Csk overexpression reduces several monokines and nitric oxide productions but enhances prostaglandin E2 production in response to lipopolysaccharide in the macrophage cell line J774A.1.

Abstract

The catalytic activity of src-family protein tyrosine kinases (src-PTK) is suppressed when a C-terminal tyrosine is phosphorylated by an intracellular PTK, C-terminal Src kinase (Csk). In the present report, to study the regulatory functions of the Csk in cells of monocyte/macrophage lineage, we transfected a eukaryotic expression vector containing rat csk cDNA in a macrophage cell line, J774A.1, and examined alterations of the response to lipopolysaccharide (LPS) in the transfectants which overexpressed Csk. Csk overexpression resulted primarily in a down-regulation of Fgr activity, an src-PTK expressed in J774A.1, and hyperphosphorylation of several cellular proteins of 35, 57, 66, 97 and 120-130 kDa. Furthermore, in these Csk transfectants, production of interleukin (IL)-1alpha, IL-6, tumor necrosis factor-alpha, and nitric oxide (NO) following LPS stimulation were reduced compared with those in parental J774A.1 or J774A.1 transfected with the vector alone. The extent of reduction paralleled the amounts of Csk proteins expressed in the Csk-transfected J774A.1. The reduced NO production in these cells was associated with low levels of mRNA of inducible NO synthetase. On the other hand, an enhancement of prostaglandin E2 production was observed in the Csk-transfected J774A.1 cells upon stimulation with LPS, which appeared to result from the high level of prostaglandin-H synthetase in the transfectants. The present findings indicate that overexpression of Csk has differential effects on the regulation of production of chemical mediators and monokines, probably via modulation of signal transduction downstream of LPS-mediated signals.