PDF(Pubmed)
Abstract：
In order to understand the coordinated proteome changes associated with differentiation of a cultured cell pluripotency model, protein expression changes induced by treatment of NT2 embryonal carcinoma cells with retinoic acid were monitored by mass spectrometry. The relative levels of over 5000 proteins were mapped across distinct cell fractions. Analysis of the chromatin fraction revealed major abundance changes among chromatin proteins and epigenetic pathways between the pluripotent and differentiated states. Protein complexes associated with epigenetic regulation of gene expression, chromatin remodelling (e.g., SWI/SNF, NuRD) and histone-modifying enzymes (e.g., Polycomb, MLL) were found to be extensively regulated. We therefore investigated histone modifications before and after differentiation, observing changes in the global levels of lysine acetylation and methylation across the four canonical histone protein families, as well as among variant histones. We identified the set of proteins with affinity to peptides housing the histone marks H3K4me3 and H3K27me3, and found increased levels of chromatin-associated histone H3 tail trimming following differentiation that correlated with increased expression levels of cathepsin proteases. We further found that inhibition of cathepsins B and D reduces histone H3 clipping. Overall, the work reveals a global reorganization of the cell proteome congruent with differentiation, highlighting the key role of multiple epigenetic pathways, and demonstrating a direct link between cathepsin B and D activity and histone modification.
摘要：
为了了解与培养细胞多能性模型的分化相关的协调蛋白质组变化，通过质谱监测维甲酸处理NT2胚胎癌细胞诱导的蛋白表达变化。超过5000种蛋白质的相对水平在不同的细胞部分上被定位。染色质分数的分析揭示了染色质蛋白之间的主要丰度变化以及多能状态和分化状态之间的表观遗传途径。与基因表达的表观遗传调控相关的蛋白质复合物，染色质重塑(例如，SWI/SNF,NuRD)和组蛋白修饰酶(例如，Polycomb,发现MLL)受到广泛调控。因此，我们研究了分化前后的组蛋白修饰，观察四个典型组蛋白家族中赖氨酸乙酰化和甲基化的全球水平的变化，以及变种组蛋白。我们鉴定了一组对含有组蛋白标记H3K4me3和H3K27me3的肽具有亲和力的蛋白质，并发现分化后染色质相关的组蛋白H3尾巴修剪水平增加，这与组织蛋白酶表达水平增加有关。我们进一步发现,组织蛋白酶B和D的抑制减少组蛋白H3剪切。总的来说,这项工作揭示了与分化一致的细胞蛋白质组的全球重组，强调多种表观遗传途径的关键作用，并证明了组织蛋白酶B和D活性与组蛋白修饰之间的直接联系。
