Association of brain protein phosphatase 1 with cytoskeletal targeting/regulatory subunits.
Protein phosphatase 1 catalytic subunit (PP1C) is highly enriched in isolated rat postsynaptic densities. Gel overlay analyses using digoxigenin (DIG)-labeled PP1C revealed four major rat brain PP1C-binding proteins (PP1bps) with molecular masses of approximately 216, 175, 134, and 75 kDa, which were (1) more abundant in brain than other rat tissues; (2) differentially expressed in microdissected brain regions; and (3) enriched in isolated cortex postsynaptic densities. PP1bp175, PP1bp134, PP1bp75, and PP1C were partially released from forebrain particulate extracts by incubation at low ionic strength, which destabilizes the actin cytoskeleton. Size-exclusion chromatography of solubilized extracts separated two main PP1 activities (approximately 600 and approximately 100 kDa). PP1bps and PP1C gamma1 were enriched in the approximately 600-kDa peak, but PP1C beta was enriched in the approximately 100-kDa peak. Furthermore, PP1bp175 and PP1bp134 exhibited lower binding of recombinant DIG-PP1C beta than recombinant DIG-PP1C gamma1 or DIG-PP1C alpha. Solubilized PP1bp175 and PP1bp134 interact with PP1C under native conditions, because they both (1) coeluted from size-exclusion and ion-exchange columns; (2) bound to microcystin-LR-Sepharose; and (3) coprecipitated using PP1C antibodies. Trypsinolysis of the approximately 600-kDa form of PP1 increased phosphorylase a phosphatase activity approximately fourfold, suggesting that interaction of PP1C with these PP1bps modulates its activity. Thus, brain PP1 activity is likely targeted to the cytoskeleton, including postsynaptic densities, by isoform-selective binding of PP1C to these targeting/regulatory subunits, contributing to the specificity of its physiological roles.
Colbran
R J
RJ
Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Nashville, Tennessee 37232-0615, U.S.A.
Bass
M A
MA
McNeill
R B
RB
Bollen
M
M
Zhao
S
S
Wadzinski
B E
BE
Strack
S
S
D000818:Animals; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003599:Cytoskeleton / Q000201:enzymology*; D007527:Isoenzymes / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D046911:Macromolecular Substances; D010749:Phosphoprotein Phosphatases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D016548:Prosencephalon / Q000201:enzymology*; D054645:Protein Phosphatase 1; D051381:Rats; D013347:Subcellular Fractions / Q000201:enzymology; D013569:Synapses / Q000201:enzymology
D007527:Isoenzymes; D046911:Macromolecular Substances; D010749:Phosphoprotein Phosphatases; D054645:Protein Phosphatase 1