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. 2024 Oct 1;15:8479. doi: 10.1038/s41467-024-52773-w

Fig. 5. Ion channel activity of TRPM7 is required for pH-sensitive virus infection.

Fig. 5

a Diagram of TRPM7 domains with annotations of pore mutant (pm) and kinase-inactive (ki) mutations. b (Left) Whole-cell patch-clamp recordings shown as current-voltage (IV) relationship of mouse TRPM7 constructs expressing SVG-A cells. The IV of non-transfected (black), wild-type (TRPM7-wt, dark gray), pore mutant (TRPM7-pm, blue), and kinase-inactive (TRPM7-ki, orange) transfected SVG-A cells are overlayed. (Right) Immunoblot of membrane fraction of non-transfected SVG-A cells (ctl) and cells transfected with mouse TRPM7-wt, -pm, and -ki constructs. Immunoblotting is done with an anti-mouse TRPM7 antibody which does not detect human TRPM7. A non-specific band seen at 45 kDa is used as a loading control. The immunoblot results were reproduced in two independent experiments. c Representative maximum projection (20 μm) images of VSV-G and VSV-pH infections (green) in SVG-A cells ectopically expressing the TRPM7 variants: wild-type (TRPM7-wt), pore mutant (TRPM7-pm), or kinase-inactive (TRPM7-ki). d Quantification of infection in SVG-A cells ectopically expressing TRPM7-wt, TRPM7-pm, and TRPM7-ki variants. Cells were infected with either parental VSV-G or VSV-pH. Percent cytosolic+ (dark green) and nuclei+ (light green) cells are shown. Averages are derived from three independent infections. P values were determined from total cells infected by one-way ANOVA with Tukey’s multiple comparison test. Mean and SEM are plotted. e Representative maximum projection (20 μm) images of VSV-G and VSV-pH infections (green) in SVG-A cells treated with TRPM7 inhibitors FTY720 and NS8593 at 5 µM or 30 µM respectively. The inhibitors were added to the cells 15 minutes prior to infection and are present throughout infection. f Quantification of VSV-G and VSV-pH infection of SVG-A cells in the presence of TRPM7 inhibitors FTY720 (5 μM) and NS8593 (30 μM). Percent cytosolic+ (dark green) and nuclei+ (light green) are shown. Averages are derived from three independent infections. P values were determined from total cells infected by one-way ANOVA with Tukey’s multiple comparison test. Mean and SEM are plotted.