B-cell lymphomas in Crebbp transgenic mice are of GCB-cell origin. (A) An example of immunohistochemical staining for the spleen of an age-matched control mouse (Cbp^WT/F) and a lymphoma-involved spleen from a Cbp^Δ/ΔxEµBcl2 mouse. The control mouse shows normal benign follicles and an expected pattern of Pax5 and Bcl6 staining. The lymphoma-involved spleen shows diffuse histology with cells that are Pax5 and Bcl6 positive, supporting a GCB-cell origin. (B) Immunoglobulin rearrangements were assessed by PCR in DNA extracted from tumor-involved spleens and a spleen from age-matched control mice (Cbp^F/FxEµBcl2). Control mice showed a laddering pattern indicative of a polyclonal B-cell population, as shown in the left-most lane. In contrast, the majority of tumor samples from Cbp^WT/Δ, Cbp^Δ/Δ, Cbp^WT/ΔxEµBcl2, and Cbp^Δ/ΔxEµBcl2 mice showed a single dominant band that is indicative of a clonal B-cell population, as seen in these examples. Eight of these bands, highlighted in red boxes, were excised and cloned for sequencing. Analysis revealed the presence of somatic hypermutation in 7/8 tumors, with an average of 2% deviation (min = 0.8%, max = 4.2%) deviation from the germ-line V-gene sequence. This provides further evidence in support of the GCB-cell origin of these tumors, or that the B cells have previously transited through the germinal center. NA, not applicable; SHM, somatic hypermutation.