The mouse Myc gene contains multiple EBF1-responsive enhancer elements. (A) UCSC Genome Browser view of the murine Myc locus and its distal interacting regions. The tracks display PAX5 (GSE126375) and EBF1 binding in 230-238 progenitor B cells, ATAC-accessibility (GSE92434), as well as a PLAC-sequencing derived virtual 4C tracks from Wt and Ebf1^−/− FL-derived pro-B cells. Myc transcriptional start site ±2.5 kb was used as the viewpoint for the virtual 4C analysis. The previously defined BENC enhancer region²⁹  is indicated by a dashed square. (B) Zoomed-in view of 3 specific regions in panel A with high PAX5 and EBF1 binding as well as ATAC accessibility in Wt pro-B cells and interaction with the Myc promoter. These regions were examined for the presence of histone modifications by reanalysis of ChIP- and CUT&RUN-sequencing data (GSE162858). The gray lines indicate the regions that are targeted for luciferase reporter activity assays. (C) Relative Firefly/Renilla (Prl0) luciferase activity obtained from reporter constructs in which the EBF1-binding regions described in panel B were cloned upstream of a basal Fos promoter in the absence (empty cDNA3) or presence of EBF1 in HeLa cells. Each dot represents one transfection, and the statistical analysis is based on a Student unpaired t test. **P < .01; ***P < .001; ****P < .0001.