Size fractionated NET-Seq reveals a conserved architecture of transcription units around yeast genes
Size fractionated NET-Seq reveals a conserved architecture of transcription units around yeast genes
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DOI：
10.1002/yea.3931
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发表时间：
2024-03-03
期刊：
影响因子：
2.6
通讯作者：
Mellor，Jane
中科院分区：
文献类型：
--
作者：
Xi，Shidong;Nguyen，Tania;Mellor，Jane
从酵母到人类的基因组都受到普遍的转录。单轮的普遍转录足以改变局部染色质构象、核小体动力学和基因表达，但很难与背景信号区分开。开发了大小分级的天然延伸转录物测序（sfNET-Seq）以精确地定位与表达水平无关的新生转录物。在文库构建之前，RNAPII相关的新生转录物被分级分离成不同的大小范围。当锚定到注释基因的转录起始位点（TSS）时，输出元基因的组合模式给出了预期的参考模式。与参考模式匹配的生物信息学模式在酿酒酵母中识别出9542个转录单位，其中47%是编码单位，53%是非编码单位。总共有3113个（33%）是未注释的非编码转录单位。将所有转录单位定位到注释基因的TSS或多聚腺苷酸化位点（PAS），揭示了相距约200 nt的不同转录本的连接对的独特结构。Reb 1转录因子仅在上游（TSS-60 nt，PAS）非编码转录单位与下游（TSS +150 nt）编码转录单位共存时富集在PAS下游30 nt处，并限制上游反义转录物的水平。广泛的转录干扰的可能性是显而易见的，从低丰度的未注释的转录单位与可变TSS（中位数-240 nt）开始在500 nt窗口上游的蛋白质编码基因的启动子，并转录。这项研究证实了一个高度交错的酵母基因组，具有不同类型的转录单位，以独特的方式改变染色质景观，具有广泛的调控潜力。
Genomes from yeast to humans are subject to pervasive transcription. A single round of pervasive transcription is sufficient to alter local chromatin conformation, nucleosome dynamics and gene expression, but is hard to distinguish from background signals. Size fractionated native elongating transcript sequencing (sfNET‐Seq) was developed to precisely map nascent transcripts independent of expression levels. RNAPII‐associated nascent transcripts are fractionation into different size ranges before library construction. When anchored to the transcription start sites (TSS) of annotated genes, the combined pattern of the output metagenes gives the expected reference pattern. Bioinformatic pattern matching to the reference pattern identified 9542 transcription units inSaccharomyces cerevisiae, of which 47% are coding and 53% are noncoding. In total, 3113 (33%) are unannotated noncoding transcription units. Anchoring all transcription units to the TSS or polyadenylation site (PAS) of annotated genes reveals distinctive architectures of linked pairs of divergent transcripts approximately 200nt apart. The Reb1 transcription factor is enriched 30nt downstream of the PAS only when an upstream (TSS −60nt with respect to PAS) noncoding transcription unit co‐occurs with a downstream (TSS +150nt) coding transcription unit and acts to limit levels of upstream antisense transcripts. The potential for extensive transcriptional interference is evident from low abundance unannotated transcription units with variable TSS (median −240nt) initiating within a 500nt window upstream of, and transcribing over, the promoters of protein‐coding genes. This study confirms a highly interleaved yeast genome with different types of transcription units altering the chromatin landscape in distinctive ways, with the potential to exert extensive regulatory control.