The Cd19 promoter binds PAX5 in B cells in vivo. DMS in vivo footprinting assays were performed at the Cd19 promoter in purified primary cells (A) and pro-B cell lines (B). Early pro-B, late pro-B, pre-B, and mature B cells were purified from bone marrow (Figure S3). G indicates in vitro DMS-treated DNA; MEF, mouse embryonic fibroblast; thy-T, THY1⁺ thymocytes; WT, wild-type pro-B cells; −/−, Pax5⁻ pro-B cells. Numbers on the left indicate the positions relative to the ATG. A vertical line on the right is indicative of a high affinity PAX5 binding site. ● and ○ represent guanine residues hyperreactive or hyporeactive to DMS, respectively; gray circles, partial footprints. (C) DNA sequence of the mouse Cd19 promoter. * Nucleotide sequences homologous to the human CD19 locus. ● and ○ represent nucleotides occupied by transcription factors as assayed by in vivo footprinting. The consensus PAX5 recognition sequence is aligned above the promoter sequence. L-shaped arrows are major transcription start sites as determined by reverse transcription transferase-dependent PCR (RT-TDPCR; data not shown). (D) Recruitment of PAX5 in primary cells. CD19⁺ bone marrow cells (BM-B), CD19⁺ spleenocytes (splenic B), and Thy-1⁺ thymocytes (thymic T) cells were purified, crosslinked, and subjected to ChIP assay using a polyclonal antibody against PAX5. Precipitated DNA was amplified with primers specific for the Cd19 promoter, +10 kb regions, and 45S rRna promoter⁵⁰  (control). Each bar represents the relative enrichment compared with the control region.