Fig. 7: Arpin binding to cell membranes increases in HIV-1 infected Jurkat T lymphocytes.

A Scheme showing the protocol of membrane flotation assay performed on infected Jurkat T cells (with the help of BioRender). B Immunoblots showing HIV-1 Gag(i)GFP, Arpin, and Tsg101 in the membrane (# 1, 2, and 3) and cytosolic (# 6, 7, and 8) fractions. The lysosomal Lamp2 protein was used as a membrane marker and the ribosomal S6 protein as a cytosol marker. C Histogram showing the % of Arpin, and Tsg101, binding to cell membranes in infected (in gray) or non-infected (in white) cells, N = 3 independent experiments, one-tailed Mann Whitney to test increase in membrane binding. D STORM/TIRF images of non-infected (upper panel) and VSV-G pseudotyped HIV-1(i)GFP∆Env infected (bottom panel) Jurkat T cells showing, at the cell surface, HIV-1(i)GFP Gag labeled with nanobody Alexa-Fluor-568 (in magenta) and Arpin labeled with anti-Arpin and anti-rabbit secondary antibody ATTO-647N (in yellow). The scale bar is 5 µm for full-cell images. The scale bar is 1 µm for zoomed images. The right graph shows the quantification of Arpin localization densities at the cell surface of non-infected (grey violin, each white dot corresponds to the number of localizations in a 1 µm² square ROI, n = 100 ROI) and of infected Jurkat T cells (black violin, each black dot corresponds to the number of localizations in a 1 µm² square ROI, n = 100 ROI). Two-tailed Mann-Whitney tests were used for localization density comparison. E Immunoblots showing the immunoprecipitation of HIV-1 Gag with the anti-Arpin antibody in transfected 293THEK model cell line. 1: non-transfected cell lysate incubated with anti-Arpin. 2: non-transfected cell lysate incubated with no antibody. 3: transfected cell lysate incubated with anti-Arpin. 4: transfected cell lysate incubated with no anti-Arpin. 5: transfected cell lysate incubated with rabbit serum. Bands showing HIV-1 Gag as well as antibody IgG (Ab) are indicated. N = 3 independent experiments. Data are presented as mean values +/- SEM, except in panel D) where data are presented as median value +/- 1^st interquartile range (IQR). Exact p-values are given in graphs. Source data are provided as Source data file.