BCL6-mediated repression of PTPROt increases tonic BCR signaling. The DLBCL cell line, DHL4, was transduced with BCL6-siRNA, SCR control oligonucleotides, or left untreated. Cells were subsequently incubated for 72 hours and stimulated with goat anti–human IgG (10 μg/mL) for 1 minute or left untreated. (A) Western analysis of SYK phosphorylation after BCL6 depletion. BCR-cross-linked cells were lysed and immunoprecipitated with anti-SYK. Immunoprecipitates were size-fractionated, blotted, and analyzed thereafter with α-pSYK Y352 antibody. The membrane was subsequently stripped and blotted with an anti-pan SYK antibody. (B) Phospho-specific flow cytometric analysis of tonic and α-Ig induced SYK Y352 and BLNK Y84 phosphorylation after BCL6 depletion. SYK Y352 and BLNK Y84 phosphorylation (top and bottom panels) was compared in cells transduced with BCL6-siRNA (red), SCR control oligonucleotides (blue), or left untreated (green) in the absence (left panel) or presence (right panel) of BCR cross-linking. Cells stained with an isotype-matched control Ig are also shown (gray dashed line). The x-axis denotes expression (log scale) and the y-axis indicates cell number.