Functional interrelationship between (Na+ + K+)-ATPase and lysolecithin acyltransferase in plasma membranes of mitogen-stimulated rabbit thymocytes.

Lymphocytes from rabbit mesenteric lymph nodes or thymus were stimulated with the mitogen concanavalin A. The cardiac glycoside, ouabain, a known inhibitor of (Na' + K+)-ATPase, dose dependently suppressed the induction of RNA and DNA synthesis.When added together with the mitogen, complete inhibition was reached at concentrations of 5 X lo-' M ouabain.When ouabain was added to stimulated lymphocytes after varying time periods, the lymphocytes continued to incorporate [3H]uridine at a rate that was reached at the time of glycoside administration, suggesting that ouabain interferes with an early event of activation.After 1 h of concanavalin A stimulation, the incorporation of ["Cloleate into lecithin of microsomal membranes, which are derived largely from the plasma membranes, was increased more than 2-fold.Preincubation of the lymphocytes (to allow optimal binding) with 5 X lo-' M ouabain completely abolished this increase.Stimulation of lymphocytes for 1 h with concanavalin A also increased in microsomal membranes the specific activities of (Na' + K+)-ATPase and lysolecithin acyltransferase which regulates the turnover of the fatty acid moieties of membrane phospholipids.Preincubation of the lymphocytes with 5 X lo-' M ouabain prevented the activation of these enzymes whereas the specific activity of unstimulated cells was not affected.Control experiments showed that the prevention of the activation of lysolecithin acyltransferase was not mediated by monovalent cation shifts or inhibition of protein synthesis.Upon subfractionation of microsomal membranes by means of affinity chromatography on concanavalin A-Sepharose, (Na' + K+)-ATPase and lysolecithin acyltransferase segregated together suggesting a close spatial association of these two enzymes.Our results suggest a functional interrelationship between (Na+ + K+)-ATPase and lysolecithin acyltransferase in the plasma membrane of lymphocytes implicated in the initiation of lymphocyte activation.Increasing evidence implicates the plasma membrane as the primary site for the initial events associated with the control