Figure 8From: Mitochondrial targeting of human NADH dehydrogenase (ubiquinone) flavoprotein 2 (NDUFV2) and its association with early-onset hypertrophic cardiomyopathy and encephalopathyThe human pathogenic NDUFV2 deletion mutant lost most of its mitochondrial targeting ability. (a) Schematic representation of the genomic structure of NDUFV2 in the first three exons. The 4-bp deletion of human pathogenic IVS2+5_+8delGTAA mutation is indicated by underlined red letters (GTAA). Dotted line represents the wild-type splicing form, and continuous line indicates the abnormal splicing form. Schematic structures of c-myc fusion proteins corresponding to the wild-type splicing form and the abnormal splicing form are also shown here. E1, E2 and E3 represent exon 1, exon 2 and exon 3, respectively. (b) The protein distribution patterns of wild-type and NDUFV2 IVS2+5_+8delGTAA mutant. The expressed proteins were labeled by an anti-c-myc-FITC antibody in transfected cells (green color), mitochondria were labeled by Mito Tracker Red (red color), and colocalization of expressed protein and mitochondria is shown as a merged image and indicated by yellow signals. The number of (+) symbols indicates that the proportion of cells exhibiting FITC fluorescence have a typical punctuated staining pattern and mitochondrial colocalization. The (++++) symbol indicates all of the FITC fluorescence signals in transfected cells are fully colocalized with mitochondria. Scale bars = 10 μm. (c) Subcellular localization of the wild-type and NDUFV2 pathogenic mutant. Western blot analyses were conducted using cytosolic (Cy) and mitochondrial (Mi) extracts from T-REx-293 cells transiently transfected with the wild-type or NDUFV2 pathogenic mutant construct. (c) The expression levels of the wild-type and the △19-40 NDUFV2 mutant proteins in the whole cell lysates. Western blot analyses were conducted using the whole cell lysates from T-REx-293 cells transiently transfected with the wild-type or the △19-40 NDUFV2 mutant construct. The expressed proteins were detected by an anti-c-myc antibody. β-tubulin was used as a cytosolic marker and ATP synthase subunit α (ATP α) was used as a mitochondrial marker.Back to article page