Hierarchical binding of DNA fragments derived from scaffold-attached regions: correlation of properties in vitro and function in vivo.
On its upstream side, the human interferon-beta gene is flanked by a 7-kb SAR (scaffold-attached region) DNA element. The core of this element is determined and subjected to in vitro reassociations with isolated scaffolds. Binding properties of SAR fragments with decreasing length are quantified and related to consensus sequences like the topoisomerase II box and an ATATTT motif. Characteristics as the stoichiometry, affinity, and cooperativity of the binding process are shown to depend on the length of SAR DNA and suggest a model involving a multiple-site attachment to protein scaffolds. We propose a rational approach for predicting the SAR mediated transcriptional enhancements in vivo from their binding properties in a standardized in vitro assay. The efficiency of this approach is demonstrated for a marker (huIFN-beta) and a selector gene (neor).
Mielke
C
C
GBF, Gesellschaft für Biotechnologische Forschung, mbH, Genetik von Eukaryonten, Braunschweig-Stöckheim, West Germany.
Kohwi
Y
Y
Kohwi-Shigematsu
T
T
Bode
J
J
D000818:Animals; D001482:Base Composition; D001483:Base Sequence; D002478:Cells, Cultured; D004247:DNA / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics* / Q000378:metabolism; D005786:Gene Expression Regulation; D005819:Genetic Markers; D006801:Humans; D007370:Interferon Type I / Q000235:genetics* / Q000378:metabolism; D051379:Mice; D008969:Molecular Sequence Data; D009154:Mutation; D012045:Regulatory Sequences, Nucleic Acid; D014158:Transcription, Genetic
D004268:DNA-Binding Proteins; D005819:Genetic Markers; D007370:Interferon Type I; D004247:DNA