Negative regulation of DNA synthesis in early erythropoietic progenitor cells (BFU-E) by a protein purified from the medium of C57BL/6 mouse marrow cells

Abstract

During studies on the influence of Fv-2 on the cycle state of the erythroid burst-forming unit (BFU-E), an activity was found in bone marrow supernatants from C57BL/6 (B6) mice that shut down DNA synthesis of the BFU-E in vitro. It acted within minutes, its action was completely reversed by a single wash, and it appeared specific to the BFU-E. The activity-causing substance, being macromolecular, heat stable, and trypsin-sensitive, was evidently a protein and was named negative regulatory protein. We purified a negative regulatory protein from a bone marrow-derived "B6 Pan" cell line with properties thus far indistinguishable from those of the negative regulatory protein obtained directly from B6 marrow. By gel filtration the protein has a Mr of approximately equal to 79,000, by cation- and anion-exchange chromatography it appears to be a neutral molecule at physiological pH, and the molecule does not bind to Con A. After five sequential chromatographic steps, we obtained a preparation active at a concentration of 25 ng/ml. Our findings are compatible with the hypothesis that quiescence of BFU-E with respect to DNA synthesis in vivo in B6 mice is mediated by negative regulatory protein molecules.