Biology subjects

Pinto, J. R.

Publications and source records attributed to Pinto, J. R..

6 recordsLinked to original sources

bioRxiv2026

Calmodulin mutation N54I causes autosomal dominant CPVT in mice

Catecholaminergic polymorphic ventricular tachycardia (CPVT) is an inherited arrhythmia syndrome characterized by stress- or catecholamine-induced ventricular arrhythmias in the absence of overt structural heart disease. Mutations in RYR2 and CASQ2 account for most genetically defined cases, although pathogenic variants in the three genes encoding calmodulin (CALM1-3) have also been linked to CPVT. Because all three CALM genes encode an identical calmodulin protein, pathogenic calmodulin variants are expected to be expressed in only a small fraction of total cellular calmodulin, raising the question of whether this limited abundance is sufficient to produce an arrhythmogenic phenotype in vivo. We generated a heterozygous mouse model carrying the human disease-associated N54I-equivalent mutation, N54I, in Calm1. Mutant calmodulin accounted for 13.7% of total cardiac calmodulin, consistent with expression from one of six Calm alleles. Under basal conditions, N54I/+ mice exhibited normal growth, survival, cardiac morphology, and surface electrocardiogram parameters. However, cardiomyocytes isolated from N54I/+ mice had increased rates of RyR2-mediated spontaneous calcium release. Following catecholaminergic challenge with isoproterenol and caffeine, N54I/+ mice exhibited significantly more premature ventricular contractions and arrhythmias than wild-type littermates. Exercise challenge in conscious mice similarly provoked ventricular ectopy and ventricular tachycardia. In addition, N54I/+ mice exhibited abnormalities of atrial and sinoatrial electrical activity, including premature atrial contractions, ectopic P waves, atrioventricular conduction slowing, and beat-to-beat variability. These findings demonstrate that expression of the N54I calmodulin variant from a single Calm1 allele is sufficient to produce a CPVT phenotype in vivo. This model provides experimental evidence linking a human disease-associated calmodulin variant to arrhythmogenesis and demonstrates the functional dominance of mutant calmodulin in the heart.

physiology↗

bioRxiv2026

Myosin Filaments of Vertebrate Skeletal and Cardiac Muscle are Highly Similar, but not Identical

Striated muscles consist of two filament types, one composed mostly of actin and the other composed mostly of myosin1,2. Actin filaments are highly similar across different muscle types and species both invertebrate and vertebrate3. Myosin filaments of vertebrate striated muscle are quite homogeneous in structure having identical lengths, governed by the giant protein titin4,5, and rotational symmetries while varying mostly in the isoforms of its proteins. Conversely, myosin filaments from invertebrate striated muscle are highly heterogeneous in multiple ways even within a single organism. Myosin filaments from vertebrate cardiac muscle have been shown to be highly similar in structure between mice and humans6-8. Conversely, thick filaments from the highly specialized insect indirect flight muscle have been shown to be highly variable in structure9-14. Here we used the drug mavacamten to stabilize a myosin head conformation known as the interacting heads motif in a fast skeletal muscle of rabbits, a highly studied model system. We show that the structure of relaxed rabbit skeletal muscle thick filaments is highly similar to those of relaxed human and mouse cardiac muscle, differing primarily in the positioning of some domains of myosin binding protein C vis-a-vis titin. In the context of the very different structures from indirect flight muscle, the result highlights different solutions to the same problems, control of muscle force and the requirements of endothermy, the internal generation of heat. In mammals, thick filaments are poised for varying levels of myosin activation15, while indirect flight muscle is poised for narrowly defined, high frequency contraction. In mammals, endothermy is a continuous problem; in insects, endothermy is primarily necessary for flight16.

biophysics↗

bioRxiv2026

The structure of the native cardiac crossbridge in the rigor state.

Cardiac contraction is driven by double-headed myosin cycling on cardiac thin filaments, where troponin-tropomyosin regulates myosin access to actin. Prior research used a single-headed myosin bound to bare actin, thereby limiting insight into coordination between myosin heads and the influence of troponin-tropomyosin on actomyosin interactions. Here, we report a high-resolution structure of the native cardiac rigor cross-bridge formed by heavy meromyosin bound to the thin filament. We show that direct communication between the two bound heads, uneven interactions between the heads and tropomyosin, and spatial constraints imposed by troponin govern myosin placement along the thin filament. Additionally, the two heads display non-equivalent motor-light chain interactions, yielding distinct lever-arm conformations indicative of asymmetric intramolecular strain. Together, these findings provide a structural framework for how the two myosin heads coordinate and how the components of the thin filament are integrated into force generation by active cross-bridges.

biophysics↗

bioRxiv2025

Desmoglein-2 deficiency results in cardiac dysfunction by compromising both Z-disc- and intercalated disc-mediated mechanotransduction

Desmoglein-2 (DSG2), a critical component of the cardiac desmosome and located at the cardiomyocyte-cardiomyocyte intercalated disc, is essential for cell-cell adhesion, cardiomyocyte mechanical stability, and electrical coupling between cells. However, its relative contribution in maintaining cardiac function at the sarcomere level remains unclear. Using 4-week-old (adolescent) and 16-week-old (adult) homozygous knock-in Dsg2-mutant (Dsg2mut/mut) mice, we found that loss of DSG2 leads to early onset chamber- and age-dependent cardiac dysfunction driven by Z-disc structural defects and increased myosin detachment rate. Interestingly, Ca{superscript 2}-activated force was markedly reduced in adolescent Dsg2mut/mut permeabilized left ventricular cardiac muscle bundles but preserved in permeabilized isolated cardiomyocytes. This disparity demonstrates that DSG2 is not only crucial for mechanical coupling between cardiomyocytes but also for force transmission within and between sarcomeres, revealing a novel role for DSG2 in maintaining contractile integrity at both the cellular and tissue levels.

biophysics↗

bioRxiv2023

Unlocking the Role of sMyBP-C: A Key Player in Skeletal Muscle Development and Growth

Skeletal muscle is the largest organ in the body, responsible for gross movement and metabolic regulation. Recently, variants in the MYBPC1 gene have been implicated in a variety of developmental muscle diseases, such as distal arthrogryposis. How MYBPC1 variants cause disease is not well understood. Here, through a collection of novel gene-edited mouse models, we define a critical role for slow myosin binding protein-C (sMyBP-C), encoded by MYBPC1, across muscle development, growth, and maintenance during prenatal, perinatal, postnatal and adult stages. Specifically, Mybpc1 knockout mice exhibited early postnatal lethality and impaired skeletal muscle formation and structure, skeletal deformity, and respiratory failure. Moreover, a conditional knockout of Mybpc1 in perinatal, postnatal and adult stages demonstrates impaired postnatal muscle growth and function secondary to disrupted actomyosin interaction and sarcomere structural integrity. These findings confirm the essential role of sMyBP-C in skeletal muscle and reveal specific functions in both prenatal embryonic musculoskeletal development and postnatal muscle growth and function.

physiology↗

bioRxiv2021

Post-translational modification patterns on β-myosin heavy chain are altered in ischemic and non-ischemic human hearts

Phosphorylation and acetylation of sarcomeric proteins are important for fine-tuning myocardial contractility. Here, we used bottom-up proteomics and label-free quantification to identify novel post-translational modifications (PTMs) on beta-myosin heavy chain ({beta}-MHC) in normal and failing human heart tissues. We report six acetylated lysines and two phosphorylated residues: K34-Ac, K58-Ac, S210-P, K213-Ac, T215-P, K429-Ac, K951-Ac, and K1195-Ac. K951-Ac was significantly reduced in both ischemic and non-ischemic failing hearts compared to non-diseased hearts. Molecular dynamics simulations show that K951-Ac may impact stability of thick filament tail interactions and ultimately myosin head positioning. K58-Ac altered the solvent exposed SH3 domain surface - known for protein-protein interactions - but did not appreciably change motor domain conformation or dynamics under conditions studied. Together, K213-Ac/T215-P altered loop 1s structure and dynamics - known to regulate ADP-release, ATPase activity, and sliding velocity. Our study suggests that {beta}-MHC acetylation levels may be influenced more by the PTM location than the type of heart disease since less protected acetylation sites are reduced in both heart failure groups. Additionally, these PTMs have potential to modulate interactions between {beta}-MHC and other regulatory sarcomeric proteins, ADP-release rate of myosin, flexibility of the S2 region, and cardiac myofilament contractility in normal and heart failure hearts.

physiology↗