Gene expression is a multi-step process that converts DNA-encoded information into proteins, involving RNA transcription, maturation, degradation, and translation. While transcriptional control is a major regulator of protein levels, the role of post-transcriptional processes such as RNA processing and degradation is less well understood due to the challenge of measuring their contributions individually. To address this challenge, we investigated the control of gene expression in Trypanosoma brucei, a unicellular parasite assumed to lack transcriptional control. Instead, mRNA levels in T. brucei are controlled by post-transcriptional processes, which enabled us to disentangle the contribution of both processes to total mRNA levels. In this study, we developed an efficient metabolic RNA labeling approach and combined ultra-short metabolic labeling with transient transcriptome sequencing (TT-seq) to confirm the long-standing assumption that RNA polymerase II transcription is unregulated in T. brucei. In addition, we established thiol (SH)-linked alkylation for metabolic sequencing of RNA (SLAM-seq) to globally quantify RNA processing rates and half-lives. Our data, combined with scRNA-seq data, indicate that RNA processing and stability independently affect total mRNA levels and contribute to the variability seen between individual cells in African trypanosomes.

中文翻译：

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SLAM-seq 揭示了 RNA 加工和稳定性对非洲锥虫基因表达的独立贡献


基因表达是一个多步骤过程，将 DNA 编码的信息转化为蛋白质，包括 RNA 转录、成熟、降解和翻译。虽然转录控制是蛋白质水平的主要调节因子，但由于单独测量其贡献的挑战，转录后过程（如 RNA 加工和降解）的作用尚不清楚。为了应对这一挑战，我们研究了布鲁氏锥虫（一种被认为缺乏转录控制的单细胞寄生虫）中基因表达的控制。相反，T. brucei 中的 mRNA 水平受转录后过程控制，这使我们能够解开这两个过程对总 mRNA 水平的贡献。在这项研究中，我们开发了一种高效的代谢 RNA 标记方法，并将超短代谢标记与瞬时转录组测序 （TT-seq） 相结合，以证实 RNA 聚合酶 II 转录在 T. brucei 中不受调节的长期假设。此外，我们建立了用于 RNA 代谢测序的巯基 （SH） 连接烷基化 （SLAM-seq），以全面量化 RNA 加工速率和半衰期。我们的数据与 scRNA-seq 数据相结合，表明 RNA 加工和稳定性独立影响总 mRNA 水平，并导致非洲锥虫中单个细胞之间的差异。