Rat basophilic leukemia cell lines defective in phospholipid methyltransferase enzymes, Ca2+ influx, and histamine release: reconstitution by hybridization.

Abstract

Variants of the rat basophilic leukemia (RBL) cell line were isolated and screened for phospholipid methyltransferase I and II activites, enzymes that convert phosphatidylethanolamine to phosphatidylcholine. Two variants were found that had decreased phospholipid methyltransferase enzyme levels and were unable to cause an influx of Ca2+ or release histamine in an IgE-mediated reaction. These cells were able to release histamine through an ionophore-induced reaction, indicating that the releasing mechanism distal to the Ca2+ channel was intact. One cell line, 1C1.B1, had low specific activity for phospholipid methyltransferase I. A 2nd variant, 2H3.B6, had reduced phospholipid methyltransferase II activity. Although both variants were unable to incorporate label from [methyl-3H]methionine or [3H]serine into phosphatidylcholine, they were able to incorporate [methyl-3H]choline and myo[2-3H(N)]inositol into phospholipids. Fusion of the 2 cell lines and isolation on selective media resulted in the growth of 8 independent hybrids. All eight had an increased number of chromosomes and normal phospholipid methyltransferase activities. Stimulation of the hybrids with IgE resulted in Ca2+ influx and histamine release. Phospholipid methylation apparently precedes and is necessary for Ca2+ influx. The hypothesis is supported that methylation is a necessary early step in the IgE-mediated histamine release reaction in RBL cells.