Hybridization-proximity labeling reveals spatially ordered interactions of nuclear RNA compartments

TY - JOUR

T1 - Hybridization-proximity labeling reveals spatially ordered interactions of nuclear RNA compartments

AU - Yap, Karen

AU - Chung, Tek Hong

AU - Makeyev, Eugene V.

N1 - Funding Information: We thank Alex Alexandrovich, Steven Lynham, and Simon Engledow for the help with protein purification, mass spectrometry, and RNA sequencing and Snezhka Oliferenko for critical comments on the manuscript. This work was supported by the Biotechnology and Biological Sciences Research Council ( BB/M001199/1 , BB/R001049/1 , and BB/V006258/1 ), the British Society for Antimicrobial Chemotherapy ( BSAC-COVID-24 ), and the European Commission ( H2020-MSCA-RISE-2016 ; project ID 734791). Publisher Copyright: © 2021 The Authors

PY - 2022/1/20

Y1 - 2022/1/20

N2 - The ability of RNAs to form specific contacts with other macromolecules provides an important mechanism for subcellular compartmentalization. Here we describe a suite of hybridization-proximity (HyPro) labeling technologies for unbiased discovery of proteins (HyPro-MS) and transcripts (HyPro-seq) associated with RNAs of interest in genetically unperturbed cells. As a proof of principle, we show that HyPro-MS and HyPro-seq can identify both known and previously unexplored spatial neighbors of the noncoding RNAs 45S, NEAT1, and PNCTR expressed at markedly different levels. Notably, HyPro-seq uncovers an extensive repertoire of incompletely processed, adenosine-to-inosine-edited transcripts accumulating at the interface between their encoding chromosomal regions and the NEAT1-containing paraspeckle compartment. At least some of these targets require NEAT1 for their optimal expression. Overall, this study provides a versatile toolkit for dissecting RNA interactomes in diverse biomedical contexts and expands our understanding of the functional architecture of the mammalian nucleus.

AB - The ability of RNAs to form specific contacts with other macromolecules provides an important mechanism for subcellular compartmentalization. Here we describe a suite of hybridization-proximity (HyPro) labeling technologies for unbiased discovery of proteins (HyPro-MS) and transcripts (HyPro-seq) associated with RNAs of interest in genetically unperturbed cells. As a proof of principle, we show that HyPro-MS and HyPro-seq can identify both known and previously unexplored spatial neighbors of the noncoding RNAs 45S, NEAT1, and PNCTR expressed at markedly different levels. Notably, HyPro-seq uncovers an extensive repertoire of incompletely processed, adenosine-to-inosine-edited transcripts accumulating at the interface between their encoding chromosomal regions and the NEAT1-containing paraspeckle compartment. At least some of these targets require NEAT1 for their optimal expression. Overall, this study provides a versatile toolkit for dissecting RNA interactomes in diverse biomedical contexts and expands our understanding of the functional architecture of the mammalian nucleus.

KW - ascorbate peroxidase

KW - digoxigenin-binding domain

KW - higher-order nuclear organization

KW - nucleolus

KW - paraspeckles

KW - perinucleolar compartment

KW - proteome

KW - proximity biotinylation

KW - RNA-containing compartments

KW - transcriptome

UR - http://www.scopus.com/inward/record.url?scp=85122994155&partnerID=8YFLogxK

U2 - 10.1016/j.molcel.2021.10.009

DO - 10.1016/j.molcel.2021.10.009

M3 - Article

C2 - 34741808

AN - SCOPUS:85122994155

SN - 1097-2765

VL - 82

SP - 463-478.e11

JO - MOLECULAR CELL

JF - MOLECULAR CELL

IS - 2

ER -