The cardiomyopathy-associated K15N mutation in tropomyosin alters actin filament pointed end dynamics

Abstract

Correct assembly of thin filaments composed of actin and actin-binding proteins is of crucial importance for properly functioning muscle cells. Tropomyosin (Tpm) mediates the binding of tropomodulin (Tmod) and leiomodin (Lmod) at the slow-growing, or pointed, ends of the thin filaments. Together these proteins regulate thin filament lengths and actin dynamics in cardiac muscle. The K15N mutation in the TPM1 gene is associated with familial dilated cardiomyopathy (DCM) but the effect of this mutation on Tpm's function is unknown. In this study, we introduced the K15N mutation in striated muscle α-Tpm (Tpm1.1) and investigated its interaction with actin, Tmod and Lmod. The mutation caused a ∼3-fold decrease in the affinity of Tpm1.1 for actin. The binding of Lmod and Tmod to Tpm1.1-covered actin filaments also decreased in the presence of the K15N mutation. Furthermore, the K15N mutation in Tpm1.1 disrupted the inhibition of actin polymerization and affected the competition between Tmod1 and Lmod2 for binding at the pointed ends. Our data demonstrate that the K15N mutation alters pointed end dynamics by affecting molecular interactions between Tpm1.1, Lmod2 and Tmod1.

Myopathic mutations in the β‐chain of tropomyosin differently affect the structural and functional properties of ββ‐ and αβ‐dimers

“…However, the physiologic manifestations of various myopathic mutations of Tpm are multifaceted, so we cannot exclude the notion that they may affect its other properties. In recent studies, it has been found that mutations in Tpm genes can lead to changes in its interaction not only with the main proteins of thin and thick filaments, i.e., with actin and myosin, but also with tropomodulin and nebulin, which can affect sarcomere structure (48,49). The investigation of the mechanisms of interaction of the ab-heterodimer bearing various myopathic mutations with different partner proteins of actin filament in muscle cells is of great importance for a more detailed understanding of the development of myopathy and muscle functioning in general.…”

Congenital myopathy‐related mutations in tropomyosin disrupt regulatory function through altered actin affinity and tropomodulin binding

“…Inhibition of G‐actin polymerization rates by Tmod1, in the presence of wild‐type and mutant Tpm3.12, was analyzed by following fluorescence increase of pyrene‐G‐actin as described in . F‐actin seeds were prepared by incubation of 3 μ m unlabeled G‐actin with 30 n m gelsolin in: 25 m m imidazole, pH 7.0, 0.1 m KCl, 2 m m MgCl 2 , 0.2 m m ATP, 1 m m DTT, 1 m m EGTA, for 1 h. Seeds were diluted five‐fold with a solution of 1.0 μ m Pyrene‐labeled (5%) G‐actin in the presence or absence of 2.5 μ m Tpm3.12, 2.5 μ m Tpm3.12‐A4V, or 3.0 μ m Tpm3.12‐R91C.…”

Thin filament dysfunctions caused by mutations in tropomyosin Tpm3.12 and Tpm1.1

“…The DCM-causing Lys15Asn decreases the affinity of Tpm1.1 to the tropomyosin-binding site of Lmod2 and both tropomyosin-binding sites of Tmod1. Defects in the interactions between the N-terminal segment of the mutant tropomyosin and the isolated tropomyosin-binding sites of Lmod2 and Tmod1 result in significantly reduced binding of full length proteins and impaired abilities to inhibit elongation of the pointed end (Colpan et al 2017). The HCMlinked mutation Arg21His causes a severe decrease in the affinity for the isolated tropomyosin binding site of Lmod2 and diminishes the ability of Lmod2 to inhibit elongation of the pointed end (Ly et al 2019).…”