Fellowship for Victoria Dunlap: Determination of the Structural Mechanism for RCAN1's Duel Regulation of Calcineurin

Regulator of Galcineurin1 (RCAN1} is an endogenous protein regulator of !he calcium-sensitive senne/threonine protein phosphatase calclneUlin. Through its regulation of calcineurin RCAN1 has been implicated in fetal cardiac and vasculature development, angiogenesis, and cardiac hypertrophy. RCAN1 can both stimulate and inhibit calclfleurin depending on its cellular concentration and phosphorylation state, however the strnctural mechanism by whiCh RCAN1 can differentially regulate calcineurin is unknown. An understanding of this mechanism is necessary to elucidate RCAN1's role in cardiac dev~opment and pathology and Its use as a possible drug target The experiments in this proposal are designed to determine RCAN1's structural mechanism for dual calclneurin regulation by investigating the thermodynamics of RCAN1 binding to calcineurin, the affect that RCAN1 binding has on calcineurin function, the conformational properties of RCAN1 both alone and bound to caicineurin, and the changes in calcineurin structure upon RCAN1 binding using a variety of biophysical techniques such as isothermal titratIon calorimetry, analytical ultracentrifugation, fluorescence, circular dichroism, and hydrogen-deuterium ex;:hange mass spectrometry. The studies will be performed using RCAN14 (isororm 4) in its unphosphorylated, singly phosphorylated. and doubly phosphorylated states.