FIGURE 7:

Functional mutagenesis of critical amino acid residues at the selectivity filter of TcMCUc and TcMCUd. (A) Western blot analysis of total protein extracts of control cells (Scr), TcMCUc-KO (MCUc KO), TcMCUc-KO plus EV (KO + EV), TcMCUc-KO plus TcMCUc (KO + MCUc), TcMCUc-KO plus TcMCUc^D157N (D157N), and TcMCUc-KO plus TcMCUc^E162Q (E162Q) epimastigotes using anti-c-Myc antibodies. Anti-tubulin (Tub) antibodies were used as a loading control. (B) Ca²⁺ uptake reconstitution in DIG-permeabilized epimastigotes. Experimental conditions were as in Figure 1D. (C) Quantification of data from three experiments as shown in B. Relative Ca²⁺ uptake at 500 s compared with controls (Scr). (D) Western blot analysis of total protein extracts of control cells (Scr), TcMCUd-KO (MCUd KO), TcMCUd-KO plus EV (KO + EV), TcMCUd-KO plus TcMCUd (KO + MCUd), TcMCUd-KO plus TcMCUd^D141N (D141N), and TcMCUd-KO plus TcMCUd^E146Q (E146Q) epimastigotes using anti-c-Myc antibodies. Anti-tubulin (Tub) antibodies were used as a loading control. (E) Ca²⁺ uptake reconstitution in DIG-permeabilized epimastigotes. Experimental conditions were as in Figure 1D. (F) Quantification of data from three experiments as shown in E. Relative Ca²⁺ uptake at 500 s compared with controls (Scr). Values in C and F are means ± SD (n = 3); ***P < 0.001; one-way ANOVA.