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. 2023 Nov 8;8(21):e167829. doi: 10.1172/jci.insight.167829

Figure 1. TSC2 is a central environmental hub that modulates mTORC1 activity and differentiation in T cells.

Figure 1

(A) Immunoblot analysis of the PI3K/AKT/TSC2/mTORC1 pathway in resting T cells upon dose dependent stimulation of different stimuli for 30 minutes. PMA: phorbol ester, αCD3, IL-2. (B) WT and TSC2–/– CD8+ T cells were stimulated via TCR and Co-Stim with agonists anti-CD3 and anti-CD28 for 1 and 3 hours, and mTORC1 and mTORC2 activity was measured by immunoblot. The 0 hour indicates baseline with no simulation. (C) Flow cytometry proliferation analysis of WT and TSC2–/– CD8+ T cells stimulated and grown in different pH level media conditions. Data are measured on day 3. (D) Different congenic naive WT P14 (gp33+) CD8+ T cells were isolated and CRISPR edited using with CAS9 and a control sgRNA (Ctrl) or 1 of 2 sgRNAs (g1, or g2) targeting TSC2. Ctrl and TSC2 sgRNA–edited CD8+ T cells were mixed at 1:1 ratio and coadoptively transferred into naive WT recipients and infected with LCMV Armstrong for acute (day 8, blood) and memory analysis of donor CD8+ T cells (spleen) via flow cytometry. n = 5/group for guide 1 (g1); n = 6/group for guide 2 (g2); *P < 0.05, **P < 0.01 paired t test. Data are representative of 3 or more independent experiments (A–C), and 2 independent exp (D).