ACTINOMYCIN BINDING PROPERTIES OF STIMULATED HUMAN LYMPHOCYTES

In cultures of human lymphocytes only a small percentage of the cells is capable of binding labelled actinomycin D (3H-AMD), and of incorporating 3H-uridine into RNA. Addition of phytohaemagglutinin (PHA) to such cultures markedly increases the frequency of cells binding 3H-AMD and of cells synthesizing RNA. There is a close parallelism between the frequency of cells capable of binding 3H-AMD and the frequency of cells synthesizing RNA. For a short period of time there is also a good correlation between the rate of RNA synthesis and the amount of 3H-AMD bound by individual cells. At later stages, however, practically all blast cells show a moderate AMD-binding, but a very strong incorporation of labelled uridine. Little 3H-AMD is lost from the cells if, after the labelling, the incubation is continued in the absence of actinomycin. Attempts to remove bound 3H-AMD by postincubation with unlabelled AMD for time periods up to 4 h did not result in any marked loss of 3H-AMD from control cells and PHA stimulated cells. It is pointed out that the quantity of AMD bound to living cells probably only represents a very small fraction of the total number of potential binding sites in the DNA. This fraction appears to vary depending on the functional state of the chromatin. Kinetic studies suggest that there are at least two different types of binding: one early and rapid form of binding and a second, slow, form of binding. The PHA-stimulated and the control cells differ with respect to the rapid type of binding but not with respect to the slower form of binding. It is suggested that the rapid type of binding reflects how large a portion of the chromatin is available for transcription. © 1969.