Activated complex of L-cells and Rickettsia prowazekii with N-ethylmaleimide-insensitive phospholipase A.
The interaction of large numbers of viable Rickettsia prowazekii cells with L-cells results in the expression of a phospholipase A activity with the concomitant release of free fatty acids and lysophosphatides from the phospholipids of the L-cell. About 50% of rickettsiae present in the suspension that was centrifuged onto an L-cell monolayer at 0 degree C to effect this interaction formed a tight L-cell-rickettsiae association from which the rickettsiae could not be removed by simple washing. Both the L-cell-rickettsiae association and the rickettsiae before association with L-cells interact with N-ethylmaleimide, so that the subsequent expression of the phospholipase A activity was inhibited (treatment of the L-cells with N-ethylmaleimide before centrifugation does not inhibit phospholipase activity). However, treatment of this association with 2,4-dinitrophenol and KCN caused much less inhibition of this phospholipase A activity than did treatment of the rickettsiae with these agents before centrifugation onto the L-cells. Incubation of the L-cell-rickettsiae association for a short time at 35 degrees C resulted in a very low level of free fatty acid formation and changed this association to an activated complex in which the phospholipase A activity was no longer sensitive to the inhibitory effects of N-ethylmaleimide. The characteristics of the association and activated complex were stable: after a 2-h incubation at 0 degrees C, the association and the activated complex retained both their basal phospholipase A activities and their characteristic responses to N-ethylmaleimide treatment. In scanning electron micrographs of the activated complexes, the rickettsiae that were initially attached were no longer visible after 45 min at 35 degrees C, and the surface of the L-cell appeared to have been etched away. These activated complexes provide a system in which modulators of the phospholipase A can be investigated without the confusion caused by the first-step receptor interaction between rickettsiae and their host cells.
D000818:Animals; D005033:Ethylmaleimide / Q000494:pharmacology; D051379:Mice; D008855:Microscopy, Electron, Scanning; D008954:Models, Biological; D010587:Phagocytosis; D010740:Phospholipases / Q000378:metabolism*; D010741:Phospholipases A / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012283:Rickettsia prowazekii / Q000201:enzymology*; D013997:Time Factors; D014438:Typhus, Epidemic Louse-Borne / Q000201:enzymology*
D010740:Phospholipases; D010741:Phospholipases A; D005033:Ethylmaleimide