Fig. 6
MIEF self-association is required for MIEF-induced mitochondrial fusion. a, b Confocal images of mitochondrial fusion from the PEG-based fusion assay in WT 293T polykaryons transfected with empty vector, MIEF1-V5, MIEF1Δ1-48-V5, or MIEF1Δ109-154-V5. Cells with stable expression of mitoRFP or mitoGFP were co-cultured, transfected with empty vector, MIEF1-V5 (WT), MIEF1Δ1-48-V5, or MIEF1Δ109-154-V5 as indicated, and then subjected to the PEG cell fusion assay. Mitochondrial fusion is indicated by co-localization of mitoRFP (red) and mitoGFP (green) (i.e., yellow mitochondria) (a, upper panels). As indicated by V5-tag staining, the MIEF1Δ1-48 mutant localizes in the cytosol, whereas MIEF1Δ109-154 is on mitochondria (a, lower panels). The extent of mitochondrial fusion in individual hybrid cells was analyzed via the Pearson’s correlation coefficient (PCC, mean ± SEM) in three independent experiments for each condition and data summarized in (b) (n represents the total number of cells analyzed for each condition). c, d The mito-PAGFP-based mitochondrial fusion assay confirmed that the cytosolic mutant MIEF1Δ1-48 and the self-association-deficient mutant MIEF1Δ109-154 lost the ability to enhance mitochondrial fusion. n represents the number of cells analyzed. ****P < 0.0001; **P < 0.01 (d)