Tropomyosin pseudo-phosphorylation results in dilated cardiomyopathy

Abstract

Edited by Velia M. FowlerPhosphorylation of cardiac sarcomeric proteins plays a major role in the regulation of the physiological performance of the heart. Phosphorylation of thin filament proteins, such as troponin I and T, dramatically affects calcium sensitivity of the myofiber and systolic and diastolic functions. Phosphorylation of the regulatory protein tropomyosin (Tpm) results in altered biochemical properties of contraction; however, little is known about the physiological effect of Tpm phosphorylation on cardiac function. To address the in vivo significance of Tpm phosphorylation, here we generated transgenic mouse lines having a phosphomimetic substitution in the phosphorylation site of ␣-Tpm (S283D). High expression of Tpm S283D variant in one transgenic mouse line resulted in an increased heart:body weight ratio, coupled with a severe dilated cardiomyopathic phenotype resulting in death within 1 month of birth. Moderate Tpm S283D mice expression in other lines caused mild myocyte hypertrophy and fibrosis, did not affect lifespan, and was coupled with decreased expression of extracellular signal-regulated kinase 1/2 kinase signaling. Physiological analysis revealed that the transgenic mice exhibit impaired diastolic function, without changes in systolic performance. Surprisingly, we observed no alterations in calcium sensitivity of the myofibers, cooperativity, or calcium-ATPase activity in the myofibers. Our experiments also disclosed that casein kinase 2 plays an integral role in Tpm phosphorylation. In summary, increased expression of pseudophosphorylated Tpm impairs diastolic function in the intact heart, without altering calcium sensitivity or cooperativity of myofibers. Our findings provide the first extensive in vivo assessment of Tpm phosphorylation in the heart and its functional role in cardiac performance.Phosphorylation plays a major role in the regulation of cardiac function by affecting numerous membrane, cytoplasmic, and sarcomeric proteins. In the heart, phosphorylation of cardiac troponin I, cardiac troponin T, myosin-binding protein C, and titin regulate myofibrillar contraction, relaxation, and cross-bridge cycling (1-3). In contrast, relatively little is known regarding the importance of tropomyosin (Tpm) 8 phosphorylation. Previous investigations established that Tpm is phosphorylated at serine 283, the penultimate amino acid of the protein (4 -8). Tpm phosphorylation in the heart is developmentally regulated, with phosphorylated ␣-Tpm being the predominant isoform during fetal and newborn stages (ϳ60 -70%); in the adult mouse heart, the level of ␣-Tpm phosphorylation is decreased to ϳ30% (3). To address this developmental decrease in cardiac Tpm phosphorylation, we previously found there is a continuous decrease after birth until 5 months, whereupon increases occur by 15 months (5). We, and others, determined there is differential Tpm phosphorylation within the four cardiac chambers, with atria having the highest levels of phosphorylated Tpm (9). 9 In vitro investigations by Heel...

Effects of cryopreservation and long-term culture on biological characteristics and proteomic profiles of human umbilical cord-derived mesenchymal stem cells

“…It is reported that ITGA6 corresponds to the activation of regeneration involving an epithelial-mesenchymal transition in adult heart [33]. TPM is a potential marker in lung cancer diagnosis [34], and the latest study showed TPM pseudophosphorylation results in dilated cardiomyopathy [35]. However, the relationship between cryopreservation of hUC-MSCs after long-term culture and diseases including cancer and cardiomyopathy remains unknown and need to be further studied.…”

The Role of Tropomyosin in Cardiac Function and Disease

“…Furthermore, the Tpm carboxyl region interacts with TnT, and work by our laboratory demonstrates this region dramatically affects cardiac function by regulating rates of contraction and relaxation, in addition to influencing myofilament calcium sensitivity [26]. The specific role that Tpm phosphorylation plays in influencing the physiological role of the carboxyl terminus and its interactions with the other contractile proteins in determining cardiac function was the subject of our investigations [14,27,28], and is being addressed in our current work [29].…”

“…that casein kinase 2 can phosphorylate the striated muscle α-Tpm isoform, and this phosphorylation is specific for the S283 amino acid residue [29]. It is possible that dependent upon conditions (i.e., fetal vs. adult stages, skeletal vs. cardiac muscle, normal physiologic conditions vs. hypertrophic stress vs. physiological stress) that different kinases are activated to phosphorylate striated muscle Tpm in a myofiberspecific manner at different developmental stages and/or physiological conditions.…”

“…As mentioned, biochemical assays, myofiber analyses, and molecular simulations indicate Tpm phosphorylation enhances the stiffness of the head-to-tail Tpm overlap region and its binding to TnT, while decreasing myofibril relaxation without significantly affecting cooperativity or altering activation kinetics [11,22,34]. Recently, we embarked on studies to address the physiological importance of Tpm phosphorylation in the heart using an in vivo model system [29]. The system we employed involved generating transgenic (TG) mice that expressed a phosphorylation mimetic specifically in the heart.…”

“…In Tpm S283D mice, there is also a significant increase in the heart:body weight ratio, and these mice usually die by 1 month of age. A morphological and histological analysis of the Tpm S283D mice that express moderate levels of the transgene shows the hearts display a mild cardiomyocyte hypertrophy with limited fibrosis [29]. This moderate phenotype does not appear to progress to a more severe condition, even after 1 year.…”