Figure 2.

MCU is required for mitochondrial calcium uniporter in HeLa cells and CGNs. (A) Two shRNA against MCU were designed to knockdown MCU in HeLa cells and the knockdown efficiency of each was detected by Western blot. (B) Knocking down MCU decreased mitochondrial Ca²⁺ uptake by assaying Mito-GCaMP3. Control or MCU knockdown HeLa cells were transfected with Mito-GCaMP3. GCaMP3 fluorescence was measured before and during exposure to histamine stimulation (100 μmol/L). (C) Representative traces Ca²⁺ response to histamine stimulation in control or MCU knockdown HeLa cells. (D) Graph shows quantification of peak Ca²⁺ intensities. (E) Representative traces of Ca²⁺ uptake in digitonin-permeabilized HeLa cells. The level of Ca²⁺ was indicated by the calcium Green-5N. (F) Graph shows relative Ca²⁺ uptake. (G) CGNs were transfected with shMCU or shvector together with Mito-GCaMP3. GCaMP3 fluorescence was measured before and after exposure to histamine (100 μmol/L). (H) Representative traces showing Ca²⁺ response after histamine stimulation over time in MCU wild type or knockdown cells were. (I) Graph shows quantification of peak Ca²⁺ intensities