Figure 4.

DO modulates DM function in a pH-dependent fashion. (A) In vitro association of biotinylated ApoB(2877–2894) to purified DR3–CLIP complexes upon CLIP dissociation was measured at different pH values in the presence of cell lysates containing equal amounts of DM or DM–DOαβ² GFP complexes (DM/DO) or lysis buffer only (⁻). Association is depicted as absorbance at 405 nm in arbitrary units (a.U.) upon saturation of the peptide exchange reaction. The data shown were derived from a representative set of experiments (n = 4). (B) Antigen-presenting capacity of p3-13–reactive T cell clone Rp15 1-1 using control (DM) and DOαβGFP-expressing cells (DM/DO) at prevention of acidification of the endosomal/lysosomal pathway. T cell proliferation is expressed as the amount of [³H]thymidine incorporation in cpm and was determined for varying amounts of M. tuberculosis sonicate with 1,500 APCs/assay well. Data shown were derived from a representative experiment performed three times; SEM <10%. T cell proliferation in response to Mel JuSo cells pulsed with HLA-DR3–restricted peptide p56-65 was not significant, and application of Mel JuSo transfected with DO without GFP tag resulted in the same effects as the DOαβGFP transfectants (data not shown). (C) Antigen-presenting capacity of p3-13–reactive T cell clone Rp15 1-1 using varying amounts of control (DM) and DOαβGFP-expressing cells (DM/DO) at prevention of acidification of the endosomal/lysosomal pathway. T cell proliferation (measured by [³H]thymidine incorporation) was determined using 100 μg/ml M. tuberculosis sonicate. Data shown were derived from a representative experiment performed three times; SEM <10%. Application of Mel JuSo transfected with DO without GFP tag yielded the same results as the DOαβGFP transfectants (data not shown). Black bars, DM + M. tuberculosis; gray bars, DM + medium; lined bars, DM-DO + M. tuberculosis; empty bars, DM-DO + medium.