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. Author manuscript; available in PMC: 2017 Jun 1.
Published in final edited form as: Mol Immunol. 2016 Apr 22;74:10–17. doi: 10.1016/j.molimm.2016.04.002

Figure 2. ERAP1 knockdown increases HLA-B27 expression.

Figure 2

(A) Whole cell extracts from U937.B27 cells stably expressing ERAP1 or scrambled (Scram) shRNA were subjected to SDS-PAGE and blotted for ERAP1 and GAPDH as a loading control. (B) ERAP1 shRNA knockdown and control (Scram) U937.B27 cells were stained with the antibodies indicated, and analyzed by flow cytometry. Relative expression is mean fluorescence intensity for ERAP1 knockdown cells compared to cells transfected with scrambled shRNA (set to 1). Data are from 2–4 independent experiments done in triplicate. (C,D) Cell surface HLA class I was IPd with W6/32 (C) or HC10 (D), treated with N-glycanase, and quantitated by IEF and Western blotting. Representative blots (left panels) and quantitative results (right panels) are shown. Longer exposures are required to better visualize the B27 band in (D). Since this results in overexposure of the B51 band, we show only the shorter exposure here. A longer exposure is shown in Supplemental Figure S1C. Relative expression is the mean (+/−SEM) of triplicate samples, with expression in scrambled shRNA cells normalized to 1. * P < 0.05; **P < 0.01.