- Email: [email protected]
Sparks of the CRISPR explosion: Applications in medicine and agriculture
Journal of Genetics and Genomics 44 (2017) 413e414
Contents lists available at ScienceDirect
Journal of Genetics and Genomics Journal homepage: www.journals.elsevier.com/journal-of-geneticsand-genomics/
Editorial
Sparks of the CRISPR explosion: Applications in medicine and agriculture Exactly 30 years ago, Nakata and coworkers published a paper analyzing the 1664-nucleotide sequence of a chromosomal DNA segment that contained the iap gene and its flanking regions in Escherichia coli (Ishino et al., 1987). At the end of their paper, the authors reported that “an unusual structure was found in the 30 -end flanking region of iap” (Ishino et al., 1987). “Five highly homologous sequences of 29 nucleotides were arranged as direct repeats with 32 nucleotides as spacing” (Ishino et al., 1987). In the following decade, similar structures have been found in many other prokaryotes. In 2002, these structures have been called clustered regularly interspaced short palindromic repeats (CRISPR) (Jansen et al., 2002). The CRISPR and CRISPR-associated (Cas) endonuclease act as a defense system against invading viruses and plasmids in many different bacterial species (Barrangou et al., 2007). In 2012, Doudna, Charpentier and colleagues applied the CRISPR/Cas system in gene targeting by bringing RNA, protein and DNA into the same locus (Jinek et al., 2012). This milestone work triggered the “CRISPR explosion” for genome engineering (Cho et al., 2013; Cong et al., 2013; Jinek et al., 2013; Mali et al., 2013; Bassett and Liu, 2014). The CRISPR technology generates mutations with very high efficiency, and is ideally suited for generating mutant alleles for functional characterization of gene function. Furthermore, homologous targeting of the endogenous locus by the CRISPR technique will also enable wider applications of defined modifications of the genome. For example, it can be used to make appropriate modifications of the endogenous locus for visualization, detection and purification of regulatory factors (Bassett et al., 2014; Tanenbaum et al., 2014; Lackner et al., 2015; Savic et al., 2015; Leonetti et al., 2016). In addition, double or multiple mutants can be easily generated to study the epistasis of two regulatory factors in a common pathway (Yang et al., 2013). The adaptation of CRISPR/Cas9 system for genome engineering has revolutionized genetic analysis by providing a targeted mutagen. The simplicity at which it can be reprogrammed to target multiple sites also enables applications of this technology in genome-wide studies, thus allowing more powerful reverse genetic analyses (Shalem et al., 2014; Wang et al., 2014; Bassett et al., 2015). The Journal of Genetics and Genomics (JGG) has published several articles related to the progress of CRISPR over the past years. Last year, JGG published a Special Issue on genome editing (Jiao and Gao, 2016), focusing on the recent progress of genome editing and the methodological improvement. Here, this JGG special issue is focusing on the applications of CRISPR in two important areas: medicine and agriculture. This special issue includes three Review Articles, one Original Research Article and three Letters to the Editor. Firstly, Haoyi Wang and colleagues provide a timely perspective
on gene editing technologies in T cell therapy (pp. 415e422). They introduce the applications of various gene editing tools in T cell therapies, with a focus on antiviral strategies and cancer immunotherapies. Then, the Review Article by Bei Yang, Jia Chen and colleagues summarizes the physiological functions and structural features of APOBECs (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like), a family of cytidine deaminases that prefer single-stranded nucleic acids as substrates (pp. 423e437). They review not only the role of APOBECs as endogeneous mutators in carcinogenesis, but also the coupling of APOBECs and CRISPR for an improved gene-editing technique at single-base precision. The advantages and limitations of the APOBEC-CRISPR system for gene editing have been discussed as well. Genetic screens have been widely used for studying gene function. However, the reverse genetic screen has been limited due to technical difficulties. Haopeng Wang, Gaofeng Fan and their colleagues contribute a timely Review Article on designing and performing a CRISPR-Cas9 based genetic screen (pp. 439e449). They summarize recent development of the CRISPR technology in genetic screening and discuss the key factors in those screens. Moreover, they provide the guidance for experimental design, potential application and future direction of CRISPR-Cas9 based genetic screens. It has been a long-term challenge to generate mouse models carrying a defined point mutation, especially disease-related point mutations. Taking advantage of the CRISPR technology and androgenetic haploid embryonic stem cells, Jinsong Li, Wen Yuan and coworkers have established an approach for efficient generation of mice carrying a precise point mutation in one step. By analyzing and pre-selecting haploid cells for generation of mice with uniform gene modifications, this strategy provides an effective procedure for the study of novel human disease mutations in mice (pp. 461e463). The sustainable agriculture requires the development of reliable and effective seed-production strategies. Using the CRISPR/Cas technology, Caixia Gao and coworkers knock out the gene thermosensitive genic male-sterile 5 (TMS5) in maize (pp. 465e468). Homozygous T1 tms5 mutants are male-sterile at a higher temperature but male-fertile at a low temperature. Since it is a point mutation with a few nucleotide changes, a big trunk of foreign DNA has not been introduced into the plant via this approach. This strategy has a great potential in agriculture. The production of guide RNAs (gRNAs) is critical for the efficiency of the CRISPR-mediated gene editing in cells. Yunge zhao and colleagues have previously harnessed the self-cleaving activities of ribozymes to generate an artificial Ribozyme-gRNA-
https://doi.org/10.1016/j.jgg.2017.09.006 1673-8527/Copyright © 2017, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, and Genetics Society of China. Published by Elsevier Limited and Science Press. All rights reserved.
414
Editorial / Journal of Genetics and Genomics 44 (2017) 413e414
Ribozyme (RGR) gene for gRNA production (Gao and Zhao, 2014). They now improve the RGR system and expand the choice of promoters for gRNA production (pp. 469e472). Furthermore, the RGR system can be applicable to other nucleases such as Cpf1 which require gRNAs with precise ends. The CRISPR technology transforms the genome engineering both in model and non-model organisms such as silkworm (Bombyx mori). To overcome the limitation of transient delivery of Cas9 and gRNAs, Qingyou Xia and colleagues describe a simple tissue-specific genome editing strategy in the silkworm (pp. 451e459). They establish two transgenic B. mori lines, one expressing Cas9 protein in specific tissues while the other expressing the gRNA constitutively. By crossing these two lines, the team produces targeted mutagenesis in the silkworm in a spatially controlled manner. This approach can be adapted in other insects. In the past five years, we witness the development of the CRISPR technology from exploration to explosion. This JGG Special Issue on Applications of CRISPR in Medicine and Agriculture provides a few sparks among such an explosion. The CRISPR technology revolution is continuing to spread out to every corner of life sciences. References Barrangou, R., Fremaux, C., Deveau, H., Richards, M., Boyaval, P., Moineau, S., Romero, D.A., Horvath, P., 2007. CRISPR provides acquired resistance against viruses in prokaryotes. Science 315, 1709e1712. Bassett, A.R., Liu, J.L., 2014. CRISPR/Cas9 and genome editing in Drosophila. J. Genet. Genomics 41, 7e19. Bassett, A.R., Kong, L., Liu, J.L., 2015. A genome-wide CRISPR library for highthroughput genetic screening in Drosophila cells. J. Genet. Genomics 42, 301e309. Bassett, A.R., Tibbit, C., Ponting, C.P., Liu, J.L., 2014. Mutagenesis and homologous recombination in Drosophila cell lines using CRISPR/Cas9. Biol. Open 3, 42e49. Cho, S.W., Kim, S., Kim, J.M., Kim, J.S., 2013. Targeted genome engineering in human cells with the Cas9 RNA-guided endonuclease. Nat. Biotechnol. 31, 230e232. Cong, L., Ran, F.A., Cox, D., Lin, S., Barretto, R., Habib, N., Hsu, P.D., Wu, X., Jiang, W., Marraffini, L.A., Zhang, F., 2013. Multiplex genome engineering using CRISPR/ Cas systems. Science 339, 819e823. Gao, Y., Zhao, Y., 2014. Self-processing of ribozyme-flanked RNAs into guide RNAs in vitro and in vivo for CRISPR-mediated genome editing. J. Integr. Plant Biol. 56, 343e349. Ishino, Y., Shinagawa, H., Makino, K., Amemura, M., Nakata, A., 1987. Nucleotide sequence of the iap gene, responsible for alkaline phosphatase isozyme conversion in Escherichia coli, and identification of the gene product. J. Bacteriol. 169,
5429e5433. Jansen, R., Embden, J.D., Gaastra, W., Schouls, L.M., 2002. Identification of genes that are associated with DNA repeats in prokaryotes. Mol. Microbiol. 43, 1565e1575. Jiao, R., Gao, C., 2016. The CRISPR/Cas9 genome editing revolution. J. Genet. Genomics 43, 227e228. Jinek, M., Chylinski, K., Fonfara, I., Hauer, M., Doudna, J.A., Charpentier, E., 2012. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337, 816e821. Jinek, M., East, A., Cheng, A., Lin, S., Ma, E., Doudna, J., 2013. RNA-programmed genome editing in human cells. eLife 2, e00471. , A., Guzzardo, P.M., Banning, C., Mangena, R., Henley, T., Lackner, D.H., Carre Oberndorfer, S., Gapp, B.V., Nijman, S.M., Brummelkamp, T.R., Bürckstümmer, T., 2015. A generic strategy for CRISPR-Cas9-mediated gene tagging. Nat. Commun. 6, 10237. Leonetti, M.D., Sekine, S., Kamiyama, D., Weissman, J.S., Huang, B., 2016. A scalable strategy for high-throughput GFP tagging of endogenous human proteins. Proc. Natl. Acad. Sci. U. S. A. 113, E3501eE3508. Mali, P., Yang, L., Esvelt, K.M., Aach, J., Guell, M., DiCarlo, J.E., Norville, J.E., Church, G.M., 2013. RNA-guided human genome engineering via Cas9. Science 339, 823e826. Savic, D., Partridge, E.C., Newberry, K.M., Smith, S.B., Meadows, S.K., Roberts, B.S., Mackiewicz, M., Mendenhall, E.M., Myers, R.M., 2015. CETCh-seq: CRISPR epitope tagging ChIP-seq of DNA-binding proteins. Genome Res. 25, 1581e1589. Shalem, O., Sanjana, N.E., Hartenian, E., Shi, X., Scott, D.A., Mikkelson, T., Heckl, D., Ebert, B.L., Root, D.E., Doench, J.G., Zhang, F., 2014. Genome-scale CRISPR-Cas9 knockout screening in human cells. Science 343, 84e87. Tanenbaum, M.E., Gilbert, L.A., Qi, L.S., Weissman, J.S., Vale, R.D., 2014. A proteintagging system for signal amplification in gene expression and fluorescence imaging. Cell 159, 635e646. Wang, T., Wei, J.J., Sabatini, D.M., Lander, E.S., 2014. Genetic screens in human cells using the CRISPR-Cas9 system. Science 343, 80e84. Yang, H., Wang, H., Shivalila, C.S., Cheng, A.W., Shi, L., Jaenisch, R., 2013. One-step generation of mice carrying reporter and conditional alleles by CRISPR/Casmediated genome engineering. Cell 154, 1370e1379.
Ji-Long Liu School of Life Science and Technology, ShanghaiTech University, Shanghai 201210, China Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT, United Kingdom E-mail addresses: [email protected], [email protected]. 16 September 2017 Available online 18 September 2017
Contents lists available at ScienceDirect
Journal of Genetics and Genomics Journal homepage: www.journals.elsevier.com/journal-of-geneticsand-genomics/
Editorial
Sparks of the CRISPR explosion: Applications in medicine and agriculture Exactly 30 years ago, Nakata and coworkers published a paper analyzing the 1664-nucleotide sequence of a chromosomal DNA segment that contained the iap gene and its flanking regions in Escherichia coli (Ishino et al., 1987). At the end of their paper, the authors reported that “an unusual structure was found in the 30 -end flanking region of iap” (Ishino et al., 1987). “Five highly homologous sequences of 29 nucleotides were arranged as direct repeats with 32 nucleotides as spacing” (Ishino et al., 1987). In the following decade, similar structures have been found in many other prokaryotes. In 2002, these structures have been called clustered regularly interspaced short palindromic repeats (CRISPR) (Jansen et al., 2002). The CRISPR and CRISPR-associated (Cas) endonuclease act as a defense system against invading viruses and plasmids in many different bacterial species (Barrangou et al., 2007). In 2012, Doudna, Charpentier and colleagues applied the CRISPR/Cas system in gene targeting by bringing RNA, protein and DNA into the same locus (Jinek et al., 2012). This milestone work triggered the “CRISPR explosion” for genome engineering (Cho et al., 2013; Cong et al., 2013; Jinek et al., 2013; Mali et al., 2013; Bassett and Liu, 2014). The CRISPR technology generates mutations with very high efficiency, and is ideally suited for generating mutant alleles for functional characterization of gene function. Furthermore, homologous targeting of the endogenous locus by the CRISPR technique will also enable wider applications of defined modifications of the genome. For example, it can be used to make appropriate modifications of the endogenous locus for visualization, detection and purification of regulatory factors (Bassett et al., 2014; Tanenbaum et al., 2014; Lackner et al., 2015; Savic et al., 2015; Leonetti et al., 2016). In addition, double or multiple mutants can be easily generated to study the epistasis of two regulatory factors in a common pathway (Yang et al., 2013). The adaptation of CRISPR/Cas9 system for genome engineering has revolutionized genetic analysis by providing a targeted mutagen. The simplicity at which it can be reprogrammed to target multiple sites also enables applications of this technology in genome-wide studies, thus allowing more powerful reverse genetic analyses (Shalem et al., 2014; Wang et al., 2014; Bassett et al., 2015). The Journal of Genetics and Genomics (JGG) has published several articles related to the progress of CRISPR over the past years. Last year, JGG published a Special Issue on genome editing (Jiao and Gao, 2016), focusing on the recent progress of genome editing and the methodological improvement. Here, this JGG special issue is focusing on the applications of CRISPR in two important areas: medicine and agriculture. This special issue includes three Review Articles, one Original Research Article and three Letters to the Editor. Firstly, Haoyi Wang and colleagues provide a timely perspective
on gene editing technologies in T cell therapy (pp. 415e422). They introduce the applications of various gene editing tools in T cell therapies, with a focus on antiviral strategies and cancer immunotherapies. Then, the Review Article by Bei Yang, Jia Chen and colleagues summarizes the physiological functions and structural features of APOBECs (apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like), a family of cytidine deaminases that prefer single-stranded nucleic acids as substrates (pp. 423e437). They review not only the role of APOBECs as endogeneous mutators in carcinogenesis, but also the coupling of APOBECs and CRISPR for an improved gene-editing technique at single-base precision. The advantages and limitations of the APOBEC-CRISPR system for gene editing have been discussed as well. Genetic screens have been widely used for studying gene function. However, the reverse genetic screen has been limited due to technical difficulties. Haopeng Wang, Gaofeng Fan and their colleagues contribute a timely Review Article on designing and performing a CRISPR-Cas9 based genetic screen (pp. 439e449). They summarize recent development of the CRISPR technology in genetic screening and discuss the key factors in those screens. Moreover, they provide the guidance for experimental design, potential application and future direction of CRISPR-Cas9 based genetic screens. It has been a long-term challenge to generate mouse models carrying a defined point mutation, especially disease-related point mutations. Taking advantage of the CRISPR technology and androgenetic haploid embryonic stem cells, Jinsong Li, Wen Yuan and coworkers have established an approach for efficient generation of mice carrying a precise point mutation in one step. By analyzing and pre-selecting haploid cells for generation of mice with uniform gene modifications, this strategy provides an effective procedure for the study of novel human disease mutations in mice (pp. 461e463). The sustainable agriculture requires the development of reliable and effective seed-production strategies. Using the CRISPR/Cas technology, Caixia Gao and coworkers knock out the gene thermosensitive genic male-sterile 5 (TMS5) in maize (pp. 465e468). Homozygous T1 tms5 mutants are male-sterile at a higher temperature but male-fertile at a low temperature. Since it is a point mutation with a few nucleotide changes, a big trunk of foreign DNA has not been introduced into the plant via this approach. This strategy has a great potential in agriculture. The production of guide RNAs (gRNAs) is critical for the efficiency of the CRISPR-mediated gene editing in cells. Yunge zhao and colleagues have previously harnessed the self-cleaving activities of ribozymes to generate an artificial Ribozyme-gRNA-
https://doi.org/10.1016/j.jgg.2017.09.006 1673-8527/Copyright © 2017, Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, and Genetics Society of China. Published by Elsevier Limited and Science Press. All rights reserved.
414
Editorial / Journal of Genetics and Genomics 44 (2017) 413e414
Ribozyme (RGR) gene for gRNA production (Gao and Zhao, 2014). They now improve the RGR system and expand the choice of promoters for gRNA production (pp. 469e472). Furthermore, the RGR system can be applicable to other nucleases such as Cpf1 which require gRNAs with precise ends. The CRISPR technology transforms the genome engineering both in model and non-model organisms such as silkworm (Bombyx mori). To overcome the limitation of transient delivery of Cas9 and gRNAs, Qingyou Xia and colleagues describe a simple tissue-specific genome editing strategy in the silkworm (pp. 451e459). They establish two transgenic B. mori lines, one expressing Cas9 protein in specific tissues while the other expressing the gRNA constitutively. By crossing these two lines, the team produces targeted mutagenesis in the silkworm in a spatially controlled manner. This approach can be adapted in other insects. In the past five years, we witness the development of the CRISPR technology from exploration to explosion. This JGG Special Issue on Applications of CRISPR in Medicine and Agriculture provides a few sparks among such an explosion. The CRISPR technology revolution is continuing to spread out to every corner of life sciences. References Barrangou, R., Fremaux, C., Deveau, H., Richards, M., Boyaval, P., Moineau, S., Romero, D.A., Horvath, P., 2007. CRISPR provides acquired resistance against viruses in prokaryotes. Science 315, 1709e1712. Bassett, A.R., Liu, J.L., 2014. CRISPR/Cas9 and genome editing in Drosophila. J. Genet. Genomics 41, 7e19. Bassett, A.R., Kong, L., Liu, J.L., 2015. A genome-wide CRISPR library for highthroughput genetic screening in Drosophila cells. J. Genet. Genomics 42, 301e309. Bassett, A.R., Tibbit, C., Ponting, C.P., Liu, J.L., 2014. Mutagenesis and homologous recombination in Drosophila cell lines using CRISPR/Cas9. Biol. Open 3, 42e49. Cho, S.W., Kim, S., Kim, J.M., Kim, J.S., 2013. Targeted genome engineering in human cells with the Cas9 RNA-guided endonuclease. Nat. Biotechnol. 31, 230e232. Cong, L., Ran, F.A., Cox, D., Lin, S., Barretto, R., Habib, N., Hsu, P.D., Wu, X., Jiang, W., Marraffini, L.A., Zhang, F., 2013. Multiplex genome engineering using CRISPR/ Cas systems. Science 339, 819e823. Gao, Y., Zhao, Y., 2014. Self-processing of ribozyme-flanked RNAs into guide RNAs in vitro and in vivo for CRISPR-mediated genome editing. J. Integr. Plant Biol. 56, 343e349. Ishino, Y., Shinagawa, H., Makino, K., Amemura, M., Nakata, A., 1987. Nucleotide sequence of the iap gene, responsible for alkaline phosphatase isozyme conversion in Escherichia coli, and identification of the gene product. J. Bacteriol. 169,
5429e5433. Jansen, R., Embden, J.D., Gaastra, W., Schouls, L.M., 2002. Identification of genes that are associated with DNA repeats in prokaryotes. Mol. Microbiol. 43, 1565e1575. Jiao, R., Gao, C., 2016. The CRISPR/Cas9 genome editing revolution. J. Genet. Genomics 43, 227e228. Jinek, M., Chylinski, K., Fonfara, I., Hauer, M., Doudna, J.A., Charpentier, E., 2012. A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity. Science 337, 816e821. Jinek, M., East, A., Cheng, A., Lin, S., Ma, E., Doudna, J., 2013. RNA-programmed genome editing in human cells. eLife 2, e00471. , A., Guzzardo, P.M., Banning, C., Mangena, R., Henley, T., Lackner, D.H., Carre Oberndorfer, S., Gapp, B.V., Nijman, S.M., Brummelkamp, T.R., Bürckstümmer, T., 2015. A generic strategy for CRISPR-Cas9-mediated gene tagging. Nat. Commun. 6, 10237. Leonetti, M.D., Sekine, S., Kamiyama, D., Weissman, J.S., Huang, B., 2016. A scalable strategy for high-throughput GFP tagging of endogenous human proteins. Proc. Natl. Acad. Sci. U. S. A. 113, E3501eE3508. Mali, P., Yang, L., Esvelt, K.M., Aach, J., Guell, M., DiCarlo, J.E., Norville, J.E., Church, G.M., 2013. RNA-guided human genome engineering via Cas9. Science 339, 823e826. Savic, D., Partridge, E.C., Newberry, K.M., Smith, S.B., Meadows, S.K., Roberts, B.S., Mackiewicz, M., Mendenhall, E.M., Myers, R.M., 2015. CETCh-seq: CRISPR epitope tagging ChIP-seq of DNA-binding proteins. Genome Res. 25, 1581e1589. Shalem, O., Sanjana, N.E., Hartenian, E., Shi, X., Scott, D.A., Mikkelson, T., Heckl, D., Ebert, B.L., Root, D.E., Doench, J.G., Zhang, F., 2014. Genome-scale CRISPR-Cas9 knockout screening in human cells. Science 343, 84e87. Tanenbaum, M.E., Gilbert, L.A., Qi, L.S., Weissman, J.S., Vale, R.D., 2014. A proteintagging system for signal amplification in gene expression and fluorescence imaging. Cell 159, 635e646. Wang, T., Wei, J.J., Sabatini, D.M., Lander, E.S., 2014. Genetic screens in human cells using the CRISPR-Cas9 system. Science 343, 80e84. Yang, H., Wang, H., Shivalila, C.S., Cheng, A.W., Shi, L., Jaenisch, R., 2013. One-step generation of mice carrying reporter and conditional alleles by CRISPR/Casmediated genome engineering. Cell 154, 1370e1379.
Ji-Long Liu School of Life Science and Technology, ShanghaiTech University, Shanghai 201210, China Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford OX1 3PT, United Kingdom E-mail addresses: [email protected], [email protected]. 16 September 2017 Available online 18 September 2017