Figure 6
Figure 6. A-I06 inhibits the synthesis of secretory μ chains, but not membrane-bound μ chains, free κ chains, and membrane-bound class I MHC heavy chains (HC); A-I06 does not affect protein transport. (A-B) Wild-type B cells were stimulated with LPS for 2 days and subsequently treated with A-106 (50μM) for an additional day. Untreated control and A-I06–treated cells were radiolabeled for 15 minutes, chased for the indicated time, and lysed. Intracellular and extracellular IgM were immunoprecipitated from (A) lysates and (B) culture media, respectively, using an anti-μ Ab. Immunoprecipitates were analyzed on a SDS-PAGE gel. Data are representative of 2 independent experiments. For each experiment, wild-type B cells were purified and pooled from 2 mouse spleens. (C-D) To reveal the effect of A-I06 on mIgM, B cells purified from μS−/− mouse spleens were stimulated with LPS for 2 days and subsequently treated with A-I06 (50μM) for an additional day. Untreated control and A-I06–treated cells were radiolabeled for 15 minutes, chased for the indicated time, and lysed. (C) Intracellular mIgM was immunoprecipitated from lysates using an anti-μ Ab. (D) Secreted free κ chains were immunoprecipitated from culture media using an anti-κ Ab. Immunoprecipitates were analyzed on an SDS-PAGE gel. *Complex-type glycan modifications. Data are representative of 2 independent experiments. For each experiment, μS−/− B cells were purified and pooled from 2 mouse spleens. (E) Similar wild-type B-cell lysates as those in panel A were immunoprecipitated using an anti-class I MHC HC Ab and analyzed by SDS-PAGE. CHO and CHO* represent high mannose-type glycans and complex-type glycans, respectively. Data are representative of 2 independent experiments. For each experiment, wild-type B cells were purified and pooled from 2 mouse spleens. (F) Similar μS−/− B-cell lysates as those in panel C were immunoprecipitated using an anti–class I MHC HC Ab and analyzed by SDS-PAGE. Data are representative of 2 independent experiments. For each experiment, μS−/− B cells were purified and pooled from 2 mouse spleens.

A-I06 inhibits the synthesis of secretory μ chains, but not membrane-bound μ chains, free κ chains, and membrane-bound class I MHC heavy chains (HC); A-I06 does not affect protein transport. (A-B) Wild-type B cells were stimulated with LPS for 2 days and subsequently treated with A-106 (50μM) for an additional day. Untreated control and A-I06–treated cells were radiolabeled for 15 minutes, chased for the indicated time, and lysed. Intracellular and extracellular IgM were immunoprecipitated from (A) lysates and (B) culture media, respectively, using an anti-μ Ab. Immunoprecipitates were analyzed on a SDS-PAGE gel. Data are representative of 2 independent experiments. For each experiment, wild-type B cells were purified and pooled from 2 mouse spleens. (C-D) To reveal the effect of A-I06 on mIgM, B cells purified from μS−/− mouse spleens were stimulated with LPS for 2 days and subsequently treated with A-I06 (50μM) for an additional day. Untreated control and A-I06–treated cells were radiolabeled for 15 minutes, chased for the indicated time, and lysed. (C) Intracellular mIgM was immunoprecipitated from lysates using an anti-μ Ab. (D) Secreted free κ chains were immunoprecipitated from culture media using an anti-κ Ab. Immunoprecipitates were analyzed on an SDS-PAGE gel. *Complex-type glycan modifications. Data are representative of 2 independent experiments. For each experiment, μS−/− B cells were purified and pooled from 2 mouse spleens. (E) Similar wild-type B-cell lysates as those in panel A were immunoprecipitated using an anti-class I MHC HC Ab and analyzed by SDS-PAGE. CHO and CHO* represent high mannose-type glycans and complex-type glycans, respectively. Data are representative of 2 independent experiments. For each experiment, wild-type B cells were purified and pooled from 2 mouse spleens. (F) Similar μS−/− B-cell lysates as those in panel C were immunoprecipitated using an anti–class I MHC HC Ab and analyzed by SDS-PAGE. Data are representative of 2 independent experiments. For each experiment, μS−/− B cells were purified and pooled from 2 mouse spleens.

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