PT  - JOURNAL ARTICLE
AU  - Charles W. Laidley
AU  - Ephraim Cohen
AU  - John E. Casida
TI  - Protein Phosphatase in Neuroblastoma Cells:  [<sup>3</sup>H]Cantharidin Binding Site in Relation to Cytotoxicity
DP  - 1997 Mar 01
TA  - Journal of Pharmacology and Experimental Therapeutics
PG  - 1152--1158
VI  - 280
IP  - 3
4099  - http://jpet.aspetjournals.org/content/280/3/1152.short
4100  - http://jpet.aspetjournals.org/content/280/3/1152.full
SO  - J Pharmacol Exp Ther1997 Mar 01; 280
AB  - Protein phosphatase 2A (PP2A) plays a central role in essential phosphorylation-dependent signal transduction pathways. It is also a principal target for many natural toxicants (cantharidin, microcystins, diarrhetic shellfish poisons) and a synthetic herbicide (endothall). This study develops a cellular model to explore the toxicology of PP2A inhibitors by use of a [3H]cantharidic acid ([3H]CA) ligand binding assay to quantify interactions at the toxicant site and cell viability to evaluate in vivotoxicity. Mouse neuroblastoma (N1E-115) cells are similar to mouse brain with respect to the affinity (12–15 nM), number (B max, 9–22 pmol/mg protein) and ligand specificity of this binding site. In addition, the competitive potency of ten analogs of CA (including endothall) and two potent diarrhetic shellfish poisons (okadaic acid and calyculin A) is correlated (r 2 = .9) with and therefore predictive of their cytotoxicity. The only exception is microcystin LR which is a potent inhibitor at the binding site but is not cytotoxic, possibly reflecting a lack of cellular uptake. ATP and several other phosphorus-containing bifunctional acids inhibit [3H]CA binding by phosphorylation-independent pathways; pyrophosphate apparently acts as a competitive inhibitor. Mn++ and five other divalent cations are also inhibitors with a unique action of Mn++ at 25 to 50 μM in increasing [3H]CA binding, which suggests a specific role in PP2A function. Neuroblastoma cells are therefore suitable to study the mechanisms by which the toxicant, ATP and Mn++ binding sites regulate PP2A activity and cell physiology. The American Society for Pharmacology and Experimental Therapeutics