α-Tropomyosin mutations Asp¹⁷⁵Asn and Glu¹⁸⁰Gly affect cardiac function in transgenic rats in different ways

Abstract

To study the mechanisms by which missense mutations in α-tropomyosin cause familial hypertrophic cardiomyopathy, we generated transgenic rats overexpressing α-tropomyosin with one of two disease-causing mutations, Asp175Asn or Glu180Gly, and analyzed phenotypic changes at molecular, morphological, and physiological levels. The transgenic proteins were stably integrated into the sarcomere, as shown by immunohistochemistry using a human-specific anti-α-tropomyosin antibody, ARG1. In transgenic rats with either α-tropomyosin mutation, molecular markers of cardiac hypertrophy were induced. Ca2+ sensitivity of cardiac skinned-fiber preparations from animals with mutation Asp175Asn, but not Glu180Gly, was decreased. Furthermore, elevated frequency and amplitude of spontaneous Ca2+ waves were detected only in cardiomyocytes from animals with mutation Asp175Asn, suggesting an increase in intracellular Ca2+ concentration compensating for the reduced Ca2+ sensitivity of isometric force generation. Accordingly, in Langendorff-perfused heart preparations, myocardial contraction and relaxation were accelerated in animals with mutation Asp175Asn. The results allow us to propose a hypothesis of the pathogenetic changes caused by α-tropomyosin mutation Asp175Asn in familial hypertrophic cardiomyopathy on the basis of changes in Ca2+ handling as a sensitive mechanism to compensate for alterations in sarcomeric structure.

Designing Heart Performance by Gene Transfer

“…Extremely low levels of E180G or D175N Tm replacement had no effect on myofilament Ca 2ϩ sensitivity or relaxation function in E180G rat myocytes, while D175N Tm desensitized the myofilaments to Ca 2ϩ and accelerated relaxation rates (955). The divergent transgenic rat phenotype could be due to expressing a human ␣Tm sequence in the rat (955), but Michele et al (564) used the human ␣Tm cDNA as the backbone for mutagenesis in the previously described acute gene transfer studies which closely paralleled the work done in transgenic mouse models.…”

“…The E180G and D175N mutant Tms have been expressed in transgenic mice (567,623,706,707) and more recently in rats (955). The direct effects of the E180G Tm mutation in transgenic mice closely paralleled that of acute gene transfer, but the compensatory changes varied between E180G transgenic mouse models.…”

Sarcomeric Proteins and Familial Hypertrophic Cardiomyopathy: Linking Mutations in Structural Proteins to Complex Cardiovascular Phenotypes

“…A number of transgenic animal models (including mouse and rat) have been generated and extensively analyzed for all five of the original cTnT and Tm human mutations originally described by Thierfelder: I79N, R92Q, del(exon15/16), D179N and E180G [76][77][78][79][80][81][82][83]. Surprisingly, despite relatively different approaches (e.g.…”

Sarcomeric Protein Mutations in Dilated Cardiomyopathy

“…Studies on the Tm structure and actin binding affinity measurements have demonstrated that the HCM associated mutations often lead to alterations in the thermal stability of the Tm protein and also a decrease in actin binding affinity [25][26][27][28]. Functional characterization of muscle fibers extracted from transgenic animal models generated with the familial HCM mutations [27,29], show increased Ca 2+ sensitivity levels, which correlate well with our results in the investigation of the effects of familial HCM mutations in Tn, the results of which are reviewed elsewhere [30].…”