Extended Data Fig. 5: Mitochondrial fragmentation is necessary for germline mitochondrial DNA selection.
From: Mitochondrial fragmentation drives selective removal of deleterious mtDNA in the germline
a–c, Stills of live images illustrating the effect that overexpressing Mitofusin (b) or reducing the expression of Drp1 (c) in the germline has on the morphology of mitochondria compared to controls (a, nos-GAL4 driving UAS-mCherry RNAi; the stills in Fig. 2c, d are higher magnifications of this image). When Mitofusin is overexpressed (nos-GAL4 driving UAS-marf47), or when the expression of Drp1 is reduced (nos-GAL4 driving UAS-Drp1.miRNA.CDS48), the mitochondria in the cysts are no longer discrete as they are in control cysts. The mitochondria (white) were labelled with a mitochondrially targeted eYFP and cell membranes (blue) were labelled with CellMask Deep Red Plasma membrane Stain. Stem cells and cysts are outlined in red. d, d′, d″, Germarium of a control heteroplasmic female (nos-GAL4 driving UAS-mCherry RNAi), raised at 29 °C, hybridized with fluorescent probes that detect either wild-type D. yakuba mtDNA (greyscale in d; green in d″) or mutant D. melanogaster mtDNA (greyscale in d′; magenta in d″). Selection for wild-type mtDNA is observed as indicated by the arrows in d and d″. e, e′, e″, f, f′, f″, Selection for wild-type mtDNA is no longer observed when Mitofusin (Mfn) is overexpressed (nos-GAL4 driving UAS-marf) or when the expression of Drp1 is reduced (nos-GAL4 driving UAS-Drp1.miRNA.CDS). Wild-type D. yakuba mtDNA, greyscale in e, f, green in e″, f″; mutant D. melanogaster mtDNA, greyscale in e′, f′, magenta in e″, f″. The dashed outlines delineate the germline. g, Scatter plot showing the percentage of mutant D. melanogaster mtDNA, as assayed by qPCR, in carcasses (carc.) and ovaries of heteroplasmic flies in which the wild-type mtDNA was either from D. yakuba or D. melanogaster. mCherry RNAi was expressed in the ovaries under control of nos-GAL4. h, Scatter plot of the amount of mutant D. melanogaster (purple) and wild-type D. yakuba (green) mtDNA, as assayed by qPCR, of young embryos laid by heteroplasmic females in which Mitofusin was overexpressed in the germline (Mfn OE, n = 4) normalized to the amount of mutant and wild-type mtDNA in young embryos laid by control heteroplasmic females (Ctrl; nos-GAL4 driving UAS-mCherry RNAi, n = 4). i, Same as g, except the analysis was performed on ovaries in which both wild-type and mutant mtDNAs were from D. melanogaster (Ctrl, n = 24; Mfn OE, n = 21). Mitofusin overexpression increases the levels of mutant mtDNA.
