Isolation and characterization of a tyrosyl phosphatase activator from rabbit skeletal muscle and Xenopus laevis oocytes

Abstract

PTPA, a specific phosphotyrosyl phosphatase activator of the PCSH2 and PCSL protein phosphatases, was purified up to apparent homogeneity from Xenopus laevis ovaries and rabbit skeletal muscle and highly purified from dog liver. PTPA appears as a 40-kDa protein in gel filtration, as well as in sucrose gradient centrifugation, and as a 37-39-kDa protein doublet in SDS-PAGE. Its estimated cellular concentration of 0.75 microM in oocytes or 0.25 microM in rabbit skeletal muscle is suggestive of an important role in the regulation of the cellular PTPase activity. The PTPase activation reaction of the PCSL phosphatase is time-dependent, ATP and Mg2+ being essential cofactors [A50(ATP) = 0.12 mM in the presence of 5 mM MgCl2]. With RCM lysozyme as substrate, the specific activity of the PTPA-activated PCSL phosphatase is 700 nmol of Pi/(min.mg). The pH optimum of the PTPase shifts from 8.5-9 in basal conditions to a neutral pH (7-7.5), and the A50 for the essential metal ion Mg2+ is decreased (3 mM). The activation is rapidly reversed in the presence of the substrate, and more slowly after removal of ATP.Mg. The PTPA-activated PCSL phosphatase represents a major PTPase activity in the cytosol of X. laevis oocytes (at least 50% of the measurable PTPase with RCM lysozyme phosphorylated on tyrosyl residues). The PTPA activation is specific for the PTPase activity of the PCSL and PCSH2 phosphatases, without affecting their phosphoseryl/threonyl phosphatase activity. However, effectors of the phosphorylase phosphatase activity, such as polycations and okadaic acid, also influence the PTPase activity. Phosphorylase alpha inhibits the activated PTPase activity (I50 = 5 microM). The PTPase activity of the other oligomeric PCS phosphatases (PCSH1 and PCSM) is not influenced, suggesting an inhibitory role for some of their subunits. This activation is compared with the recently described PTPase stimulation of the PCS phosphatases by ATP/PPi [Goris, J., Pallen, C. J., Parker, P. J., Hermann, J., Waterfield, M. D., & Merlevede, W. (1988) Biochem. J. 256, 1029-1034] and by tubulin [Jessus, C., Goris, J., Cayla, X., Hermann, J., Hendrix, P., Ozon, R., & Merlevede, W. (1989) Eur. J. Biochem. 180, 15-22].

Protein phosphatase 2A interacts with the Src kinase substrate p130CAS

“…PP2A has been reported to possess tyrosine phosphatase activity in the presence of a speci®c activator (Cayla et al, 1990). However, we do not observe any evidence for tyrosine dephosphorylation of Cas in our system (Figure 7a).…”

Critical Role for Protein Phosphatase 2A Heterotrimers in Mammalian Cell Survival

“…Most PP2A catalytic subunits in the cell form heterotrimeric complexes followed by the AC core dimer and, finally, numerous lower abundance complexes (22)(23)(24)(25)(26)(27). Highlighting the role of PP2A in growth control, several viruses encode proteins that subvert PP2A activity by binding to either dimeric or trimeric forms of the phosphatase (1).…”

Functional analysis of the promoter region of the human phosphotyrosine phosphatase activator gene: Yin Yang 1 is essential for core promoter activity

“…( [6][7][8]. In itro the PTPase activity of PP2A D can be upregulated specifically by a protein called PTPA (PTPase activator) [9][10][11]. PTPA can modulate PP2A D activity only in the presence of ATP\Mg# + .…”

“…PTPA can modulate PP2A D activity only in the presence of ATP\Mg# + . The exact mechanism of activation is not clear, although phosphorylation can be ruled out [9,12]. PTPA has been cloned from rabbit and human cDNA libraries [12], and homologues have been found in Xenopus, Drosophila, Saccharomyces cere isiae and Schizosaccharomyces pombe [13].…”