PKCδ deficiency in patients’ EBV-B cells. (A) RT-qPCR for PRKCD with a probe spanning the junction between exons 3 and 4 (left) and a probe spanning the junction between exons 17 and 18 (right), in EBV-B cells from healthy controls (Ctrl; n = 10) and patients. GUSB was used for normalization (n = 3; mean ± SD). (B) Western blot of total protein extracts from EBV-B cells from healthy controls (C1, C2) and patients (left). PKCδ was detected with a polyclonal anti-PKCδ antibody. The asterisk indicates nonspecific bands. Quantification of the PKCδ protein normalized against GAPDH (right; n = 3; mean ± SD). The results shown are representative of three independent experiments. (C) Western blot of total protein extract from the EBV-B cells of controls or patients. PKCδ was detected with a polyclonal anti-PKCδ antibody. The autophosphorylation of PKCδ was detected with antibodies against the T505 (upper panel) and S643 (lower panel) phosphorylation sites. The results shown are representative of three independent experiments. (D) Apoptosis of EBV-B cells from healthy controls (Ctrl; n = 10) and patients (n = 8) after 24 h of stimulation with APO-1-1 (APO; 1 µg/ml) or PMA (100 ng/ml). The percentages of live cells were normalized against the number of nonstimulated (NS) cells, after acquisition by flow cytometry over a constant time (n = 3; mean ± SD). A linear mixed model was used to determine whether survival rates were the same for PKCδ-deficient patients and controls after APO or PMA stimulation (n.s., not significant; ***, P < 0.001).