Programmable RNA editing with compact CRISPR–Cas13 systems from uncultivated microbes

Makarova, K. S. et al. An updated evolutionary classification of CRISPR-Cas systems. Nat. Rev. Microbiol. 13, 722–736 (2015).

Fonfara, I. et al. Phylogeny of Cas9 determines functional exchangeability of dual-RNA and Cas9 among orthologous type II CRISPR-Cas systems. Nucleic Acids Res. 42, 2577–2590 (2014).

Harrington, L. B. et al. Programmed DNA destruction by miniature CRISPR-Cas14 enzymes. Science 362, 839–842 (2018).

Pausch, P. et al. CRISPR-CasΦ from huge phages is a hypercompact genome editor. Science 369, 333–337 (2020).

Wang, D., Zhang, F. & Gao, G. CRISPR-based therapeutic genome editing: strategies and in vivo delivery by AAV vectors. Cell 181, 136–150 (2020).

Shmakov, S. et al. Discovery and functional characterization of diverse class 2 CRISPR-Cas systems. Mol. Cell 60, 385–397 (2015).

Abudayyeh, O. O. et al. C2c2 is a single-component programmable RNA-guided RNA-targeting CRISPR effector. Science 353, aaf5573 (2016).

Smargon, A. A., Shi, Y. J. & Yeo, G. W. RNA-targeting CRISPR systems from metagenomic discovery to transcriptomic engineering. Nat. Cell Biol. 22, 143–150 (2020).

Markowitz, V. M. et al. IMG: the integrated microbial genomes database and comparative analysis system. Nucleic Acids Res. 40, D115–D122 (2012).

O’Connell, M. R. Molecular mechanisms of RNA targeting by Cas13-containing type VI CRISPR-Cas systems. J. Mol. Biol. 431, 66–87 (2019).

Makarova, K. S. et al. Evolutionary classification of CRISPR-Cas systems: a burst of class 2 and derived variants. Nat. Rev. Microbiol. 18, 67–83 (2020).

Lynch, K. H., Stothard, P. & Dennis, J. J. Genomic analysis and relatedness of P2-like phages of the Burkholderia cepacia complex. BMC Genomics 11, 599 (2010).

Saridaki, A. et al. Wolbachia prophage DNA adenine methyltransferase genes in different Drosophila-Wolbachia associations. PLoS ONE 6, e19708 (2011).

Sternberg, N. & Coulby, J. Cleavage of the bacteriophage P1 packaging site (pac) is regulated by adenine methylation. Proc. Natl Acad. Sci. USA 87, 8070–8074 (1990).

Konermann, S. et al. Transcriptome engineering with RNA-targeting type VI-D CRISPR effectors. Cell 173, 665–676.e614 (2018).

Smargon, A. A. et al. Cas13b is a type VI-B CRISPR-associated RNA-guided RNase differentially regulated by accessory proteins Csx27 and Csx28. Mol. Cell 65, 618–630.e617 (2017).

Abudayyeh, O. O. et al. RNA targeting with CRISPR-Cas13. Nature 550, 280–284 (2017).

Wang, Q. et al. The CRISPR-Cas13a gene-editing system induces collateral cleavage of RNA in glioma cells. Adv. Sci. (Weinh.) 6, 1901299 (2019).

Abbott, T. R. et al. Development of CRISPR as an antiviral strategy to combat SARS-CoV-2 and influenza. Cell https://doi.org/10.1016/j.cell.2020.04.020 (2020).

Freije, C. A. et al. Programmable inhibition and detection of RNA viruses using Cas13. Mol. Cell 76, 826–837.e811 (2019).

Nguyen, T. M., Zhang, Y. & Pandolfi, P. P. Virus against virus: a potential treatment for 2019-nCov (SARS-CoV-2) and other RNA viruses. Cell Res. 30, 189–190 (2020).

Hulo, C. et al. ViralZone: a knowledge resource to understand virus diversity. Nucleic Acids Res. 39, D576–D582 (2011).

de Wit, E. et al. Prophylactic and therapeutic remdesivir (GS-5734) treatment in the rhesus macaque model of MERS-CoV infection. Proc. Natl Acad. Sci. USA 117, 6771–6776 (2020).

Ruch, T. R. & Machamer, C. E. The coronavirus E protein: assembly and beyond. Viruses 4, 363–382 (2012).

Harding, A. T., Heaton, B. E., Dumm, R. E. & Heaton, N. S. Rationally designed influenza virus vaccines that are antigenically stable during growth in eggs. mBio https://doi.org/10.1128/mBio.00669-17 (2017).

Abudayyeh, O. O. et al. A cytosine deaminase for programmable single-base RNA editing. Science 365, 382–386 (2019).

Kushawah, G. et al. CRISPR-Cas13d induces efficient mRNA knockdown in animal embryos. Dev. Cell 54, 805–817 (2020).

He, B. et al. Modulation of metabolic functions through Cas13d-mediated gene knockdown in liver. Protein Cell https://doi.org/10.1007/s13238-020-00700-2 (2020).

Zhou, H. et al. Glia-to-neuron conversion by CRISPR-CasRx alleviates symptoms of neurological disease in mice. Cell https://doi.org/10.1016/j.cell.2020.03.024 (2020).

Zhou, C. et al. CasRx-mediated RNA targeting prevents choroidal neovascularization in a mouse model of age-related macular degeneration. Natl Sci. Rev. https://doi.org/10.1093/nsr/nwaa033 (2020).

Mehta, A. & Merkel, O. M. Immunogenicity of Cas9 protein. J. Pharm. Sci. 109, 62–67 (2020).

East-Seletsky, A. et al. Two distinct RNase activities of CRISPR-C2c2 enable guide-RNA processing and RNA detection. Nature 538, 270–273 (2016).

East-Seletsky, A., O’Connell, M. R., Burstein, D., Knott, G. J. & Doudna, J. A. RNA targeting by functionally orthogonal type VI-A CRISPR-Cas enzymes. Mol. Cell 66, 373–383.e373 (2017).

Cox, D. B. T. et al. RNA editing with CRISPR-Cas13. Science 358, 1019–1027 (2017).

Edgar, R. C. PILER-CR: fast and accurate identification of CRISPR repeats. BMC Bioinformatics 8, 18 (2007).

Hyatt, D. et al. Prodigal: prokaryotic gene recognition and translation initiation site identification. BMC Bioinformatics 11, 119 (2010).

Katoh, K. & Standley, D. M. MAFFT multiple sequence alignment software version 7: improvements in performance and usability. Mol. Biol. Evolution 30, 772–780 (2013).

Kumar, S., Stecher, G. & Tamura, K. MEGA7: Molecular Evolutionary Genetics Analysis version 7.0 for bigger datasets. Mol. Biol. Evolution 33, 1870–1874 (2016).

Zhang, Y. I-TASSER server for protein 3D structure prediction. BMC Bioinformatics 9, 40 (2008).

Biswas, A., Gagnon, J. N., Brouns, S. J., Fineran, P. C. & Brown, C. M. CRISPRTarget: bioinformatic prediction and analysis of crRNA targets. RNA Biol. 10, 817–827 (2013).

Martin, M. Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet J. https://doi.org/10.14806/ej.17.1.200 (2011)

Clement, K. et al. CRISPResso2 provides accurate and rapid genome editing sequence analysis. Nat. Biotechnol. 37, 224–226 (2019).

Kluesner, M. G. et al. EditR: a method to quantify base editing from Sanger sequencing. CRISPR J. 1, 239–250 (2018).

Cox, M. P., Peterson, D. A. & Biggs, P. J. SolexaQA: at-a-glance quality assessment of Illumina second-generation sequencing data. BMC Bioinformatics 11, 485 (2010).

Kim, D., Langmead, B. & Salzberg, S. L. HISAT: a fast spliced aligner with low memory requirements. Nat. Methods 12, 357–360 (2015).

Anders, S., Pyl, P. T. & Huber, W. HTSeq—a Python framework to work with high-throughput sequencing data. Bioinformatics 31, 166–169 (2015).

Love, M. I., Huber, W. & Anders, S. Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15, 550 (2014).

Picardi, E. & Pesole, G. REDItools: high-throughput RNA editing detection made easy. Bioinformatics 29, 1813–1814 (2013).

Smigielski, E. M., Sirotkin, K., Ward, M. & Sherry, S. T. dbSNP: a database of single nucleotide polymorphisms. Nucleic Acids Res. 28, 352–355 (2000).

Gootenberg, J. S. et al. Nucleic acid detection with CRISPR-Cas13a/C2c2. Science 356, 438–442 (2017).

Tambe, A., East-Seletsky, A., Knott, G. J., Doudna, J. A. & O’Connell, M. R. RNA binding and HEPN-nuclease activation are decoupled in CRISPR-Cas13a. Cell Rep. 24, 1025–1036 (2018).