CRISPRi; MT: RNA/DNA Editing; gene silencing; gene therapy; myotonic dystrophy type 1; neuromuscular disease; Molecular Medicine; Drug Discovery
Abstract :
[en] Myotonic dystrophy type 1 (DM1) is a neuromuscular disease that originates from an expansion of CTG microsatellites in the 3' untranslated region of the DMPK gene, thus leading to the expression of transcripts containing expanded CUG repeats (CUGexp). The pathophysiology is explained by a toxic RNA gain of function where CUGexp RNAs form nuclear aggregates that sequester and alter the function of MBNL splicing factors, triggering splicing misregulation linked to the DM1 symptoms. There is currently no cure for DM1, and most therapeutic strategies aim at eliminating CUGexp-DMPK transcripts. Here, we investigate a DMPK-promoter silencing strategy using CRISPR interference as a new alternative approach. Different sgRNAs targeting the DMPK promoter are evaluated in DM1 patient muscle cells. The most effective guides allowed us to reduce the level of DMPK transcripts and CUGexp-RNA aggregates up to 80%. The CUGexp-DMPK repression corrects the overall transcriptome, including spliceopathy, and reverses a physiological parameter in DM1 muscle cells. Its action is specific and restricted to the DMPK gene, as confirmed by genome-wide expression analysis. Altogether, our findings highlight DMPK-promoter silencing by CRISPRi as a promising therapeutic approach for DM1.
Disciplines :
Neurology
Author, co-author :
Porquet, Florent ; Université de Liège - ULiège > Département des sciences biomédicales et précliniques > Pharmacologie ; Sorbonne Université, INSERM, Institut de Myologie, Centre de Recherche en Myologie, 75013 Paris, France
Weidong, Lin; Laboratory of Molecular and Cellular Epigenetics, GIGA-Cancer, ULiège, 4000 Liège, Belgium
Jehasse, Kevin ; Université de Liège - ULiège > GIGA > GIGA Neurosciences - Neurophysiology
Gazon, Hélène ; Université de Liège - ULiège > GIGA > GIGA Cancer - Cellular and Molecular Epigenetics
Kondili, Maria; Sorbonne Université, INSERM, Institut de Myologie, Centre de Recherche en Myologie, 75013 Paris, France
Blacher, Silvia ; Université de Liège - ULiège > Département des sciences biomédicales et précliniques > Biologie cellulaire et moléculaire
Massotte, Laurent ; Université de Liège - ULiège > Département des sciences biomédicales et précliniques
Furling, Denis; Sorbonne Université, INSERM, Institut de Myologie, Centre de Recherche en Myologie, 75013 Paris, France
Gillet, Nicolas ; Université de Liège - ULiège > Agronomie, Bio-ingénierie et Chimie (AgroBioChem) > Microbial technologies ; Namur Research Institute for Life Sciences (NARILIS), Integrated Veterinary Research Unit (URVI), University of Namur, 5000 Namur, Belgium
Klein, Arnaud François; Sorbonne Université, INSERM, Institut de Myologie, Centre de Recherche en Myologie, 75013 Paris, France
Seutin, Vincent ; Université de Liège - ULiège > Département des sciences biomédicales et précliniques > Pharmacologie
Willems, Luc ; Université de Liège - ULiège > GIGA > GIGA Cancer - Cellular and Molecular Epigenetics
The authors thank the human cell immortalization facility of the Myology Institute (Paris, France) as well as the GIGA technological platforms for support. In particular, the authors are grateful to François Giroulle, Alexandre Hego, Latifa Karim, Sandra Ormenese, Wouter Coppieters, Alexandra Revnic, Arnaud Lavergne, Raafat Stephan, and Stephen Freeman for their technical expertise and engagement, as well as Mario Gomes-Pereira, Marc Bitoun, and Geneviève Gourdon for their advice. This work was supported by the Fonds National de la Recherche Scientifique (FNRS, FRIA grant), The Myotonic Dystrophy Foundation and Wyck Foundations (2018 PhD trainee fellowship in Myotonic Dystrophy), the Association Belge contre les Maladies Neuro-Musculaires (ABMM; call 2019–2020), and the Fondation Léon Fredericq (FLF). F.P. was supported by fellowships of the Fonds National de la Recherche Scientifique , The Myotonic Dystrophy Foundation , and Wyck Foundations . V.S. is a full professor at ULiege, while L. Willems is a research director at the FNRS and A.F.K. is a research scientist at INSERM.The authors thank the human cell immortalization facility of the Myology Institute (Paris, France) as well as the GIGA technological platforms for support. In particular, the authors are grateful to François Giroulle, Alexandre Hego, Latifa Karim, Sandra Ormenese, Wouter Coppieters, Alexandra Revnic, Arnaud Lavergne, Raafat Stephan, and Stephen Freeman for their technical expertise and engagement, as well as Mario Gomes-Pereira, Marc Bitoun, and Geneviève Gourdon for their advice. This work was supported by the Fonds National de la Recherche Scientifique (FNRS, FRIA grant), The Myotonic Dystrophy Foundation and Wyck Foundations (2018 PhD trainee fellowship in Myotonic Dystrophy), the Association Belge contre les Maladies Neuro-Musculaires (ABMM; call 2019–2020), and the Fondation Léon Fredericq (FLF). F.P. was supported by fellowships of the Fonds National de la Recherche Scientifique, The Myotonic Dystrophy Foundation, and Wyck Foundations. V.S. is a full professor at ULiege, while L. Willems is a research director at the FNRS and A.F.K. is a research scientist at INSERM. Conceptualization, F.P.; methodology, F.P.; software, S.B. and M.K.; formal analysis, F.P. and K.J.; investigation, F.P. L. Weidong, A.F.K. H.G. and L.M.; resources, E.D.V. D.F. and A.F.K.; writing – original draft, F.P.; writing – review & editing, F.P. A.F.K. D.F. V.S. and L. Willems; visualization, F.P. and A.F.K.; supervision, A.F.K. N.A.G. L. Willems, and V.S.; project administration, V.S. and L. Willems; funding acquisition, F.P. The authors declare no competing interests.
Gourdon, G., Meola, G., Myotonic dystrophies: state of the art of new therapeutic developments for the CNS. Front. Cell. Neurosci. 11 (2017), 101–114, 10.3389/fncel.2017.00101.
Theadom, A., Rodrigues, M., Roxburgh, R., Balalla, S., Higgins, C., Bhattacharjee, R., Jones, K., Krishnamurthi, R., Feigin, V., Prevalence of muscular dystrophies : a systematic literature Review. Neuroepidemiology 43 (2014), 259–268, 10.1159/000369343.
Johnson, N.E., Butterfield, R.J., Mayne, K., Newcomb, T., Imburgia, C., Dunn, D., Duval, B., Feldkamp, M.L., Weiss, R.B., Population-based prevalence of myotonic dystrophy type 1 using genetic analysis of statewide blood screening program. Neurology 96 (2021), e1045–e1053, 10.1212/WNL.0000000000011425.
Brook, J.D., McCurrach, M.E., Harley, H.G., Buckler, A.J., Church, D., Aburatani, H., Hunter, K., Stanton, V.P., Thirion, J.-P., Hudson, T., et al. Molecular basis of myotonic dystrophy: expansion of a trinucleotide (CTG) repeat at the 3’ end of a transcript encoding a protein kinase family member. Cell 69 (1992), 385–808, 10.1016/0092-8674(92)90418-C.
De Antonio, M., Dogan, C., Hamroun, D., Mati, M., Zerrouki, S., Eymard, B., Katsahian, S., Bassez, G., French Myotonic Dystrophy Clinical Network. Unravelling the myotonic dystrophy type 1 clinical spectrum: a systematic registry-based study with implications for disease classification. Rev. Neurol. 172 (2016), 572–580, 10.1016/j.neurol.2016.08.003.
Taneja, K.L., McCurrach, M., Schalling, M., Housman, D., Singer, R.H., Foci of trinucleotide repeat transcripts in nuclei of myotonic dystrophy cells and tissues. J. Cell Biol. 128 (1995), 995–1002, 10.1083/jcb.128.6.995.
Jain, A., Vale, R.D., RNA phase transitions in repeat expansion disorders. Nature 546 (2017), 243–247, 10.1038/nature22386.
Goers, E.S., Purcell, J., Voelker, R.B., Gates, D.P., Berglund, J.A., MBNL1 binds GC motifs embedded in pyrimidines to regulate alternative splicing. Nucleic Acids Res. 38 (2010), 2467–2484, 10.1093/nar/gkp1209.
Osborne, R.J., Lin, X., Welle, S., Sobczak, K., O'Rourke, J.R., Swanson, M.S., Thornton, C.A., Transcriptional and post-transcriptional impact of toxic RNA in myotonic dystrophy. Hum. Mol. Genet. 18 (2009), 1471–1481, 10.1093/hmg/ddp058.
Kanadia, R.N., Johnstone, K.A., Mankodi, A., Lungu, C., Thornton, C.A., Esson, D., Timmers, A.M., Hauswirth, W.W., Swanson, M.S., A muscleblind knockout model for myotonic dystrophy. Science 302 (2003), 1978–1980, 10.1126/science.1088583.
Lee, K.Y., Li, M., Manchanda, M., Batra, R., Charizanis, K., Mohan, A., Warren, S.A., Chamberlain, C.M., Finn, D., Hong, H., et al. Compound loss of muscleblind-like function in myotonic dystrophy. EMBO Mol. Med. 5 (2013), 1887–1900, 10.1002/emmm.201303275.
Thomas, J.D., Sznajder, Ł.J., Bardhi, O., Aslam, F.N., Anastasiadis, Z.P., Scotti, M.M., Nishino, I., Nakamori, M., Wang, E.T., Swanson, M.S., Disrupted prenatal RNA processing and myogenesis in congenital myotonic dystrophy. Genes Dev. 31 (2017), 1122–1133, 10.1101/gad.300590.117.
Charizanis, K., Lee, K.Y., Batra, R., Goodwin, M., Zhang, C., Yuan, Y., Shiue, L., Cline, M., Scotti, M.M., Xia, G., et al. Muscleblind-like 2-mediated alternative splicing in the developing brain and dysregulation in myotonic dystrophy. Neuron 75 (2012), 437–450, 10.1016/j.neuron.2012.05.029.
Wang, E.T., Treacy, D., Eichinger, K., Struck, A., Estabrook, J., Olafson, H., Wang, T.T., Bhatt, K., Westbrook, T., Sedehizadeh, S., et al. Transcriptome alterations in myotonic dystrophy skeletal muscle and heart. Hum. Mol. Genet. 28 (2019), 1312–1321, 10.1093/hmg/ddy432.
Rau, F., Lainé, J., Ramanoudjame, L., Ferry, A., Arandel, L., Delalande, O., Jollet, A., Dingli, F., Lee, K.-Y., Peccate, C., et al. Abnormal splicing switch of DMD's penultimate exon compromises muscle fibre maintenance in myotonic dystrophy. Nat. Commun., 6, 2015, 7205, 10.1038/ncomms8205.
Fugier, C., Klein, A.F., Hammer, C., Vassilopoulos, S., Ivarsson, Y., Toussaint, A., Tosch, V., Vignaud, A., Ferry, A., Messaddeq, N., et al. Misregulated alternative splicing of BIN1 is associated with T tubule alterations and muscle weakness in myotonic dystrophy. Nat. Med. 17 (2011), 720–725, 10.1038/nm.2374.
Freyermuth, F., Rau, F., Kokunai, Y., Linke, T., Sellier, C., Nakamori, M., Kino, Y., Arandel, L., Jollet, A., Thibault, C., et al. Splicing misregulation of SCN5A contributes to cardiac-conduction delay and heart arrhythmia in myotonic dystrophy. Nat. Commun., 7, 2016, 11067, 10.1038/ncomms11067.
Savkur, R.S., Philips, A.V., Cooper, T.A., Aberrant regulation of insulin receptor alternative splicing is associated with insulin resistance in myotonic dystrophy. Nat. Genet. 29 (2001), 40–47, 10.1038/ng704.
Charlet-B, N., Savkur, R.S., Singh, G., Philips, A.V., Grice, E.A., Cooper, T.A., Loss of the muscle-specific chloride channel in type 1 myotonic dystrophy due to misregulated alternative splicing. Mol. Cell 10 (2002), 45–53, 10.1016/S1097-2765(02)00572-5.
Mankodi, A., Takahashi, M.P., Jiang, H., Beck, C.L., Bowers, W.J., Moxley, R.T., Cannon, S.C., Thornton, C.A., Expanded CUG repeats trigger aberrant splicing of ClC-1 chloride channel pre-mRNA and hyperexcitability of skeletal muscle in myotonic dystrophy. Mol. Cell 10 (2002), 35–44, 10.1016/S1097-2765(02)00563-4.
Dirksen, R.T., Lueck, J.D., Swanson, M.S., Wheeler, T.M., Thornton, C.A., Swanson, M.S., Dirksen, R.T., Thornton, C.A., Correction of ClC-1 splicing eliminates chloride channelopathy and myotonia in mouse models of myotonic dystrophy Find the latest version : correction of ClC-1 splicing eliminates chloride channelopathy and myotonia in mouse models of myotonic dystrophy. J. Clin. Invest. 117 (2007), 3952–3957, 10.1172/JCI33355.3952.
Ashizawa, T., Gagnon, C., Groh, W.J., Gutmann, L., Johnson, N.E., Meola, G., Moxley, R., Pandya, S., Rogers, M.T., Simpson, E., et al. Consensus-based care recommendations for adults with myotonic dystrophy type 1. Neurol. Clin. Pract. 8 (2018), 507–520, 10.1212/CPJ.0000000000000531.
Thomas, J.D., Oliveira, R., Sznajder, Ł.J., Swanson, M.S., Myotonic dystrophy and developmental regulation of RNA processing. Compr. Physiol. 8 (2018), 509–553, 10.1002/cphy.c170002.
Pinto, B.S., Saxena, T., Oliveira, R., Méndez-Gómez, H.R., Cleary, J.D., Denes, L.T., McConnell, O., Arboleda, J., Xia, G., Swanson, M.S., Wang, E.T., Impeding transcription of expanded microsatellite repeats by deactivated Cas9 impeding transcription of expanded microsatellite repeats by deactivated Cas9. Mol. Cell 68 (2017), 479–490.e5, 10.1016/j.molcel.2017.09.033.
Batra, R., Nelles, D.A., Pirie, E., Blue, S.M., Marina, R.J., Wang, H., Chaim, I.A., Thomas, J.D., Zhang, N., Nguyen, V., et al. Elimination of toxic microsatellite repeat expansion article elimination of toxic microsatellite repeat expansion RNA by RNA-targeting Cas9. Cell 170 (2017), 899–912.e10, 10.1016/j.cell.2017.07.010.
Thakore, P.I., D'Ippolito, A.M., Song, L., Safi, A., Shivakumar, N.K., Kabadi, A.M., Reddy, T.E., Crawford, G.E., Gersbach, C.A., Highly specific epigenome editing by CRISPR-Cas9 repressors for silencing of distal regulatory elements. Nat. Methods 12 (2015), 1143–1149, 10.1038/nmeth.3630.
Gilbert, L.A., Horlbeck, M.A., Adamson, B., Villalta, J.E., Chen, Y., Whitehead, E.H., Guimaraes, C., Panning, B., Ploegh, H.L., Bassik, M.C., et al. Genome-scale CRISPR-mediated control of gene repression and activation. Cell 159 (2014), 647–661, 10.1016/j.cell.2014.09.029.
Pickar-Oliver, A., Gersbach, C.A., The next generation of CRISPR–Cas technologies and applications. Nat. Rev. Mol. Cell Biol. 20 (2019), 490–507, 10.1038/s41580-019-0131-5.
Himeda, C.L., Jones, T.I., Jones, P.L., CRISPR/dCas9-mediated transcriptional inhibition ameliorates the epigenetic dysregulation at D4Z4 and represses DUX4-fl in FSH muscular dystrophy. Mol. Ther. 24 (2016), 527–535, 10.1038/mt.2015.200.
André, L.M., van Cruchten, R.T.P., Willemse, M., Bezstarosti, K., Demmers, J.A.A., van Agtmaal, E.L., Wansink, D.G., Wieringa, B., Recovery in the myogenic program of congenital myotonic dystrophy myoblasts after excision of the expanded (CTG)n repeat. Int. J. Mol. Sci., 20, 2019, 5685, 10.3390/ijms20225685.
Gudde, A.E., González-Barriga, A., van den Broek, W.J., Wieringa, B., Wansink, D.G., A low absolute number of expanded transcripts is involved in myotonic dystrophy type 1 manifestation in muscle. Hum. Mol. Genet. 25 (2016), 1648–1662, 10.1093/hmg/ddw042.
Furling, D., Lemieux, D., Taneja, K., Puymirat, J., Decreased levels of myotonic dystrophy protein kinase ( DMPK ) and delayed differentiation in human myotonic dystrophy myoblasts. Neuromuscul. Disord. 11 (2001), 728–735.
Arandel, L., Polay-Espinosa, M., Matloka, M., Bazinet, A., De Dea Diniz, D., Naouar, N., Rau, F., Jollet, A., Edom-Vovard, F., Mamchaoui, K., et al. Immortalized human myotonic dystrophy muscle cell lines to assess therapeutic compounds. Dis Model Mech, dmm., 2017, 027367, 10.1242/dmm.027367.
Jauvin, D., Chrétien, J., Pandey, S.K., Martineau, L., Revillod, L., Bassez, G., Lachon, A., MacLeod, A.R., Gourdon, G., Wheeler, T.M., et al. Targeting DMPK with antisense oligonucleotide improves muscle strength in myotonic dystrophy type 1 mice. Mol. Ther. Nucleic Acids 7 (2017), 465–474, 10.1016/j.omtn.2017.05.007.
Klinck, R., Fourrier, A., Thibault, P., Toutant, J., Durand, M., Lapointe, E., Caillet-Boudin, M.L., Sergeant, N., Gourdon, G., Meola, G., et al. RBFOX1 cooperates with MBNL1 to control splicing in muscle, including events altered in myotonic dystrophy type 1. PLoS One 9 (2014), 18–21, 10.1371/journal.pone.0107324.
Nakamori, M., Sobczak, K., Puwanant, A., Welle, S., Eichinger, K., Pandya, S., Dekdebrun, J., Heatwole, C.R., Mcdermott, M.P., Chen, T., et al. Splicing biomarkers of disease severity in myotonic dystrophy. Ann. Neurol. 74 (2013), 862–872, 10.1002/ana.23992.
Trapnell, C., Williams, B.A., Pertea, G., Mortazavi, A., Kwan, G., Van Baren, M.J., Salzberg, S.L., Wold, B.J., Pachter, L., Transcript assembly and quantification by RNA-Seq reveals unannotated transcripts and isoform switching during cell differentiation. Nat. Biotechnol. 28 (2010), 511–515, 10.1038/nbt.1621.
Zhang, H., Wen, J., Bigot, A., Chen, J., Shang, R., Mouly, V., Bi, P., Human Myotube Formation Is Determined by MyoD-Myomixer/Myomaker axis. Sci. Adv., 2020, 10.1126/sciadv.abc4062.
Spruston, N., Johnston, D., Perforated patch-clamp analysis of the passive membrane properties of three classes of hippocampal neurons. J. Neurophysiol. 67 (1992), 508–529, 10.1152/jn.1992.67.3.508.
François, V., Klein, A.F., Beley, C., Jollet, A., Lemercier, C., Garcia, L., Furling, D., Selective silencing of mutated mRNAs in DM1 by using modified hU7-snRNAs. Nat. Struct. Mol. Biol. 18 (2010), 85–87, 10.1038/nsmb.1958.
Klein, A.F., Varela, M.A., Arandel, L., Holland, A., Naouar, N., Arzumanov, A., Seoane, D., Revillod, L., Bassez, G., Ferry, A., et al. Peptide-conjugated oligonucleotides evoke long-lasting myotonic dystrophy correction in patient-derived cells and mice. J. Clin. Invest. 129 (2019), 4739–4744, 10.1172/JCI128205.
Arandel, L., Matloka, M., Klein, A.F., Rau, F., Sureau, A., Ney, M., Cordier, A., Kondili, M., Polay-Espinoza, M., Naouar, N., et al. Reversal of RNA toxicity in myotonic dystrophy via a decoy RNA-binding protein with high affinity for expanded CUG repeats. Nat. Biomed. Eng. 6 (2022), 207–220, 10.1038/s41551-021-00838-2.
Lo Scrudato, M., Poulard, K., Sourd, C., Tomé, S., Klein, A.F., Corre, G., Huguet, A., Furling, D., Gourdon, G., Buj-Bello, A., Genome editing of expanded CTG repeats within the human DMPK gene reduces nuclear RNA foci in the muscle of DM1 mice. Mol. Ther. 27 (2019), 1372–1388, 10.1016/j.ymthe.2019.05.021.
Benders, A.A., Timmermans, J.A., Oosterhof, A., Ter Laak, H.J., van Kuppevelt, T.H., Wevers, R.A., Veerkamp, J.H., Deficiency of Na+/K+-ATPase and sarcoplasmic reticulum Ca2+-ATPase in skeletal muscle and cultured muscle cells of myotonic dystrophy patients. Biochem. J. 293 (1993), 269–274, 10.1042/bj2930269.
Benders, A.A., Li, J., Lock, R.A., Bindels, R.J., Bonga, S.E., Veerkamp, J.H., Copper toxicity in cultured human skeletal muscle cells: the involvement of Na+/K+-ATPase and the Na+/Ca2+-exchanger. Pflügers Archiv 428 (1994), 461–467, 10.1007/BF00374566.
Behrens, M.I., Jalil, P., Serani, A., Vergara, F., Alvarez, O., Possible role of apamin-sensitive K+ channels in myotonic dystrophy. Muscle Nerve 17 (1994), 1264–1270.
Tajhya, R.B., Hu, X., Tanner, M.R., Huq, R., Kongchan, N., Neilson, J.R., Rodney, G.G., Horrigan, F.T., Timchenko, L.T., Beeton, C., Functional KCa1.1 channels are crucial for regulating the proliferation, migration and differentiation of human primary skeletal myoblasts. Cell Death Dis. 7 (2016), 1–12, 10.1038/cddis.2016.324.
Nurowska, E., Constanti, A., Dworakowska, B., Mouly, V., Furling, D., Lorenzon, P., Pietrangelo, T., Dołowy, K., Ruzzier, F., Potassium currents in human myogenic cells from healthy and congenital myotonic dystrophy foetuses. Cell. Mol. Biol. Lett. 14 (2009), 336–346, 10.2478/s11658-009-0006-4.
Franke, C., Hatt, H., Iaizzo, P.A., Lehmann-Horn, F., Characteristics of Na+ channels and Cl- conductance in resealed muscle fibre segments from patients with myotonic dystrophy. J. Physiol. 425 (1990), 391–405.
Waryah, C.B., Moses, C., Arooj, M., Blancafort, P., Zinc fingers, TALEs, and CRISPR systems: a comparison of tools for epigenome editing. Epigenome Editing, 2018, Methods in Molecular Biology, 19–64.
Wegmann, S., DeVos, S.L., Zeitler, B., Marlen, K., Bennett, R.E., Perez-Rando, M., MacKenzie, D., Yu, Q., Commins, C., Bannon, R.N., et al. Persistent Repression of Tau in the Brain Using Engineered Zinc Finger Protein Transcription Factors. 2021. Sci. Adv., 2021, 10.1126/sciadv.abe1611.
Kim, D., Luk, K., Wolfe, S.A., Kim, J.-S., Evaluating and enhancing target specificity of gene-editing nucleases and deaminases. Annu. Rev. Biochem. 88 (2019), 191–220, 10.1146/annurev-biochem-013118-111730.
Leroux, A.E., Schulze, J.O., Biondi, R.M., AGC kinases, mechanisms of regulation and innovative drug development. Semin. Cancer Biol. 48 (2018), 1–17, 10.1016/j.semcancer.2017.05.011.
Jansen, G., Groenen, P.J., Bächner, D., Jap, P.H., Coerwinkel, M., Oerlemans, F., Van Den Broek, W., Gohlsch, B., Pette, D., Plomp, J.J., et al. Abnormal myotonic dystrophy protein kinase levels produce only mild myopathy in mice. Nat. Genet. 13 (1996), 316–324, 10.1038/ng0796-316.
Reddy, S., Smith, D.B., Rich, M.M., Leferovich, J.M., Reilly, P., Davis, B.M., Tran, K., Rayburn, H., Bronson, R., Cros, D., et al. Mice lacking the myotonic dystrophy protein kinase develop a late onset progressive myopathy. Nat. Genet. 13 (1996), 325–335, 10.1038/ng0796-325.
Berul, C.I., Maguire, C.T., Gehrmann, J., Reddy, S., Progressive atrioventricular conduction block in a mouse myotonic dystrophy model. J. Intervent. Card Electrophysiol. 4 (2000), 351–358, 10.1023/A:1009842114968.
Berul, C.I., Maguire, C.T., Aronovitz, M.J., Greenwood, J., Miller, C., Gehrmann, J., Housman, D., Mendelsohn, M.E., Reddy, S., DMPK dosage alterations result in atrioventricular conduction abnormalities in a mouse myotonic dystrophy model. J. Clin. Invest. 103 (1999), R1–R7, 10.1172/JCI5346.
Carrell, S.T., Carrell, E.M., Auerbach, D., Pandey, S.K., Bennett, C.F., Dirksen, R.T., Thornton, C.A., Dmpk gene deletion or antisense knockdown does not compromise cardiac or skeletal muscle function in mice. Hum. Mol. Genet. 25 (2016), 4328–4338, 10.1093/hmg/ddw266.
Pandey, S.K., Wheeler, T.M., Justice, S.L., Kim, A., Younis, H.S., Gattis, D., Jauvin, D., Puymirat, J., Swayze, E.E., Freier, S.M., et al. Identification and characterization of modified antisense oligonucleotides targeting DMPK in mice and nonhuman primates for the treatment of myotonic dystrophy type 1. J. Pharmacol. Exp. Therapeut. 355 (2015), 329–340.
Thornton, C.A., Wang, E., Carrell, E.M., Myotonic dystrophy: approach to therapy. Curr. Opin. Genet. Dev. 44 (2017), 135–140, 10.1016/j.gde.2017.03.007.
Angelbello, A.J., Rzuczek, S.G., Mckee, K.K., Chen, J.L., Olafson, H., Cameron, M.D., Moss, W.N., Wang, E.T., Disney, M.D., Precise small-molecule cleavage of an r ( CUG ) repeat expansion in a myotonic dystrophy mouse model. Proc. Natl. Acad. Sci. USA 116 (2019), 7799–7804, 10.1073/pnas.1901484116.
Tomé, S., Gourdon, G., DM1 phenotype variability and triplet repeat instability: challenges in the development of new therapies. Int. J. Mol. Sci., 21, 2020, 457, 10.3390/ijms21020457.
Zu, T., Gibbens, B., Doty, N.S., Gomes-Pereira, M., Huguet, A., Stone, M.D., Margolis, J., Peterson, M., Markowski, T.W., Ingram, M.A.C., et al. Non-ATG-initiated translation directed by microsatellite expansions. Proc. Natl. Acad. Sci. USA 108 (2011), 260–265, 10.1073/pnas.1013343108.
Colella, P., Ronzitti, G., Mingozzi, F., Emerging issues in AAV-mediated in vivo gene therapy. Mol. Ther. Methods Clin. Dev. 8 (2018), 87–104, 10.1016/j.omtm.2017.11.007.
Thorley, M., Duguez, S., Mazza, E.M.C., Valsoni, S., Bigot, A., Mamchaoui, K., Harmon, B., Voit, T., Mouly, V., Duddy, W., et al. Skeletal muscle characteristics are preserved in hTERT/cdk4 human myogenic cell lines. Skeletal Muscle 6 (2016), 1–12, 10.1186/s13395-016-0115-5.
Mamchaoui, K., Trollet, C., Bigot, A., Negroni, E., Chaouch, S., Wolff, A., Kandalla, P.K., Marie, S., Di Santo, J., St Guily, J.L., et al. Immortalized pathological human myoblasts : towards a universal tool for the study of neuromuscular disorders. Skeletal Muscle, 1, 2011, 34.
Bigot, A., Klein, A.F., Gasnier, E., Jacquemin, V., Ravassard, P., Butler-Browne, G., Mouly, V., Furling, D., Large CTG repeats trigger p16-dependent premature senescence in myotonic dystrophy type 1 muscle precursor cells. Am. J. Pathol. 174 (2009), 1435–1442, 10.2353/ajpath.2009.080560.
Radzisheuskaya, A., Shlyueva, D., Müller, I., Helin, K., Optimizing sgRNA position markedly improves the efficiency of CRISPR/dCas9-mediated transcriptional repression. Nucleic Acids Res. 44 (2016), 1–13, 10.1093/nar/gkw583.
Horlbeck, M.A., Gilbert, L.A., Villalta, J.E., Adamson, B., Pak, R.A., Chen, Y., Fields, A.P., Park, C.Y., Corn, J.E., Kampmann, M., Weissman, J.S., Compact and highly active next-generation libraries for CRISPR-mediated gene repression and activation. Elife 5 (2016), 1197600–1197620, 10.7554/eLife.19760.
Mandegar, M.A., Huebsch, N., Frolov, E.B., Shin, E., Truong, A., Olvera, M.P., Chan, A.H., Miyaoka, Y., Holmes, K., Spencer, C.I., et al. CRISPR interference efficiently induces specific and reversible gene silencing in human iPSCs. Cell Stem Cell 18 (2016), 541–553, 10.1016/j.stem.2016.01.022.
Heigwer, F., Kerr, G., Boutros, M., E-CRISP: fast CRISPR target site identification. Nat. Methods 11 (2014), 122–123, 10.1038/nmeth.2812.
Liu, H., Wei, Z., Dominguez, A., Li, Y., Wang, X., Qi, L.S., CRISPR-ERA: a comprehensive design tool for CRISPR-mediated gene editing, repression and activation. Bioinformatics 31 (2015), 3676–3678, 10.1093/bioinformatics/btv423.
Labun, K., Montague, T.G., Gagnon, J.A., Thyme, S.B., Valen, E., CHOPCHOP v2: a web tool for the next generation of CRISPR genome engineering. Nucleic Acids Res. 44 (2016), W272–W276, 10.1093/nar/gkw398.
Stemmer, M., Thumberger, T., Del Sol Keyer, M., Wittbrodt, J., Mateo, J.L., CCTop: an intuitive, flexible and reliable CRISPR/Cas9 target prediction tool. PLoS One 10 (2015), 1–11, 10.1371/journal.pone.0124633.
Doench, J.G., Fusi, N., Sullender, M., Hegde, M., Vaimberg, E.W., Donovan, K.F., Smith, I., Tothova, Z., Wilen, C., Orchard, R., et al. Optimized sgRNA design to maximize activity and minimize off-target effects of CRISPR-Cas9. Nat. Biotechnol. 34 (2016), 184–191, 10.1038/nbt.3437.
Sanson, K.R., Hanna, R.E., Hegde, M., Donovan, K.F., Strand, C., Sullender, M.E., Vaimberg, E.W., Goodale, A., Root, D.E., Piccioni, F., Doench, J.G., Optimized libraries for CRISPR-Cas9 genetic screens with multiple modalities. Nat. Commun. 9 (2018), 5416–5515, 10.1038/s41467-018-07901-8.
Sanjana, N.E., Shalem, O., Zhang, F., Improved vectors and genome-wide libraries for CRISPR screening. Nat. Methods 11 (2014), 783–784, 10.1038/nmeth.3047.
Emi, N., Friedmann, T., Yee, J.K., Pseudotype formation of murine leukemia virus with the G protein of vesicular stomatitis virus. J. Virol. 65 (1991), 1202–1207, 10.1128/jvi.65.3.1202-1207.1991.
Klein, A.F., Arandel, L., Marie, J., Furling, D., FISH protocol for myotonic dystrophy type 1 cells. Methods Mol. Biol. 2056 (2020), 203–215, 10.1007/978-1-4939-9784-8_13.
Otsu, N., A threshold selection method from gray-level histograms. IEEE Trans. Syst. Man Cybern. 9 (1979), 62–66.
Chujo, T., Yamazaki, T., Kawaguchi, T., Kurosaka, S., Takumi, T., Nakagawa, S., Hirose, T., Unusual semi-extractability as a hallmark of nuclear body-associated architectural noncoding RNAs. EMBO J. 36 (2017), 1447–1462, 10.15252/embj.201695848.
Nutter, C.A., Bubenik, J.L., Oliveira, R., Ivankovic, F., Sznajder, Ł.J., Kidd, B.M., Pinto, B.S., Otero, B.A., Carter, H.A., Vitriol, E.A., et al. Cell-type-specific dysregulation of RNA alternative splicing in short tandem repeat mouse knockin models of myotonic dystrophy. Genes Dev. 33 (2019), 1635–1640, 10.1101/gad.328963.119.
Poitras, E., Houde, A., La PCR en temps réel: principes et applications. Reviews in Biology and Biotechnology 2 (2002), 2–11.
Stern-Straeter, J., Bonaterra, G.A., Hörmann, K., Kinscherf, R., Goessler, U.R., Identification of valid reference genes during the differentiation of human myoblasts. BMC Mol. Biol. 10 (2009), 1–9, 10.1186/1471-2199-10-66.
Hubaux, R., Vandermeers, F., Cosse, J.P., Crisanti, C., Kapoor, V., Albelda, S.M., Mascaux, C., Delvenne, P., Hubert, P., Willems, L., Valproic acid improves second-line regimen of small cell lung carcinoma in preclinical models. ERJ Open Res 1 (2015), 00028–02015.
Eriksson, M., Real-time RT-PCR for CTG repeat-containing genes. Methods Mol. Biol. 277 (2004), 77–84, 10.1385/1-59259-804-8:077.
Martin, M., Cutadapt removes adapter sequences from high-throughput sequencing reads. EMBnet 17 (2011), 10–12.
Anders, S., Pyl, P.T., Huber, W., HTSeq-A Python framework to work with high-throughput sequencing data. Bioinformatics 31 (2015), 166–169, 10.1093/bioinformatics/btu638.
Shen, S., Park, J.W., Lu, Z.X., Lin, L., Henry, M.D., Wu, Y.N., Zhou, Q., Xing, Y., rMATS: robust and flexible detection of differential alternative splicing from replicate RNA-Seq data. Proc. Natl. Acad. Sci. USA 111 (2014), E5593–E5601, 10.1073/pnas.1419161111.
Love, M.I., Huber, W., Anders, S., Moderated estimation of fold change and dispersion for RNA-seq data with DESeq2. Genome Biol. 15 (2014), 550–621, 10.1186/s13059-014-0550-8.
Babicki, S., Arndt, D., Marcu, A., Liang, Y., Grant, J.R., Maciejewski, A., Wishart, D.S., Heatmapper: web-enabled heat mapping for all. Nucleic Acids Res. 44 (2016), W147–W153, 10.1093/NAR/GKW419.
Liao, Y., Wang, J., Jaehnig, E.J., Shi, Z., Zhang, B., WebGestalt 2019: gene set analysis toolkit with revamped UIs and APIs. Nucleic Acids Res. 47 (2019), W199–W205, 10.1093/nar/gkz401.
Aken, B.L., Ayling, S., Barrell, D., Clarke, L., Curwen, V., Fairley, S., Fernandez Banet, J., Billis, K., García Girón, C., Hourlier, T., et al. The Ensembl gene annotation system. Database 2016 (2016), 1–19, 10.1093/database/baw093.
Liao, Y., Smyth, G.K., Shi, W., FeatureCounts: an efficient general purpose program for assigning sequence reads to genomic features. Bioinformatics 30 (2014), 923–930, 10.1093/bioinformatics/btt656.
Noble, W.S., How does multiple testing correction work?. Nat. Biotechnol. 27 (2009), 1135–1137, 10.1038/nbt1209-1135.
