Fig. 4: AID-downregulated human CLL cells upregulate the ER stress response and primary IgHV-unmutated human CLL cells express higher levels of XBP1s mRNA; loss of secretory IgM leads to downregulation of XBP1s production in CLL cells.

A WaC3 human CLL cells were not transduced, transduced with a non-targeting control shRNA, or transduced with an AID-targeting shRNA, and lysates were immunoblotted for the indicated proteins. B Control shRNA- or AID-targeting shRNA-transduced WaC3 cells were treated with 10 μg/mL tunicamycin for the indicated times, and lysates were immunoblotted for the indicated proteins. C Control shRNA- or AID-targeting shRNA-transduced WaC3 cells were treated with indicated concentrations of B-I09 for 3 days and subjected to XTT assays. Percentage growth was determined by comparing cells treated with B-I09 with those that were untreated and are shown as the means ± SD. D IC₅₀ values from three independent XTT experiments comparing control shRNA- or AID-targeting shRNA-transduced WaC3 cells treated with B-I09 for 3 days. E Primary human CLL cells purified from the peripheral blood of patients with IgHV-mutated (n = 17) and IgHV-unmutated (n = 17) CLL were lysed for purification of RNA. The mRNA levels of XBP1s were measured by quantitative RT-PCR and data were normalized to HPRT1 (a house-keeping gene). F B cells purified from the spleens of AID^−/− and AID^−/−/μS^−/− mice were stimulated with 20 μg/mL LPS for 3 days and lysates were immunoblotted for the indicated proteins. G CLL cells purified from the spleens of Eμ-TCL1 and μS^−/−/Eμ-TCL1 mice were stimulated with 20 μg/mL LPS for 3 days and lysates were immunoblotted for the indicated proteins. H CLL cells purified from the spleens of 4 μS^−/−/Eμ-TCL1, 4 Eμ-TCL1, and 4 AID^−/−/Eμ-TCL1 mice were directly lysed and immunoblotted for the indicated proteins.