Figure 4. Defective immunoproteasome complexes in JASL cells.

(A) PSMB8 mRNA expression in transformed B cells from 1-1M, 1-2ES, and 1-3PT were evaluated by real-time PCR. Data are representative of 4 experiments. (B) Expression of PSMB8 and β-actin in EBV-transformed B cells from 1-1M, 1-2ES, 1-3PT, and an unrelated control, evaluated by Western blotting. (C) Cell lysates from transformed B cells of 1-1M (lane 1), 1-2ES (lane 2), 1-3PT (lane 3), and unrelated healthy control (lane 4) were subjected to native PAGE and immunoblotted with anti-Ump1, anti-α6, and anti-β1i polyclonal antibodies. Arrows denote bands corresponding to the 20S or 26S proteasome, as well as intermediate complexes. Asterisks denote nonspecific bands. (D) Glycerol gradient centrifugation was performed by using extracts of transformed B cells from 1-3PT and an unrelated healthy control. Fractions were immunoblotted with anti-α6, anti-β1i, and anti-PSMB8 antibody. Arrowheads denote assembly intermediates as well as 20S and 26S immunoproteasomes. PSMB8 genotypes are shown. All data are representative of at least 4 experiments.