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J. Clin. Invest., Dec 17 2024, e183353 | https://doi.org/10.1172/JCI183353
Regulation of sarcomere formation and function in the healthy heart requires a titin intronic enhancer
Yuri Kim1,2,^,*, Seong Won Kim1,^, David Saul2,^, Meraj Neyazi1,3,^, Manuel Schmid1,4,^, Hiroko Wakimoto1,^, Neil Slaven5, Joshua H. Lee6, Olivia Layton1, Lauren K. Wasson1, Justin H. Letendre6, Feng Xiao7, Jourdan K. Ewoldt6, Konstantinos Gkatzis8, Peter Sommer8, BénédicteGobert8, Nicolas Wiest-Daesslé8, Quentin McAfee9, Nandita Singhal2, Mingyue Lun1, Joshua M. Gorham1, Zolt Arany9, Arun Sharma10, Christopher N. Toepfer11,12, Gavin Y. Oudit13,14, William T. Pu7,15, Diane E. Dickel5,16, Len A. Pennacchio5,17,18, Axel Visel5,17,19, Christopher S. Chen6, J. G. Seidman1,#, Christine E. Seidman1,2,20,#,*
1Department of Genetics, Harvard Medical School, Boston, MA, USA
2Division of Cardiovascular Medicine, Brigham and Women’s Hospital, Boston, MA, USA
3Department of Cardiology, University Heart and Vascular Center Hamburg, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
4German Heart Center, Technical University of Munich, Munich, Germany
5Environmental Genomics and Systems Biology Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
6Department of Biomedical Engineering, Boston University, Boston, MA, USA
7Department of Cardiology, Boston Children's Hospital, Harvard Medical School, Boston, MA, USA
8Ksilink, 67000, Strasbourg, France
9Cardiovascular Institute, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
10Smidt Heart Institute, Cedars-Sinai Medical Center, Los Angeles, CA, USA
11Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, UK
12Wellcome Centre for Human Genetics, University of Oxford, Oxford, UK
13Department of Medicine, University of Alberta, Edmonton, Alberta, Canada
14Mazankowski Alberta Heart Institute, Edmonton, Alberta, Canada
15Harvard Stem Cell Institute, Cambridge, MA, USA
16Currently affiliated with Octant Inc., Emeryville, CA, USA
17U.S. Department of Energy Joint Genome Institute, One Cyclotron Road, Berkeley, CA, USA
18Comparative Biochemistry Program, University of California, Berkeley, CA, USA
19School of Natural Sciences, University of California, Merced, CA, USA
20Howard Hughes Medical Institute, Chevy Chase, MD, USA
^,#These authors contributed equally
*Corresponding authors
1Department of Genetics, Harvard Medical School, Boston, MA, USA
2Division of Cardiovascular Medicine, Brigham and Women’s Hospital, Boston, MA, USA
3Department of Cardiology, University Heart and Vascular Center Hamburg, University Medical Center Hamburg-Eppendorf, Hamburg, Germany
4German Heart Center, Technical University of Munich, Munich, Germany
5Environmental Genomics and Systems Biology Division, Lawrence Berkeley National Laboratory, Berkeley, CA, USA
6Department of Biomedical Engineering, Boston University, Boston, MA, USA
7Department of Cardiology, Boston Children's Hospital, Harvard Medical School, Boston, MA, USA
8Ksilink, 67000, Strasbourg, France
9Cardiovascular Institute, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA.
10Smidt Heart Institute, Cedars-Sinai Medical Center, Los Angeles, CA, USA
11Division of Cardiovascular Medicine, Radcliffe Department of Medicine, University of Oxford, Oxford, UK
12Wellcome Centre for Human Genetics, University of Oxford, Oxford, UK
13Department of Medicine, University of Alberta, Edmonton, Alberta, Canada
14Mazankowski Alberta Heart Institute, Edmonton, Alberta, Canada
15Harvard Stem Cell Institute, Cambridge, MA, USA
16Currently affiliated with Octant Inc., Emeryville, CA, USA
17U.S. Department of Energy Joint Genome Institute, One Cyclotron Road, Berkeley, CA, USA
18Comparative Biochemistry Program, University of California, Berkeley, CA, USA
19School of Natural Sciences, University of California, Merced, CA, USA
20Howard Hughes Medical Institute, Chevy Chase, MD, USA
^,#These authors contributed equally
*Corresponding authors
Abstract
Heterozygous truncating variants in the sarcomere protein titin (TTN) are the most common genetic cause of heart failure. To understand mechanisms that regulate abundant cardiomyocyte TTN expression we characterized highly conserved intron 1 sequences that exhibited dynamic changes in chromatin accessibility during differentiation of human cardiomyocytes from induced pluripotent stem cells (hiPSC-CMs). Homozygous deletion of these sequences in mice caused embryonic lethality while heterozygous mice demonstrated allele-specific reduction in Ttn expression. A 296 bp fragment of this element, denoted E1, was sufficient to drive expression of a reporter gene in hiPSC-CMs. Deletion of E1 downregulated TTN expression, impaired sarcomerogenesis, and decreased contractility in hiPSC-CMs. Site-directed mutagenesis of predicted NKX2-5- and MEF2-binding sites within E1 abolished its transcriptional activity. Embryonic mice expressing E1 reporter gene constructs validated in vivo cardiac-specific activity of E1 and the requirement for NKX2-5 and MEF2 binding sequences. Moreover, isogenic hiPSC-CMs containing a rare E1 variant in the predicted MEF2 binding motif that was identified in a patient with unexplained DCM showed reduced TTN expression. Together these discoveries define an essential, functional enhancer that regulates TTN expression. Manipulation of this element may advance therapeutic strategies to treat DCM caused by TTN haploinsufficiency.
논문정보
- Research article
- 2024년 12월 (BRIC 등록일 2024-12-23)
- 국외 연구진
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