The NORAD lncRNA assembles a topoisomerase complex critical for genome stability

• Published in: Nature (London) 2018-09, Vol.561 (7721), p.132-136
• Main Authors: Munschauer, Mathias, Nguyen, Celina T., Sirokman, Klara, Hartigan, Christina R., Hogstrom, Larson, Engreitz, Jesse M., Ulirsch, Jacob C., Fulco, Charles P., Subramanian, Vidya, Chen, Jenny, Schenone, Monica, Guttman, Mitchell, Carr, Steven A., Lander, Eric S.
• Format: Article
• Language: English
• Subjects: 13/109; 13/31; 14/19; 38/32; 42/89; 631/337/384/2568; 631/45/612/1230; 82/58; Antisense RNA; Binding Sites; Biochemistry; Bioinformatics; Cell Cycle; Cell Cycle Proteins - metabolism; Cell Nucleus - metabolism; Cell Survival; Chromosome Segregation; Damage; Defects; Deoxyribonucleic acid; DNA; DNA biosynthesis; DNA Damage; DNA Repair; DNA Repair Enzymes - metabolism; DNA Replication; DNA topoisomerase; DNA Topoisomerases, Type I - metabolism; Experiments; Gene expression; Genomes; Genomic Instability; Genomics; Heterogeneous-Nuclear Ribonucleoproteins - metabolism; Human genome; Humanities and Social Sciences; Humans; Instability; Letter; Localization; Mammals; Mass Spectrometry; Mass spectroscopy; Methods; multidisciplinary; Multiprotein Complexes - chemistry; Multiprotein Complexes - metabolism; Non-coding RNA; Nuclear Proteins - metabolism; Nuclei (cytology); Phenotypes; Physiological aspects; Protein Binding; Protein purification; Proteins; Proteomics; Replication; Ribonucleic acid; Ribonucleoproteins - metabolism; RNA; RNA sequencing; RNA Splicing Factors - metabolism; RNA, Long Noncoding - genetics; RNA, Long Noncoding - metabolism; RNA-binding protein; RNA-Binding Proteins - metabolism; Science; Science (multidisciplinary); Scientific imaging; Software; Spectroscopy; Stability; Suppressors; Topoisomerases; Transcription Factors - metabolism
• ISSN: 0028-0836; 1476-4687
• DOI: 10.1038/s41586-018-0453-z

The human genome contains thousands of long non-coding RNAs 1 , but specific biological functions and biochemical mechanisms have been discovered for only about a dozen 2 – 7 . A specific long non-coding RNA—non-coding RNA activated by DNA damage ( NORAD )—has recently been shown to be required for maintaining genomic stability 8 , but its molecular mechanism is unknown. Here we combine RNA antisense purification and quantitative mass spectrometry to identify proteins that directly interact with NORAD in living cells. We show that NORAD interacts with proteins involved in DNA replication and repair in steady-state cells and localizes to the nucleus upon stimulation with replication stress or DNA damage. In particular, NORAD interacts with RBMX, a component of the DNA-damage response, and contains the strongest RBMX-binding site in the transcriptome. We demonstrate that NORAD controls the ability of RBMX to assemble a ribonucleoprotein complex—which we term NORAD -activated ribonucleoprotein complex 1 (NARC1)—that contains the known suppressors of genomic instability topoisomerase I (TOP1), ALYREF and the PRPF19–CDC5L complex. Cells depleted for NORAD or RBMX display an increased frequency of chromosome segregation defects, reduced replication-fork velocity and altered cell-cycle progression—which represent phenotypes that are mechanistically linked to TOP1 and PRPF19–CDC5L function. Expression of NORAD in trans can rescue defects caused by NORAD depletion, but rescue is significantly impaired when the RBMX-binding site in NORAD is deleted. Our results demonstrate that the interaction between NORAD and RBMX is important for NORAD function, and that NORAD is required for the assembly of the previously unknown topoisomerase complex NARC1, which contributes to maintaining genomic stability. In addition, we uncover a previously unknown function for long non-coding RNAs in modulating the ability of an RNA-binding protein to assemble a higher-order ribonucleoprotein complex. The long non-coding RNA NORAD interacts with proteins involved in DNA replication and repair, and controls the ability of RBMX to form a ribonucleoprotein complex that helps to maintain genomic stability.