Article Figures & Data
Figures
- FIGURE 1.
Effect of TRAF5 status on TLR-mediated cytokine production by B lymphocytes. Splenic B cell isolation and detection of cytokine production were performed as described in Materials and Methods. (A) Splenic B cells were isolated from TRAF5 KO and TRAF5 WT mice and cultured with the indicated TLR ligands for 48 h. Cytokine production from TRAF5 KO B cells (▪) was compared with TRAF5 WT B cells (□). (B) CH12.LX B cells were cultured for 20 h with IPTG to induce overexpression of TRAF5. Cytokine production after 48 h of TLR stimulation from these cells (▪) was compared with CH12.LX cells that were not induced with IPTG (□). Data represent the mean values ± SD of three experiments. Statistical analysis was conducted using Student t test. *p < 0.05.
- FIGURE 2.
Effect of TRAF5 deficiency on TLR-mediated cytokine production by BMDC. BMDC (A) and BMM (B) differentiation was performed as described in Materials and Methods. BMDC and BMM derived from TRAF5 WT and TRAF5 KO mice were cultured with the indicated TLR ligands for 24 h. Cytokine production from TRAF5 KO BMDC or BMM (▪) was compared with TRAF5 WT BMDC or BMM (□). Data represent the mean values ± SD of three experiments. Statistical analysis was conducted using Student t test. *p < 0.05.
- FIGURE 3.
Effect of TRAF5 status on TLR-induced Ab production. Ab production was measured as described in Materials and Methods. (A) CH12.LX B cells were cultured for 20 h with IPTG to induce overexpression of TRAF5 (CH12–TRAF5). IgM production after TLR ligation from these cells (▪) was compared with CH12.LX cells that were not induced with IPTG (□). (B) Splenic B cells were isolated from TRAF5 KO and TRAF5 WT mice and cultured with the indicated TLR ligands for 5 d. IgM production from TRAF5 KO B cells (▪) was compared with TRAF5 WT B cells (□). Data represent the mean values ± SD of three experiments. Statistical analysis was conducted using Student t test. *p < 0.05.
- FIGURE 4.
TLR-induced MAPK activation in TRAF5-deficient B lymphocytes. Splenic B cells were isolated from TRAF5 KO and WT mice as described in Materials and Methods and stimulated with TLR7 agonist R848 for the indicated times. Preparation of whole-cell lysates and Western blotting were as described in Materials and Methods. Western blots shown are representative of three experiments, whereas the quantitation provided combines three separate experiments.
- FIGURE 5.
TLR-induced NF-κB activation in TRAF5-deficient B lymphocytes. Splenic B cells were isolated from TRAF5 KO and WT mice as described in Materials and Methods and stimulated with TLR7 agonist R848 for the indicated times. Preparation of whole-cell lysates and Western blotting were as described in Materials and Methods. Western blots shown are representative of three experiments, whereas the quantitation provided combines three separate experiments.
- FIGURE 6.
TLR-induced association of TRAF5 with TLR signaling proteins. CH12.LX B cells (A, B, D) were cultured for 20 h with IPTG to induce expression of FLAG-tagged TRAF5. Splenic B cell isolation was performed as described in Materials and Methods (C). Cells were stimulated with TLR7 agonist R848 for the indicated times. Immunoprecipitation of FLAG (A), MyD88 (B), and TRAF6 (C, D) and Western blotting were performed as described in Materials and Methods. Quantitation in (C) and (D) is shown below the blots as percent of maximum association.
Additional Files
Data Supplement
Files in this Data Supplement:
- Supplemental Figures 1-3 (PDF, 3,079 Kb) -
Description:
Figure S1. Effect of TRAF5 deficiency on B cell proliferation, survival, and cytokine production.
Figure S2. Effect of TRAF5 deficiency on B cell subset cytokine production.
Figure S3. Effect of TRAF5 status on the marginal zone B cell compartment.
- Supplemental Figures 1-3 (PDF, 3,079 Kb) -
Description:
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