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Fig. 1 | BMC Biology

Fig. 1

From: Determinants of the cytosolic turnover of mitochondrial intermembrane space proteins

Fig. 1

Diverse cytosolic stability of IMS proteins revealed by the tandem fluorescent protein timer approach. a Schematic illustration of a tandem fluorescent protein timer (tFT) fusion. The protein of interest is tagged with two fluorescent proteins with different maturation kinetics. The fluorescent signal ratio of slowly maturing mCherry protein and the rapidly maturing sfGFP protein reflects the half-life of the entire protein fusion. b Tenfold dilutions of WT cells that expressed the indicated plasmid-borne tFT fusions or an empty vector control were spotted on selective medium agar plates with either glucose or glycerol as the main carbon source. c Live confocal imaging of WT cells that expressed the indicated plasmid-borne tFT fusions and an empty vector control. Yeast were grown in minimal selective media with 3% glycerol at 24 °C. Prior to imaging, cells were stained with MitoTracker Deep Red FM and calcofluor white to label mitochondria and the cell wall, respectively. d Ratio of mCherry and sfGFP fluorescent signals measured in WT cells that expressed the indicated plasmid-borne tFT fusions. Cells were cultured in liquid selective media with 2% glucose at 28 °C. The ratio for the tFT alone was set to 1. The data are expressed as mean ± SEM. n = 6. ev, empty vector; N-deg, N-terminal degron; tFT, tandem fluorescent protein timer; WT, wild-type

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