In vivo structure of the cell cycle-regulated human cdc25C promoter.
The cdc25C promoter is regulated during the cell cycle by the transcriptional repressor CDF-1 that inhibits the activation function of upstream transcriptional activators, most notably the nuclear factor Y/CAAT box binding factor (NF-Y/CBF). In this report a detailed analysis of the in vivo structure of the cdc25C promoter was made. Micrococcus nuclease and methidiumpropyl-EDTA footprinting strongly suggest that the proximal promoter encompassing the cell cycle-dependent element/cell cycle genes homology region and the upstream NF-Y sites is organized in a positioned nucleosome throughout the cell cycle. Furthermore, structural perturbations were detected by DNase I, phenanthroline copper, and KMnO(4) footprinting at the NF-Y binding sites in vivo, which is in agreement with the reported property of NF-Y to bend DNA in vitro. Similar results were obtained with the structurally and functionally related cyclin A promoter. The structural perturbations seen in DNase I and phenanthroline copper footprints were less pronounced in G(0) cells when compared with cycling cells. This presumably reflects a weakened in vivo interaction of NF-Y with its cognate DNA element in G(0). It is likely that these structural perturbations, together with the reported ability of NF-Y to recruit histone acetyl transferase activity, contribute to an opened chromatin structure as a prerequisite for optimal regulation through activation and repression.
Körner
K
K
Institute of Molecular Biology and Tumor Research, Philipps-University Marburg, Emil-Mannkopff-Strasse 2, D-35033 Marburg, Germany.
Müller
R
R
D001483:Base Sequence; D002453:Cell Cycle / Q000502:physiology*; D018797:Cell Cycle Proteins / Q000235:genetics*; D002460:Cell Line; D019925:Cyclin A / Q000235:genetics; D018983:DNA Footprinting; D017931:DNA Primers; D006801:Humans; D008836:Micrococcal Nuclease / Q000378:metabolism; D011196:Potassium Permanganate; D011401:Promoter Regions, Genetic / Q000502:physiology*; D020687:cdc25 Phosphatases / Q000235:genetics*
D018797:Cell Cycle Proteins; D019925:Cyclin A; D017931:DNA Primers; D011196:Potassium Permanganate; C517009:CDC25C protein, human; D020687:cdc25 Phosphatases; D008836:Micrococcal Nuclease