Biophysical Society Thematic Meeting | Canterbury 2023

Towards a More Perfect Union: Multi-Scale Models of Muscle and Their Experimental Validation

Poster Abstracts

8-POS Board 8 TRUNCATED TITIN INCORPORATES INTO DCM PATIENT MYOCARDIAL SARCOMERE AND TRANSMITS FORCE ACROSS THE HALF-SARCOMERE Quentin McAfee 1 ; Matt Caporizzo 2 ; Keita Uchida 2 ; Kenneth C Bedi, Jr. 1 ; Kenneth B Margulies 1 ; Zolt Arany 1 ; Benjamin Prosser 2 ; 1 University of Pennsylvania, Cardiovascular Institute, Philadelphia, PA, USA 2 University of Pennsylvania, Department of Physiology, Philadelphia, PA, USA Truncated titin protein (TTNtv) has recently been shown by multiple groups to be present in the myocardium of patients with dilated cardiomyopathy caused by heterozygous truncations in TTN. However, it has remained unclear whether truncated titin incorporates into the myocardial sarcomere, possibly acting in a dominant negative fashion. To specifically detect truncated titin in the human TTNtv + DCM myocardium, we raised an antibody against a patient specific frameshift antigen, a proteome-unique sequence of amino acids appended to the C-terminus of a single patient’s A-band truncation site. By western blot, this antibody specifically detects truncated titin at the expected molecular weight. Staining skinned cardiomyocytes from this patient with this antibody yields twin stripes 240nm from the M line, a location predicted by the truncation site, demonstrating truncated titin binds to the thick filament. To determine whether truncated titin was able to integrate with the Z-disc and transmit force across the stretched sarcomere, we stretched skinned cardiomyocyte fragments from this patient that were labeled with this antibody. When stretched from normal through supraphysiological sarcomere lengths, truncated titin remained attached to the thick filament. To determine if truncated titin was attached to the Z-disc and capable of transmitting force across the half-sarcomere, we treated stretched sarcomeres bearing TTNtv with 400mM KCl to disrupt the thick filament. This caused truncated titin to detach from the thick filament and retract to the Z-disk, consistent with titin’s behavior as an entropic spring attached at one end to the Z-disk. These data demonstrate that truncated titin is incorporated into the myocardial sarcomere, and in binding to both the Z-disk and thick filament, transmits force across the half sarcomere. Broadly, these data support the hypothesis that truncated titin may act at the sarcomere in a dominant negative mechanism to promote the development of dilated cardiomyopathy.

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