My interests lie in elucidating post-translational mechanisms of acute and chronic regulation of the 20S proteasome within the context of cardiac hypertrophy. During the pathogenesis of hypertrophy, it is evident that net protein synthesis is increased, allowing the addition of de novo sarcomeres in parallel (hypertrophy) or in series (dilatation). While considerable advances have been made towards understanding pathways involved in stimulating hypertrophic growth via transcription or protein synthesis, the counter phenomenon, protein degradation, which may malfunction during pathologic hypertrophic growth, has been vastly understudied. Protein degradation catalyzed by 20S proteasomes, has been assumed a promiscuous, minimally-regulated process at the mercy of the ubiquitin-dependent 19S recognition system. Specifically, susceptibility of cardiac 20S proteasomes to acute or chronic post-translational regulation has been ignored. As protein-protein interactions are governed by the relative level of synthesis and degradation, the ratio of free versus assembled, and the proximity of associating partners, we hypothesize that phosphorylation of 20S proteolytic subunits may acutely regulate complex formation and proteolytic activity required for transduction and amplification of signaling pathways. Furthermore, under chronic pressure overload, we hypothesize that disrupted signaling to proteasomes may, in part, contribute to the pathogenesis of hypertrophy and downstream progression of cardiac failure.
Importantly, post-translational modifications are dynamic in nature, thus the tools we use to study these phenomena must be capable of quantitatively capturing these events on a physiologic timescale. A large portion of my project involves developing and implementing novel mass spectrometric approaches to quantify acute and chronic changes in the kinetics of post-translational profiles of cardiac 20S proteasomes. Additionally, my project includes a variety of traditional biochemical and classic physiology techniques, including immuno-precipitation, protein purification (HPLC, FPLC), 2DE, 2D-DIGE, and whole animal as well as cultured primary myocyte physiological measurements. Marrying functional, biochemical and proteomic profiles will allow for comprehensive assessment of phenotype and an integrated understanding of signaling to 20S proteasomes in cardiac cells.