Induced cardiomyocyte maturation: Cardiac transcription factors are necessary but not sufficient
Autoři:
Sophie Dal-Pra aff001; Conrad P. Hodgkinson aff001; Victor J. Dzau aff001
Působiště autorů:
Mandel Center for Hypertension and Atherosclerosis Research, Duke University, Durham, North Carolina, United States of America
aff001; Cardiovascular Research Center, Duke University Medical Center, Durham, North Carolina, United States of America
aff002
Vyšlo v časopise:
PLoS ONE 14(10)
Kategorie:
Research Article
prolekare.web.journal.doi_sk:
https://doi.org/10.1371/journal.pone.0223842
Souhrn
The process by which fibroblasts are directly reprogrammed into cardiomyocytes involves two stages; initiation and maturation. Initiation represents the initial expression of factors that induce fibroblasts to transdifferentiate into cardiomyocytes. Following initiation, the cell undergoes a period of maturation before becoming a mature cardiomyocyte. We wanted to understand the role of cardiac development transcription factors in the maturation process. We directly reprogram fibroblasts into cardiomyocytes by a combination of miRNAs (miR combo). The ability of miR combo to induce cardiomyocyte-specific genes in fibroblasts was lost following the knockdown of the cardiac transcription factors Gata4, Mef2C, Tbx5 and Hand2 (GMTH). To further clarify the role of GMTH in miR combo reprogramming we utilized a modified CRISPR-Cas9 approach to activate endogenous GMTH genes. Importantly, both miR combo and the modified CRISPR-Cas9 approach induced comparable levels of GMTH expression. While miR combo was able to reprogram fibroblasts into cardiomyocyte-like cells, the modified CRISPR-Cas9 approach could not. Indeed, we found that cardiomyocyte maturation only occurred with very high levels of GMT expression. Taken together, our data indicates that while endogenous cardiac transcription factors are insufficient to reprogram fibroblasts into mature cardiomyocytes, endogenous cardiac transcription factors are necessary for expression of maturation genes.
Klíčová slova:
Small interfering RNAs – Transcription factors – Transfection – MicroRNAs – DNA transcription – Fibroblasts – Guide RNA
Zdroje
1. Chan SS, Shi X, Toyama A, Arpke RW, Dandapat A, Iacovino M, et al. Mesp1 patterns mesoderm into cardiac, hematopoietic, or skeletal myogenic progenitors in a context-dependent manner. Cell Stem Cell. 2013;12(5):587–601. doi: 10.1016/j.stem.2013.03.004 23642367; PubMed Central PMCID: PMC3646300.
2. Bondue A, Lapouge G, Paulissen C, Semeraro C, Iacovino M, Kyba M, et al. Mesp1 acts as a master regulator of multipotent cardiovascular progenitor specification. Cell Stem Cell. 2008;3(1):69–84. doi: 10.1016/j.stem.2008.06.009 18593560.
3. Klattenhoff CA, Scheuermann JC, Surface LE, Bradley RK, Fields PA, Steinhauser ML, et al. Braveheart, a long noncoding RNA required for cardiovascular lineage commitment. Cell. 2013;152(3):570–83. doi: 10.1016/j.cell.2013.01.003 23352431; PubMed Central PMCID: PMC3563769.
4. Kelly RG. How Mesp1 makes a move. J Cell Biol. 2016;213(4):411–3. doi: 10.1083/jcb.201604121 27185831; PubMed Central PMCID: PMC4878095.
5. Chiapparo G, Lin X, Lescroart F, Chabab S, Paulissen C, Pitisci L, et al. Mesp1 controls the speed, polarity, and directionality of cardiovascular progenitor migration. J Cell Biol. 2016;213(4):463–77. doi: 10.1083/jcb.201505082 27185833; PubMed Central PMCID: PMC4878090.
6. Lindsley RC, Gill JG, Murphy TL, Langer EM, Cai M, Mashayekhi M, et al. Mesp1 coordinately regulates cardiovascular fate restriction and epithelial-mesenchymal transition in differentiating ESCs. Cell Stem Cell. 2008;3(1):55–68. doi: 10.1016/j.stem.2008.04.004 18593559; PubMed Central PMCID: PMC2497439.
7. Yang X, Pabon L, Murry CE. Engineering adolescence: maturation of human pluripotent stem cell-derived cardiomyocytes. Circ Res. 2014;114(3):511–23. doi: 10.1161/CIRCRESAHA.114.300558 24481842; PubMed Central PMCID: PMC3955370.
8. Lassar AB, Paterson BM, Weintraub H. Transfection of a DNA locus that mediates the conversion of 10T1/2 fibroblasts to myoblasts. Cell. 1986;47(5):649–56. doi: 10.1016/0092-8674(86)90507-6 2430720.
9. Weintraub H, Tapscott SJ, Davis RL, Thayer MJ, Adam MA, Lassar AB, et al. Activation of muscle-specific genes in pigment, nerve, fat, liver, and fibroblast cell lines by forced expression of MyoD. Proc Natl Acad Sci U S A. 1989;86(14):5434–8. doi: 10.1073/pnas.86.14.5434 2748593; PubMed Central PMCID: PMC297637.
10. Qian L, Huang Y, Spencer CI, Foley A, Vedantham V, Liu L, et al. In vivo reprogramming of murine cardiac fibroblasts into induced cardiomyocytes. Nature. 2012;485(7400):593–8. doi: 10.1038/nature11044 22522929; PubMed Central PMCID: PMC3369107.
11. Song K, Nam YJ, Luo X, Qi X, Tan W, Huang GN, et al. Heart repair by reprogramming non-myocytes with cardiac transcription factors. Nature. 2012;485(7400):599–604. doi: 10.1038/nature11139 22660318; PubMed Central PMCID: PMC3367390.
12. Ieda M, Fu JD, Delgado-Olguin P, Vedantham V, Hayashi Y, Bruneau BG, et al. Direct reprogramming of fibroblasts into functional cardiomyocytes by defined factors. Cell. 2010;142(3):375–86. doi: 10.1016/j.cell.2010.07.002 20691899; PubMed Central PMCID: PMC2919844.
13. Vacchi-Suzzi C, Hahne F, Scheubel P, Marcellin M, Dubost V, Westphal M, et al. Heart structure-specific transcriptomic atlas reveals conserved microRNA-mRNA interactions. PLoS One. 2013;8(1):e52442. doi: 10.1371/journal.pone.0052442 23300973; PubMed Central PMCID: PMC3534709.
14. Jayawardena TM, Egemnazarov B, Finch EA, Zhang L, Payne JA, Pandya K, et al. MicroRNA-mediated in vitro and in vivo direct reprogramming of cardiac fibroblasts to cardiomyocytes. Circ Res. 2012;110(11):1465–73. doi: 10.1161/CIRCRESAHA.112.269035 22539765; PubMed Central PMCID: PMC3380624.
15. Wang X, Hodgkinson CP, Lu K, Payne AJ, Pratt RE, Dzau VJ. Selenium Augments microRNA Directed Reprogramming of Fibroblasts to Cardiomyocytes via Nanog. Sci Rep. 2016;6:23017. doi: 10.1038/srep23017 26975336; PubMed Central PMCID: PMC4792153.
16. Li Y, Dal-Pra S, Mirotsou M, Jayawardena TM, Hodgkinson CP, Bursac N, et al. Tissue-engineered 3-dimensional (3D) microenvironment enhances the direct reprogramming of fibroblasts into cardiomyocytes by microRNAs. Sci Rep. 2016;6:38815. doi: 10.1038/srep38815 27941896; PubMed Central PMCID: PMC5150639.
17. Dal-Pra S, Hodgkinson CP, Mirotsou M, Kirste I, Dzau VJ. Demethylation of H3K27 Is Essential for the Induction of Direct Cardiac Reprogramming by miR Combo. Circ Res. 2017;120(9):1403–13. doi: 10.1161/CIRCRESAHA.116.308741 28209718; PubMed Central PMCID: PMC5409871.
18. Hodgkinson CP, Gomez JA, Baksh SS, Payne A, Schmeckpeper J, Pratt RE, et al. Insights from molecular signature of in vivo cardiac c-Kit(+) cells following cardiac injury and beta-catenin inhibition. J Mol Cell Cardiol. 2018;123:64–74. doi: 10.1016/j.yjmcc.2018.08.024 30171847; PubMed Central PMCID: PMC6192834.
19. Jayawardena TM, Finch EA, Zhang L, Zhang H, Hodgkinson CP, Pratt RE, et al. MicroRNA induced cardiac reprogramming in vivo: evidence for mature cardiac myocytes and improved cardiac function. Circ Res. 2015;116(3):418–24. doi: 10.1161/CIRCRESAHA.116.304510 25351576; PubMed Central PMCID: PMC4312531.
20. Liu Z, Chen O, Zheng M, Wang L, Zhou Y, Yin C, et al. Re-patterning of H3K27me3, H3K4me3 and DNA methylation during fibroblast conversion into induced cardiomyocytes. Stem Cell Res. 2016;16(2):507–18. doi: 10.1016/j.scr.2016.02.037 26957038; PubMed Central PMCID: PMC4828257.
21. Furtado MB, Costa MW, Pranoto EA, Salimova E, Pinto AR, Lam NT, et al. Cardiogenic genes expressed in cardiac fibroblasts contribute to heart development and repair. Circ Res. 2014;114(9):1422–34. doi: 10.1161/CIRCRESAHA.114.302530 24650916; PubMed Central PMCID: PMC4083003.
22. Jayawardena T, Mirotsou M, Dzau VJ. Direct reprogramming of cardiac fibroblasts to cardiomyocytes using microRNAs. Methods Mol Biol. 2014;1150:263–72. doi: 10.1007/978-1-4939-0512-6_18 24744005.
23. Chavez A, Scheiman J, Vora S, Pruitt BW, Tuttle M, E PRI, et al. Highly efficient Cas9-mediated transcriptional programming. Nat Methods. 2015;12(4):326–8. doi: 10.1038/nmeth.3312 25730490; PubMed Central PMCID: PMC4393883.
24. Perez-Pinera P, Kocak DD, Vockley CM, Adler AF, Kabadi AM, Polstein LR, et al. RNA-guided gene activation by CRISPR-Cas9-based transcription factors. Nat Methods. 2013;10(10):973–6. doi: 10.1038/nmeth.2600 23892895; PubMed Central PMCID: PMC3911785.
25. Horlbeck MA, Gilbert LA, Villalta JE, Adamson B, Pak RA, Chen Y, et al. Compact and highly active next-generation libraries for CRISPR-mediated gene repression and activation. Elife. 2016;5. doi: 10.7554/eLife.19760 27661255; PubMed Central PMCID: PMC5094855.
26. Mohamed TM, Stone NR, Berry EC, Radzinsky E, Huang Y, Pratt K, et al. Chemical Enhancement of In Vitro and In Vivo Direct Cardiac Reprogramming. Circulation. 2017;135(10):978–95. doi: 10.1161/CIRCULATIONAHA.116.024692 27834668; PubMed Central PMCID: PMC5340593.
27. Zhou H, Morales MG, Hashimoto H, Dickson ME, Song K, Ye W, et al. ZNF281 enhances cardiac reprogramming by modulating cardiac and inflammatory gene expression. Genes Dev. 2017;31(17):1770–83. doi: 10.1101/gad.305482.117 28982760; PubMed Central PMCID: PMC5666675.
28. Addis RC, Ifkovits JL, Pinto F, Kellam LD, Esteso P, Rentschler S, et al. Optimization of direct fibroblast reprogramming to cardiomyocytes using calcium activity as a functional measure of success. J Mol Cell Cardiol. 2013;60:97–106. doi: 10.1016/j.yjmcc.2013.04.004 23591016; PubMed Central PMCID: PMC3679282.
29. Hodgkinson CP, Dzau VJ. Conserved microRNA program as key to mammalian cardiac regeneration: insights from zebrafish. Circ Res. 2015;116(7):1109–11. doi: 10.1161/CIRCRESAHA.115.305852 25814680.
30. Black JB, Adler AF, Wang HG, D'Ippolito AM, Hutchinson HA, Reddy TE, et al. Targeted Epigenetic Remodeling of Endogenous Loci by CRISPR/Cas9-Based Transcriptional Activators Directly Converts Fibroblasts to Neuronal Cells. Cell Stem Cell. 2016;19(3):406–14. doi: 10.1016/j.stem.2016.07.001 27524438; PubMed Central PMCID: PMC5010447.
31. Wang L, Liu Z, Yin C, Asfour H, Chen O, Li Y, et al. Stoichiometry of Gata4, Mef2c, and Tbx5 influences the efficiency and quality of induced cardiac myocyte reprogramming. Circ Res. 2015;116(2):237–44. doi: 10.1161/CIRCRESAHA.116.305547 25416133; PubMed Central PMCID: PMC4299697.
32. Pikkarainen S, Tokola H, Kerkela R, Ruskoaho H. GATA transcription factors in the developing and adult heart. Cardiovasc Res. 2004;63(2):196–207. doi: 10.1016/j.cardiores.2004.03.025 15249177.
33. Srivastava D, Thomas T, Lin Q, Kirby ML, Brown D, Olson EN. Regulation of cardiac mesodermal and neural crest development by the bHLH transcription factor, dHAND. Nat Genet. 1997;16(2):154–60. doi: 10.1038/ng0697-154 9171826.
34. Horb ME, Thomsen GH. Tbx5 is essential for heart development. Development. 1999;126(8):1739–51. 10079235.
35. Lin Q, Schwarz J, Bucana C, Olson EN. Control of mouse cardiac morphogenesis and myogenesis by transcription factor MEF2C. Science. 1997;276(5317):1404–7. doi: 10.1126/science.276.5317.1404 9162005; PubMed Central PMCID: PMC4437729.
36. Takeuchi JK, Bruneau BG. Directed transdifferentiation of mouse mesoderm to heart tissue by defined factors. Nature. 2009;459(7247):708–11. doi: 10.1038/nature08039 19396158; PubMed Central PMCID: PMC2728356.
37. Sepulveda JL, Vlahopoulos S, Iyer D, Belaguli N, Schwartz RJ. Combinatorial expression of GATA4, Nkx2-5, and serum response factor directs early cardiac gene activity. J Biol Chem. 2002;277(28):25775–82. doi: 10.1074/jbc.M203122200 11983708.
38. Ow JR, Palanichamy Kala M, Rao VK, Choi MH, Bharathy N, Taneja R. G9a inhibits MEF2C activity to control sarcomere assembly. Sci Rep. 2016;6:34163. doi: 10.1038/srep34163 27667720; PubMed Central PMCID: PMC5036183.
39. Al-Maqtari T, Hong KU, Vajravelu BN, Moktar A, Cao P, Moore JBt, et al. Transcription factor-induced activation of cardiac gene expression in human c-kit+ cardiac progenitor cells. PLoS One. 2017;12(3):e0174242. doi: 10.1371/journal.pone.0174242 28355297; PubMed Central PMCID: PMC5371315.
40. Abad M, Hashimoto H, Zhou H, Morales MG, Chen B, Bassel-Duby R, et al. Notch Inhibition Enhances Cardiac Reprogramming by Increasing MEF2C Transcriptional Activity. Stem Cell Reports. 2017;8(3):548–60. doi: 10.1016/j.stemcr.2017.01.025 28262548; PubMed Central PMCID: PMC5355682.
41. Bai F, Ho Lim C, Jia J, Santostefano K, Simmons C, Kasahara H, et al. Directed Differentiation of Embryonic Stem Cells Into Cardiomyocytes by Bacterial Injection of Defined Transcription Factors. Sci Rep. 2015;5:15014. doi: 10.1038/srep15014 26449528; PubMed Central PMCID: PMC4598736.
42. Kathiriya IS, Nora EP, Bruneau BG. Investigating the transcriptional control of cardiovascular development. Circ Res. 2015;116(4):700–14. doi: 10.1161/CIRCRESAHA.116.302832 25677518; PubMed Central PMCID: PMC4332409.
43. Ratz M, Testa I, Hell SW, Jakobs S. CRISPR/Cas9-mediated endogenous protein tagging for RESOLFT super-resolution microscopy of living human cells. Sci Rep. 2015;5:9592. doi: 10.1038/srep09592 25892259; PubMed Central PMCID: PMC4402611.
44. Delfosse K, Wozny MR, Jaipargas EA, Barton KA, Anderson C, Mathur J. Fluorescent Protein Aided Insights on Plastids and their Extensions: A Critical Appraisal. Front Plant Sci. 2015;6:1253. doi: 10.3389/fpls.2015.01253 26834765; PubMed Central PMCID: PMC4719081.
45. Lisenbee CS, Karnik SK, Trelease RN. Overexpression and mislocalization of a tail-anchored GFP redefines the identity of peroxisomal ER. Traffic. 2003;4(7):491–501. 12795694.
46. Veerman CC, Kosmidis G, Mummery CL, Casini S, Verkerk AO, Bellin M. Immaturity of human stem-cell-derived cardiomyocytes in culture: fatal flaw or soluble problem? Stem Cells Dev. 2015;24(9):1035–52. doi: 10.1089/scd.2014.0533 25583389.
47. Bedada FB, Wheelwright M, Metzger JM. Maturation status of sarcomere structure and function in human iPSC-derived cardiac myocytes. Biochim Biophys Acta. 2016;1863(7 Pt B):1829–38. doi: 10.1016/j.bbamcr.2015.11.005 26578113; PubMed Central PMCID: PMC4864165.
48. Koivumaki JT, Naumenko N, Tuomainen T, Takalo J, Oksanen M, Puttonen KA, et al. Structural Immaturity of Human iPSC-Derived Cardiomyocytes: In Silico Investigation of Effects on Function and Disease Modeling. Front Physiol. 2018;9:80. doi: 10.3389/fphys.2018.00080 29467678; PubMed Central PMCID: PMC5808345.
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