p53-independent expression of p21^CIP1/WAF1 in plasmacytic cells during G₂ cell cycle arrest induced by Actinobacillus actinomycetemcomitans cytolethal distending toxin

The cytolethal distending toxin (CDT) from Actinobacillus actinomycetemcomitans has been shown to induce cell cycle arrest in the G₂/M phase in HeLa cells. In the present study, the mechanism of CDT-induced cell cycle arrest was investigated by using HS-72 cells, a murine B-cell hybridoma cell line. Using flow cytometric analysis, we found that the recombinant CDT (rCDT) from A. actinomycetemcomitans induced G₂ cell cycle arrest in HS-72 cells and that rCDT upregulated expression of the cyclin-dependent kinase inhibitor p21^CIP1/WAF1 and the tumor suppressor protein p53. HS-72 cells transfected with the E6/E7 gene of human papillomavirus type 16, which lacked rCDT-induced accumulation of p53, exhibited expression of p21^CIP1/WAF1 or G₂ cell cycle arrest upon exposure to rCDT. Furthermore, ectopic expression of a dominant negative p53 mutant did not inhibit rCDT-mediated p21^CIP1/WAF1 expression or G₂ cell cycle arrest in HS-72 cells. These results suggest that the CDT from A. actinomycetemcomitans induces p21^CIP1/WAF1 expression and G₂ cell cycle arrest in B-lineage cells by p53-independent pathways. Together with additional observations made with HeLa cells and COS-1 cells cultured with the rCDT from A. actinomycetemcomitans, the results of this study indicate that CDT-induced p53 accumulation may not be required for G₂ cell cycle arrest and that an increased level of p21^CIP1/WAF1 may be important for sustaining G₂ cell cycle arrest in several mammalian cells.