Fig. 3

Hox cofactors Exd and Hth alter the binding profile of Ubx. a The pMT-Hth2AGFPUbx bicistronic expression vector used to co-express Hth and Ubx-GFP in Kc167 cells. The construct contains the Drosophila metallothionein (MT) promoter, Hth cDNA isoform A, Thosea asigna 2A self-cleaving peptide (T2A), enhanced Green Fluorescent Protein (eGFP) fused to Ubx cDNA isoform E, C-terminal peptide (containing V5 and polyhistidine tags and SV40 polyadenylation signal), and an ampicillin resistance gene. b Exd immunolabelling (red) was used to confirm that transfection of Kc167 cells with pMT-Hth2AGFPUbx results in the expression of functional Hth with Hth-dependant recruitment of Exd into the nucleus. Left: In non-transfected cells (Hth−), Exd is excluded from the nucleus (arrowhead). Middle: In transfected cells (Hth+), Hth induces nuclear accumulation of Exd protein (arrowhead). Right: Same as ‘Middle’ but also showing Ubx-GFP (green). Hth +/− cells were separated by FACS following transfection (Additional file 1: Figure S1). c Comparison of binding profiles of Ubx and Ubx in the presence of Hth (Experiment 2). Examples of cofactor-dependent binding are highlighted in grey. d Venn diagram showing the overlap of binding peaks (q-value 1e–10) between Ubx and Ubx + Hth. 51 % of the Ubx + Hth peaks are novel. e Venn diagram showing the overlap of binding peaks between Ubx + Hth and DNase1 (q-value 1e–10 for Ubx + Hth and 1e–2 for DNase1). 17 % of the Ubx + Hth peaks are in DNase1 inaccessible chromatin