Fig. 4: Disrupting sialic-acid/Siglec interactions enhances the susceptibility of HIV-infected CD4⁺ T cells to innate-immune-mediated cytotoxicity. | Nature Communications

Fig. 4: Disrupting sialic-acid/Siglec interactions enhances the susceptibility of HIV-infected CD4⁺ T cells to innate-immune-mediated cytotoxicity.

From: HIV-induced sialoglycans on infected CD4+ T cells promote immune evasion from myeloid cell-mediated killing

Fig. 4

a Schematic representation of the experimental workflow for generating CRISPR/Cas9-mediated knockout of GALNT6, NANP, and B4GALNT1 in primary CD4⁺ T cells infected with HIV. Some of the images in panel a were created in BioRender. Singh, S. (2025) https://BioRender.com/8d3lee4. b qRT-PCR analysis of GALNT6, NANP, and B4GALNT1 expression in gene-edited, HIV-infected primary CD4⁺ T cells. Statistical analysis was performed using two-sided ratio paired t tests. Each dot represents samples from an individual donor, run in triplicate and averaged. N = 4 donors. c NTC-targeted or gene-edited HIV-infected primary CD4⁺ T cells were co-cultured with autologous monocytes for 16 h (E:T ratio = 10:1). After overnight incubation, the co-cultures were stained with Aqua Live/Dead (BV510) and antibodies against CD3 and intracellular HIV p24. Representative contour plots (left panel) show a reduction in p24⁺ cells in gene-edited HIV-infected primary CD4⁺ T cells upon co-culture with autologous monocytes compared to NTC-targeted cells. The right panels depict normalized cytotoxicity, calculated based on either the percentage of p24⁺ cells or the reduction in p24 MFI. Two-sided Paired t tests. Each dot represents samples from an individual donor. N = 5 donors. d–f HIV-infected primary CD4⁺ T cells were co-cultured with autologous monocytes (E:T ratio = 10:1) in the presence of anti-Siglec-3 antibody or isotype control. After overnight incubation, the co-cultures were stained with Aqua Live/Dead (BV510) and antibodies against CD3 and intracellular HIV p24. d Representative flow cytometry plots. e Percentage of p24⁺ cells and cytotoxicity calculated based on % p24. f p24 MFI and cytotoxicity calculated based on p24 MFI. Statistical analysis was performed using paired one-way ANOVA, corrected by the two-stage step-up procedure of Benjamini, Krieger, and Yekutieli. Error bars represent mean ± standard deviation; n = 3 donors.

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