Protein phosphatases 1 and 2A dephosphorylate B-50 in presynaptic plasma membranes from rat brain.
The protein B-50 is dephosphorylated in rat cortical synaptic plasma membranes (SPM) by protein phosphatase type 1 and 2A (PP-1 and PP-2A)-like activities. The present studies further demonstrate that B-50 is dephosphorylated not only by a spontaneously active PP-1-like enzyme, but also by a latent form after pretreatment of SPM with 0.2 mM cobalt/20 micrograms of trypsin/ml. The activity revealed by cobalt/trypsin was inhibited by inhibitor-2 and by high concentrations (microM) of okadaic acid, identifying it as a latent form of PP-1. In the presence of inhibitor-2 to block PP-1, histone H1 (16-64 micrograms/ml) and spermine (2 mM) increased B-50 dephosphorylation. This sensitivity to polycations and the reversal of their effects on B-50 dephosphorylation by 2 nM okadaic acid are indicative of PP-2A-like activity. PP-1- and PP-2A-like activities from SPM were further displayed by using exogenous phosphorylase alpha and histone H1 as substrates. Both PP-1 and PP-2A in rat SPM were immunologically identified with monospecific antibodies against the C-termini of catalytic subunits of rabbit skeletal muscle PP-1 and PP-2A. Okadaic acid-induced alteration of B-50 phosphorylation, consistent with inhibition of protein phosphatase activity, was demonstrated in rat cortical synaptosomes after immunoprecipitation with affinity-purified anti-B-50 immunoglobulin G. These results provide further evidence that SPM-bound PP-1 and PP-2A-like enzymes that share considerable similarities with their cytosolic counterparts may act as physiologically important phosphatases for B-50.
Han
Y F
YF
Departments of Biochemistry, Medical College of Ohio, Toledo 43699-0008.
Wang
W
W
Schlender
K K
KK
Ganjeizadeh
M
M
Dokas
L A
LA
D000818:Animals; D001921:Brain / Q000378:metabolism*; D002412:Cations / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D003035:Cobalt / Q000494:pharmacology; D004988:Ethers, Cyclic / Q000494:pharmacology; D019922:GAP-43 Protein; D007158:Immunologic Techniques; D008297:Male; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009419:Nerve Tissue Proteins / Q000037:antagonists & inhibitors / Q000378:metabolism*; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D010766:Phosphorylation; D051381:Rats; D011919:Rats, Inbred Strains; D013569:Synapses / Q000378:metabolism*; D013574:Synaptosomes / Q000378:metabolism; D014357:Trypsin / Q000494:pharmacology
D002412:Cations; D004988:Ethers, Cyclic; D019922:GAP-43 Protein; D008562:Membrane Glycoproteins; D009419:Nerve Tissue Proteins; D019319:Okadaic Acid; D003035:Cobalt; D010749:Phosphoprotein Phosphatases; D014357:Trypsin