Panel a shows a Western blot analysis with five rows of protein bands. The rows are labeled Transferrin, STARD7, YME1L, VDAC, and SDHA. The columns are labeled DMSO and Fato with molecular weights 8k, 12k, and 40k. Panel b contains two bar graphs for HeLa cells. The left graph shows the percentage of Mito CoQ with DMSO and IPP treatments. The right graph shows the percentage of non-Mito CoQ with DMSO and IPP treatments. Panel c contains two bar graphs for Hepg2 cells. The left graph shows the percentage of Mito CoQ with DMSO and IPP treatments. The right graph shows the percentage of non-Mito CoQ with DMSO and IPP treatments.
CoQ distribution following SCAP inhibition and IPP supplementation. (a) WT HeLa cells were treated with DMSO or fatostatin for 24 h, and mitochondria were extracted using differential centrifugation. The cells were centrifuged at different speeds (8,000g, 12,000g, and 40,000g) to separate mitochondrial fractions. Western blotting was performed to detect markers specific to mitochondrial (VDAC, SDHA, and YME1L) and PM (transferrin) fractions. (b and c) HeLa (b) and HepG2 (c) cells were treated with either DMSO or IPP for 24 h. Cells were collected, and mitochondrial and non-mitochondrial fractions were isolated using differential centrifugation (n = 3 biological replicates). The pellets from each fraction were sonicated in MTBE buffer, and lipids were extracted. The lipid phase was then analyzed by mass spectrometry to measure CoQ levels. MTBE, methyl-tert-butyl ether. Source data are available for this figure: SourceData FS3.