Cardiac hypertrophy is the primary response of the heart to an increased hemodynamic load. In this project, we will determine how an increase in load is transferred by integrins to the modulation of intracellular signals that are responsible for hypertrophic growth. During mechanical stimulation, integrins are known to transduce biochemical signals and therefore, can be anticipated to play a significant role. Our recent studies revealed cytoskeletal recruitment (focal complex assembly) of several signaling proteins, including beta3-integrin, c-Src and focal adhesion kinase (FAK) in 4 to 48 h pressure overloaded myocardium. To improve upon the standard two-dimensional (2D) laminin model, we developed a cell culture system in which adult feline cardiocytes were embedded three-dimensionally (3D) in type I collagen and stimulated integrins using a synthetic peptide bearing an Arg-Gly-Asp (RGD) motif. Similar to the pressure overload model, RGD-stimulation of integrins caused focal complex assembly in this model. Further, we were able to mimic the activation of a critical kinase, p70/85 S6 kinase (S6K1) that was observed in pressure overloaded myocardium, with RGD stimulation of cardiocytes in both types of cell culture models. Whereas we established simple models and found a possible link between integrin activation and hypertrophic growth, two fundamental questions remain unanswered: (i) what is the molecular mechanism for the focal adhesion complex formation during integrin stimulation in adult cardiocytes and (ii) what specific role do integrin activation and focal adhesion complex play for mediating hypertrophic growth? These two primary questions lay the foundation to this proposal and will be addressed through two Specific Aims: (i) using a 3D model system, determine the role of key signaling proteins in the integrin-mediated focal complex formation observed in pressure overloaded myocardium; and (ii) determine the importance of integrin activation and focal complex formation in the cellular redistribution and regulation of ribosomal components for hypertrophic growth. As part of the First Specific Aim, we will identify the involvement of specific integrin subtypes and explore the role played by small GTPases (RhoA and Rac1) and non-receptor tyrosine kinases (FAK and c-Src) in focal complex formation. In the Second Specific Aim, we will study the importance of integrin signaling for cellular redistribution of ribosomal proteins, S6K1 activation and selective mobilization of 5'-TOP mRNA into polysome fractions. These studies of molecular mechanisms underlying integrin signaling and translational upregulation will contribute significantly to our understanding of load-induced hypertrophic growth.
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