E2F mediates induction of the Sp1-controlled promoter of the human DNA polymerase varepsilon B-subunit gene POLE2

Abstract

The B-subunits of replicative DNA polymerases from Archaea to humans belong to the same protein family, suggesting that they share a common fundamental function. We report here the gene structure for the B-subunit of human DNA polymerase epsilon (POLE2), whose expression and transcriptional regulation is typical for replication proteins with some unique features. The 75 bp core promoter region, located within exon 1, contains an Sp1 element that is a critical determinant of promoter activity as shown by the luciferase reporter, electrophoretic mobility shift and DNase I footprinting assays. Two overlapping E2F elements adjacent to the Sp1 element are essential for full promoter activity and serum response. Binding sites for E2F1 and NF-1 reside immediately downstream from the core promoter region. Our results suggest that human POLE2 is regulated by two E2F-pocket protein complexes, one associated with Sp1 and the other with NF-1. So far, only one replicative DNA polymerase B-subunit gene promoter, POLA2 encoding the B-subunit of DNA polymerase alpha, has been characterized. Mitogenic activation of the POLE2 promoter by an E2F-mediated mechanism resembles that of POLA2, but the regulation of basal promoter activity is different between these two genes.

Genetic analysis of two Arabidopsis DNA polymerase epsilon subunits during early embryogenesis

“…In yeast and human it has been shown that some DNA polymerase ɛ subunits are transcriptionally regulated at the G 1 /S phase transition. For instance, POLE2 is regulated by two E2F–pocket protein complexes (Huang et al. , 2001).…”

Genome-Wide In Silico Identification of Transcriptional Regulators Controlling the Cell Cycle in Human Cells

“…Full lists of genes whose promoters were found to contain high scoring sites for the enriched TFs are provided in Supplemental Tables B1-B8 (available online at www.genome.org.) Chang et al 2001;Huang et al 2001;Nishikawa et al 2001;Parisi et al 2002). As for E2F and NF-Y, co-occurrence of functional binding sites for both TFs was reported in several promoters, including Cdc2, TK, POLA, Cyclin A, and several histone genes (Matuoka and Yu Chen 1999).…”

Regulation of transcription of the human MRP7 gene

“…The contribution of these Sp1C and D to overall promoter activity is minimal in the presence of the interacting E2F and Sp1A and B sites. E2F-Sp1 interactions important in regulating transcription have also been reported (Lin et al, 1996;Rotheneder et al, 1999;Huang et al, 2001) for other promoters. Moreover, similar to that demonstrated in the case of the cdc25A gene (Chen and Prywes, 1999), on the basis of the supershift obtained in an EMSA using antibody directed against the E2F pocket protein, p107, we were able to more definitively conclude that an interaction between an E2F factor and a noncanonical E2F sequence element occurs in the MRP7 promoter.…”

“…Annealing of ss oligomers and electrophoresis conditions were as reported elsewhere (Huang et al, 2001). Ds oligomers were 32 P-end-labeled using the Gel Shift Assay Systems kit from Promega.…”