Quantitative Proteomics Uncovers Novel Factors Involved in Developmental Differentiation of Trypanosoma brucei

Developmental differentiation is a universal biological process that allows cells to adapt to different environments to perform specific functions. African trypanosomes progress through a tightly regulated life cycle in order to survive in different host environments when they shuttle between an insect vector and a vertebrate host. Transcriptomics has been useful to gain insight into RNA changes during stage transitions; however, RNA levels are only a moderate proxy for protein abundance in trypanosomes. We quantified 4270 protein groups during stage differentiation from the mammalian-infective to the insect form and provide classification for their expression profiles during development. Our label-free quantitative proteomics study revealed previously unknown components of the differentiation machinery that are involved in essential biological processes such as signaling, posttranslational protein modifications, trafficking and nuclear transport. Furthermore, guided by our proteomic survey, we identified the cause of the previously observed differentiation impairment in the histone methyltransferase DOT1B knock-out strain as it is required for accurate karyokinesis in the first cell division during differentiation. This epigenetic regulator is likely involved in essential chromatin restructuring during developmental differentiation, which might also be important for differentiation in higher eukaryotic cells. Our proteome dataset will serve as a resource for detailed investigations of cell differentiation to shed more light on the molecular mechanisms of this process in trypanosomes and other eukaryotes.

发育分化是一类普遍存在的生物学过程，可使细胞适应不同环境并执行特定功能。非洲锥虫（African trypanosomes）在昆虫媒介与脊椎动物宿主之间穿梭时，需经历严格调控的生命周期以在不同宿主环境中存活。转录组学（Transcriptomics）曾被用于解析阶段转换过程中的RNA变化，但在锥虫中，RNA水平仅能适度反映蛋白质丰度。本研究对从感染哺乳动物的阶段向昆虫寄生阶段的分化过程中的4270个蛋白质群组进行了定量分析，并对其发育过程中的表达谱进行了分类。我们的无标记定量蛋白质组学（label-free quantitative proteomics）研究揭示了分化机制中此前未知的组分，这些组分参与信号传导、蛋白质翻译后修饰、物质运输及核转运等核心生物学过程。此外，依托本蛋白质组学研究的结果，我们明确了组蛋白甲基转移酶DOT1B（histone methyltransferase DOT1B）敲除菌株中此前观测到的分化缺陷的成因：该酶是锥虫分化过程中首次细胞分裂时精准完成核分裂（karyokinesis）所必需的。该表观遗传调控因子可能参与发育分化过程中的核心染色质重塑，这一过程或许对高等真核生物的细胞分化同样具有重要意义。本研究的蛋白质组数据集将为细胞分化的深入研究提供宝贵资源，以进一步阐明锥虫及其他真核生物细胞分化过程的分子机制。