Journal: The EMBO Journal

Article Title: STIM1-containing contact sites promote direct calcium flux from the endoplasmic reticulum to mitochondria

doi: 10.1038/s44318-026-00700-8

Figure Lengend Snippet: ( A ) Wild-type HEK293 and STIM1-KO cells were subjected to MAM fractionation, and TH, CM, MAM, and ER fractions were analyzed by immunoblotting (5 μg protein/lane). ACSL4 was used as a loading control for MAM fraction. ( B – D ) Enrichment of IP3Rs in the MAM fraction was quantified relative to total ER levels (MAM + ER signals). Data were normalized to the WT condition and plotted as mean ± S.D. from n = 5 biological replicates for IP3R3 and IP3R2, and n = 3 biological replicates for IP3R1/2/3. Statistical analysis with unpaired t-test in all cases. ( E ) GRP75 and VDAC1 proteins levels were analyzed in the MAM fractions from WT HEK293 and STIM1-KO HEK293 cells. ACSL4 was used as a loading control. ( F , G ) Data were normalized to the WT condition and plotted as mean ± S.D. from n = 4 biological replicates. Statistical analysis with unpaired t-test, p = 0.0288 in ( F ), p = 0.0452 in ( G ). ( H ) Methanol-fixed HEK293 cells were incubated with the indicated primary antibodies (rabbit anti-VDAC1 and/or mouse anti-IP3R1/2/3) as well as the DNA probes rabbit-PLUS and mouse-MINUS. As negative controls, cells were incubated with individual primary antibodies. Representative images for all the conditions are shown. Scale bar = 10 μm. ( I ) Quantification of the assay from ( H ), with the number of cells analyzed indicated in parentheses. The black line represents the mean of the data. Statistical analysis with unpaired t-test, **** p < 0.0001, * p = 0.0139. .

Article Snippet: Rabbit anti-VDAC1 (1:300 in blocking buffer, for IF) , Proteintech , 55259-1-AP.

Techniques: Fractionation, Western Blot, Control, Incubation