Promoter and enhancer elements containing a conserved sequence motif are recognized by nuclear factor III, a protein stimulating adenovirus DNA replication

Abstract

Nuclear factor III (NFIII) is a protein from HeLa cells that stimulates the initiation of adenovirus type 2 (Ad2) DNA replication by binding to a specific nucleotide sequence in the origin, adjacent to the nuclear factor I recognition site. DNA sequences sharing a high degree of homology to the NFIII binding site in Ad2 were found in a number of transcription regulatory elements, all containing the octanucleotide sequence ATGCAAAT. We have analysed the interaction between NFIII and the octamer-containing sequences in a histone H2B promoter, immunoglobulin light and heavy chain promoters, an immunoglobulin heavy chain enhancer, a U2 snRNA enhancer and the SV40 enhancer as well as Ad4. All sequences were recognized by NFIII as indicated by gel retardation assays, DNase I footprinting and methylation protection experiments. A comparison of the relative binding affinities using competition assays indicated that mutations in the octanucleotide sequence reduced the binding affinity considerably. Small but significant differences in affinity were also observed depending on the sequences bordering the conserved octanucleotide. The methylation protection patterns indicate that both major and minor groove contacts are involved in NFIII binding. The data suggest that NFIII could function both in adenovirus DNA replication and in the transcriptional control of several groups of genes sharing the octanucleotide sequence.