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HOTAIR Lifts Noncoding RNAs to New Levels

2007; Cell Press; Volume: 129; Issue: 7 Linguagem: Inglês

10.1016/j.cell.2007.06.014

1097-4172

Caroline J. Woo, Robert E. Kingston,

RNA modifications and cancer

It is not clear to what extent noncoding RNAs regulate the homeobox (HOX) genes that encode key regulators of development in the embryo. In this issue, Rinn et al., 2007Rinn J.L. Kertesz M. Wang J.K. Squazzo S.L. Xu X. Brugman S.A. Goodnough H. Helms J.A. Farnham P.J. Segal E. Chang H.Y. Cell. 2007; (this issue)PubMed Google Scholar characterize noncoding RNAs that regulate HOX genes and discover one, HOTAIR, that unexpectedly regulates a HOX gene cluster on a different chromosome than the HOX cluster that encodes it. It is not clear to what extent noncoding RNAs regulate the homeobox (HOX) genes that encode key regulators of development in the embryo. In this issue, Rinn et al., 2007Rinn J.L. Kertesz M. Wang J.K. Squazzo S.L. Xu X. Brugman S.A. Goodnough H. Helms J.A. Farnham P.J. Segal E. Chang H.Y. Cell. 2007; (this issue)PubMed Google Scholar characterize noncoding RNAs that regulate HOX genes and discover one, HOTAIR, that unexpectedly regulates a HOX gene cluster on a different chromosome than the HOX cluster that encodes it. “A thing long expected takes the form of the unexpected when at last it comes,” Mark Twain once wrote. A striking example of this notion is provided in this issue by Rinn et al., 2007Rinn J.L. Kertesz M. Wang J.K. Squazzo S.L. Xu X. Brugman S.A. Goodnough H. Helms J.A. Farnham P.J. Segal E. Chang H.Y. Cell. 2007; (this issue)PubMed Google Scholar. It has long been thought that HOX gene clusters, key players in embryonic development, are regulated by cis-acting noncoding RNAs (ncRNAs). In their new work, Rinn et al. use a high-resolution tiled microarray to systematically identify and characterize hundreds of ncRNAs transcribed from the HOX gene cluster of 11 human fibroblast cell lines isolated from distinct positions along the anterior-posterior and proximal-distal body axes. They home in on an intriguingly positioned ncRNA they call HOTAIR and make the surprising discovery that this ncRNA acts in trans to regulate a HOX gene cluster on a completely different chromosome, not the HOX gene cluster that encodes it. The HOX gene cluster serves as a classic model of gene regulation during embryonic development. The 39 human HOX genes are contained within four clusters, called HOXA–D, on four independent chromosomes. The clusters are organized similarly which strongly suggests that they have arisen through gene duplication. During development, the expression of a gene within the linear organization of each HOX cluster correlates with body position, an unusual situation that suggests conservation of the regulatory mechanisms of the four clusters. Proteins responsible for establishing epigenetic modifications are known to regulate HOX gene expression, and it has long been proposed that ncRNAs guide the epigenetic regulation of these genes (Rank et al., 2002Rank G. Prestel M. Paro R. Mol. Cell. Biol. 2002; 22: 8026-8034Crossref PubMed Scopus (125) Google Scholar, Petruk et al., 2006Petruk S. Sedkov Y. Riley K.M. Hodgson J. Schweisguth F. Hirose S. Jaynes J.B. Brock H.W. Mazo A. Cell. 2006; 127: 1209-1221Abstract Full Text Full Text PDF PubMed Scopus (207) Google Scholar and references therein). The HOX proteins themselves are master regulators of other transcription factors, and their overall expression profiles are tightly regulated spatially and temporally. Functionally, the HOX genes are critical for correct patterning of the axes during development. The significance of correct HOX gene expression was first observed in Drosophila. Since then, HOX gene clusters have been observed in all metazoans and have similar roles in axial development (reviewed in Lemons and McGinnis, 2006Lemons D. McGinnis W. Science. 2006; 313: 1918-1922Crossref PubMed Scopus (235) Google Scholar). Differential HOX gene expression is established early on in the developing embryo. Once established, the HOX gene expression profile needs to be maintained even as the embryo continues through development. The Polycomb group (PcG) and Trithorax (TrxG) group protein complexes maintain this form of cellular memory as regions of silenced or active transcription, respectively (Klymenko and Muller, 2004Klymenko T. Muller J. EMBO Rep. 2004; 5: 373-377Crossref PubMed Scopus (193) Google Scholar). The PcG complexes are associated with trimethylated K27 on histone H3, a modification frequently required for this epigenetic silencing. The TrxG complex is associated with trimethylated K4 on histone H3 which is associated with genes that are actively transcribed. In addition to the HOX gene expression profiles of the human adult fibroblast cell lines originating from different axial positions, Rinn et al., 2007Rinn J.L. Kertesz M. Wang J.K. Squazzo S.L. Xu X. Brugman S.A. Goodnough H. Helms J.A. Farnham P.J. Segal E. Chang H.Y. Cell. 2007; (this issue)PubMed Google Scholar describe the discovery of many ncRNAs expressed from these fibroblasts. Previously detailed transcriptional analyses of human HOX clusters revealed the presence of ncRNAs, many of which have been implicated in transcriptional regulation of neighboring HOX genes (Mainguy et al., 2007Mainguy G. Koster J. Woltering J. Hansen H. Durston A. PLoS ONE. 2007; 2: e356https://doi.org/10.1371/journal.pone.0000356Crossref PubMed Scopus (65) Google Scholar, Sessa et al., 2007Sessa L. Breiling A. Lavorgna G. Silvestri L. Casari G. Orlando V. RNA. 2007; 13: 223-239Crossref PubMed Scopus (103) Google Scholar). By combining the powers of a high-throughput high-resolution tiled microarray and bioinformatics algorithms, Rinn et al., 2007Rinn J.L. Kertesz M. Wang J.K. Squazzo S.L. Xu X. Brugman S.A. Goodnough H. Helms J.A. Farnham P.J. Segal E. Chang H.Y. Cell. 2007; (this issue)PubMed Google Scholar identified nearly 30 kb of new ncRNAs. Similar to the ncRNAs found by other groups, the majority of these transcripts are intergenic and are transcribed in the direction opposite to the HOX genes. By having such a deep database, however, these investigators were able to demonstrate that the ncRNAs are also differentially expressed in different cell types according to their original position along the body axis, and, moreover, sequence motifs could be assigned to unique cell types. Given the flurry of studies on HOX to map potential epigenetic regulatory marks on histones (Bernstein et al., 2006Bernstein B.E. Mikkelsen T.S. Xie X. Kamal M. Huebert D.J. Cuff J. Fry B. Meissner A. Wernig M. Plath K. et al.Cell. 2006; 125: 315-326Abstract Full Text Full Text PDF PubMed Scopus (3738) Google Scholar, Lee et al., 2006Lee T.I. Jenner R.G. Boyer L.A. Guenther M.G. Levine S.S. Kumar R.M. Chevalier B. Johnstone S.E. Cole M.F. Isono K. et al.Cell. 2006; 125: 301-313Abstract Full Text Full Text PDF PubMed Scopus (1791) Google Scholar, Boyer et al., 2006Boyer L.A. Plath K. Zeitlinger J. Brambrink T. Medeiros L.A. Lee T.I. Levine S.S. Wernig M. Tajonar A. Ray M.K. et al.Nature. 2006; 441: 349-353Crossref PubMed Scopus (1930) Google Scholar), the authors asked whether there were any correlations between epigenetic marks and the location of ncRNAs. Chromatin immunoprecipitation (ChIP)-microarray experiments revealed, not surprisingly, that RNA polymerase II and trimethylated H3K4 (a mark of active transcription) occupied the same regions as the ncRNAs. These were mutually exclusive on the HOX clusters with the localization of Suz12 and trimethylated H3K27, a mark of repressed chromatin. This was elegantly shown at the HOXA cluster, where a boundary between HOXA7 and HOXA9 delineates a clear breakpoint between oppositely acting covalent marks in two distinctly regulated fibroblasts from two different axial positions. The surprise came, however, when Rinn et al., 2007Rinn J.L. Kertesz M. Wang J.K. Squazzo S.L. Xu X. Brugman S.A. Goodnough H. Helms J.A. Farnham P.J. Segal E. Chang H.Y. Cell. 2007; (this issue)PubMed Google Scholar characterized one well-placed ncRNA in this region. This 2158 nucleotide RNA, termed HOTAIR, was localized to a regulatory boundary in the HOXC cluster and was expressed in both distal and posterior fibroblasts. If HOTAIR was similar to other previously characterized ncRNAs, it might have had a proximal regulatory role (Sessa et al., 2007Sessa L. Breiling A. Lavorgna G. Silvestri L. Casari G. Orlando V. RNA. 2007; 13: 223-239Crossref PubMed Scopus (103) Google Scholar). However, knockdown of HOTAIR showed no changes in the HOXC cluster, but instead showed a loss of transcriptional repression from a 40 kb region of the HOXD cluster. In addition, the repressive mark, trimethylated histone H3K27, and the regulatory complex that produces this mark, the PcG complex PRC2, were no longer present at that particular region of the HOXD cluster when HOTAIR was depleted. Pull-down experiments of PRC2 components demonstrated a direct and specific interaction with HOTAIR. The observation that this ncRNA binds to an epigenetic regulatory complex and changes methylation patterns on a different HOX cluster on a different chromosome is unexpected. There are many questions that remain concerning how HOTAIR silences the 40 kb region of the HOXD locus. Is this silencing effect direct or indirect? The interaction of HOTAIR with PRC2 is consistent with a direct effect but does not rule out indirect effects. Would ectopic expression of HOTAIR in another fibroblast be sufficient to suppress the HOXD locus? The association of HOTAIR with PRC2 also suggests that the mechanism is epigenetic, but this remains an open question. Furthermore, nature tends to use mechanisms more than once: are there other trans-acting ncRNAs from the HOX cluster or elsewhere that have not yet been found? The characterization of HOTAIR raises intriguing questions about how ncRNAs work and adds to the functional complexity of this class of regulatory RNAs.