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. 2024 Nov 23;15:10163. doi: 10.1038/s41467-024-54344-5

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© The Author(s) 2024

Open Access This article is licensed under a Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License, which permits any non-commercial use, sharing, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if you modified the licensed material. You do not have permission under this licence to share adapted material derived from this article or parts of it. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by-nc-nd/4.0/.

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Fig. 1 — A, B B cells were stimulated with LPS/IL-4/BAFF and cultured with no amino acids (No AA), essential amino acids (EAA), or full amino acids (Full AA) (A), or indicated concentrations of amino acids (B) overnight. CD98, CD86, p-4EBP1, p-S6, and FSC-A levels were measured by flow cytometry. For CD98, CD86, p-S6, and FSC-A levels, n = 5 for each group. n = 3 for each group in p-4EBP1 expression. C–F Tamoxifen was administered to animals intraperitoneally daily for 4 consecutive days. Splenic B cells were purified 7 days after the last tamoxifen injection and stimulated with LPS/IL-4/BAFF. C Expression of p-4EBP1, p-S6, CD98, and FSC-A was measured by flow cytometry after overnight activation. Cre^ER control (WT) (n = 4), Cre^ERRraga^fl/flRragb^fl/fl (n = 4), Cre^ERRptor^fl/fl (n = 4). D Expression of p-4EBP1, p-S6, p-S6K, AID, and LAMP1 was measured by immunoblot. β-actin was used as the loading control. Arrow indicates non-specific bands. Data represents 3 independent experiments. E B cells were labeled with CellTrace violet (CTV) and stimulated with indicated stimuli for 72 h. CTV dilution was measured by flow cytometry. F Expression of IgG1 and CTV dilution were examined by flow cytometry. Right, summary of the percentages of divided cells and IgG1⁺ B cells. WT (n = 8), Cre^ERRraga^fl/flRragb^fl/fl (n = 8), Cre^ERRptor^fl/fl (n = 4). G Rapamycin was added at 24 h after B cell activation with LPS/IL-4/BAFF. IgG1 expression was examined by flow cytometry at 72 h after activation. Right, summary of the percentages of IgG1⁺ B cells. The numbers indicate the fold differences of average IgG1⁺ percentages between WT and RagA/RagB deficient B cells. WT (n = 5), Cre^ERRraga^fl/flRragb^fl/fl (n = 4). Error bars represent mean ± SEM. ns, not significant. *p < 0.05, **p < 0.01, ***p < 0.001, and ****p < 0.0001, one-way ANOVA (A, B, and F), two-tailed/unpaired Student’s t-test (G), or two-way ANOVA (C). Source data are provided as a Source Data file.