The dynamic process of transcription termination produces transient RNA intermediates that are difficult to distinguish from each other via short-read sequencing methods. Here, we use single-molecule nascent RNA sequencing to characterize the various forms of transient RNAs during termination at genome-wide scale in wildtype Arabidopsis and in atxrn3, fpa, and met1 mutants. Our data reveal a wide range of termination windows among genes, ranging from ~ 50 nt to over 1000 nt. We also observe efficient termination before downstream tRNA genes, suggesting that chromatin structure around the promoter region of tRNA genes may block pol II elongation. 5′ Cleaved readthrough transcription in atxrn3 with delayed termination can run into downstream genes to produce normally spliced and polyadenylated mRNAs in the absence of their own transcription initiation. Consistent with previous reports, we also observe long chimeric transcripts with cryptic splicing in fpa mutant; but loss of CG DNA methylation has no obvious impact on termination in the met1 mutant. Our method is applicable to establish a comprehensive termination landscape in a broad range of species.

中文翻译：

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单分子新生RNA测序揭示拟南芥转录终止情况


转录终止的动态过程会产生短暂的 RNA 中间体，这些中间体很难通过短读长测序方法相互区分。在这里，我们使用单分子新生 RNA 测序来表征野生型拟南芥以及 atxrn3、fpa 和 met1 突变体在全基因组范围内终止过程中各种形式的瞬时 RNA。我们的数据揭示了基因之间的广泛终止窗口，范围从约 50 nt 到超过 1000 nt。我们还观察到下游 tRNA 基因之前的有效终止，这表明 tRNA 基因启动子区域周围的染色质结构可能会阻止 pol II 延伸。 atxrn3 中具有延迟终止的 5' 剪切通读转录可以进入下游基因，在缺乏自身转录起始的情况下产生正常剪接和聚腺苷酸化的 mRNA。与之前的报道一致，我们还在 fpa 突变体中观察到具有隐秘剪接的长嵌合转录本；但CG DNA甲基化的缺失对met1突变体的终止没有明显影响。我们的方法适用于在广泛的物种中建立全面的终止景观。