蛋白质被注释为具有转录因子(TF)活性的蛋白质。六个TF(RREB1、SRCAP、在ATRA治疗后的早期时间点(3-12小时),CCDC124,TRIM24,BRD7和BUD31)下调,三个TFsEWSR1,ENO1和FUS上调。生物信息学注释表明HL-60核蛋白质组参与RUNX1触发途径的DNA损伤识别,在p53调节途径中。通过使用稳定的同位素标记的肽标准(MRM/SIS)进行预定的多反应监测,我们发现以下蛋白质的含量持续增加:PRAM1,CEPBP,RBPJ,ATRA诱导的粒细胞分化过程中HL-60细胞核部分中的HIC1。在STAT1,CASP3,PARP1和PRKDC蛋白的情况下,在ATRA治疗后的早期时间点(3-12小时)观察到它们的含量短暂增加.获得的粒细胞分化过程中核蛋白质组组成和动力学的数据可能有利于开发具有突变p53基因的白血病的新治疗方法。
Studies of induced granulocytic differentiation help to reveal molecular mechanisms of cell maturation. The nuclear proteome represents a rich source of regulatory molecules, including transcription factors (TFs). It is important to have an understanding of molecular perturbations at the early stages of the differentiation processes. By applying the proteomic quantitative profiling using isobaric labeling, we found that the contents of 214, 319, 376, 426, and 391 proteins were altered at 3, 6, 9, 12, and 72 h, respectively, compared to 0 h in the HL-60 cell nuclear fraction under all-trans-retinoid acid (ATRA) treatment. From 1860 identified nuclear proteins, 231 proteins were annotated as proteins with transcription factor (TF) activity. Six TFs (RREB1, SRCAP, CCDC124, TRIM24, BRD7, and BUD31) were downregulated and three TFs EWSR1, ENO1, and FUS were upregulated at early time points (3-12 h) after ATRA treatment. Bioinformatic annotation indicates involvement of the HL-60 nuclear proteome in DNA damage recognition in the RUNX1-triggered pathway, and in the p53-regulation pathway. By applying scheduled multiple reaction monitoring using stable isotopically labeled peptide standards (MRM/SIS), we found a persistent increase in the content of the following proteins: PRAM1, CEPBP, RBPJ, and HIC1 in the HL-60 cell nuclear fraction during ATRA-induced granulocytic differentiation. In the case of STAT1, CASP3, PARP1, and PRKDC proteins, a transient increase in their content was observed at early time points (3-12 h) after the ATRA treatment. Obtained data on nuclear proteome composition and dynamics during granulocytic differentiation could be beneficial for the development of new treatment approaches for leukemias with the mutated p53 gene.