THE SERINE THREONINE PHOSPHATASE INHIBITOR, CALYCULIN-A, INHIBITS ANDDISSOCIATES MACROPHAGE RESPONSES TO LIPOPOLYSACCHARIDE

LPS-stimulated macrophages (M phi) produce inflammatory mediators that are largely responsible for the pathophysiology associated with septi c shock. M phi respond to LPS with rapid protein phosphorylation and d ephosphorylation on serine, threonine, and tyrosine residues. If these events are critical for the cellular response to LPS, the kinases and /or phosphatases involved may be vulnerable targets for pharmacologic intervention. Recent studies demonstrated that tyrosine kinase inhibit ors block LPS-induced tyrosine phosphorylation of MAP kinases as well as TNF-alpha and IL-1 beta production. To investigate a role for serin e/threonine phosphatases, we evaluated the effect of calyculin A, a po tent serine/threonine phosphatase inhibitor, on LPS stimulation of mur ine M phi. Pretreatment of M phi with calyculin A inhibited LPS-induce d expression of six immediate-early genes: TNF-alpha, IL-1 beta, IFN-b eta IP-10, IRF-1, and TNFR-2. Calyculin A added 1.5 h after LPS treatm ent greatly reduced accumulation of IP-10, IRF-1, and TNFR-2 mRNA, but not TNF-alpha, IL-1 beta, and IFN-beta mRNA. Calyculin A, in the abse nce or presence of LPS, resulted in sustained tyrosine phosphorylation of the MAP kinases. These findings suggest that an ''early'' serine/t hreonine phosphatase activity is essential for LPS stimulation of M ph i and that the activation of MAP kinases is not sufficient for the ind uction of these immediate-early genes. The requirement for a ''late'' phosphatase activity for expression of a subset of LPS-inducible genes dissociates at least two regulatory pathways in LPS signal transducti on.