Extended Data Fig. 7: Impact of Asn on CD8+ T cell metabolism and TCR signalling.
From: Asparagine enhances LCK signalling to potentiate CD8+ T-cell activation and anti-tumour responses
a and b, Mouse naive CD8+ T cells were left untreated (naive) or activated in Asn/Asp-free medium (Ctrl), or medium supplied with Asn or Asp for indicated times. All medium was added with 10% dialysed FBS. The abundance of cellular metabolites at different time points were analysed by LC-MS/MS. The heat map shows the relative abundance of metabolites over time in CD8+ T cells stimulated in Ctrl, Asn- or Asp-medium. Log2 fold changes (FC) are relative to relative to cells stimulated in Asn/Asp-free (Ctrl) medium at corresponding time points. Data are representative of three independent experiments. Structural isomers were condensed and counted only once as the employed technology cannot distinguish metabolites with identical molecular weight. c, TCR signalling that initiates T cell activation. d, Related to Fig. 4e. Scheme of OT-I adoptive transfer and OVA-peptide treatment experiment. e, Western blot analysis of mouse CD8+ T cells activated in medium containing increasing amounts of Asn in the absence or presence of PP2 (10 µM). Data are representative of three independent experiments. f, Related to Fig. 5e–k. CD45.1 C57BL/6 mice maintained on an Asn-free diet were intraperitoneally injected with Asn or PBS (Ctrl), or PP2 (30 µg/mouse) or GDC-0032 (100 µg/mouse) every two days. 8 days later, mice were adoptively transferred with naive CD45.2+ OT-I CD8+ T cells and injected with OVA peptide as indicated. g, Related to Fig. 5l–o, Scheme of Asn-free diet CD45.1 C57BL/6 mice i.p. with Asn and/or PP2 and then adoptively transferred with naive CD45.2+ OT-I CD8+ T cells and infected with LmOVA as indicated. Uncropped blots for e and numerical source data for a, b, are provided.
