Mao, Y. S., Sunwoo, H., Zhang, B., Spector, D. L. (2011) Direct visualization of the co-transcriptional assembly of a nuclear body by noncoding RNAs. Nature Cell Biology, 13 (1). pp. 95-9101. ISSN 1465-7392
Abstract
The cell nucleus is a highly compartmentalized organelle harbouring a variety of dynamic membraneless nuclear bodies. How these subnuclear domains are established and maintained is not well understood. Here, we investigate the molecular mechanism of how one nuclear body, the paraspeckle, is assembled and organized. Paraspeckles are discrete ribonucleoprotein bodies found in mammalian cells and implicated in nuclear retention of hyperedited mRNAs. We developed a live-cell imaging system that allows for the inducible transcription of Men ɛ/β (also known as Neat1; ref. 12) noncoding RNAs (ncRNAs) and the direct visualization of the recruitment of paraspeckle proteins. Using this system, we demonstrate that Men ɛ/β ncRNAs are essential to initiate the de novo assembly of paraspeckles. These newly formed structures effectively harbour nuclear-retained mRNAs confirming that they are bona fide functional paraspeckles. By three independent approaches, we show that it is the act of Men ɛ/β transcription, but not ncRNAs alone, that regulates paraspeckle maintenance. Finally, fluorescence recovery after photobleaching (FRAP) analyses supported a critical structural role for Men ɛ/β ncRNAs in paraspeckle organization. This study establishes a model in which Men ɛ/β ncRNAs serve as a platform to recruit proteins to assemble paraspeckles.
| Item Type: | Paper |
|---|---|
| Uncontrolled Keywords: | Animals Cell Line Cell Nucleus Fluorescence Recovery After Photobleaching Humans In Situ Hybridization, Fluorescence Intranuclear Inclusion Bodies Kinetics Luminescent Proteins Mice Microscopy, Fluorescence Nuclear Proteins RNA, Messenger RNA, Untranslated RNA-Binding Proteins Reverse Transcriptase Polymerase Chain Reaction Transcription, Genetic |
| Subjects: | bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification bioinformatics > genomics and proteomics > genetics & nucleic acid processing > DNA, RNA structure, function, modification > transcription bioinformatics > genomics and proteomics > genetics & nucleic acid processing |
| CSHL Authors: | |
| Communities: | CSHL Cancer Center Shared Resources > DNA Sequencing Service CSHL Cancer Center Shared Resources > Microscopy Service CSHL labs > Spector lab CSHL Cancer Center Program > Gene Regulation and Cell Proliferation |
| Depositing User: | Matt Covey |
| Date: | 2011 |
| Date Deposited: | 10 Dec 2012 20:03 |
| Last Modified: | 13 Oct 2015 14:59 |
| PMCID: | PMC3007124 |
| Related URLs: | |
| URI: | https://repository.cshl.edu/id/eprint/26370 |
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