Jerković, IvanaIbrahim, Daniel MuradAndrey, GuillaumeHaas, Stefan A.Hansen, PeterJanetzki, CatrinGonzález Navarrete, Irene, 1983-Robinson, Peter N.Hecht, JochenMundlos, Stefan2018-07-042018-07-042017Jerković I, Ibrahim DM, Andrey G, Haas S, Hansen P, Janetzki C et al. Genome-Wide Binding of Posterior HOXA/D Transcription Factors Reveals Subgrouping and Association with CTCF. PLoS Genet. 2017 Jan 19;13(1):e1006567. DOI: 10.1371/journal.pgen.10065671553-7390http://hdl.handle.net/10230/35027Homeotic 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.application/pdfengThis is an open access article, free of all copyright, and may be freely reproduced, distributed, transmitted, modified, built upon, or otherwise used by anyone for any lawful purpose. The work is made available under the Creative Commons CC0 public domain dedication (http://creativecommons.org/publicdomain/zero/1.0/).Genome-wide binding of posterior HOXA/D transcription factors reveals subgrouping and association with CTCFinfo:eu-repo/semantics/articlehttp://dx.doi.org/10.1371/journal.pgen.1006567Gene expression regulationHomeoboxBinding analysisCell differentiationSequence motif analysisProtein bindingChondrogenesisChromatinMesoderminfo:eu-repo/semantics/openAccess