Fig. 1
From: MICU1 controls cristae junction and spatially anchors mitochondrial Ca2+ uniporter complex
Super-resolution SIM microscopy localizes MICU1 to the IBM. a Cells were transiently transfected with MICU1-YFP (green), then stained with Mitotracker Red FM (MTR) (magenta) and examined using simultaneous dual-color 3D-SIM either under resting conditions, or 4 min after stimulation with 100 µM histamine. The upper panels provide an overall view of the mitochondria, and the dashed squares indicate the regions shown magnified below. The figures show merges of MICU1-YFP and MTR, along with MICU1-YFP (MICU1) and IMM (MTR) signals alone. No changes in the distribution of MICU1-YFP (nor IMM staining) were observed upon stimulation with histamine. b Line plots of MICU1 (green line) and the IMM staining with MTR (magenta line) of regions marked with dashed white lines in a are shown. A clear sub mitochondrial localization of MICU1 to the IBM is visible. c HeLa cells transfected with MICU1-YFP (green) and mCherry-TOM22 (magenta) were imaged using simultaneous dual-color 3D-SIM. Left panels show representative images. Right panels show line plots through mitochondria at locations indicated by the white dashed lines. By subjecting both channels (TOM22-m (magenta line), MICU1-m (green line)) to a double Gaussian-fit for each individual line (TOM22-fit (dotted magenta line), MICU1-fit (dotted green line) (right panels), the peaks of both distributions could be determined with sub-pixel accuracy. d HeLa cells transfected with MICU1-YFP and mCherry-TOM22 or MICU1-mCherry were analyzed regarding the relative distance between the YFP and mCherry distributions like shown in c. Data are shown as dot plots with the mean ± SD as red middle line and whiskers, respectively (nmCherry-TOM22 = 105, nMICU1-mCherry) = 80). *P < 0.05 vs. respective control conditions carried out with unpaired double-sided T-test. e Schematic illustration of the mitochondrial inner boundary membrane and outer membrane showing the determined spacing between MICU1 and TOM22 of 21 ± 3 nm (mean ± SD). This value matches published distances between the OMM and IMM (lower right). Images and analyses were obtained from 2 to 5 mitochondria in at least 3 cells in each of 8 independent experiments (nmCherry-TOM22 = 8/24/105, nMICU1-mCherry) = 8/40/80). Source data are provided as a Source Data file
