1、RepurposingGeneration of telomerase RNAGeneration of circular RNAs.Exon 6Exon 7Exon 8hnRNP A1/A2ISSSMN2.Exon 6Exon 7Exon 8hnRNP A1/A2SMN2ASOSmall molecules and antisense oligonucleotides.Exon 1 Exon 433.Exon 1A.Exon skippingB.Intron pairing.Exon 1Exon 2Exon 2Exon 2Exon 2Exon 2Exon 2Exon 2Exon 3Exon
2、3Exon 3Exon 3Exon 3Exon 3Exon 3Exon 4BPBPBP.Forked product(terminated)Intron 2 lariat3Double lariatIntra-lariat splicingCircular RNA12.Exon 1Exon 1.Step 2Step 1.Active telomeraseMature TER1(telomerase RNA)XSMN2.Exon 7U1NVS-SM1Exon 8Intron 7Exon 7 inclusionSMN2-Centromeric survival of motor neuron 2
3、geneU2.U2SSAAp14Decoy-BPS.5353BPSExonExonSSA-spliceostatin ADMD-Duchenne muscular dystrophy geneDantrolene.Exon 48Exon 51Exon 52DMDmutASOIntron 48/50.Exon 48Exon 51Exon 52Stop.Exon 48Exon 51Exon 52DMDmutIntron 48/50C-terminally truncated,non-functionaldystrophin protein.Exon 48 Exon 52Internally del
4、eted,partlyfunctional dystrophin proteinIndirectly viaCa2+signaling?.U2SF3b155p1453U2SF3b155p1453Prp5ATPE7107ABPSDisease linksU2AF65U2AF35U2 snRNAU2 snRNPSF3b155p14AExon 1Exon 2Myelodysplastic syndromesChronic lymphocytic leukaemiaChanges leading to haplo-insuffciencyor dominant negative effectsSpin
5、al muscular atrophySmproteinssnRNASMNSm siteU6atacU4atacTaybi-Linder syndrome(MOPD I)U2AF65U2AF35SF1/BBPExon 1AExon 2Myelodysplastic syndromesU1U2APPTSRSF135U2AF65.Chronic myelomonocytic leukemiahPrp8hBrr2hPrp6hPrp31hPrp3hPAP-1Retinitis pigmentosaRRMRRMRRMRRMISSU6 U4U5SF3b155SF3b155OOAcONHOOOOOOOOHH
6、NNNNNON+OHNNNOHONNOOHOHOOOOHOHSee online version for legends and references.690 Cell 162,July 30,2015 2015 Elsevier Inc.DOI http:/dx.doi.org/10.1016/j.cell.2015.07.033SnapShot:Spliceosome Dynamics IIIMarkus C.Wahl1 and Reinhard Lhrmann21Laboratory of Structural Biochemistry,Freie Universitt Berlin,T
7、akustrae 6,14195 Berlin,Germany2Department of Cellular Biochemistry,Max Planck Institute for Biophysical Chemistry,Am Fassberg 11,37077 Gttingen,Germany690.e1 Cell 162,July 30,2015 2015 Elsevier Inc.DOI http:/dx.doi.org/10.1016/j.cell.2015.07.033SnapShot:Spliceosome Dynamics IIIMarkus C.Wahl1 and Re
8、inhard Lhrmann21Laboratory of Structural Biochemistry,Freie Universitt Berlin,Takustrae 6,14195 Berlin,Germany2Department of Cellular Biochemistry,Max Planck Institute for Biophysical Chemistry,Am Fassberg 11,37077 Gttingen,GermanyThe complex compositional and conformational dynamics of spliceosomes
9、 required for regulated splicing are prone to malfunction when mutations affect splicing factors or cis-acting regulatory sequences.Indeed,many such mutations have been linked to heritable diseases or malignancies in humans.Small molecule modulators and antisense oligonucleotides or analogs harbor g
10、reat potential for therapies and several substances that can modulate splicing events have entered clinical trials.The intricate functional and regulatory dynamics of spliceosomes are very susceptible to small changes in the functionalities or levels of the associated splicing factors or the involve
11、d cis-acting elements on pre-mRNAs.Thus not surprisingly,up to 60%of disease-causing mutations in humans are thought to affect splicing or alternative splicing(Wang and Cooper,2007).For example,mutations that lead to amino acid changes in the U4/U6U5 tri-snRNP proteins Prp8,Brr2,Prp6,Prp31,Prp3,or P
12、AP-1 have been linked to severe forms of autosomal-dominant retinitis pigmentosa,a major cause of human blindness.Changes in early spliceosome assembly factors,U2 snRNP-associated protein SF3B1(SF3b155)or the splicing regulatory SR protein SRSF1(ASF/SF2),have been pinpointed as causes or modifiers o
13、f various hematologic disorders.Furthermore,changes in minor spliceosome components have also been linked to disease,such as nucleotide exchanges in U4atac snRNA in the case of Taybi-Linder syndrome(MOPD I).Mutations can also affect the splicing machinery indirectly by disturbing the functions of sp
14、licing-related proteins.For example,the survival of motor neuron protein(SMN)acts as an assembly chaperone for snRNP core particles and reduced amounts of functional SMN lead to spinal muscular atrophy(SMA),possibly due to reduced levels of active splicing machinery(Sleeman,2013).It is believed that
15、 SMN-linked SMA itself roots in a disturbed interplay of splicing regulatory mechanisms.In humans,the SMN protein is encoded by two genes,telomeric SMN1 and centromeric SMN2,which differ by only a few,translationally silent nucleotide exchanges.Yet,while SMN1 gives rise predominantly to functional,f
16、ull-length SMN protein,a single,silent nucleotide exchange in SMN2 exon 7 leads to the dysfunction of an exonic splicing enhancer(ESE)element that in SMN1 is recognized by the SRSF1(ASF/SF2)splicing regulator.As a consequence,in SMN2 pre-mRNA,SRSF1 cannot efficiently counteract hnRNP A1/A2-specific
17、intronic splicing silencer(ISS)elements in intron 7,thus leading to 90%of transcripts lacking exon 7(Hua et al.,2008).The resulting truncated version of SMN is not functional and is quickly degraded.SMN2 therefore cannot compensate for reduced SMN levels in cases where SMN1 is mutated.In some other
18、disease scenarios,functional and structural studies have also suggested molecular disease principles(Mozaffari-Jovin et al.,2013 and references therein),but often the actual molecular-mechanistic bases of splicing-factor-related diseases remain unknown.In particular,it is still largely unclear how d
19、efects in a ubiquitous cellular machinery,such as the spliceosome,can lead to tissue-specific disorders.Analysis of the molecular disease principles,including those that underlie SMN-linked SMA,is complicated,as some of the affected splicing factors are also involved in other cellular functions(Slee
20、man,2013).Only in recent years have several small molecule modulators of the spliceosome been discovered(Bonnal et al.,2012),including bacterial metabolites that exhibit antitumor activities and that often target components of the U2 snRNP-associated SF3b multi-protein complex.Such findings might in
21、dicate that the pathogenicity of some bacteria involves inhibition of the host splicing machinery.Small molecule modulators as well as antisense oligonucleotides(ASOs,predominantly based on nucleotide analogs)harbor great potential for the development of treatment strategies for splicing-factor-rela
22、ted disorders(Kole et al.,2012).For example,a small molecule substance,NVS-SM1,has recently been reported to counteract SMN2 exon 7 skipping by reinforcing the U1 snRNP-5 splice site interaction(Palacino et al.,2015).NVS-SM1 and another SMN2 splice-modifying small molecule(Naryshkin et al.,2014)as w
23、ell as ASOs that modulate splicing of the SMN2 gene or the Duchenne muscular dystrophy gene(DMD)(Kole et al.,2012)are presently in clinical trials(https:/www.clinicaltrials.gov/).Much remains to be learned for a full understanding of the conformational and compositional dynamics of spliceosomes,how
24、they are exploited for regulated splicing,and how their malfunction leads to disease.Recently,it has even been demonstrated that spliceosomes can be“repurposed”to serve other functions,possibly by mechanisms similar to those that regulate splicing events themselves.For example,regulated termination
25、of splicing after step 1 is used to mature telomerase RNA in Schizosaccharomyces pombe(Box et al.,2008)and splicing-related processes seem to give rise to exonic circular RNAs(Jeck and Sharpless,2014),whose functions are still poorly understood.Small molecule modulators will undoubtedly serve as pow
26、erful tools in the future to further dissect functional and regulatory dynamics of the spliceosome.RefeRencesBonnal,S.,Vigevani,L.,and Valcrcel,J.(2012).Nat.Rev.Drug Discov.11,847859.Box,J.A.,Bunch,J.T.,Tang,W.,and Baumann,P.(2008).Nature 456,910914.Hua,Y.,Vickers,T.A.,Okunola,H.L.,Bennett,C.F.,and
27、Krainer,A.R.(2008).Am.J.Hum.Genet.82,834848.Jeck,W.R.,and Sharpless,N.E.(2014).Nat.Biotechnol.32,453-461.Kole,R.,Krainer,A.R.,and Altman,S.(2012).Nat.Rev.Drug Discov.11,125140.Mozaffari-Jovin,S.,Wandersleben,T.,Santos,K.F.,Will,C.L.,Lhrmann,R.,and Wahl,M.C.(2013).Science 341,8084.Naryshkin,N.A.,Weet
28、all,M.,Dakka,A.,Narasimhan,J.,Zhao,X.,Feng,Z.,Ling,K.K.,Karp,G.M.,Qi,H.,Woll,M.G.,et al.(2014).Science 345,688693.Palacino,J.,Swalley,S.E.,Song,C.,Cheung,A.K.,Shu,L.,Zhang,X.,Van Hoosear,M.,Shin,Y.,Chin,D.N.,Keller,C.G.,et al.(2015).Nat.Chem.Biol.11,511517.Sleeman,J.(2013).Biochem.Soc.Trans.41,871875.Wang,G.S.,and Cooper,T.A.(2007).Nat.Rev.Genet.8,749761.