Rinn, John L

Ule, Jernej

Genome Biology

John L Rinn

Jernej Ule

Crossref

'Oming in on RNA–protein interactions

Rinn, John

Harvard University - DASH

 'Oming in on RNA–protein interactions  (Article begins on next page)The Harvard community has made this article openly available.Please share how this access benefits you. Your story matters.Citation Rinn, John L., and Jernej Ule. 2014. “'Oming in on RNA–proteininteractions.” Genome Biology 15 (1): 401. doi:10.1186/gb4158.http://dx.doi.org/10.1186/gb4158.Published Version doi:10.1186/gb4158Accessed February 16, 2015 10:13:04 AM ESTCitable Link http://nrs.harvard.edu/urn-3:HUL.InstRepos:12406530Terms of Use This article was downloaded from Harvard University's DASHrepository, and is made available under the terms and conditionsapplicable to Other Posted Material, as set forth athttp://nrs.harvard.edu/urn-3:HUL.InstRepos:dash.current.terms-of-use#LAAEDITORIAL‘Oming in on RNA–protein interactionsJohn L Rinn1,2,3* and Jernej Ule4*Welcome to the RBPomeRNA has consistently broken dogmas, owing to itsmultitude of unexpected functions. However, RNA doesfaithfully adhere to one rule: it always functions throughinteractions with proteins. The studies in this issue focuson the rapidly expanding repertoire of diverse RNA–protein interactions [1] and their functional roles andphysiological consequences, both from the perspectiveof RNA-binding proteins (RBPs) and from the vantagepoint of RNAs - coding and noncoding - that interfacewith RBPs.RNA-protein interactions are fascinating for manyreasons, one being their role in evolution - from the earliestlife forms to the most complex organisms (examplesreviewed in [2-4]). For example, the interactions betweenpre-mRNA and proteins fine-tune alternative splicing in amanner that can gradually create new protein functional-ities without the need to create additional genes andwithout affecting existing proteins [4-6]. Moreover, therehas been an emergence of numerous noncanonical RBPs(that is, proteins not previously thought to function asRBPs) that are influenced by interactions with RNAtranscripts coding and noncoding alike [7,8]. In fact,genome-wide footprinting articles in this issue [9-11]demonstrate a vast and diverse landscape of RNA–RBPcomplexes that play key regulatory roles.With recent advances in technology, together withpowerful combined experimental and computationaldevelopments, we have witnessed unprecedented newinsights into the diverse and dynamic interactions thatoccur between RNA and proteins. These range fromnew functions of well-established RBPs to the molecularsequences and structures harbored in RNA that driveinteractions with proteins. Despite this great progress,the present issue of Genome Biology demonstrates thatthere are still unresolved aspects of RBP biology.Technology opening up new horizons of theRBPomeA major challenge in understanding RNA–protein inter-actions has been to remove non-specific abundant RNAswhen mapping protein-binding sites in low-abundantRNAs. For instance, in order to understand how tissuespecificity of splicing is determined, it was important toidentify protein-binding sites on pre-mRNAs withoutcontamination from mRNAs or ribosomal RNAs. Thiswas achieved with the UV crosslinking and immuno-precipitation (CLIP) method, which employs stringentpurification steps to identify specific binding sites to theexclusion of non-specific events [12].As advances were made in CLIP technology [13,14],integrative computational approaches were developed tocombine CLIP with genome-wide studies of alternativesplicing to define regulatory maps for specific RBPs.These maps unraveled the position-dependent principlesthat were capable of predicting the functional bindingsites of RBPs [15-17]. Moreover, the regulatory elementspositioned around alternative exons were used to derivea splicing code with a capacity to predict tissue-specificsplicing with a reasonable accuracy [18,19].Technology developed to better understand RNA–protein interactions is not limited to CLIP, but is alsomaturing on several additional fronts [11,14]. Indeed,presented in this issue are a number of novel genome-wide experimental and analysis techniques that yieldnew insights into RBPology [9,10,20-25]. RNA pull-downs (also known as RIP-seq) and RNA-footprintingmethods are revealing key RBP interactions that do notfit the mold of classic RBPs. Technologies will thereforeneed to advance further if we are to better understandthe interplay of noncoding transcripts and RBPs. Whyare long noncoding RNA (lncRNA) transcripts alter-natively spliced by RBPs like their translated mRNAcounterparts? Can lncRNAs serve as decoys, scaffolds orallosteric effectors of RBPs [26]?Future studies will need to focus on additional tech-nologies in order to identify the specific sequences andstructures of RBP–RNA complexes on a larger scale. Forexample, new methods that determine the genome-wide* Correspondence: john_rinn@harvard.edu; j.ule@ucl.ac.uk1Department of Stem Cell and Regenerative Biology, Harvard University,Cambridge, MA 02138, USA4Department of Molecular Neuroscience, UCL Institute of Neurology, QueenSquare, London WC1N 3BG, UKFull list of author information is available at the end of the article© 2014 BioMed Central Ltd.Rinn and Ule Genome Biology 2014, 15:401http://genomebiology.com/2014/15/1/401structure of RNA molecules in vivo will be instrumentalin achieving this goal [27,28]. The many other keyaspects of the RBPome that still remain to be solvedinclude understanding the combinatorial interactions ofproteins on a given RNA substrate. Can multiple inter-actions provide combinatorial control? Methods tounderstand the structure and interactions of RBPs onfull-length RNAs will be needed in the future.New RNA species detected by high-throughputsequencingThe ability to detect low-abundant RNAs by usinghigh-throughput sequencing has led to the discovery ofthousands of noncoding RNAs (discussed in [29]). Onefacet of the noncoding transcriptome that has been of in-tensive research focus is lncRNAs. As perhaps expected,lncRNAs have been found to break the rules and formnumerous noncanonical interactions with proteins such aschromatin regulatory complexes, cohesins and transcrip-tion factors (see, for example, [30-32]). Moreover, lncRNAshave been shown to influence the regulatory dynamics ofsmall RNAs through sponging [33] and other mechanisms.However, it is not known whether lncRNA–protein inter-actions are generally required to mediate the functions oflncRNAs or, vice versa, whether lncRNAs modulate RBPs.The advent of new techniques that can identify RNA–DNA and RNA–protein interactions are revealing a newregulatory layer of lncRNAs. These techniques, whichinclude CHART [34], RAP [35] and ChOP [36], are con-ceptually similar to ChIP. Instead of mapping DNA–protein interactions, however, these methods determinethe localization of RNA on DNA and, moreover, enablethe study of proteins that interact with RNA at thesesites. Experiments of this nature will provide missingclues into the regulatory principles of how lncRNAsarrive at their target sites, the proteins they interact withto get there and which sequences are specificallyrequired. Collectively, lncRNA–protein interactionsperhaps herald a new code for RNA localization aroundthe genome and how it influences local and distalepigenetic aspects of nuclear architecture through theseinteractions. This is an area certain to be of intensedevelopment and research focus in the future.New RNA modifications detected by high-throughput sequencingModified ribonucleic bases have been an area of activeinterest for over half a century. Yet recent genomicsequencing technologies have expanded the modifiedRNA space beyond classic tRNA and rRNA modifi-cations to incorporate mRNA and lncRNA transcripts aswell (reviewed in [37,38]). The field of epitranscrip-tomics is in its infancy, and many questions remain tobe answered. Why are these modifications so prevalent?How do they influence RNA-binding interactions andRNA structure? What is their function? Moreover, thesesame questions also apply to the regulatory layersformed by RNA editing [39]. Specifically, Levanon andcolleagues demonstrate that only a very small fraction ofsites edited by ADAR are conserved.Understanding disease-causing mutationsImpact upon human disease is perhaps the most import-ant new frontier in the study of the RBPome. We needto use classic genetic approaches and population studiesto identify mutations in both RBPs and RNA substratesthemselves. Great progress has been made in findingmutations in RBPs associated with disease risk, such asthe RBPs FUS and TDP-43 in amyotrophic lateral scler-osis (reviewed in [40]). Yet more work will be needed tounderstand what changes occur to RNA substrates indisease and their effect on structure and function, al-though progress in this area is already underway(reviewed in this issue in [41]).Several studies in this issue have taken on the challengeof examining the intersection between the RBPome andhuman disease. Mort, Mooney and colleagues identifysingle-base mutations that affect the alternative splicingof key regulatory mRNAs [25]. Kechavarzi and Jangaexplore the dysregulation of RBP-encoding transcripts incancer and the resulting changes in protein interactionnetworks [42]. Finally, Tuschl, Wessels and colleaguesuse PAR-CLIP data to identify differential microRNAtargeting that is correlated with breast cancer subtypes[43]. These types of studies are archetypical for nume-rous future studies to understand the influence of theRBPome on human health and disease.The next challenge will be to integrate DNA and RNAbiology to understand how the various transcriptionaland post-transcriptional mechanisms cooperate to or-chestrate gene expression. This knowledge will be cru-cial in order to benchmark the mutations that causedisease by changing gene expression.Cometh the hour, cometh the special issueThe field’s rapid diversification and growth, togetherwith an increasing impact on human health, makes forperfect timing for a Genome Biology issue focused onthe RBPome!Author details1Department of Stem Cell and Regenerative Biology, Harvard University,Cambridge, MA 02138, USA.2Broad Institute, Cambridge, MA 02142, USA.3Beth Israel Deaconess Medical Center, Boston, MA 02215, USA.4Departmentof Molecular Neuroscience, UCL Institute of Neurology, Queen Square,London WC1N 3BG, UK.Published: 31 January 2014Rinn and Ule Genome Biology 2014, 15:401 Page 2 of 3http://genomebiology.com/2014/15/1/401References1. Attar N: The RBPome: where the brains meet the brawn. Genome Biol2014, 15:402.2. 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'Oming in on RNA–protein interactions

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