Approaches for Understanding the Mechanisms of Long Noncoding RNA Regulation of Gene Expression.

SUMMARYMammalian genomes encode tens of thousands of long noncoding RNAs (lncRNAs) that have been implicated in a diverse array of biological processes and human diseases. In recent years, the development of new tools for studying lncRNAs has enabled important progress in defining the mechanisms by which Xist and other lncRNAs function. This collective work provides a framework for how to define the mechanisms by which lncRNAs act. This includes defining lncRNA function, identifying and characterizing lncRNA-protein interactions, and lncRNA localization in the cell. In this review, we discuss various experimental approaches for deciphering lncRNA mechanisms and discuss issues and limitations in interpreting these results. We explore what these data can reveal about lncRNA function and mechanism as well as emerging insights into lncRNA biology that have been derived from these studies.

[1]  Polycomb-Group Protein , 2020, Definitions.

[2]  B. Tabak,et al.  Higher-Order Inter-chromosomal Hubs Shape 3D Genome Organization in the Nucleus , 2018, Cell.

[3]  J. Mendell,et al.  Functional Classification and Experimental Dissection of Long Noncoding RNAs , 2018, Cell.

[4]  John L Rinn,et al.  Spatiotemporal allele organization by allele-specific CRISPR live-cell imaging: SNP-CLING , 2017, Nature Structural & Molecular Biology.

[5]  C. Bond,et al.  Paraspeckles: Where Long Noncoding RNA Meets Phase Separation. , 2017, Trends in biochemical sciences.

[6]  N. Brockdorff,et al.  hnRNPK Recruits PCGF3/5-PRC1 to the Xist RNA B-Repeat to Establish Polycomb-Mediated Chromosomal Silencing , 2017, Molecular cell.

[7]  M. Hentze,et al.  Specific RNP capture with antisense LNA/DNA mixmers , 2017, RNA.

[8]  S. Crooke,et al.  RNase H1-Dependent Antisense Oligonucleotides Are Robustly Active in Directing RNA Cleavage in Both the Cytoplasm and the Nucleus , 2017, Molecular therapy : the journal of the American Society of Gene Therapy.

[9]  N. Brockdorff,et al.  PCGF3/5–PRC1 initiates Polycomb recruitment in X chromosome inactivation , 2017, Science.

[10]  R. Batey,et al.  Intrinsically disordered RGG/RG domains mediate degenerate specificity in RNA binding , 2017, Nucleic acids research.

[11]  K. Hall Faculty Opinions recommendation of Chromosomes. A comprehensive Xist interactome reveals cohesin repulsion and an RNA-directed chromosome conformation. , 2017 .

[12]  Edith Heard,et al.  Novel players in X inactivation: insights into Xist-mediated gene silencing and chromosome conformation , 2017, Nature Structural &Molecular Biology.

[13]  R. Young,et al.  A Phase Separation Model for Transcriptional Control , 2017, Cell.

[14]  R. Margueron,et al.  PRC2 is dispensable for HOTAIR‐mediated transcriptional repression , 2017, The EMBO journal.

[15]  Howard Y. Chang,et al.  NONCODING RNA: CRISPRi‐based genome‐scale identification of functional long noncoding RNA loci in human cells , 2017 .

[16]  E. Lander,et al.  Local regulation of gene expression by lncRNA promoters, transcription and splicing , 2016, Nature.

[17]  Noah Ollikainen,et al.  Xist recruits the X chromosome to the nuclear lamina to enable chromosome-wide silencing , 2016, Science.

[18]  Jesse M. Engreitz,et al.  Long non-coding RNAs: spatial amplifiers that control nuclear structure and gene expression , 2016, Nature Reviews Molecular Cell Biology.

[19]  Sharon R Grossman,et al.  Systematic mapping of functional enhancer–promoter connections with CRISPR interference , 2016, Science.

[20]  Samie R. Jaffrey,et al.  m6A RNA methylation promotes XIST-mediated transcriptional repression , 2016, Nature.

[21]  J. Rinn,et al.  In Vivo Characterization of Linc-p21 Reveals Functional cis-Regulatory DNA Elements. , 2016, Cell reports.

[22]  Tsung-Cheng Chang,et al.  Noncoding RNA NORAD Regulates Genomic Stability by Sequestering PUMILIO Proteins , 2016, Cell.

[23]  P. Avner,et al.  Quantitative predictions of protein interactions with long noncoding RNAs , 2016, Nature Methods.

[24]  A. Ruthenburg,et al.  Nuclear Fractionation Reveals Thousands of Chromatin-Tethered Noncoding RNAs Adjacent to Active Genes. , 2015, Cell reports.

[25]  Jeannie T. Lee,et al.  Chromosomes. A comprehensive Xist interactome reveals cohesin repulsion and an RNA-directed chromosome conformation. , 2015, Science.

[26]  N. Brockdorff,et al.  A Pooled shRNA Screen Identifies Rbm15, Spen, and Wtap as Factors Required for Xist RNA-Mediated Silencing , 2015, Cell reports.

[27]  S. Thore,et al.  Identification of Spen as a Crucial Factor for Xist Function through Forward Genetic Screening in Haploid Embryonic Stem Cells , 2015, Cell reports.

[28]  J. Rinn,et al.  Multiplexable, locus-specific targeting of long RNAs with CRISPR-Display , 2015, Nature Methods.

[29]  Qiangfeng Cliff Zhang,et al.  Systematic Discovery of Xist RNA Binding Proteins , 2015, Cell.

[30]  Michael J. Sweredoski,et al.  The Xist lncRNA directly interacts with SHARP to silence transcription through HDAC3 , 2015, Nature.

[31]  J. Rinn,et al.  Localization and abundance analysis of human lncRNAs at single-cell and single-molecule resolution , 2015, Genome Biology.

[32]  A. Hyman,et al.  Liquid-liquid phase separation in biology. , 2014, Annual review of cell and developmental biology.

[33]  Michael Y Tolstorukov,et al.  The long noncoding RNAs NEAT1 and MALAT1 bind active chromatin sites. , 2014, Molecular cell.

[34]  Sharon R Grossman,et al.  RNA-RNA Interactions Enable Specific Targeting of Noncoding RNAs to Nascent Pre-mRNAs and Chromatin Sites , 2014, Cell.

[35]  Justin Demmerle,et al.  Spatial separation of Xist RNA and polycomb proteins revealed by superresolution microscopy , 2014, Proceedings of the National Academy of Sciences.

[36]  M. Guttman,et al.  Methods for comprehensive experimental identification of RNA-protein interactions , 2014, Genome Biology.

[37]  David R. Kelley,et al.  Topological Organization of Multi-chromosomal Regions by Firre , 2014, Nature structural & molecular biology.

[38]  D. Reinberg,et al.  Jarid2 Is Implicated in the Initial Xist-Induced Targeting of PRC2 to the Inactive X Chromosome. , 2014, Molecular cell.

[39]  Michael Morse,et al.  Multiple knockout mouse models reveal lincRNAs are required for life and brain development , 2013, eLife.

[40]  Sarah K. Bowman,et al.  High-resolution Xist binding maps reveal 2-step spreading during X-inactivation , 2013, Nature.

[41]  T. Cech,et al.  Promiscuous RNA binding by Polycomb Repressive Complex 2 , 2013, Nature Structural &Molecular Biology.

[42]  Michael Y Tolstorukov,et al.  The CLAMP protein links the MSL complex to the X chromosome during Drosophila dosage compensation. , 2013, Genes & development.

[43]  E. Lander,et al.  The Xist lncRNA Exploits Three-Dimensional Genome Architecture to Spread Across the X Chromosome , 2013, Science.

[44]  Nicholas T. Ingolia,et al.  Ribosome Profiling Provides Evidence that Large Noncoding RNAs Do Not Encode Proteins , 2013, Cell.

[45]  Craig D Wenger,et al.  RNA–protein analysis using a conditional CRISPR nuclease , 2013, Proceedings of the National Academy of Sciences.

[46]  David G. Knowles,et al.  The GENCODE v7 catalog of human long noncoding RNAs: Analysis of their gene structure, evolution, and expression , 2012, Genome research.

[47]  Howard Y. Chang,et al.  Genome regulation by long noncoding RNAs. , 2012, Annual review of biochemistry.

[48]  Giacomo Cavalli,et al.  Polycomb: a paradigm for genome organization from one to three dimensions. , 2012, Current opinion in cell biology.

[49]  Howard Y. Chang,et al.  Genomic maps of long noncoding RNA occupancy reveal principles of RNA-chromatin interactions. , 2011, Molecular cell.

[50]  Cole Trapnell,et al.  Integrative annotation of human large intergenic noncoding RNAs reveals global properties and specific subclasses. , 2011, Genes & development.

[51]  J. Rinn,et al.  lincRNAs act in the circuitry controlling pluripotency and differentiation , 2011, Nature.

[52]  Howard Y. Chang,et al.  Long noncoding RNAs and human disease. , 2011, Trends in cell biology.

[53]  Howard Y. Chang,et al.  A long noncoding RNA maintains active chromatin to coordinate homeotic gene expression , 2011, Nature.

[54]  M. Dundr,et al.  Nucleation of nuclear bodies by RNA , 2011, Nature Cell Biology.

[55]  D. Spector,et al.  Direct Visualization of the Co-transcriptional Assembly of a Nuclear Body by Noncoding RNAs , 2010, Nature Cell Biology.

[56]  A. Lamond,et al.  Nuclear speckles. , 2011, Cold Spring Harbor perspectives in biology.

[57]  Toshiro K. Ohsumi,et al.  Genome-wide identification of polycomb-associated RNAs by RIP-seq. , 2010, Molecular cell.

[58]  B. Blencowe,et al.  The nuclear-retained noncoding RNA MALAT1 regulates alternative splicing by modulating SR splicing factor phosphorylation. , 2010, Molecular cell.

[59]  N. Brockdorff,et al.  The matrix protein hnRNP U is required for chromosomal localization of Xist RNA. , 2010, Developmental cell.

[60]  Robert B Darnell,et al.  HITS‐CLIP: panoramic views of protein–RNA regulation in living cells , 2010, Wiley interdisciplinary reviews. RNA.

[61]  J. Rinn,et al.  A Large Intergenic Noncoding RNA Induced by p53 Mediates Global Gene Repression in the p53 Response , 2010, Cell.

[62]  J. Ule,et al.  iCLIP reveals the function of hnRNP particles in splicing at individual nucleotide resolution , 2010, Nature Structural &Molecular Biology.

[63]  J. Rinn,et al.  Many human large intergenic noncoding RNAs associate with chromatin-modifying complexes and affect gene expression , 2009, Proceedings of the National Academy of Sciences.

[64]  John N. Hutchinson,et al.  An architectural role for a nuclear noncoding RNA: NEAT1 RNA is essential for the structure of paraspeckles. , 2009, Molecular cell.

[65]  P. Zamore,et al.  Small silencing RNAs: an expanding universe , 2009, Nature Reviews Genetics.

[66]  Michael F. Lin,et al.  Chromatin signature reveals over a thousand highly conserved large non-coding RNAs in mammals , 2009, Nature.

[67]  Jennifer A. Mitchell,et al.  The Air Noncoding RNA Epigenetically Silences Transcription by Targeting G9a to Chromatin , 2008, Science.

[68]  Antoine H. F. M. Peters,et al.  Polycomb group proteins Ezh2 and Rnf2 direct genomic contraction and imprinted repression in early mouse embryos. , 2008, Developmental cell.

[69]  Jeannie T. Lee,et al.  Polycomb Proteins Targeted by a Short Repeat RNA to the Mouse X Chromosome , 2008, Science.

[70]  C. Barreau,et al.  Tethering of proteins to RNAs by bacteriophage proteins , 2008, Biology of the cell.

[71]  E. Heard,et al.  A novel role for Xist RNA in the formation of a repressive nuclear compartment into which genes are recruited when silenced. , 2006, Genes & development.

[72]  T. Jenuwein,et al.  Recruitment of PRC1 function at the initiation of X inactivation independent of PRC2 and silencing , 2006, The EMBO journal.

[73]  T. Magnuson,et al.  The Polycomb Group Protein EED Is Dispensable for the Initiation of Random X-Chromosome Inactivation , 2006, PLoS genetics.

[74]  J. Steitz,et al.  Evidence for reassociation of RNA-binding proteins after cell lysis: implications for the interpretation of immunoprecipitation analyses. , 2004, RNA.

[75]  Anton Wutz,et al.  A Chromosomal Memory Triggered by Xist Regulates Histone Methylation in X Inactivation , 2004, PLoS biology.

[76]  Hengbin Wang,et al.  Role of Histone H3 Lysine 27 Methylation in X Inactivation , 2003, Science.

[77]  D. Engelke,et al.  RNA affinity tags for purification of RNAs and ribonucleoprotein complexes. , 2002, Methods.

[78]  Rudolf Jaenisch,et al.  Chromosomal silencing and localization are mediated by different domains of Xist RNA , 2002, Nature Genetics.

[79]  N. Brockdorff,et al.  Xist RNA exhibits a banded localization on the inactive X chromosome and is excluded from autosomal material in cis. , 1999, Human molecular genetics.

[80]  Alan Ashworth,et al.  Xist has properties of the X-chromosome inactivation centre , 1997, Nature.

[81]  Carolyn J. Brown,et al.  Evidence that heteronuclear proteins interact with the XIST RNA in vitro , 1996, Somatic cell and molecular genetics.

[82]  R. Jaenisch,et al.  A 450 kb Transgene Displays Properties of the Mammalian X-Inactivation Center , 1996, Cell.

[83]  J. Mcneil,et al.  XIST RNA paints the inactive X chromosome at interphase: evidence for a novel RNA involved in nuclear/chromosome structure , 1996, The Journal of cell biology.

[84]  S. Rastan,et al.  Requirement for Xist in X chromosome inactivation , 1996, Nature.

[85]  A. Ashworth,et al.  Conservation of position and exclusive expression of mouse Xist from the inactive X chromosome , 1991, Nature.

[86]  Carolyn J. Brown,et al.  A gene from the region of the human X inactivation centre is expressed exclusively from the inactive X chromosome , 1991, Nature.