Dataset Information

HMGB binding to DNA: single and double box motifs.

ABSTRACT: High mobility group (HMG) proteins are nuclear proteins believed to significantly affect DNA interactions by altering nucleic acid flexibility. Group B (HMGB) proteins contain HMG box domains known to bind to the DNA minor groove without sequence specificity, slightly intercalating base pairs and inducing a strong bend in the DNA helical axis. A dual-beam optical tweezers system is used to extend double-stranded DNA (dsDNA) in the absence as well as presence of a single box derivative of human HMGB2 [HMGB2(box A)] and a double box derivative of rat HMGB1 [HMGB1(box A+box B)]. The single box domain is observed to reduce the persistence length of the double helix, generating sharp DNA bends with an average bending angle of 99+/-9 degrees and, at very high concentrations, stabilizing dsDNA against denaturation. The double box protein contains two consecutive HMG box domains joined by a flexible tether. This protein also reduces the DNA persistence length, induces an average bending angle of 77+/-7 degrees , and stabilizes dsDNA at significantly lower concentrations. These results suggest that single and double box proteins increase DNA flexibility and stability, albeit both effects are achieved at much lower protein concentrations for the double box. In addition, at low concentrations, the single box protein can alter DNA flexibility without stabilizing dsDNA, whereas stabilization at higher concentrations is likely achieved through a cooperative binding mode.

SUBMITTER: McCauley MJ

PROVIDER: S-EPMC2117627 | biostudies-literature | 2007 Dec

REPOSITORIES: biostudies-literature

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HMGB binding to DNA: single and double box motifs.

McCauley Micah J MJ Zimmerman Jeff J Maher L James LJ Williams Mark C MC

Journal of molecular biology 20070929 4

High mobility group (HMG) proteins are nuclear proteins believed to significantly affect DNA interactions by altering nucleic acid flexibility. Group B (HMGB) proteins contain HMG box domains known to bind to the DNA minor groove without sequence specificity, slightly intercalating base pairs and inducing a strong bend in the DNA helical axis. A dual-beam optical tweezers system is used to extend double-stranded DNA (dsDNA) in the absence as well as presence of a single box derivative of human H ...[more]

PMID: 17964600

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Project description:BackgroundNeuroblastoma, a cancer derived from precursor cells of the sympathetic nervous system, is a major cause of childhood cancer related deaths. The single most important prognostic indicator of poor clinical outcome in this disease is genomic amplification of MYCN, a member of a family of oncogenic transcription factors.MethodologyWe applied MYCN chromatin immunoprecipitation to microarrays (ChIP-chip) using MYCN amplified/non-amplified cell lines as well as a conditional knockdown cell line to determine the distribution of MYCN binding sites within all annotated promoter regions.ConclusionAssessment of E-box usage within consistently positive MYCN binding sites revealed a predominance for the CATGTG motif (p<0.0016), with significant enrichment of additional motifs CATTTG, CATCTG, CAACTG in the MYCN amplified state. For cell lines over-expressing MYCN, gene ontology analysis revealed enrichment for the binding of MYCN at promoter regions of numerous molecular functional groups including DNA helicases and mRNA transcriptional regulation. In order to evaluate MYCN binding with respect to other genomic features, we determined the methylation status of all annotated CpG islands and promoter sequences using methylated DNA immunoprecipitation (MeDIP). The integration of MYCN ChIP-chip and MeDIP data revealed a highly significant positive correlation between MYCN binding and DNA hypermethylation. This association was also detected in regions of hemizygous loss, indicating that the observed association occurs on the same homologue. In summary, these findings suggest that MYCN binding occurs more commonly at CATGTG as opposed to the classic CACGTG E-box motif, and that disease associated over expression of MYCN leads to aberrant binding to additional weaker affinity E-box motifs in neuroblastoma. The co-localization of MYCN binding and DNA hypermethylation further supports the dual role of MYCN, namely that of a classical transcription factor affecting the activity of individual genes, and that of a mediator of global chromatin structure.

Dataset Information

HMGB binding to DNA: single and double box motifs.

Publications

HMGB binding to DNA: single and double box motifs.

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