REGULATION OF INSP3 RECEPTOR FUNCTION BY MAPK

Yale University, New Haven CT

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Abstract

This project will focus on how growth factors and the activation of the MARK pathway control intracellular Ca2+ in intact cells and will suggest a molecular basis for the specific roles for nuclear and cytosolic Ca2+ signaling and the importance of the regulation of these effects on the growth and function of hepatocytes. Three isoforms of the InsPSR have been identified. The cellular localization of each receptor isoform may be correlated with its functional properties. In hepatocytes the InsPSR type 1 isoform (lnsP3R-1) is cytosolic whereas the type 2 isoform (lnsP3R-2) is found throughout the cell, but is predominantly found in the nucleus and the canalicular region. We hypothesize that in hepatocytes MKP-1 differentially regulates the function and distribution of the InsPSR. In this project the interplay between the InsPSR and MAP kinases will be tested on the activity of single InsPSR channels, the ability of isolated cells to generate transient changes in intracellular Ca2+, and the dynamics of InsPSR distribution. An understanding of these complex interactions is necessary to explain the molecular mechanisms of Ca2+ signaling and thus the regeneraton of liver. The hypotheses to be tested include 1) Does activation of MAPK alter the function of the InsPSR and intracellular Ca2+ signaling? 2) Does MKP-1 inactivation of MAPK alter the function of the InsPSR and intracellular Ca2+ signaling? and 3) Does MAPK phosphorylation modulate the InsPSR isoform distribution within the cell? The preliminary results presented here show for the first time that the InsPSR of hepatocytes are regulated by the MAPK pathway The experiments outlined in this project will investigate the functional of this regulation of the InsPSR at the single channel level and will correlate the channel properties with cell and organ function. The results to be obtained will identify regulatory factors that determine isoform-specific Ca2+ signaling responses, how the cell regulates the channel isoforms to optimize cellular responses, and how this regulation goes awry in pathophysiological situations, such as liver degeneration and regeneration. A long term goal will be to use the molecular information obtained in these studies to suggest useful treatments for individuals affected with liver disease.