title: The B''/PR72 subunit mediates Ca2+-dependent dephosphorylation of DARPP-32 by protein phosphatase 2A.
abstract: In dopaminoceptive neurons, dopamine- and cAMP-regulated phosphoprotein of 32 kDa (DARPP-32) plays a central role in integrating the effects of dopamine and other neurotransmitters. Phosphorylation of DARPP-32 at Thr-34 by protein kinase A results in inhibition of protein phosphatase 1 (PP1), and phosphorylation at Thr-75 by Cdk5 (cyclin-dependent kinase 5) results in inhibition of protein kinase A. Dephosphorylation at Thr-34 involves primarily the Ca(2+)-dependent protein phosphatase, PP2B (calcineurin), whereas dephosphorylation of Thr-75 involves primarily PP2A, the latter being subject to control by both cAMP- and Ca(2+)-dependent regulatory mechanisms. In the present study, we have investigated the mechanism of Ca(2+)-dependent regulation of Thr-75 by PP2A. We show that the PR72 (or B'' or PPP2R3A) regulatory subunit of PP2A is highly expressed in striatum. Through the use of overexpression and down-regulation by using RNAi, we show that PP2A, in a heterotrimeric complex with the PR72 subunit, mediates Ca(2+)-dependent dephosphorylation at Thr-75 of DARPP-32. The PR72 subunit contains two Ca(2+) binding sites formed by E and F helices (EF-hands 1 and 2), and we show that the former is necessary for the ability of PP2A activity to be regulated by Ca(2+), both in vitro and in vivo. Our studies also indicate that the PR72-containing form of PP2A is necessary for the ability of glutamate acting at alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid and NMDA receptors to regulate Thr-75 dephosphorylation. These studies further our understanding of the complex signal transduction pathways that regulate DARPP-32. In addition, our studies reveal an alternative intracellular mechanism whereby Ca(2+) can activate serine/threonine phosphatase activity.
authors.affiliation: Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University, New York, NY 10021, USA.
authors.forename: Jung-Hyuck
authors.initials: JH
authors.lastname: Ahn
authors.forename: Jee Young
authors.initials: JY
authors.lastname: Sung
authors.forename: Thomas
authors.initials: T
authors.lastname: McAvoy
authors.forename: Akinori
authors.initials: A
authors.lastname: Nishi
authors.forename: Veerle
authors.initials: V
authors.lastname: Janssens
authors.forename: Jozef
authors.initials: J
authors.lastname: Goris
authors.forename: Paul
authors.initials: P
authors.lastname: Greengard
authors.forename: Angus C
authors.initials: AC
authors.lastname: Nairn
mesh_terms: D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D051937:Dopamine and cAMP-Regulated Phosphoprotein 32 / Q000378:metabolism*; D006801:Humans; D015151:Immunoblotting; D047468:Immunoprecipitation; D009474:Neurons / Q000378:metabolism*; D009841:Oligonucleotides; D010766:Phosphorylation; D054648:Protein Phosphatase 2 / Q000378:metabolism*; D021122:Protein Subunits / Q000378:metabolism; D034622:RNA Interference
chemical_list: D051937:Dopamine and cAMP-Regulated Phosphoprotein 32; D009841:Oligonucleotides; C497928:PPP1R1B protein, human; D021122:Protein Subunits; D054648:Protein Phosphatase 2; D002118:Calcium