ABSTRACT

mRNA homoeostasis is favoured by crosstalk between transcription and degradation machineries. Both the Ccr4-Not and the Xrn1-decaysome complexes have been described to influence transcription. While Ccr4-Not has been shown to directly stimulate transcription elongation, the information available on how Xrn1 influences transcription is scarce and contradictory. In this study we have addressed this issue by mapping RNA polymerase II (RNA pol II) at high resolution, using CRAC and BioGRO-seq techniques in Saccharomyces cerevisiae. We found significant effects of Xrn1 perturbation on RNA pol II profiles across the genome. RNA pol II profiles at 5ʹ exhibited significant alterations that were compatible with decreased elongation rates in the absence of Xrn1. Nucleosome mapping detected altered chromatin configuration in the gene bodies. We also detected accumulation of RNA pol II shortly upstream of polyadenylation sites by CRAC, although not by BioGRO-seq, suggesting higher frequency of backtracking before pre-mRNA cleavage. This phenomenon was particularly linked to genes with poorly positioned nucleosomes at this position. Accumulation of RNA pol II at 3ʹ was also detected in other mRNA decay mutants. According to these and other pieces of evidence, Xrn1 seems to influence transcription elongation at least in two ways: by directly favouring elongation rates and by a more general mechanism that connects mRNA decay to late elongation.

摘要

转录和降解机制之间的串扰有利于 mRNA 稳态。Ccr4-Not 和 Xrn1-decaysome 复合物都被描述为影响转录。虽然 Ccr4-Not 已被证明可以直接刺激转录延伸，但关于 Xrn1 如何影响转录的可用信息很少且相互矛盾。在这项研究中，我们通过在酿酒酵母中使用 CRAC 和 BioGRO-seq 技术以高分辨率绘制 RNA 聚合酶 II (RNA pol II) 来解决这个问题. 我们发现 Xrn1 扰动对整个基因组的 RNA pol II 谱有显着影响。5'处的 RNA pol II 谱显示出显着的变化，这与在没有 Xrn1 的情况下降低的延伸率相一致。核小体作图检测到基因体中染色质构型的改变。我们还通过 CRAC 检测到 RNA pol II 在聚腺苷酸化位点上游附近的积累，尽管不是通过 BioGRO-seq，这表明在前 mRNA 切割之前回溯的频率更高。这种现象与在该位置具有不良定位的核小体的基因特别相关。在其他 mRNA 衰变突变体中也检测到 RNA pol II 在 3' 处的积累。根据这些和其他证据，Xrn1 似乎至少以两种方式影响转录延伸：