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

Abstract

Nuclear factor HI (NFI) 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 orign, adjacent to the nuclear factor I recognition site. DNA sequences sharing a high degree of homology to the NFm 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 NF[II 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 NlI 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 signifit 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 NFEL could function both in adenovirus DNA replication and in the transcriptional control of several groups of genes sharing the octanucleotide sequence.

Genomic organization of the KTX2 gene, encoding a ‘short’ scorpion toxin active on K+ channels

“…proximal upstream region was searched for putative cw-regulatory sequences: Octamer-1 and -4 (Oct-1 and Oct-4) binding sites [11] and a ssARS-T binding site [12] were found (Fig. 2).…”

Identification of a key regulatory element for the basal activity of the human insulin-like growth factor II gene promoter P3

“…EMSA binding conditions were determined as described [17]. Briefly, binding reactions were performed by mixing DNA probes, ds competitor DNA and binding buffer [20 mM Hepes\KOH (pH 7.6)\1 mM MgCl # \75 mM KCl\1 µg of poly(dI-dC) (Pharmacia, Uppsala, Sweden)\1 mM dithiothreitol\0.018 % Nonidet P40], after which protein extract was added.…”

“…DNase I footprinting reactions were performed as described previously [17]. The probe for DNase I footprinting contained an EcoRI\SalI P3 fragment from k290 to j134 labelled in the upper strand.…”

A protein involved in minichromosome maintenance in yeast binds a transcriptional enhancer conserved in eukaryotes.

“…Inspection of nucleotide sequences of ARSs reveals that there are multiple MCE-like elements in certain ARSs (V. Chang, unpubl.). Many examples of site-specific DNA-binding proteins playing dual roles in transcriptional activation and replication initiation in eukaryotes have been described (Dean et al 1987;Jones et al 1987;Mecas and Studgen 1987;Pruijn et al 1987;Stenlund et al 1987;O'Neill and Kelly 1988). A recent report by Santoro et al (1988) convincingly showed that a family of CCAAT-box binding proteins serves both as promoter-selective transcription activators and as initiation factors for DNA replication in mammalian in vitro systems.…”