The cardiocyte responds to increased hemodynamic loading by augmented cell mass and by changes in specific gene expression that result in contractile dysfunction. In early pressure overload cardiac hypertrophy this contractile dysfunction may be primarily due to the increased microtubule formation that increases the internal resistance to sarcomere motion. In late hypertrophy and failure, contractile function is further compromised in the cardiomyocyte by changes in Ca2+ homeostasis. We determined that the rapid upregulation of Na-Ca exchanger message is regulated at the transcriptional level, remains up-regulated throughout the period of hypertrophic growth and results in increased exchanger protein and activity. We have identified the cis elements which are required for expression of the ncxl gene in the heart. Importantly, we have also discovered a novel element that is not only required for cardiac expression but is important for the exchanger up-regulation. Here we are in an excellent position to examine the molecular mechanisms which mediate exchanger expression in response to hemodynamic load. In addition, we can directly address, using transgenic mice, the question of whether up- regulation of the Na-Ca exchange is part of a recapitulation of embryonic expression triggered by hypertrophy or whether its up-regulation is triggered by changes in Ca2+ homeostasis brought about by the drop in SR Ca2+- ATPase expression and activity. Lastly, given that the exchanger is the predominant mechanism for Ca2+ efflux, it is surprising that so little is known about how the exchanger activity is regulated. We have discovered that exchanger activity is dramatically affected in adult cardiocytes by inhibition of protein phosphatases. Preliminary data indicate that this regulation of the exchanger may be mediated by interaction with cytoskeletal elements. What are the cellular factors that regulate Ca-Ca exchanger activity in the adult cardiocyte and how do they mediate exchanger activity in the normal and hypertrophic heart? The specific objectives of the proposal are: 1) characterize the cis-regulatory elements and 2) identify and characterize the trans-acting factors responsible for cardiac-specific and load-induced regulation of the nxc1 gene, and 3) begin to characterize the regulation of Na-Ca exchanger activity in cardiac hypertrophy. This work presents a unique opportunity to gain insight into the transcriptional regulation of a gene whose product is critical to calcium homeostasis and understand how exchanger activity is regulated in the normal and hypertrophied heart.
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