Cell cycle analysis of p26-BCL-2 protein levels in proliferating lymphoma and leukemia cell lines

The BCL-2 (B-cell lymphoma/leukemia-2) gene is frequently involved in t(14;18) translocations in non-Hodgkin's lymphomas and encodes a 26-kDa intracellular, membrane-associated protein. Expression of the BCL-2 gene has previously been correlated with cellular proliferation in normal and neoplastic lymphoid cells under a variety of experimental conditions. To examine the regulation of p26-BCL-2 protein levels during the cell cycle, we utilized the method of counterflow centrifugal elutriation to enrich for cells in various phases of the cell cycle. Relative levels of p26-BCL-2 protein were measured by immunoblotting, and comparisons were made with a cell cycle-regulated protein, p62-CYCLIN-A, and a protein whose levels are constant throughout the cell cycle, p36-PCNA (DNA polymerase-delta auxiliary factor). Relative levels of p26-BCL-2 and p36-PCNA did not vary among cell fractions enriched for specific phases of the cell cycle, whereas p62-CYCLIN-A was elevated in late S- and G2/M-phase cells. Similar results were obtained with lymphoma and leukemia cell lines that have either normal or translocated BCL-2 genes. These results obtained by elutriation were confirmed by pharmacologically inducing cell cycle arrest in proliferating lymphoid cell lines with hydroxyurea, quercetin, and nocodazole which blocked cells at S, G2, and M phases, respectively. Taken together, the data indicate that p26-BCL-2 is not a true cell cycle-regulated protein, although its levels can fluctuate in connection with changes in rates of cellular proliferation under some circumstances.