Abstract 1618: Temporally Induced Expression of Active Protein Phosphatase Inhibitor-1 in the Adult Mouse Heart Enhances Cardiac Function
Protein Phosphatase Inhibitor-1 (I-1) is a critical regulator of cardiac contractility. Upon phosphorylation at threonine-35 by PKA, I-1 potently inhibits type-1 protein phosphatase (PP1) and augments cardiac contractility. To define the potential benefits of acute I-1 activation in the adult heart in the absence of early developmental compensatory responses, we generated a double transgenic mouse model with cardiac-specific and temporally regulated expression of active I-1 (T35D). This Tet-off system requires two transgenes: a cardiac-specific promoter driving expression of the tetracycline controlled transactivator gene (TG1); and a TetO-attenuated promoter driving expression of the I-1 cDNA (TG2). Thus, the double transgenic (DTG) mice are expected to express I-1 once doxycycline is removed from their diet. Indeed, a time course of doxycycline withdrawal revealed that four weeks were sufficient to elicit maximal contractile responses, which were similar to those exhibited by DTGs fed a doxycycline free diet upon birth. The rates of contraction and relaxation (±dP/dTmax) were increased by 22% and 31%, respectively, while Tau was decreased by 33% in the DTG group compared to the wild-types (WT), as assessed in Langendorf perfused hearts. TG2 mice exhibited similar parameters to WTs, indicating no leakage by the TG2 promoter in the absence of the TG1 promoter. Accordingly, acute infection of rat cardiomyocytes with an adenovirus encoding the constitutively active I-1, induced a significant increase of basal contractility, compared to the I-1 wild-type. Further proteomics and biochemical studies in the DTG hearts with induced I-1 expression showed no alterations in the protein expression levels. However, the phosphorylation of myosin light chain 2a (identified by 2D-gels followed by MALDI-TOF MS), and p-Ser16-PLN was significantly increased. Our results indicate that acute inhibitor-1 activation in the adult heart enhances cardiac function, which is partially mediated through increased phospholamban and myosin light chain 2a phosphorylation. Future studies will determine the potential therapeutic value of inhibitor-1 after the onset of cardiac remodeling, either during the compensated or decompensated states of hypertrophy.