Treatment with inhibitors of PP-1, PP-2A, or PP-2B activity did not result in the generation of detectable levels of phospho-Ser65 inhibitor-1 in acute striatal slices (data not shown). may serve mainly because an integrative signaling mechanism that governs the responsiveness of inhibitor-1 to cAMP-dependent protein kinase activation. Intracellular transmission transduction in eukaryotic organisms relies on the precise rules of protein phosphorylation by protein kinases and protein phosphatases. These enzymes are (S)-Mapracorat broadly classified based on their specificity for serine, threonine, or tyrosine residues. Approximately equal numbers of protein-tyrosine kinases and phosphatases are encoded by most eukaryotic genomes (1, 2). In contrast, merely a handful of protein serine/threonine phosphatases look like required for reversing the actions of a much larger cohort of protein serine/threonine kinases (3C5), raising the query of how protein serine/threonine phosphatase specificity is definitely accomplished. To counter this numerical disparity, protein serine/threonine phosphatases rely on a rich array of regulatory subunits that control the localization, activity, and substrate specificity of protein phosphatase catalytic subunits. In the (S)-Mapracorat case of protein phosphatase 1 (PP-1),2 one of the major eukaryotic protein serine/threonine phosphatases, nearly 60 actual and putative regulator proteins have been recognized (S)-Mapracorat to day (6, 7). Most of these regulators are involved in the focusing on of PP-1 to specific subcellular locations, whereas several modulate its catalytic activity. Historically, protein phosphatase inhibitor-1 (inhibitor-1, or I-1) was the 1st such endogenous molecule found to regulate protein phosphatase activity (8). This 19-kDa (S)-Mapracorat protein has a highly conserved primary sequence in vertebrates ranging from fish to mammals (9, 10). It mainly lacks elements of secondary structure (11), probably explaining why it is unusually stable to warmth, acidity, detergents, and organic solvents (12). Phosphorylation at Thr35 by cAMP-dependent protein kinase (PKA) converts the inactive protein into a selective and highly potent inhibitor of the catalytic subunit of PP-1 (IC50 1 nM) (8, 13). This site is dephosphorylated from the type-2 protein serine/threonine phosphatases, PP-2A and PP-2B (Ca2+/calmodulin-dependent protein phosphatase, or calcineurin) (14C16). PP-2B activity predominates in the presence of high intracellular Ca2+, placing inhibitor-1 rules under the opposing influences of cAMP and Ca2+ signaling. Downstream from inhibitor-1, a mechanism for transmission amplification is provided by the substrate specificity of PP-1. PP-1 dephosphorylates a broad spectrum of phosphoproteins targeted by an array of protein serine/threonine kinases, including PKA as well as others (14). Therefore, the inhibition of PP-1 by phospho-Thr35 inhibitor-1 results in the amplification of PKA-dependent signaling cascades and has the potential to impose cAMP rules upon the phosphorylation state of cellular substrates phosphorylated by additional protein kinases. Inhibitor-1 is definitely highly indicated in the brain, adipose cells, kidney, and skeletal muscle mass (17), with lower levels happening in the heart and lung (9). It has been shown to play a particularly important role like a PP-1 inhibitor in excitable cells like mind and cardiac muscle mass, where it has emerged as a key player in models of synaptic plasticity (16, 18) and cardiomyocyte contractility (19C21), respectively. Within the brain, inhibitor-1 is especially enriched in the cerebral cortex, striatum, and dentate gyrus of the hippocampal Rabbit Polyclonal to OR89 formation (22). In the heart, nearly 80% of inhibitor-1 is found in the sarcoplasmic reticulum (23), consistent with a role for inhibitor-1 in the rules of PP-1 substrates associated with this compartment in cardiomyocytes. Protein phosphatase regulatory subunits like inhibitor-1 can be controlled by multiple protein kinases and (S)-Mapracorat phosphatases, thus providing as potential points of integration for disparate transmission transduction cascades. For instance, DARPP-32, a homologous PP-1 inhibitor highly enriched in the striatum, is definitely phosphorylated at different serine/threonine residues by PKA (24), casein kinases 1 and 2 (25, 26), and cyclin-dependent kinase 5 (Cdk5) (27). Consistent with this notion, inhibitor-1 isolated from rabbit skeletal muscle mass in earlier studies was found to be greatly phosphorylated at Ser67 (28). This residue was later on characterized as a site of phosphorylation by mitogen-activated protein kinase (MAPK), cyclin-dependent kinase 1 (Cdk1), and Cdk5 and in.