Titin A-band truncation in mice causes stress-induced dilated cardiomyopathy
Ida G. Lunde1,2,3, Hiroko Wakimoto1, Jianming Jiang1, Michael A. Burke1,4, Victor Soukoulis1, Andreas
Unger5, David Conner1, Joshua Gorham1, Steve DePalma1, Geir Christensen2,3, Wolfgang A. Linke5,
Jonathan G. Seidman1,*, Christine E. Seidman3,4,*
1Department
of Genetics, Harvard Medical School, Boston, MA, USA.
for Experimental Medical Research, Oslo University Hospital and University of Oslo, Norway.
3KG Jebsen Cardiac Research Center and Center for Heart Failure Research, University of Oslo,
Oslo, Norway.
4Division of Cardiology, Brigham and Women´s Hospital, Boston, MA, USA.
5Department of Cardiovascular Physiology, Ruhr University Bochum, Bochum, Germany.
* authors contributed equally
2Institute
20% of dilated cardiomyopathy (DCM) patients carry heterozygous truncating mutations in the giant
protein titin (TTNΔ). Titin spans the cardiomyocyte sarcomere and is important for assembly,
contraction, relaxation and signaling. Truncating mutations are overrepresented in A-band. We
assessed the cardiac phenotype of mice with titin A-band truncation (TTNΔA).
To generate TTNΔA mice we introduced lox-P sites flanking exons 276-277 and crossed with
EIIa-Cre mice, causing a premature stop codon in the A-band. Heterozygous intercrosses produced
no homozygous TTNΔA pups. Genotyping revealed homozygous embryos at E8.5-E10.5, with fetal
demise at E10.5. Heterozygous mice (age 6-80 weeks) were viable, fertile and not different from WT in
appearance, activity, or echocardiographic phenotype. TTNΔA hearts showed mutant and WT mRNA,
while blots detected no mutant protein. TTNΔA and WT mice were stressed for ten weeks by voluntary
cage-wheel running and two weeks of isoproterenol infusion, evoking no difference in
echocardiographic phenotypes. Compound TTNΔA/LMNA, TTNΔA/MYH6F764L and TTNΔA/PLNR9C
mutation mice showed no exacerbation of DCM compared to LMNA, MYH6F764L and PLNR9C mice,
respectively. TTNΔA mice receiving two weeks of ANGII infusion showed hypertrophy with
exacerbated diastolic dysfunction. Thoracic aortic constriction (TAC) exacerbated DCM in TTNΔA,
with increased left atrial diameter, lung weight and BNP. RNA sequencing of TAC TTNΔA showed
differential expression of 1465 transcripts. Pathway analyses implicated TGFβ as upstream regulator
and identified cardiotoxic categories hypertrophy, cell death, infarction, dilation and damage. Finally,
AAV9-mediated RNAi knock-down of TTN (70% down) in mice produced severe DCM, mortality and
disruption of sarcomere and tissue structure.
Transcripts encoding a titin A-band truncation are expressed but do not yield detectable
mutant protein. Homozygous mice are embryonic lethal, while heterozygous mice show DCM and
congestive heart failure upon pressure overload. The recapitulation of DCM in mice suggests that titin
A-band truncation causally underlie the late-onset DCM seen in patients.