Protein phosphatase 2A inhibitors, I(PP2A)/1 and I(PP2A)/2, associate with and modify the substrate specificity of protein phosphatase 1

Abstract

Recombinant I(PP2A)1 and I(PP2A)2 did not affect the activity of the catalytic subunit of protein phosphatase 1 (PP1(c)) with 32P-labeled myelin basic protein, histone H1, and phosphorylase when assayed in the absence of divalent cations. However, in the presence of Mn2+, I(PP2A)/1 and I(PP2A)/2 stimulated PP1(c) activity by 15-20-fold with myelin basic protein and histone H1 but not phosphorylase. Half-maximal stimulation occurred at 2 and 4 nM I(PP2A)/1 and I(PP2A)/2, respectively. Moreover, I(PP2A)/1 and I(PP2A)/2 reduced the Mn2+ requirement by about 30-fold to 10 μM. In contrast, PP1(c) activity was unaffected by I(PP2A)/1 and I(PP2A)/2 in the presence of Co3+ (0.1 mM), Mg2+ (2 mM), Ca2+ (0.5 mM), and Zn2+ (0.1 mM). Following gel filtration chromatography on Sephacryl S-200 in the presence of Mn2+, PP1(c) coeluted with I(PP2A)1 and I(PP2A)2 in the void volume. However, when I(PP2A)/1 and 1(PP2A)/2 or Mn2+ were omitted, PP1(c) emerged with a V(e)/V(o) of ~1.6. The results demonstrate that I(PP2A)/1 and I(PP2A)/2 associate with and modify the substrate specificity of PP1(c) in the presence of physiological concentrations of Mn2+. A novel role is suggested for I(PP2A)/1 and I(PP2A)/2 in the reciprocal regulation of two major mammalian serine/threonine phosphatases, PP1 and PP2A.