Intracellular signalling of initiation of DNA synthesis: Effect of cell‐free extracts from anti‐receptor‐stimulated B lymphocytes on spleen cell nuclei

Abstract

To study the relatively late intracellular signals involved in the proliferative response of B lymphocytes to antibodies specific for surface membrane immunoglobulins, extracts from antibody activated cells were mixed with Xenopus laevis splenic nuclei, and the incorporation of thymidine 5'-triphosphate into DNA was assessed. The slight incorporation observed with either nuclei or extract alone was markedly enhanced upon mixing the two entities when the extract was derived from cells cultured with but not without anti-receptor antibody. The appearance of active extract correlated well with the culture requirements necessary for the induction of B lymphocyte proliferation and, as revealed by time course studies, the active component arises relatively late in the activation process. Moreover, the appearance of active extracts is independent of DNA synthesis but is dependent on protein synthesis as judged from studies with metabolic inhibitors. Appropriate homogenization of activated cells yielded nuclei and cytoplasm with 85% of the activity confined to nuclei. In addition, purified active extracts exhibited DNA binding although the active component was readily distinguishable from polymerase alpha by chromatographic techniques. It is tentatively concluded that the active component represents either some replication protein other than polymerase or some earlier signal necessary to induce the formation or utilization of replicating proteins.