Skip to main content
. Author manuscript; available in PMC: 2020 May 1.
Published in final edited form as: Virology. 2019 Feb 21;531:192–202. doi: 10.1016/j.virol.2019.02.011

Figure 1: Nef-mediated inhibition of NFAT signalling.

Figure 1:

NFAT-driven luciferase activity was quantified in Nef-transfected Jurkat cells following TCR stimulation, to measure the ability of Nef to inhibit NFAT induction by TCR signalling. Nef was expressed in a pSELECT-GFPzeo plasmid co-expressing green fluorescent protein (GFP). (A) Flow cytometry was performed to assess the percentage of live Jurkat cells (indicated by the circular gate) following transfection. (B) Transfection efficiency of each Nef clone was assessed by flow cytometry to detect the percentage of GFP positive cells (indicated by the square gate), which represent Nef-transfected cells. We confirmed that cellular toxicity and transfection efficiency were similar across clones within an experiment and thus did not confound the assay interpretation. (C) Absolute light units measured 6 hours after TCR stimulation are shown for Jurkat cells transfected with SF2 Nef, G2A Nef and 3 patient-derived Nef clones (NC1-3). SF2 and G2A Nef clones were included as positive and negative controls in each experiment. (D) The ability of each patient-derived Nef clone to inhibit NFAT signalling was normalised to the controls such that SF2 Nef represented 100% activity while G2A Nef represented zero activity. In panels C and D, the mean and standard deviation of two independent experiments (each with triplicate measurements) is shown.