Control of the human cell cycle by a bacterial protein, gapstatin

The oral Gram-negative bacterium Actinobacillus actinomycetemcomitans is a major pathogen in human periodontal disease. Saline extraction releases a r ange of surface-associated components from this bacterium, including one wh ich exhibits potent anti-proliferative activity as assessed by its capacity to inhibit DNA synthesis by human and other mammalian cells. Cultures incu bated with this bacterial fraction for a prolonged period comprise a high p roportion of cells containing a 4n level of DNA. Studies using hydroxyurea- synchronized cultures shelved that cells treated with the surface-associate d fraction were arrested in the G(2) phase of the cell cycle and did not en ter mitosis. This G(2)/M blockade was observed only when the bacterial frac tion was added to the cells during early S phase. Our data also suggest tha t the active bacterial component hinds to surface receptors expressed by th e human cells and may act by a novel mechanism which involves down-regulati on of cyclin B1 expression. The anti-proliferative activity of the bacterial fraction, purified by a co mbination of ammonium sulphate precipitation, HPLC anion exchange and gel f iltration, has been shown to be an 8 kDa protein, which we have called gaps tatin. Purified gapstatin was shown to be responsible for the the inhibitor y effects of the surface-associated fraction on mammalian cells.