title: Genome-Wide Binding of Posterior HOXA/D Transcription Factors Reveals Subgrouping and Association with CTCF.
abstract: Homeotic genes code for key transcription factors (HOX-TFs) that pattern the animal body plan. During embryonic development, Hox genes are expressed in overlapping patterns and function in a partially redundant manner. In vitro biochemical screens probing the HOX-TF sequence specificity revealed largely overlapping sequence preferences, indicating that co-factors might modulate the biological function of HOX-TFs. However, due to their overlapping expression pattern, high protein homology, and insufficiently specific antibodies, little is known about their genome-wide binding preferences. In order to overcome this problem, we virally expressed tagged versions of limb-expressed posterior HOX genes (HOXA9-13, and HOXD9-13) in primary chicken mesenchymal limb progenitor cells (micromass). We determined the effect of each HOX-TF on cellular differentiation (chondrogenesis) and gene expression and found that groups of HOX-TFs induce distinct regulatory programs. We used ChIP-seq to determine their individual genome-wide binding profiles and identified between 12,721 and 28,572 binding sites for each of the nine HOX-TFs. Principal Component Analysis (PCA) of binding profiles revealed that the HOX-TFs are clustered in two subgroups (Group 1: HOXA/D9, HOXA/D10, HOXD12, and HOXA13 and Group 2: HOXA/D11 and HOXD13), which are characterized by differences in their sequence specificity and by the presence of cofactor motifs. Specifically, we identified CTCF binding sites in Group 1, indicating that this subgroup of HOX-proteins cooperates with CTCF. We confirmed this interaction by an independent biological assay (Proximity Ligation Assay) and demonstrated that CTCF is a novel HOX cofactor that specifically associates with Group 1 HOX-TFs, pointing towards a possible interplay between HOX-TFs and chromatin architecture.
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, Berlin, Germany.|Berlin-Brandenburg School for Regenerative Therapies, Berlin, Germany.
authors.forename: Ivana
authors.identifier: 0000-0001-9707-8303
authors.initials: I
authors.lastname: Jerković
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, Berlin, Germany.|Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, Berlin, Germany.|Berlin Institute of Health, Berlin, Germany.
authors.forename: Daniel M
authors.identifier: 0000-0002-0019-3354
authors.initials: DM
authors.lastname: Ibrahim
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, Berlin, Germany.
authors.forename: Guillaume
authors.identifier: 0000-0002-0911-4907
authors.initials: G
authors.lastname: Andrey
authors.affiliation: Department of Computational Molecular Biology, Max-Planck-Institute for Molecular Genetics, Berlin, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Haas
authors.affiliation: Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, Berlin, Germany.|Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Peter
authors.identifier: 0000-0001-5535-2845
authors.initials: P
authors.lastname: Hansen
authors.affiliation: Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Catrin
authors.initials: C
authors.lastname: Janetzki
authors.affiliation: Genomics Unit, Centre for Genomic Regulation, Barcelona, Spain.|Universitat Pompeu Fabra (UPF), Barcelona, Spain.
authors.forename: Irene
authors.initials: I
authors.lastname: González Navarrete
authors.affiliation: Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, Berlin, Germany.|Department of Computational Molecular Biology, Max-Planck-Institute for Molecular Genetics, Berlin, Germany.|Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Peter N
authors.identifier: 0000-0002-0736-9199
authors.initials: PN
authors.lastname: Robinson
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, Berlin, Germany.|Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, Berlin, Germany.|Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Berlin, Germany.|Genomics Unit, Centre for Genomic Regulation, Barcelona, Spain.|Universitat Pompeu Fabra (UPF), Barcelona, Spain.
authors.forename: Jochen
authors.identifier: 0000-0002-6996-5279
authors.initials: J
authors.lastname: Hecht
authors.affiliation: Max Planck Institute for Molecular Genetics, RG Development & Disease, Berlin, Germany.|Institute for Medical and Human Genetics, Charité Universitätsmedizin Berlin, Berlin, Germany.|Berlin Institute of Health, Berlin, Germany.|Berlin-Brandenburg Center for Regenerative Therapies (BCRT), Charité Universitätsmedizin Berlin, Berlin, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Mundlos
mesh_terms: D000818:Animals; D000076246:CCCTC-Binding Factor; D002645:Chickens; D020219:Chondrogenesis; D002843:Chromatin / Q000378:metabolism; D018507:Gene Expression Regulation, Developmental; D016678:Genome; D018398:Homeodomain Proteins / Q000378:metabolism*; D008648:Mesoderm / Q000378:metabolism; D011485:Protein Binding; D012097:Repressor Proteins / Q000378:metabolism*; D015533:Transcriptional Activation
chemical_list: D000076246:CCCTC-Binding Factor; D002843:Chromatin; D018398:Homeodomain Proteins; D012097:Repressor Proteins