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FOXM1 binds directly to non-consensus sequences in the human genome.



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Sanders, Deborah A 
Gormally, Michael V 
Marsico, Giovanni 
Beraldi, Dario 


BACKGROUND: The Forkhead (FKH) transcription factor FOXM1 is a key regulator of the cell cycle and is overexpressed in most types of cancer. FOXM1, similar to other FKH factors, binds to a canonical FKH motif in vitro. However, genome-wide mapping studies in different cell lines have shown a lack of enrichment of the FKH motif, suggesting an alternative mode of chromatin recruitment. We have investigated the role of direct versus indirect DNA binding in FOXM1 recruitment by performing ChIP-seq with wild-type and DNA binding deficient FOXM1. RESULTS: An in vitro fluorescence polarization assay identified point mutations in the DNA binding domain of FOXM1 that inhibit binding to a FKH consensus sequence. Cell lines expressing either wild-type or DNA binding deficient GFP-tagged FOXM1 were used for genome-wide mapping studies comparing the distribution of the DNA binding deficient protein to the wild-type. This shows that interaction of the FOXM1 DNA binding domain with target DNA is essential for recruitment. Moreover, analysis of the protein interactome of wild-type versus DNA binding deficient FOXM1 shows that the reduced recruitment is not due to inhibition of protein-protein interactions. CONCLUSIONS: A functional DNA binding domain is essential for FOXM1 chromatin recruitment. Even in FOXM1 mutants with almost complete loss of binding, the protein-protein interactions and pattern of phosphorylation are largely unaffected. These results strongly support a model whereby FOXM1 is specifically recruited to chromatin through co-factor interactions by binding directly to non-canonical DNA sequences.



Base Sequence, Binding Sites, Cell Line, Chromatin Immunoprecipitation, Consensus Sequence, DNA, Forkhead Box Protein M1, Forkhead Transcription Factors, Gene Expression Regulation, Genome, Human, HEK293 Cells, HeLa Cells, Humans, Mutation, Promoter Regions, Genetic, Protein Binding, Protein Structure, Tertiary, Proteomics, Sequence Analysis, DNA

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Genome Biol

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Springer Science and Business Media LLC
Cancer Research UK (CB4330)
Cancer Research UK (C14303/A17197)
We would like to acknowledge the Genomics and bioinformatics core at the CRUK Research Institute for the Illumina sequencing and the Proteomics core for the LC/MS-MS protein analysis for the RIME experiments. We acknowledge the support from The University of Cambridge and Cancer Research UK. The Balasubramanian Laboratory is supported by core funding from Cancer Research UK (C14303/A17197). SB is a Wellcome Trust Principle Investigator.