Lmod2 is necessary for effective skeletal muscle contraction.

Muscle contraction is a regulated process driven by the sliding of actin-thin filaments over myosin-thick filaments. Lmod2 is an actin filament length regulator and essential for life since human mutations and complete loss of Lmod2 in mice lead to dilated cardiomyopathy and death. To study the little-known role of Lmod2 in skeletal muscle, we created a mouse model with Lmod2 expressed exclusively in the heart but absent in skeletal muscle. Loss of Lmod2 in skeletal muscle results in decreased force production in fast- and slow-twitch muscles. Soleus muscle from rescued Lmod2 knockout mice have shorter thin filaments, increased Lmod3 levels, and present with a myosin fiber type switch from fast myosin heavy chain (MHC) IIA to the slower MHC I isoform. Since Lmod2 regulates thin-filament length in slow-twitch but not fast-twitch skeletal muscle and force deficits were observed in both muscle types, this work demonstrates that Lmod2 regulates skeletal muscle contraction, independent of its role in thin-filament length regulation.

A nemaline myopathy-linked mutation inhibits the actin-regulatory functions of tropomodulin and leiomodin.

Actin is a highly expressed protein in eukaryotic cells and is essential for numerous cellular processes. In particular, efficient striated muscle contraction is dependent upon the precise regulation of actin-based thin filament structure and function. Alterations in the lengths of actin-thin filaments can lead to the development of myopathies. Leiomodins and tropomodulins are members of an actin-binding protein family that fine-tune thin filament lengths, and their dysfunction is implicated in muscle diseases. An Lmod3 mutation [G326R] was previously identified in patients with nemaline myopathy (NM), a severe skeletal muscle disorder; this residue is conserved among Lmod and Tmod isoforms and resides within their homologous leucine-rich repeat (LRR) domain. We mutated this glycine to arginine in Lmod and Tmod to determine the physiological function of this residue and domain. This G-to-R substitution disrupts Lmod and Tmod's LRR domain structure, altering their binding interface with actin and destroying their abilities to regulate thin filament lengths. Additionally, this mutation renders Lmod3 nonfunctional in vivo. We found that one single amino acid is essential for folding of Lmod and Tmod LRR domains, and thus is essential for the opposing actin-regulatory functions of Lmod (filament elongation) and Tmod (filament shortening), revealing a mechanism underlying the development of NM.

Whole-Exome Sequencing Identifies Homozygote Nonsense Variants in LMOD2 Gene Causing Infantile Dilated Cardiomyopathy.

As an essential component of the sarcomere, actin thin filament stems from the Z-disk extend toward the middle of the sarcomere and overlaps with myosin thick filaments. Elongation of the cardiac thin filament is essential for normal sarcomere maturation and heart function. This process is regulated by the actin-binding proteins Leiomodins (LMODs), among which LMOD2 has recently been identified as a key regulator of thin filament elongation to reach a mature length. Few reports have implicated homozygous loss of function variants of LMOD2 in neonatal dilated cardiomyopathy (DCM) associated with thin filament shortening. We present the fifth case of DCM due to biallelic variants in the LMOD2 gene and the second case with the c.1193G>A (p.W398*) nonsense variant identified by whole-exome sequencing. The proband is a 4-month male infant of Hispanic descent with advanced heart failure. Consistent with previous reports, a myocardial biopsy exhibited remarkably short thin filaments. However, compared to other cases of identical or similar biallelic variants, the patient presented here has an unusually late onset of cardiomyopathy during infancy. Herein, we present the phenotypic and histological features of this variant, confirm the pathogenic impact on protein expression and sarcomere structure, and discuss the current knowledge of LMOD2-related cardiomyopathy.

EVL is a novel focal adhesion protein involved in the regulation of cytoskeletal dynamics and vascular permeability.

Increases in lung vascular permeability is a cardinal feature of inflammatory disease and represents an imbalance in vascular contractile forces and barrier-restorative forces, with both forces highly dependent upon the actin cytoskeleton. The current study investigates the role of Ena-VASP-like (EVL), a member of the Ena-VASP family known to regulate the actin cytoskeleton, in regulating vascular permeability responses and lung endothelial cell barrier integrity. Utilizing changes in transendothelial electricial resistance (TEER) to measure endothelial cell barrier responses, we demonstrate that EVL expression regulates endothelial cell responses to both sphingosine-1-phospate (S1P), a vascular barrier-enhancing agonist, and to thrombin, a barrier-disrupting stimulus. Total internal reflection fluorescence demonstrates that EVL is present in endothelial cell focal adhesions and impacts focal adhesion size, distribution, and the number of focal adhesions generated in response to S1P and thrombin challenge, with the focal adhesion kinase (FAK) a key contributor in S1P-stimulated EVL-transduced endothelial cell but a limited role in thrombin-induced focal adhesion rearrangements. In summary, these data indicate that EVL is a focal adhesion protein intimately involved in regulation of cytoskeletal responses to endothelial cell barrier-altering stimuli. Keywords: cytoskeleton, vascular barrier, sphingosine-1-phosphate, thrombin, focal adhesion kinase (FAK), Ena-VASP like protein (EVL), cytoskeletal regulatory protein.

Disruption of cardiac thin filament assembly arising from a mutation in LMOD2: A novel mechanism of neonatal dilated cardiomyopathy.

Neonatal heart failure is a rare, poorly-understood presentation of familial dilated cardiomyopathy (DCM). Exome sequencing in a neonate with severe DCM revealed a homozygous nonsense variant in leiomodin 2 (LMOD2, p.Trp398*). Leiomodins (Lmods) are actin-binding proteins that regulate actin filament assembly. While disease-causing mutations in smooth (LMOD1) and skeletal (LMOD3) muscle isoforms have been described, the cardiac (LMOD2) isoform has not been previously associated with human disease. Like our patient, Lmod2-null mice have severe early-onset DCM and die before weaning. The infant's explanted heart showed extraordinarily short thin filaments with isolated cardiomyocytes displaying a large reduction in maximum calcium-activated force production. The lack of extracardiac symptoms in Lmod2-null mice, and remarkable morphological and functional similarities between the patient and mouse model informed the decision to pursue cardiac transplantation in the patient. To our knowledge, this is the first report of aberrant cardiac thin filament assembly associated with human cardiomyopathy.