We assessed the expression and distribution of p21/Waf-1 in TLM1 melanoma cells that exhibit contact inhibition and require serum for growth. The growth stage of cells stimulated to enter the mitotic cell cycle synchronously and grow to confluence was characterized by distinct, yet consistent levels and patterns of distribution of p21/Waf-1. Significantly, sustained accumulation of p21/Waf-1 in the nuclear compartment was seen only after 4 days in culture when cell-to-cell contacts were established, leading to a diminished rate of cell growth. Overexpression of wild-type waf-1 in melanoma cells reduced growth of subconfluent cells, decreased Cdk4 activity with a concomitant increase in hypophosphorylated Rb, and promoted cell death by apoptosis. The data support the premise that cell-to-cell contacts provide signals that mediate sustained nuclear localization of p21/Waf-1 leading to cell growth arrest; furthermore, an elevation in the activity of this protein can lead to apoptosis. (C) 2000 Elsevier Science Ireland Ltd.
Bibliographical noteFunding Information:
The authors gratefully acknowledge Dr David Beach for providing the plasmid encoding wild-type human waf-1. We also wish to thank Cheryl Morales and Maren Fuentes for technical assistance, John B. Roths for assistance with image analysis, and Drs Stuart Helfand, Gheorge Stoica, Tom Welsh, Jennifer Thomas, Carol Chitko-McKown, and Peter Nowell for reading the manuscript and for helpful discussions. This work was supported in part by NIH Clinical Investigator Development Award K08-HL03130, Grant 1626 from the Canine Health Foundation of the American Kennel Club and Grant 98-CA36 from Morris Animal Foundation to J.F.M.
- Cell cycle
- Contact inhibition
- Cyclin-dependent kinase inhibitor