Department of Biochemistry, Purdue University, BCHM A343, 175 S. University Street, West Lafayette, IN 47907-2063, USA.
Purdue University Institute for Cancer Research, Purdue University, Hansen Life Sciences Research Building, Room 141, 201 S. University Street West Lafayette, IN 47907-2064, USA.
Nucleic Acids Res. 2023 Aug 25;51(15):7900-7913. doi: 10.1093/nar/gkad579.
PHO84 is a budding yeast gene reported to be negatively regulated by its cognate antisense transcripts both in cis and in trans. In this study, we performed Transient-transcriptome sequencing (TT-seq) to investigate the correlation of sense/antisense pairs in a dbp2Δ strain and found over 700 sense/antisense pairs, including PHO84, to be positively correlated, contrasting the prevailing model. To define what mechanism regulates the PHO84 gene and how this regulation could have been originally attributed to repression by the antisense transcript, we conducted a series of molecular biology and genetics experiments. We now report that the 3' untranslated region (3'UTR) of PHO84 plays a repressive role in sense expression, an activity not linked to the antisense transcripts. Moreover, we provide results of a genetic screen for 3'UTR-dependent repression of PHO84 and show that the vast majority of identified factors are linked to negative regulation. Finally, we show that the PHO84 promoter and terminator form gene loops which correlate with transcriptional repression, and that the RNA-binding protein, Tho1, increases this looping and the 3'UTR-dependent repression. Our results negate the current model for antisense non-coding transcripts of PHO84 and suggest that many of these transcripts are byproducts of open chromatin.
PHO84 是一个芽殖酵母基因，据报道，其在顺式和反式都受到与其同源反义转录本的负调控。在这项研究中，我们进行了瞬时转录组测序 (TT-seq) 来研究 dbp2Δ 菌株中正义/反义对的相关性，发现包括 PHO84 在内的超过 700 个正义/反义对呈正相关，与流行模型相反。为了确定什么机制调节 PHO84 基因，以及这种调节最初如何归因于反义转录本的抑制，我们进行了一系列分子生物学和遗传学实验。我们现在报告 PHO84 的 3'非翻译区 (3'UTR) 在正义表达中发挥抑制作用，这种活性与反义转录本无关。此外，我们提供了 PHO84 3'UTR 依赖性抑制的遗传筛选结果，并表明大多数鉴定出的因子都与负调控有关。最后，我们表明 PHO84 启动子和终止子形成与转录抑制相关的基因环，并且 RNA 结合蛋白 Tho1 增加了这种环化和 3'UTR 依赖性抑制。我们的结果否定了 PHO84 反义非编码转录本的当前模型，并表明这些转录本中的许多是开放染色质的副产物。
