Transcription factors and the down-regulation of G1/S boundary genes in human diploid fibroblasts during senescence.

The hallmark of cellular aging is the failure of senescent cells to initiate the DNA synthesis during the progression of cell cycle. Since most, if not all, of the G1/S genes exhibit a significant down-regulation during aging, an alteration of gene regulation at late G1/S boundary could be a major contributing factor for the loss of dividing potential during cell senescence. The underlying cause for the apparent global attenuation of gene expression at late G1/S boundary is not clear. Since we have shown that thymidine kinase (TK) and dihydrofolate reductase (DHFR) are transcriptionally regulated during aging, we suspect that a similar mechanism may be operative in the age-dependent down-regulation of other G1/S genes. DNA binding activities using Y-box containing sequence in TK promoter or E2F containing sequence in DHFR promoter show prominent serum-responsiveness in low passage cells and dramatic attenuation in senescent cells. Promoter analysis using GCG program reveals striking similarities in promoter organization of twelve age-dependent G1/S genes. Specifically, these genes can be divided into two groups, one group contains tandem multiple CCAAT element, similar to that in TK promoter and the other contains E2F site, similar to that in DHFR promoter. Further analysis shows that the promoter of transcription factor, NF-Y, which recognizes CBP/tk site contains a tandem, two Y-box motif, similar to that in TK promoter and that the promoter of E2F1 contains four E2F motifs and two tandem CCAAT elements. Thus, these two important transcription factors could undergo autoregulatory control themselves. It is possible that regulation of only a few of transcription factors such as CBP/tk (NF-Y) and E2F1 may be sufficient to cause a global attenuation of most of G1/S genes in human diploid fibroblasts during senescence.