Specific NEMO mutations impair CD40-mediated c-Rel activation and B cell terminal differentiation
Ashish Jain, … , Steven Holland, Jonathan M.J. Derry
Ashish Jain, … , Steven Holland, Jonathan M.J. Derry
Published December 1, 2004
Citation Information: J Clin Invest. 2004;114(11):1593-1602. https://doi.org/10.1172/JCI21345.
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Article Immunology

Specific NEMO mutations impair CD40-mediated c-Rel activation and B cell terminal differentiation

Abstract

Hypomorphic mutations in the zinc finger domain of NF-κB essential modulator (NEMO) cause X-linked hyper-IgM syndrome with ectodermal dysplasia (XHM-ED). Here we report that patient B cells are characterized by an absence of Ig somatic hypermutation (SHM) and defective class switch recombination (CSR) despite normal induction of activation-induced cytidine deaminase (AID) and Iε-Cε transcripts. This indicates that AID expression alone is insufficient to support neutralizing antibody responses. Furthermore, we show that patient B cells stimulated with CD40 ligand are impaired in both p65 and c-Rel activation, and whereas addition of IL-4 can enhance p65 activity, c-Rel activity remains deficient. This suggests that these NF-κB components have different activation requirements and that IL-4 can augment some but not all NEMO-dependent NF-κB signaling. Finally, using microarray analysis of patient B cells we identified downstream effects of impaired NF-κB activation and candidate factors that may be necessary for CSR and SHM in B cells.

Authors

Ashish Jain, Chi A. Ma, Eduardo Lopez-Granados, Gary Means, William Brady, Jordan S. Orange, Shuying Liu, Steven Holland, Jonathan M.J. Derry

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IL-4 “rescues” the transcriptional regulation of many genes in B cells o...
IL-4 “rescues” the transcriptional regulation of many genes in B cells of XHM-ED patients. B cells from XHM and XHM-ED patients were stimulated with CD40L alone or CD40L + IL-4 for 6 hours or 24 hours. Each row represents the ratio of expression in stimulated vs. unstimulated cells for each gene. Each column represents the data from an independent stimulation experiment. A color bar shows the magnitude of gene expression changes as a ratio of expression: significantly induced (red), unchanged (black), or repressed (green). (A) Genes regulated by CD40L stimulation in XHM. Inclusion in the analysis required at least a 3-fold change (6 hours vs. 0 hours, or 24 hours vs. 0 hours) with a P value < 0.01 for at least 3 comparisons in 2 different XHM patients in whom CD40 signaling is normal; 1,270 probe sets met this criteria. (B) Genes not regulated by CD40L stimulation in XHM-ED. Of the 1,270 probe sets that showed regulation in XHM B cells, 275 were identified that showed no significant regulation in the B cells from 2 independent XHM-ED patients in response to CD40 stimulation. (C) Genes induced with CD40L + IL-4 stimulation. The expression of the 275 probe sets that have impaired expression in XHM-ED B cells in response to CD40L stimulation alone were reclustered with data from XHM and XHM-ED B cells stimulated with CD40L and IL-4. Note the restored expression of many genes in XHM-ED patients in response to CD40L + IL-4 stimulation compared with CD40L alone. Gene identities can be seen in Supplemental Table 1.