Abstract 754: Expression Analysis and Functional Significance of a Novel Cardiac Specific Human Tropomyosin in Normal and Cardiomyopathy Patients
Tropomyosins (TM) comprise a family of actin binding proteins that are central to the control of calcium-regulated striated muscle contraction. Although α-TM has been identified as a major striated muscle isoform, the precise TM protein composition in human hearts is unknown. Thus, the objective of the current study was to identify and characterize the striated TM isoforms in human hearts. Previous work using human RNA identified a novel tropomyosin isoform designated as TPM1κ that is specific to cardiac tissue. However, the presence of TPM1κ protein had not been confirmed. To address this, we developed a TPM1κ specific antibody and quantified the levels of TPM1κ protein in the hearts of normal and various cardiomyopathy patients. We now report for the first time that TPM1κ protein is expressed and subsequently incorporated into organized myofibrils in human heart. In acute cardiomyopathy patients, there is a reduction in the TPM1κ levels with a concomitant increase in the β-TM isoform, while in chronic cardiomyopathy patients, an increase in the levels of TPM1κ is observed. To investigate the role of TPM1κ in sarcomeric function and myofilament calcium sensitivity, we generated transgenic (TG) mice overexpressing TPM1κ in the hearts. Results show that TPM1κ protein is expressed at moderate to high levels in various lines of TG mice. Western blot analysis shows that the total amount of TM protein remains unchanged between TG and control mice; however, as the levels of TPM1κ increase, there is a decrease in endogenous α-TM. Immunohistochemical studies using the TPM1κ antibody show the incorporation of the TPM1κ protein in the myofilaments but no significant pathological alterations are observed in these TG mice. Hearts that express TPM1κ protein exhibit significant decreases in rates of contraction and relaxation when assessed by ex vivo work-performing cardiac analyses. Studies on skinned fiber bundles demonstrate decreased myofilament calcium sensitivity with no change in maximum developed tension. In conclusion, results from our TG mouse studies indicate that overexpression of TPM1κ lead to physiological alterations in hearts and provide a possible mechanistic explanation for the isoform switch that is observed in cardiomyopathy patients.