A binding site for the transcription factor grainyhead/nuclear transcription factor-1 contributes to regulation of the Drosophila proliferating cell nuclear antigen gene promoter

The Drosophila proliferating cell nuclear antigen promoter contains multipl e transcriptional regulatory elements, including upstream regulatory elemen t (URE), DNA replication-related element, E2F recognition sites, and three common regulatory factor for DNA replication and DNA replication-related el ement-binding factor genes recognition sites. In nuclear extracts of Drosop hila embryos, we detected a protein factor, the URE-binding factor (UREF), that recognizes the nucleotide sequence 5'-AAACCAGTTGGCA located within URE , Analyses in Drosophila Kc cells and transgenic flies revealed that the UR EF-binding site plays an important role in promoter activity both in cultur ed cells and in living flies. A yeast one-hybrid screen using URE as a bait allowed isolation of a cDNA encoding a transcription factor, Grainyhead/nu clear transcription factor-1 (GRH/NTF-1). The nucleotide sequence required for binding to GRH was indistinguishable from that for UREF detected in emb ryo nuclear extracts, Furthermore, a specific antibody to GRH reacted with UREF in embryo nuclear extracts, From these results we conclude that GRH is identical to UREF, Although GRH has been thought to be involved in regulat ion of differentiation-related genes, this study demonstrates, for the firs t time, involvement of a GRH-binding site in regulation of the DNA replicat ion-related proliferating cell nuclear antigen gene.