Figure 3.

Autophagy-mediated proteolysis is required for optimal IL21 production in Mem from EC. Autophagy-mediated proteolysis and IL21 production were both determined in activated Mem under specific BECN1 gene silencing (siRNA IDs: 137,198), or PRKAA1 (compound C), lysosomal (BaF or Chloro.), and protease (E64d-PepA) inactivation. (A) Levels of BECN1 were determined on Mem after 24 h of cell transfection with negative or BECN1 siRNAs by flow cytometry. i. Representative histograms of BECN1 expression in transfected Mem from EC. ii. Percentages of BECN1⁺ cells in Mem with or without specific BECN1 silencing for all study groups. % of BECN1 decrease was also indicated in bold for the EC’ and HIV^neg’s groups. (B) Autophagy-mediated proteolysis assessed in polyclonally activated Mem with or without specific BECN1 gene silencing or chemical inhibitors. Levels of IL21 production in Mem that have then been either (C) i. polyclonally or (D) HIV-1-specifically activated for 6 h with or without specific BECN1 gene silencing or chemical inhibitors. (C) ii. Representative histograms of IL21 expression in polyclonally-activated Mem from EC with or without autophagy blockade. iii. Confirmation of PRKAA1 inhibition by compound C was determined by western blotting using polyclonally activated Mem from EC (representative blots of 3 independent confirmations). N = 6. The error bars indicate standard deviations from the means. β, symbol used for paired t-test (comparison between treated Mem and untreated control). *, symbol used for Mann-Whitney test (comparison between study groups).