Fig. 2. Nuclear GSK3 suppresses serine/one-carbon metabolism.

Cell lines were established with pTRIPZ-EV, pTRIPZ-HA-GSK3-(WT), pTRIPZ-HA-NES-GSK3β, pTRIPZ-HA-NLS-GSK3β, or pTRIPZ-HA-NLS-GSK3β-(KD) in NCI-H1299 cells. WT, wild type; HA, hemagglutinin. (A and B) Cells were exposed to 1 μM doxycycline for 24 hours, and (A) whole-cell lysate was subjected to immunoblot analysis with the indicated antibodies, or (B) total RNA was extracted and subjected to reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis with indicated primers. (C and D) EV, NES-GSK3, or NLS-GSK3 cells were exposed to 1 μM doxycycline for 24 hours before U-¹³C-Serine or U-¹³C-Glucose labeling. Cells were fed with U-13C-Serine or U-13C-Glucose for 4 hours, and metabolites were subsequently extracted and subjected to liquid chromatography–mass spectrometry (LC-MS) analysis. *P < 0.05, **P < 0.01.