Effects ofP16DNA Methylation on Proliferation, Senescence, and Lifespan of Human Fibroblasts

Abstract

ABSTRACTThe aim is to study the effects ofP16DNA methylation on lifespan of normal cells. An expression-controllable pTRIPZ vector expressing P26-specific zinc finger binding protein-based methyltransferase (P16-Dnmt) was used to induceP16methylation in primary CCD-I8C0 fibroblasts via stable transfection. Long-term dynamic IncuCyte analysis showed that CCD-I8C0 fibroblasts expressing baseline P16-Dnmt continued proliferating until passage-26 in the 53thpost-transfection week, while vector control cells stopped proliferating at passage-6 and completely died 2 weeks later. The proliferation rate of baseline P16-Dnmt cells was significantly higher than that of vector control cells. The proportion of P-galactosidase-positive staining cells was significantly decreased in baseline P16-Dnmt cells compared to vector control cells. The P16 expression was lost in baseline P16-Dnmt cells at and after passage-6. The average telomere length in baseline P16-Dnmt cells also gradually decreased. In conclusion,P16methylation could prevent senescence, promote proliferation, and expand lifespan of human fibroblasts, which may play a role in cancer development.SummaryA zinc finger protein-based DNA methyltransferase (P16-Dnmt) expressed at the baseline level could specifically methylateP16promoter CpG islands.P16methylation induced by baseline P16-Dnmt could significantly prevent senescence, promote proliferation, and expand lifespan of primary human fibroblasts.