Investigation of human lymphocyte plasma membrane associated nucleic acid

Abstract

Plasma membranes were prepared from the human lymphocyte cell line WIL23A by hypotonic swelling, Dounce homogenization, differential and equilibrium centrifugation. The resulting vesiculated membrane fragments were found to have densities of 1.10 and 1.17 g/ml, and were defined by lactoperoxidase mediated whole cell iodination, L-[3H] fucose incorporation, 5'-nucleotidase activity (EC 3.1.3.5) and electron micrographic visualization. Recovery of plasma membrane from whole cell homogenates was estimated to be approximately 30-35% as judged by the recovery of 125I-labeled cell surface protein. When plasma membranes were prepared from cells which had been incubated for 18 h in the presence of 0.5 muCi/ml [3H] thymidine such that greater than 10(9) acid insoluble counts could be demonstrated in the whole cell homogenates, no [3H] thymidine label and presumably, therefore, no DNA, could be shown to be coincident with either the 1.10 or 1.17 density. Similar experiments with [3H] uridine suggested that 90% of the plasma membranes did not contain RNA, while 10% remained questionable.