Zentrum für Molekulare Biologie Heidelberg, DKFZ-ZMBH Allianz, Heidelberg, Germany.
PLoS One. 2011 Mar 31;6(3):e18425. doi: 10.1371/journal.pone.0018425.
Progression of the eukaryotic cell cycle requires the regulation of hundreds of genes to ensure that they are expressed at the required times. Integral to cell cycle progression in yeast and animal cells are temporally controlled, progressive waves of transcription mediated by cell cycle-regulated transcription factors. However, in the kinetoplastids, a group of early-branching eukaryotes including many important pathogens, transcriptional regulation is almost completely absent, raising questions about the extent of cell-cycle regulation in these organisms and the mechanisms whereby regulation is achieved. Here, we analyse gene expression over the Trypanosoma brucei cell cycle, measuring changes in mRNA abundance on a transcriptome-wide scale. We developed a "double-cut" elutriation procedure to select unperturbed, highly synchronous cell populations from log-phase cultures, and compared this to synchronization by starvation. Transcriptome profiling over the cell cycle revealed the regulation of at least 430 genes. While only a minority were homologous to known cell cycle regulated transcripts in yeast or human, their functions correlated with the cellular processes occurring at the time of peak expression. We searched for potential target sites of RNA-binding proteins in these transcripts, which might earmark them for selective degradation or stabilization. Over-represented sequence motifs were found in several co-regulated transcript groups and were conserved in other kinetoplastids. Furthermore, we found evidence for cell-cycle regulation of a flagellar protein regulon with a highly conserved sequence motif, bearing similarity to consensus PUF-protein binding motifs. RNA sequence motifs that are functional in cell-cycle regulation were more widespread than previously expected and conserved within kinetoplastids. These findings highlight the central importance of post-transcriptional regulation in the proliferation of parasitic kinetoplastids.
真核细胞周期的进展需要调控数百个基因，以确保它们在适当的时间表达。在酵母和动物细胞的细胞周期进展中，至关重要的是受细胞周期调控的转录因子介导的、具有时间控制的渐进转录波。然而，在动基体生物（一组早期分支的真核生物，包括许多重要的病原体）中，转录调控几乎完全缺失，这引发了关于这些生物中细胞周期调控的程度以及实现调控的机制的问题。在这里，我们分析了在锥虫细胞周期中的基因表达情况，在转录组范围内测量了 mRNA 丰度的变化。我们开发了一种“双切割”淘选程序，从对数期培养物中选择未受干扰的高度同步细胞群体，并将其与饥饿同步进行比较。细胞周期的转录组分析揭示了至少 430 个基因的调控。虽然只有少数与酵母或人类的已知细胞周期调控转录物同源，但它们的功能与表达高峰时发生的细胞过程相关。我们在这些转录物中搜索 RNA 结合蛋白的潜在靶标，这些靶标可能会导致它们的选择性降解或稳定。在几个共调控转录物组中发现了过表达的序列基序，并且在其他动基体生物中保守。此外，我们发现了一个鞭毛蛋白调控子的细胞周期调控的证据，该调控子具有高度保守的序列基序，与 PUF 蛋白结合基序的共识相似。在细胞周期调控中具有功能的 RNA 序列基序比以前预期的更为广泛，并且在动基体生物中保守。这些发现强调了转录后调控在寄生动基体生物增殖中的核心重要性。
