title: High-affinity binding of tumor-suppressor protein p53 and HMGB1 to hemicatenated DNA loops.
abstract: We have recently observed that chromatin architectural protein HMGB1 (previously reported to be involved in numerous biological processes such as DNA replication, recombination, repair, tumor growth, and metastasis) could bind with extremely high affinity (K(d) < 1 pM) to a novel DNA structure that forms a DNA loop maintained at its base by a hemicatenane (hcDNA). The loop of hcDNA contains a track of repetitive sequences derived from CA-microsatellites. Here, we report using a gel-retardation assay that tumor-suppressor protein p53 can also bind to hcDNA. p53 is a crucial molecule protecting cells from malignant transformation by regulating cell-cycle progression, apoptosis, and DNA repair by activation or repression of transcription of its target genes by binding to specific p53 DNA-binding sites and/or certain types of DNA lesions or alternative DNA structures. The affinity of p53 for hcDNA (containing sequences with no resemblance to the p53 DNA consensus sequence) is >40-fold higher (K(d) approximately 0.5 nM) than that for its natural specific binding sites within its target genes (Mdm2 promoter). Binding of p53 to hcDNA remains detectable in the presence of up to approximately 4 orders of magnitude of mass excess of competitor linear DNA, suggesting a high specificity of the interaction. p53 displays a higher affinity for hcDNA than for DNA minicircles (lacking functional p53-specific binding sequence) with a size similar to that of the loop within the hcDNA, indicating that the extreme affinity of p53 for hcDNA is likely due to the binding of the protein to the hemicatenane. Although binding of p53 to hcDNA occurs in the absence of the nonspecific DNA-binding extreme carboxy-terminal regulatory domain (30-C, residues 363-393), the isolated 30-C domain (but not the sequence-specific p53 "core domain", residues 94-312) can also bind hcDNA. Only the full-length p53 can form stable ternary complexes with hcDNA and HMGB1. The possible biological relevance of p53 and HMGB1 binding to hemicatenanes is discussed.
authors.affiliation: Laboratory of Analysis of Chromosomal Proteins, Academy of Sciences of the Czech Republic, Institute of Biophysics, 612 65 Brno, Czech Republic. stros@ibp.cz
authors.forename: Michal
authors.initials: M
authors.lastname: Stros
authors.forename: Eva
authors.initials: E
authors.lastname: Muselíková-Polanská
authors.forename: Sárka
authors.initials: S
authors.lastname: Pospísilová
authors.forename: François
authors.initials: F
authors.lastname: Strauss
mesh_terms: D000818:Animals; D001665:Binding Sites; D002417:Cattle; D043864:DNA, Catenated / Q000737:chemistry* / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D005982:Glutathione Transferase / Q000235:genetics / Q000378:metabolism; D024243:HMGB1 Protein / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D009690:Nucleic Acid Conformation; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011485:Protein Binding / Q000502:physiology; D017434:Protein Structure, Tertiary / Q000502:physiology; D051381:Rats; D013379:Substrate Specificity; D013950:Thymus Gland / Q000166:cytology; D015533:Transcriptional Activation; D016159:Tumor Suppressor Protein p53 / Q000737:chemistry* / Q000378:metabolism; D025521:Tumor Suppressor Proteins / Q000378:metabolism
chemical_list: D043864:DNA, Catenated; D024243:HMGB1 Protein; D016159:Tumor Suppressor Protein p53; D025521:Tumor Suppressor Proteins; D005982:Glutathione Transferase