황태영 (Lieber Institute for Brain Development and Johns Hopkins University School of Medicine)

한빛사논문

조회240

Lieber Institute for Brain Development and Johns Hopkins University School of Medicine

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Nucleic Acids Res., May 10 2024, gkae334 | https://doi.org/10.1093/nar/gkae334 Massively parallel dissection of RNA in RNA-protein interactions in vivo

Yu Hsuan Lee^1,†, Evan P. Hass^2,†, Will Campodonico², Yong Kyu Lee¹, Erika Lasda², Jaynish S. Shah², John L. Rinn^2,*,‡ and Taeyoung Hwang^1,3,4,*,‡

¹Lieber Institute for Brain Development, Johns Hopkins Medical Campus, Baltimore, MD 21205, USA

²Department of Biochemistry and BioFrontiers Institute, University of Colorado, Boulder, CO 80309, USA

³Department of Neurology, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA

⁴Solomon H. Snyder Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, MD 21205, USA

^*To whom correspondence should be addressed. : Taeyoung Hwang

Correspondence may also be addressed to John L. Rinn.

^†The first two authors should be regarded as Joint First Authors.

^‡These authors jointly supervised this work: John L. Rinn and Taeyoung Hwang.

Abstract

Many of the biological functions performed by RNA are mediated by RNA-binding proteins (RBPs), and understanding the molecular basis of these interactions is fundamental to biology. Here, we present massively parallel RNA assay combined with immunoprecipitation (MPRNA-IP) for in vivo high-throughput dissection of RNA-protein interactions and describe statistical models for identifying RNA domains and parsing the structural contributions of RNA. By using custom pools of tens of thousands of RNA sequences containing systematically designed truncations and mutations, MPRNA-IP is able to identify RNA domains, sequences, and secondary structures necessary and sufficient for protein binding in a single experiment. We show that this approach is successful for multiple RNAs of interest, including the long noncoding RNA NORAD, bacteriophage MS2 RNA, and human telomerase RNA, and we use it to interrogate the hitherto unknown sequence or structural RNA-binding preferences of the DNA-looping factor CTCF. By integrating systematic mutation analysis with crosslinking immunoprecipitation, MPRNA-IP provides a novel high-throughput way to elucidate RNA-based mechanisms behind RNA-protein interactions in vivo.

논문정보

형식Research article
게재일2024년 05월 (BRIC 등록일 2024-05-14)
연구진국외 연구진
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