Kinetics of antibody production by single cells. II. The action of transcription and translation inhibitors upon the metabolism of haemolysin-secreting cells

Abstract

The plaque-forming cell (PFC) population from spleens of mice at the peak of the primary immune response against sheep red blood cells has been analysed using a plaque-assay technique and automatic photographic equipment which recorded plaque formation and growth within a carboxymethyl-cellulose gel. Preliminary biochemical studies, using tritiated uridine and leucine as markers, and actinomycin D and puromycin as inhibitors of transcription and translation respectively, were performed with normal spleen cells. The effect of each of these inhibitors was then tested by means of the plaque-assay technique.

The formation and subsequent growth of a plaque within a normal gel are processes in which both transcription and translation are required for haemolysin secretion. It was found that in PFC populations, the functional lifespan of messenger RNA (m-RNA) specific for haemolysin translation has a mean value of 4 hours, while the functional lifespan of total RNA in whole spleen cell populations is not more than 2 hours. The pool of haemolysin stored in a PFC has a mean secretion time of 1.5 hours.

There was considerable heterogeneity with regard to the storage of haemolysin and to the functional lifespan of the haemolysin-specific m-RNA from one PFC to another within the spleen population. Furthermore, some PFC which displayed a high secretion rate of haemolysin were unaffected by actinomycin D, while others were inhibited within a few hours. The short-term action of each inhibitor has been studied. Puromycin was found to have a reversible action on PFC expression, while a 1-hour duration of action of actinomycin D was sufficient to inhibit irreversibly the subsequent PFC expression.