Multiomic analysis reveals a key BCAT1 role in mTOR activation by B cell receptor and TLR9
Rui Guo, Yizhe Sun, Matthew Y. Lim, Hardik Shah, Joao A. Paulo, Rahaman A. Ahmed, Weixing Li, Yuchen Zhang, Haopeng Yang, Liang Wei Wang, Daniel Strebinger, Nicholas A. Smith, Meng Li, Merrin Man Long Leong, Michael Lutchenkov, Jin Hua Liang, Zhixuan Li, Yin Wang, Rishi Puri, Ari Melnick, Michael R. Green, John M. Asara, Adonia E. Papathanassiu, Duane R. Wesemann, Steven P. Gygi, Vamsi K. Mootha, Benjamin E. Gewurz
Rui Guo, Yizhe Sun, Matthew Y. Lim, Hardik Shah, Joao A. Paulo, Rahaman A. Ahmed, Weixing Li, Yuchen Zhang, Haopeng Yang, Liang Wei Wang, Daniel Strebinger, Nicholas A. Smith, Meng Li, Merrin Man Long Leong, Michael Lutchenkov, Jin Hua Liang, Zhixuan Li, Yin Wang, Rishi Puri, Ari Melnick, Michael R. Green, John M. Asara, Adonia E. Papathanassiu, Duane R. Wesemann, Steven P. Gygi, Vamsi K. Mootha, Benjamin E. Gewurz
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Research Article Cell biology Metabolism

Multiomic analysis reveals a key BCAT1 role in mTOR activation by B cell receptor and TLR9

Abstract

B lymphocytes play major adaptive immune roles, producing antibodies and driving T cell responses. However, how immunometabolism networks support B cell activation and differentiation in response to distinct receptor stimuli remains incompletely understood. To gain insights, we systematically investigated acute primary human B cell transcriptional, translational, and metabolomic responses to B cell receptor (BCR), TLR9, CD40-ligand (CD40L), IL-4, or combinations thereof. T cell–independent BCR/TLR9 costimulation, which drives malignant and autoimmune B cell states, highly induced transaminase branched chain amino acid transaminase 1 (BCAT1), which localized to lysosomal membranes to support branched chain amino acid synthesis and mTORC1 activation. BCAT1 inhibition blunted BCR/TLR9, but not CD40L/IL-4–triggered B cell proliferation, IL-10 expression, and BCR/TLR pathway–driven lymphoma xenograft outgrowth. These results provide a valuable resource, reveal receptor-mediated immunometabolism remodeling to support key B cell phenotypes, and identify BCAT1 as an activated B cell therapeutic target.

Authors

Rui Guo, Yizhe Sun, Matthew Y. Lim, Hardik Shah, Joao A. Paulo, Rahaman A. Ahmed, Weixing Li, Yuchen Zhang, Haopeng Yang, Liang Wei Wang, Daniel Strebinger, Nicholas A. Smith, Meng Li, Merrin Man Long Leong, Michael Lutchenkov, Jin Hua Liang, Zhixuan Li, Yin Wang, Rishi Puri, Ari Melnick, Michael R. Green, John M. Asara, Adonia E. Papathanassiu, Duane R. Wesemann, Steven P. Gygi, Vamsi K. Mootha, Benjamin E. Gewurz

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Figure 4

BCR/TLR9 stimulation drives BCAA synthesis in human primary B cells.

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BCR/TLR9 stimulation drives BCAA synthesis in human primary B cells. (A)...
(A) Isotope tracing schematic. [13C]-L-leucine_m+6 was used to trace BCAT1 BCAA catabolism to a-keto-isocaproic acid (KIC); [15N]-glutamine_m+2 was used to monitor BCAT1 BCAA biosynthesis from KIC, a-keto-isovaleric acid (KIV), or a-keto b-methylvaleric acid (KMV). Gln, glutamine. Glu, glutamate. (B) Isotope tracing experimental design. Primary B cells from n = 4 donors were pretreated with vehicle or ERG245(100 μM) for 1 hour and then stimulated by αIgM + CpG for 24 hours in the presence of vehicle or ERG245. Cells were washed with PBS 3 times and resuspended in glutamine/leucine-free media supplemented with 381 mM 13C6-leucine and 2.054 mM 15N2-glutamine + 10% dialyzed FBS. ERG245(100 μM) and αIgM + CpG stimulants were also refreshed at this time point. Intracellular metabolites were profiled at 8, 24, and 48 hours later. (C) Ion intensities of m+6-labeled and -unlabeled KIC at the indicated times in cells treated with vehicle control or ERG245. (D) Ion intensities of labeled and unlabeled leucine (Leu) at the indicated times in cells treated with vehicle control or ERG245. (E) Ion intensities of labeled and unlabeled isoleucine (Ile) at the indicated times in cells treated with vehicle control or ERG245. (F) Ion intensities of labeled and unlabeled valine (Val) at the indicated times in cells treated with vehicle control or ERG245. (G) Heatmap analysis of metabolite z scores in primary B cells treated with vehicle or ERG245 (100 μM) and stimulated as indicated for 24 hours. (H) Volcano plot visualization of –log10 (P value statistical significance) and log2 (fold-change metabolite abundance) from metabolomic analysis of ERG245-treated versus vehicle-treated primary B cells stimulated by αIgM + CpG for 24 hours from n = 4 replicates. (I) MetaboAnalyst pathway enrichment analysis of metabolites diminished by ERG245 treatment in αIgM + CpG–stimulated B cells at 24 hours. P values were calculated by 2-way ANOVA followed by Tukey’s multiple-comparison test (C–F).