Identification and characterization of novel isoforms of human DP-1 -: DP-1α regulates the transcriptional activity of E2F1 as well as cell cycle progression in a dominant-negative manner

The cell cycle-regulating transcription factors DP-1 and E2F form a heterodimeric complex and play a central role in cell cycle progression. Two different DP subunits (DP-1 and DP-2) exist in humans. In this study, we identified two novel DP-1 isoforms (DP-1 alpha and DP-1 beta) and characterized their structure and function. DP-1 alpha is composed of 278 amino acids and lacks a portion of the C-terminal heterodimerization domain, whereas DP-1 alpha is composed of 357 amino acids with a frameshift that causes truncation of the C-terminal domain. Yeast two-hybrid and immunoprecipitation assays demonstrated that DP-1 alpha binding to E2F1 was significantly reduced as compared with that of wild-type DP-1 or DP-1 beta. Immunofluorescence analysis revealed that the subcellular localization of both DP-1 isoforms changed from the cytoplasm to the nucleus in HEK 293 cells cotransfected with E2F1 and wild-type DP-1 or DP-1 beta. However, such a translocation for DP-1 alpha was barely observed. Reverse transcription-PCR results showed that the three DP-1 isoforms are expressed ubiquitously at equal levels in several normal human tissues. We also demonstrated the expression of these isoforms at the protein level by Western blotting. Interestingly, we observed a significant decrease in transcriptional activity, a marked delay of cell cycle progression, and an inhibition of cell proliferation in DP-1 alpha-transfected HEK 293 cells. Together, the results of the present study suggest that DP-1 alpha is a novel isoform of DP-1 that acts as a dominant-negative regulator of cell cycle progression.