Fig 2: In vitro HIV-1-infected CD4 T cells display increased surface expression of the TRAIL receptors DR4/5Primary human CD4 T cells from eight different donors (n = 8) were separately infected with lab-adapted and primary HIV-1 strains. Combined expression of DR4 and DR5 was assessed by flow cytometry 4 days after infection. Uninfected CD4 T cells were determined as CD4-positive and HIV-1 p24-negative, infected cells as CD4-negative and p24-positive. Histograms (overlay) of one representative donor displaying combined DR4/5 surface expression on CD4 T cells previously incubated with NL4-3, JR-CSF, WITO, CH198, or CH236. Expression is displayed as fluorescence intensity (x-axis).Comparison of the combined surface expression of DR4/5 between mock (white circles), uninfected (grey circles), and infected CD4 T cells (orange circles), from left to right: Mock: n = 8; NL4-3: n = 8; JR-CSF: n = 8; WITO: n = 7; CH198: n = 7; CH236: n = 8 (7–8 different donors per condition). Expression is displayed as relative fluorescence intensity (RFI) after normalization to the respective secondary AB-only control (y-axis).Data information: Wilcoxon signed-rank test. Values for non-infected and infected cells of the same donor are connected with lines. Source data are available online for this figure.

Fig 3: Antibody-mediated blocking of TRAIL inhibits NK cell degranulationDegranulation of primary human NK cells after co-culture with various target cells in the presence or absence of aTRAIL or aDR4/5. Comparison of CD107a expression after co-culture with 721.221 target cells with either 10 µg/ml aTRAIL or isotype control (Iso) using flow cytometry (n = 10 different donors per condition). Each data point represents the mean of two technical replicates. Effector:target ratio was 1:1. Left panel: Concatenated density plot depicting CD107a expression as fluorescence intensity for one representative donor. Right panel: Box plots showing relative frequency of CD107a+ NK cells (y-axis).Box plots display inhibition of degranulation (y-axis) after co-culture with 721.221 target cells in presence of either isotype or aTRAIL as relative reduction compared to no antibody (n = 10 different donors per condition).Comparison of CD107a expression after co-culture with autologous HIV-I-infected CD4 T cells with either 10 µg/ml aTRAIL or isotype control (Iso) using flow cytometry (n = 9 different donors per condition). Each data point represents the mean of two technical replicates. Left panel: Concatenated density plot depicting CD107a expression as fluorescence intensity for one representative donor. Right panel: Box plots showing relative frequency of CD107a+ NK cells (y-axis).Box plots display inhibition of degranulation (y-axis) after co-culture with autologous HIV-I-infected CD4 T cells in the presence of either isotype or aTRAIL as relative reduction compared to no antibody (n = 9 different donors per condition).Representative histograms (overlay, left panel) and bar graphs (n = 3 independent experiments, right panel) showing the individual and combined surface expression of DR4 and DR5 on 721.221 cells. Each data point represents the mean of three technical replicates.Comparison of CD107a expression after co-culture with 721.221 target cells in the presence of either aDR4/5 (10 µg/ml each) or 20 µg/ml isotype control (Iso) using flow cytometry (n = 9 different donors per condition). Effector:target ratio was 1:1. Box plots showing relative frequency of CD107a+ NK cells (y-axis).Box plots display inhibition of degranulation after co-culture with 721.221 target cells in the presence of either isotype or aDR4/5 as relative reduction compared to no antibody (n = 9 different donors per condition).Data information: Wilcoxon signed-rank test. Adjustment for multiple comparisons was performed using Bonferroni. Box plots represent the median and 25%/75% percentile. Whiskers indicate minimum and maximum data points. Bar graphs represent the mean and the associated whiskers display the SD. Source data are available online for this figure.

Fig 4: TRAIL contributes to NK-cell-mediated killing of target cellsLysis of different target cells in co-culture with NK cells was quantified in various cytotoxicity assays. Left panel: Representative contour plots showing depletion of 721.221 target cells in the presence of NK cells. Middle panel: Percentage of target cells remaining (y-axis) after co-culture with NK cells in the presence of either aTRAIL or isotype control, in reference to target cells kept alone. Right panel: Box plots displaying difference in target cells remaining (y-axis) between aTRAIL and isotype conditions displayed as p.p. (n = 12 different donors). Each data point represents the mean of at least two technical replicates.Left panel: Representative contour plots showing the percentage of .221-DR4/5KO (control) and .221-Cas9 cells (target) in the presence or absence of NK cells. Middle panel: Ratio between .221-DR4/5KO and .221-Cas9 cells (y-axis) in the presence or absence of NK cells. Right panel: Specific lysis of .221-Cas9 cells displayed as percent (n = 12 different donors). Each data point represents the mean of at least three technical replicates.Left panel: Representative contour plots showing the percentage of Raji-pSIP (control) and Raji-DR5++ (target) in the presence or absence of NK cells. Middle panel: Ratio between Raji-pSIP and Raji-DR5++ (y-axis) in the presence or absence of NK cells. Right panel: Specific lysis of Raji-DR5++ cells displayed as % (n = 12 different donors). Each data point represents the mean of at least three technical replicates.Data information: Wilcoxon signed-rank test. Experiments were performed in four batches with three different donors each. “No NK” control samples served as a reference for all donors in each batch. Lines connect each data value of the NK cell condition with their designated “No NK” control. Box plots represent the median and 25%/75% percentile. Whiskers indicate minimum and maximum data points. Source data are available online for this figure.