Figure 2.

(A) Naïve B cells from XBP-1^WT, XBP-1^WT/MD4 and XBP-1^KO/MD4 mice were cultured in the presence of LPS for 4 days and then labeled with [³⁵S]-methionine/cysteine for 4 h. Triton X-114 lysis and separation were performed. Both the pellet and soluble fractions were precipitated with an antibody against μ, or with biotin or biotinylated-HEL. The pellet fraction containing μM is shown. (B-C) Cell lysates were prepared as in panel A. Lysates were split into 2 fractions, one of which was immunoprecipitated with an anti-μ antibody, and the other of which was subjected to five sequential rounds of precipitation with biotinylated-HEL, followed by a subsequent immunoprecipitation using the anti-μ antibody. For each cell type (XBP-1^WT/MD4 and XBP-1^KO/MD4), the amount of μ recovered from the first fraction by immunoprecipitation using the anti-μ antibody was designated as the total μ. By comparing to the total μ, the percentage of μ recovered from the second fraction was designated for each of the five sequential biotinylated-HEL recovery steps and for the final recovery with the anti-μ antibody. The quantitation of μS (B) was performed on fractions taken from the Triton X-114 supernatant, and the μM quantitation (C) was performed on the Triton X-114 pellet fractions. The results shown are an average of two separate experiments.