From Adhesions to Z-discs: Regulation at the Cardiac Sarcomere by Distinct Paralogs of Alpha-Actinin and Myosin II

Abstract

Gene duplications have endowed the vertebrate heart with multiple gene paralogs encoding key cytoskeletal proteins, enabling precise regulation of cardiac muscle function. This dissertation explores how distinct paralogs of myosin II and alpha-actinin contribute to the assembly, contractility, and function of the cardiac sarcomere, the minimal contractile unit in cardiac muscle. Experimental evidence suggests α- and β-myosin II-associated heavy chains (encoded by MYH6 and MYH7) exhibit distinct, non-uniform distributions within cardiac sarcomeres in both human iPSC-derived cardiomyocytes (hiCMs) and in vivo models. Signal against β-myosin II heavy chain extended further from the sarcomere center than signal against α-myosin II heavy chains, suggesting specific biomechanical tasks are partitioned between each heavy chain during cardiac contraction-relaxation. Selective inhibition of β-myosin II disrupted sarcomere assembly in hiCMs while depletion of β-myosin II alone did not, suggesting functional redundancy during assembly between β-myosin II and α-myosin II. α- and β-myosin heavy chain ratio within a cardiac myocyte influenced β-myosin II inhibitor sensitivity, suggesting atrial and ventricular myocytes may be disparately impacted by pharmacological inhibition. Next, evidence is presented concerning distinct alpha-actinin proteins encoded by gene paralogs ACTN1 and ACTN4. In vitro assays support the essentiality of ACTN1 in anchoring early sarcomere precursors at cardiac myocyte focal adhesions, ensuring proper sarcomere assembly. By contrast, evidence suggests another closely-related “non-muscle” alpha-actinin, ACTN4, localizes to the mature cardiac Z-disc (the sarcomere border) and destabilizes sarcomeric actin cross-links. Depletion of ACTN4 increased sarcomere length, elevated contractile force, and triggered cellular hypertrophy in cardiac myocytes, plus contractility-driven biomechanical chamber remodeling in zebrafish embryos—each without classical markers of disease. Collectively, these findings underscore how gene duplications enable nuanced regulation of the heart’s contractile apparatus.