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. Author manuscript; available in PMC: 2018 Jul 6.
Published in final edited form as: Mol Cell. 2017 Jun 8;67(1):84–95.e5. doi: 10.1016/j.molcel.2017.05.020

Figure 2. Autophagy promotes Glut1 recycling to the plasma membrane.

Figure 2

(A and B) Indicated cell types transiently expressing Flag-Glut1 incubated with α-Flag, washed and cultured at 37°C for indicated times prior to staining with Alexa 488 secondary antibody to determine the fraction of surface label remaining by flow cytometry (mean ± SEM, n=3 independent experiments). (C and D) Cells transiently expressing Flag-Glut1 incubated with α-Flag at 37°C in full media, washed and chased in glucose-free media for indicated times, followed by staining with Alexa 488 secondary antibody to determine the kinetics of surface Flag-Glut1 re-emergence by flow cytometry (mean ± SEM, n=3). (E) Cells stained with Alexa Fluor 488-conjugated transferrin (Tfn) at 37°C in serum-free media, and chased in full media (mean ± SEM, n=3). P-values calculated using unpaired t-test. *P<0.05 (F) Confocal live-cell images of atg5+/+ and atg5−/− MEFs stably expressing GFP-Glut1 following 2h of glucose starvation for the indicated times. Insets: Magnification of the boxed regions. In top insets, arrows delineate GFP-Glut1 containing vesicular structures migrating towards the plasma membrane in atg5+/+ MEFs. Scale bar, 10μm. Related to Movie S1 and S2.