Proliferating cell nuclear antigen decreases in normal human keratinocytes with differentiation stimuli but not in an...

Proliferating cell nuclear antigen (PCNA) is a co-factor for DNA polymerase delta, which replicates genomic DNA during cell growth and division. Using a monoclonal antibody to PCNA (PC10) and conventional immunofluorescent techniques, we have compared the effect of differentiation stimuli on PCNA expression in normal and HPV immortalised keratinocytes. Two positive nuclear staining patterns were observed, a strong speckled form characteristic of proliferating cells and a weaker diffuse form. Strong nuclear staining was present in 44 +/- 4% (mean +/- SEM) of normal keratinocytes proliferating as a monolayer in 70 microM calcium serum-free medium but decreased to 13 +/- 3% after the differentiation stimulus of 2 mM calcium medium for 2 days. An even greater reduction was observed following other differentiation agents, 1,25 dihydroxyvitamin D3, the phorbol ester TPA and the non-specific protein kinase C inhibitor staurosporine. Transforming growth factor-beta, which slows keratinocyte growth without inducing differentiation, reduced strong staining to 17 +/- 3% of cells, but with an increase in the diffuse pattern of staining from 39 +/- 4 to 57 +/- 3%. HPV immortalised cells were resistant to the above agents except staurosporine, which inhibited growth and reduced the strong nuclear staining from 44 +/- 5% to 15 +/- 2%.