Scholarlink

<p>Cotranscriptional RNA folding pathways typically involve the sequential formation of folding intermediates. Existing methods for cotranscriptional RNA structure probing map the structure of nascent RNA in the context of a terminally arrested transcription elongation complex. Consequently, the rearrangement of RNA structures as nucleotides are added to the transcript can be inferred but is not assessed directly. Here, we describe <Emphasis Type="Underline">l</Emphasis>inked-<Emphasis Type="Underline">m</Emphasis>ultipoint <Emphasis Type="Underline">T</Emphasis>ranscription <Emphasis Type="Underline">E</Emphasis>longation <Emphasis Type="Underline">C</Emphasis>omplex RNA structure <Emphasis Type="Underline">prob</Emphasis>ing (TECprobe-LM), which assesses the cotranscriptional rearrangement of RNA structures by sequentially positioning <i>E. coli</i> RNAP at two or more points within a DNA template so that nascent RNA can be chemically probed. We validate TECprobe-LM by measuring known folding events that occur within the <i>E. coli</i> signal recognition particle RNA, <i>Clostridium beijerinckii pfl</i> ZTP riboswitch, and <i>Bacillus cereus crcB</i> fluoride riboswitch folding pathways. Our findings establish TECprobe-LM as a strategy for observing cotranscriptional RNA folding events directly using chemical probing.</p>

错误：搜索内容不能为空，请输入英文关键词
错误：关键词超出字数限制，请精简
高级检索

Sequential structure probing of cotranscriptional RNA folding intermediates

  • 期刊论文

  • 出版模式：

    开放获取

  • 发表日期：

    2025年6月1日

查看全文
  • Courtney E. Szyjka,
  • Skyler L. Kelly,
  • Eric J. Strobel

摘要

Cotranscriptional RNA folding pathways typically involve the sequential formation of folding intermediates. Existing methods for cotranscriptional RNA structure probing map the structure of nascent RNA in the context of a terminally arrested transcription elongation complex. Consequently, the rearrangement of RNA structures as nucleotides are added to the transcript can be inferred but is not assessed directly. Here, we describe linked-multipoint Transcription Elongation Complex RNA structure probing (TECprobe-LM), which assesses the cotranscriptional rearrangement of RNA structures by sequentially positioning E. coli RNAP at two or more points within a DNA template so that nascent RNA can be chemically probed. We validate TECprobe-LM by measuring known folding events that occur within the E. coli signal recognition particle RNA, Clostridium beijerinckii pfl ZTP riboswitch, and Bacillus cereus crcB fluoride riboswitch folding pathways. Our findings establish TECprobe-LM as a strategy for observing cotranscriptional RNA folding events directly using chemical probing.