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Studies on Early Events in the Differentiation of Antibody-Producting Cells

I. Patterns of Deoxyribonucleic Acid Replication in Antigen-Stimulated Mouse Spleen Cells¹

From the Departments of Microbiology and Zoology, Tel Aviv University, Tel Aviv, Israel

Abstract

The incorporation rates of tritium-labeled thymidine, uridine, and amino acids into macromolecules, and the appearance of plaque-forming cells (PFC) were studied at 4-hr intervals, in spleen cells obtained from C₅₇BL mice after a secondary intraperitoneal injection of sheep red blood cells. The incorporation rates displayed, over the period studied, clear maxima and minima occurring in the following order; a peak of uridine incorporation at 40 hr after immunization, 4 hr later a peak of thymidine incorporation, and at 60 hr a peak of amino acid incorporation. The number of PFC increased stepwise, the first and the second increases occurring at 60 and 76 hr after immunization, respectively. A similar sequence of biosynthetic events was observed in mouse spleen cells restimulated with sheep red blood cells in vitro. These results were interpreted as reflecting an antigen-induced, synchronized metabolic activity of the cells.

In an attempt to detect possible changes induced in the replication pattern of deoxyribonucleic acid (DNA) synthesized in response to antigenic stimulation, we compared the reassociation rate of DNA newly synthesized in spleen cells 47 to 48 hr after the antigenic stimulation with that of DNA from normal cells. The newly synthesized DNA displayed, between C₀t 1000 and C₀t 10,000 a faster reassociation rate than the bulk DNA obtained from the same cells or DNA obtained from normal cells. This suggests a temporary selective synthesis of that part of the genome during the pulse labeling. Further support for this assumption was provided by the changes in specific activity of single stranded "immune" DNA along the C₀t curve.

Footnotes

¹ This study was partially supported by a grant from the Bat-Sheva de Rothschild Fund for The Advancement of Science and Technology.

² Present address: Department of Cell Biology, The Weizmann Institute of Science, Rehovot, Israel.

³ To whom correspondence should be addressed.