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<p>RNA-protein interactions critically regulate gene expression and cellular processes, yet their comprehensive mapping remains challenging due to their structural diversity. We introduce PRIM-seq (protein-RNA interaction mapping by sequencing), a method for concurrent de novo identification of RNA-binding proteins and their associated RNAs. PRIM-seq generates unique chimeric DNA sequences by proximity ligation of RNAs with protein-linked DNA barcodes, which are subsequently decoded through sequencing. We apply PRIM-seq to two human cell lines and construct a human RNA-protein association network (HuRPA), encompassing &gt;350,000 associations involving ~7,000 RNAs and ~11,000 proteins, including 2,610 proteins that each interact with at least 10 distinct RNAs. We experimentally validate the tumorigenesis-associated lincRNA LINC00339, the RNA with the highest number of protein associations in HuRPA, as a protein-associated RNA. We further validate the RNA-associating abilities of chromatin-conformation regulators SMC1A, SMC3 and RAD21, as well as the metabolic enzyme PHGDH. PRIM-seq enables systematic discovery and prioritization of RNA-binding proteins and their targets without gene- or protein-specific reagents.</p>

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Genome-wide mapping of RNA-protein associations through sequencing

  • 期刊论文

  • 发表日期：

    2025年9月9日

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  • Zhijie Qi,
  • Shuanghong Xue,
  • Junchen Chen,
  • Wenxin Zhao,
  • Kara Johnson,
  • Xingzhao Wen,
  • John Lalith Charles Richard,
  • Pei Lin,
  • Sheng Zhong

摘要

RNA-protein interactions critically regulate gene expression and cellular processes, yet their comprehensive mapping remains challenging due to their structural diversity. We introduce PRIM-seq (protein-RNA interaction mapping by sequencing), a method for concurrent de novo identification of RNA-binding proteins and their associated RNAs. PRIM-seq generates unique chimeric DNA sequences by proximity ligation of RNAs with protein-linked DNA barcodes, which are subsequently decoded through sequencing. We apply PRIM-seq to two human cell lines and construct a human RNA-protein association network (HuRPA), encompassing >350,000 associations involving ~7,000 RNAs and ~11,000 proteins, including 2,610 proteins that each interact with at least 10 distinct RNAs. We experimentally validate the tumorigenesis-associated lincRNA LINC00339, the RNA with the highest number of protein associations in HuRPA, as a protein-associated RNA. We further validate the RNA-associating abilities of chromatin-conformation regulators SMC1A, SMC3 and RAD21, as well as the metabolic enzyme PHGDH. PRIM-seq enables systematic discovery and prioritization of RNA-binding proteins and their targets without gene- or protein-specific reagents.