CRISPR/Cas9技术及其在非编码RNA编辑中的应用
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摘要
基因的定点修饰是研究基因功能的重要手段之一,而基因定点修饰技术的发展也经历了从早期的基因打靶到三代人工核酸内切酶的过程。第三代人工核酸内切酶规律成簇间隔短回文重复序列及其相关蛋白(CRTSPR/Cas)因其操作简便、成本低廉、作用高效,被迅速应用于生物学研究的各个领域。非编码RNA(ncRNAs)不具编码蛋白功能,直接以RNA的形式参与机体的几乎所有生理、病理过程。本文对CRISPR/Cas9技术的结构、发展和应用进行综述,并着重介绍其在miRNA和lncRNA编辑中的应用。
Genetic modification is one of the most important means for studying the function of genes.The development of genetic modification technique has undergone a process from gene targeting to the third generation of artificial endonuclease.Recently,the third generation genetic engineering tool called Clustered Regularly Interspaced Short Palindromic Repeats(CRISPR)/CRISPR-associated(Cas)system has been widely applied to biomedical research due to its simple operation,low cost and high efficiency.Noncoding RNAs(ncRNAs)have no protein-coding capability and they participate in almost all physiological and pathological processes of the form of RNA directly.This article briefly reviews the structure,development and application of CRISPR/Cas9 technology,and then highlights its application in the editing of miRNAs and lncRNAs.
Genetic modification is one of the most important means for studying the function of genes.The development of genetic modification technique has undergone a process from gene targeting to the third generation of artificial endonuclease.Recently,the third generation genetic engineering tool called Clustered Regularly Interspaced Short Palindromic Repeats(CRISPR)/CRISPR-associated(Cas)system has been widely applied to biomedical research due to its simple operation,low cost and high efficiency.Noncoding RNAs(ncRNAs)have no protein-coding capability and they participate in almost all physiological and pathological processes of the form of RNA directly.This article briefly reviews the structure,development and application of CRISPR/Cas9 technology,and then highlights its application in the editing of miRNAs and lncRNAs.
引文
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[3]Joung J K,Sander J D.TALENs:a widely applicable technology for targeted genome editing[J].Nature Reviews Molecular Cell Biology,2013,14(1):49-55.
[4]Mussolino C,Cathomen T.RNA guides genome engineering[J].Nature Biotechnology,2013,31(3):208-209.
[5]Grissa I,Vergnaud G,Pourcel A C.The CRISPRdb database and tools to display CRISPRs and to generate dictionaries of spacers and repeats[J].Bmc Bioinformatics,2007,8(1):172.
[6]Makarova K S,Haft D H,Barrangou R,et al.Evolution and classification of the CRISPR-Cas systems[J].Nature Reviews Microbiology,2011,9(6):467-477.
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[8]Mojica F J,Diez-Villasenor C,Garcia-Martinez J,et al.Short motif sequences determine the targets of the prokaryotic CRISPR defence system[J].Microbiology,2009,155(Pt 3):733.
[9]Brouns S J J,Jore M M,Lundgren M,et al.Small CRISPRRNAs Guide Antiviral Defense in Prokaryotes[J].Science,2008,321(5891):960-964.
[10]Pougach K,Semenova EE,Datsenko KA,et al.Transcription,processing and function of CRISPR cassettes in Escherichia coli[J].Molecular Microbiology,2010,77(6):1367-1379.
[11]Umit Pul,Wurm R,Arslan Z,et al.Identification and characterization of E.coli CRISPR-cas promoters and their silencing by H-NS[J].Molecular Microbiology,2010,75(6):1495-1512.
[12]Garneau JE,Dupuis ME,Villion M,et al.The CRISPR/Cas bacterial immune system cleaves bacteriophage and plasmid DNA[J].Nature,2010,468(7320):67.
[13]Deltcheva E,Chylinski K,Sharma C M,et al.CRISPR RNAmaturation by trans-encoded small RNA and host factor RNase III[J].Nature,2011,471(7340):602-607.
[14]Cong L,Ran F A,Cox D,et al.Multiplex genome engineering using CRISPR/Cas systems.[J].Trends in Genetics Tig,2013,8(11):819-823.
[15]Mali P,Yang L,Esvelt K M,et al.RNA-Guided Human Genome Engineering via Cas9[J].Science,2013,339(6121):823.
[16]Ha S J,Kim J S,Sung Y H,et al.Highly efficient gene knockout in mice and zebrafish with RNA-guided endonucleases[J].Genome Research,2014,24(1):125-131.
[17]Zhou J,Shen B,Zhang W,et al.One-step generation of different immunodeficient mice with multiple gene modifications by CRISPR/Cas9mediated genome engineering[J].Int J Biochem Cell Biol,2014,46(1):49-55.
[18]Zhao P,Zhang Z,Ke H,et al.Oligonucleotide-based targeted gene editing in C elegans via the CRISPR/Cas9system[J].Cell Research,2014,24(2):247-250.
[19]Fu Y,Foden J A,Khayter C,et al.High-frequency off-target mutagenesis induced by CRISPR-Cas nucleases in human cells[J].Nature Biotechnology,2013,31(9):822-826.
[20]Ran F A,Hsu P D,Lin C Y,et al.Double nicking by RNA-guided CRISPR Cas9for enhanced genome editing specificity[J].Cell,2013,154(6):1380-1389.
[21]Kim K,Ryu S M,Kim S T,et al.Highly efficient RNA-guided base editing in mouse embryos[J].Nature Biotechnology,2017,35(5):435.
[22]Liao H K,Hatanaka F,Araoka T,et al.In Vivo Target Gene Activation via CRISPR/Cas9-Mediated Trans-epigenetic Modulation[J].Cell,2017,171(7):1495.
[23]Harrington L B,Doxzen K W,Ma E,et al.A Broad-Spectrum Inhibitor of CRISPR-Cas9[J].Cell,2017,170(6):1224.
[24]Nayak D D,Metcalf W W.Cas9-mediated genome editing in the methanogenic archaeon Methanosarcina acetivorans.[J].Proceedings of the National Academy of Sciences of the United States of America,2017,114(11):2976.
[25]Jones D L,Leroy P,Unoson C,et al.Kinetics of dCas9target search in Escherichia coli[J].Science,2017,357(6358):1420-1424.
[26]Paix A,Folkmann A,Goldman D H,et al.Precision genome editing using synthesis-dependent repair of Cas9-induced DNA breaks[J].Proceedings of the National Academy of Sciences of the United States of America,2017,114(50):10745.
[27]Gao X,Tao Y,Lamas V,et al.Treatment of autosomal dominant hearing loss by in vivo delivery of genome editing agents[J].Nature,2017:217-221.
[28]Ma H,Martigutierrez N,Park SW,et al.Correction of a pathogenic gene mutation in human embryos[J].Nature,2017,548(7668):413.
[29]Wright MW,Bruford EA.Naming'junk':Human non-protein coding RNA(ncRNA)gene nomenclature[J].Human Genomics,2011,5(2):1-9.
[30]Guttman M,Amit I,Garber M,et al.Chromatin signature reveals over a thousand highly conserved large non-coding RNAs in mammals.[J].Nature,2009,458(7235):223.
[31]Ha M,Kim VN.Regulation of microRNA biogenesis[J].Nature Reviews Molecular Cell Biology,2014,15(8):509-524.
[32]Bushati N,Cohen SM,Stark A,et al.Animal MicroRNAs confer robustness to gene expression and have a significant impact on 3'UTR evolution[J].Cell,2005,123(6):1133-1146.
[33]Lim LP,Lau NC,Garrett-Engele P,et al.Microarray analysis shows that some microRNAs downregulate large numbers of target mRNAs[J].Nature,2005,433(7027):769.
[34]Yoon JH,Abdelmohsen K,Gorospe M.Functional interactions among microRNAs and long noncoding RNAs[J].Seminars in Cell&Developmental Biology,2014,34(4):9-14.
[35]Xiao A,Wang Z,Hu Y,et al.Chromosomal deletions and inversions mediated by TALENs and CRISPR/Cas in zebrafish[J].Nucleic Acids Research,2013,41(14):141.
[36]Ho TT,Zhou N,Huang J,et al.Targeting non-coding RNAs with the CRISPR/Cas9system in human cell lines[J].Nucleic Acids Research,2015,43(3):17.
[37]Chang H,Yi B,Ma R,et al.CRISPR/cas9,a novel genomic tool to knock down microRNAin vitroandin vivo[J].Sci Rep,2016,6:22312.
[38]Zhang Z,Ursin R,Mahapatra S,et al.CRISPR/CAS9ablation of individual miRNAs from a miRNA family reveals their individual efficacies for regulating cardiac differentiation.[J].Mechanisms of Development,2018,150:10-20.
[39]Kurata JS,Lin R J.MicroRNA-focused CRISPR-Cas9Library Screen Reveals Fitness-Associated miRNAs[J].Rna-a Publication of the Rna Society,2018,24(7):118.
[40]Wang KC,Chang HY.Molecular Mechanisms of Long Noncoding RNAs[J].Molecular Cell,2011,43(6):904.
[41]Han J,Zhang J,Chen L,et al.Efficient in vivo deletion of a large imprinted lncRNA by CRISPR/Cas9[J].Rna Biology,2014,11(7):829-835.
[42]Peng L,Pan P,Chen J,et al.A tetracycline-inducible CRISPR/Cas9 system,targeting two long non-coding RNAs,suppresses the malignant behavior of bladder cancer cells[J].Oncology Letters,2018,16(4):4309-4316.
[43]Stafford DA,Dichmann DS,Chang JK,et al.Deletion of the sclerotome-enriched lncRNA PEAT augments ribosomal protein expression[J].Proc Natl Acad Sci U S A,2017,114(1):101-106.
[44]Leisegang MS,Fork C,Josipovic I,et al.Long Noncoding RNA MANTIS Facilitates Endothelial Angiogenic Function.[J].Circulation,2017,136(1):65-79.
[45]Li R,Fu D,Zhu B,et al.CRISPR/Cas9‐mediated mutagenesis of lncRNA1459alters tomato fruit ripening[J].Plant Journal,2018,94(3):513-524.
[46]Goyal A,Myacheva K,Grob M,et al.Challenges of CRISPR/Cas9applications for long non-coding RNA genes.[J].Nucleic Acids Research,2016,11(19):6883-6894.
[2]Li T,Huang S,Zhao X,et al.Modularly assembled designer TAL effector nucleases for targeted gene knockout and gene replacement in eukaryotes[J].Nucleic Acids Research,2011,39(14):6315.
[3]Joung J K,Sander J D.TALENs:a widely applicable technology for targeted genome editing[J].Nature Reviews Molecular Cell Biology,2013,14(1):49-55.
[4]Mussolino C,Cathomen T.RNA guides genome engineering[J].Nature Biotechnology,2013,31(3):208-209.
[5]Grissa I,Vergnaud G,Pourcel A C.The CRISPRdb database and tools to display CRISPRs and to generate dictionaries of spacers and repeats[J].Bmc Bioinformatics,2007,8(1):172.
[6]Makarova K S,Haft D H,Barrangou R,et al.Evolution and classification of the CRISPR-Cas systems[J].Nature Reviews Microbiology,2011,9(6):467-477.
[7]Terns M P,Terns R M.CRISPR-based adaptive immune systems.[J].Current Opinion in Microbiology,2011,14(3):321-327.
[8]Mojica F J,Diez-Villasenor C,Garcia-Martinez J,et al.Short motif sequences determine the targets of the prokaryotic CRISPR defence system[J].Microbiology,2009,155(Pt 3):733.
[9]Brouns S J J,Jore M M,Lundgren M,et al.Small CRISPRRNAs Guide Antiviral Defense in Prokaryotes[J].Science,2008,321(5891):960-964.
[10]Pougach K,Semenova EE,Datsenko KA,et al.Transcription,processing and function of CRISPR cassettes in Escherichia coli[J].Molecular Microbiology,2010,77(6):1367-1379.
[11]Umit Pul,Wurm R,Arslan Z,et al.Identification and characterization of E.coli CRISPR-cas promoters and their silencing by H-NS[J].Molecular Microbiology,2010,75(6):1495-1512.
[12]Garneau JE,Dupuis ME,Villion M,et al.The CRISPR/Cas bacterial immune system cleaves bacteriophage and plasmid DNA[J].Nature,2010,468(7320):67.
[13]Deltcheva E,Chylinski K,Sharma C M,et al.CRISPR RNAmaturation by trans-encoded small RNA and host factor RNase III[J].Nature,2011,471(7340):602-607.
[14]Cong L,Ran F A,Cox D,et al.Multiplex genome engineering using CRISPR/Cas systems.[J].Trends in Genetics Tig,2013,8(11):819-823.
[15]Mali P,Yang L,Esvelt K M,et al.RNA-Guided Human Genome Engineering via Cas9[J].Science,2013,339(6121):823.
[16]Ha S J,Kim J S,Sung Y H,et al.Highly efficient gene knockout in mice and zebrafish with RNA-guided endonucleases[J].Genome Research,2014,24(1):125-131.
[17]Zhou J,Shen B,Zhang W,et al.One-step generation of different immunodeficient mice with multiple gene modifications by CRISPR/Cas9mediated genome engineering[J].Int J Biochem Cell Biol,2014,46(1):49-55.
[18]Zhao P,Zhang Z,Ke H,et al.Oligonucleotide-based targeted gene editing in C elegans via the CRISPR/Cas9system[J].Cell Research,2014,24(2):247-250.
[19]Fu Y,Foden J A,Khayter C,et al.High-frequency off-target mutagenesis induced by CRISPR-Cas nucleases in human cells[J].Nature Biotechnology,2013,31(9):822-826.
[20]Ran F A,Hsu P D,Lin C Y,et al.Double nicking by RNA-guided CRISPR Cas9for enhanced genome editing specificity[J].Cell,2013,154(6):1380-1389.
[21]Kim K,Ryu S M,Kim S T,et al.Highly efficient RNA-guided base editing in mouse embryos[J].Nature Biotechnology,2017,35(5):435.
[22]Liao H K,Hatanaka F,Araoka T,et al.In Vivo Target Gene Activation via CRISPR/Cas9-Mediated Trans-epigenetic Modulation[J].Cell,2017,171(7):1495.
[23]Harrington L B,Doxzen K W,Ma E,et al.A Broad-Spectrum Inhibitor of CRISPR-Cas9[J].Cell,2017,170(6):1224.
[24]Nayak D D,Metcalf W W.Cas9-mediated genome editing in the methanogenic archaeon Methanosarcina acetivorans.[J].Proceedings of the National Academy of Sciences of the United States of America,2017,114(11):2976.
[25]Jones D L,Leroy P,Unoson C,et al.Kinetics of dCas9target search in Escherichia coli[J].Science,2017,357(6358):1420-1424.
[26]Paix A,Folkmann A,Goldman D H,et al.Precision genome editing using synthesis-dependent repair of Cas9-induced DNA breaks[J].Proceedings of the National Academy of Sciences of the United States of America,2017,114(50):10745.
[27]Gao X,Tao Y,Lamas V,et al.Treatment of autosomal dominant hearing loss by in vivo delivery of genome editing agents[J].Nature,2017:217-221.
[28]Ma H,Martigutierrez N,Park SW,et al.Correction of a pathogenic gene mutation in human embryos[J].Nature,2017,548(7668):413.
[29]Wright MW,Bruford EA.Naming'junk':Human non-protein coding RNA(ncRNA)gene nomenclature[J].Human Genomics,2011,5(2):1-9.
[30]Guttman M,Amit I,Garber M,et al.Chromatin signature reveals over a thousand highly conserved large non-coding RNAs in mammals.[J].Nature,2009,458(7235):223.
[31]Ha M,Kim VN.Regulation of microRNA biogenesis[J].Nature Reviews Molecular Cell Biology,2014,15(8):509-524.
[32]Bushati N,Cohen SM,Stark A,et al.Animal MicroRNAs confer robustness to gene expression and have a significant impact on 3'UTR evolution[J].Cell,2005,123(6):1133-1146.
[33]Lim LP,Lau NC,Garrett-Engele P,et al.Microarray analysis shows that some microRNAs downregulate large numbers of target mRNAs[J].Nature,2005,433(7027):769.
[34]Yoon JH,Abdelmohsen K,Gorospe M.Functional interactions among microRNAs and long noncoding RNAs[J].Seminars in Cell&Developmental Biology,2014,34(4):9-14.
[35]Xiao A,Wang Z,Hu Y,et al.Chromosomal deletions and inversions mediated by TALENs and CRISPR/Cas in zebrafish[J].Nucleic Acids Research,2013,41(14):141.
[36]Ho TT,Zhou N,Huang J,et al.Targeting non-coding RNAs with the CRISPR/Cas9system in human cell lines[J].Nucleic Acids Research,2015,43(3):17.
[37]Chang H,Yi B,Ma R,et al.CRISPR/cas9,a novel genomic tool to knock down microRNAin vitroandin vivo[J].Sci Rep,2016,6:22312.
[38]Zhang Z,Ursin R,Mahapatra S,et al.CRISPR/CAS9ablation of individual miRNAs from a miRNA family reveals their individual efficacies for regulating cardiac differentiation.[J].Mechanisms of Development,2018,150:10-20.
[39]Kurata JS,Lin R J.MicroRNA-focused CRISPR-Cas9Library Screen Reveals Fitness-Associated miRNAs[J].Rna-a Publication of the Rna Society,2018,24(7):118.
[40]Wang KC,Chang HY.Molecular Mechanisms of Long Noncoding RNAs[J].Molecular Cell,2011,43(6):904.
[41]Han J,Zhang J,Chen L,et al.Efficient in vivo deletion of a large imprinted lncRNA by CRISPR/Cas9[J].Rna Biology,2014,11(7):829-835.
[42]Peng L,Pan P,Chen J,et al.A tetracycline-inducible CRISPR/Cas9 system,targeting two long non-coding RNAs,suppresses the malignant behavior of bladder cancer cells[J].Oncology Letters,2018,16(4):4309-4316.
[43]Stafford DA,Dichmann DS,Chang JK,et al.Deletion of the sclerotome-enriched lncRNA PEAT augments ribosomal protein expression[J].Proc Natl Acad Sci U S A,2017,114(1):101-106.
[44]Leisegang MS,Fork C,Josipovic I,et al.Long Noncoding RNA MANTIS Facilitates Endothelial Angiogenic Function.[J].Circulation,2017,136(1):65-79.
[45]Li R,Fu D,Zhu B,et al.CRISPR/Cas9‐mediated mutagenesis of lncRNA1459alters tomato fruit ripening[J].Plant Journal,2018,94(3):513-524.
[46]Goyal A,Myacheva K,Grob M,et al.Challenges of CRISPR/Cas9applications for long non-coding RNA genes.[J].Nucleic Acids Research,2016,11(19):6883-6894.