Downregulation of DNA topoisomerase I in old versus young human diploid fibroblasts

Abstract

DNA topoisomerase I (Topo I) is an enzyme that alters the superhelicity of DNA. It has been implicated in such critical cellular functions as transcription, DNA replication, and recombination. Roles for Topo I in DNA repair following DNA damage have also been studied extensively. In the present investigation, we examined the regulation of Topo I expression and activity during cellular replicative senescence. We found that the capacity of Topo I to relax supercoiled DNA molecules is significantly decreased in senescent diploid fibroblasts when compared to young (early passage) fibroblasts. We also found that the steady-state expression level of Topo I mRNA is correlated with enzyme activity, i.e., decreased in early vs. late passage cells. We also treated early and late passage cells with agents that may modulate the process of cellular senescence: UV light, retinoic acid, and interleukin-1β. We found that all three agents decreased the activity of Topo I in young fibroblasts and increased the activity of Topo I in senescent fibroblasts. This effect was most striking following exposure of the cells to retinoic acid, so to analyze this effect, we postulated an age-dependent kinetics of Topo I mRNA induction in response to retinoic acid. Consistent with this postulate, we found that whereas exposure of early passage cells to retinoic acid results, in a matter of hours, in a decrease in the expression of Topo I mRNA, exposure of the senescent cells to retinoic acid results in an increased expression. These observations suggest that processes that are altered in senescent fibroblasts, such as DNA replication and repair, may be due, in part, to alteration in the expression and activity of DNA Topo I.