Figure 3.
Mitochondrial respiration is activated in region 2B and is essential for mtDNA selective inheritance. (A) Expression of mtDNA- and nuclear-encoded ETCs genes in the germarium. The spatial patterns of nuclear and mitochondrial encoded mRNAs were revealed by smFISH assay. Fluorescently labeled probes targeted mtDNA-encoded ND4 and CoI transcripts and the nuclear-encoded ND-SDGH and cox5A transcripts. Scale bar, 10 µm. (B) Quantification of total immunofluorescence intensity per volume in regions 2A and 2B (n = 6). The expression of all RNAs was markedly increased at region 2B. Data presented are means ± SD. **, P < 0.005; ***, P < 0.001. (C) Mitochondria membrane potential staining demonstrated by the mitochondrial membrane potential indicator TMRM (red) and mitochondrial fluorescent dye MitoTracker (green). The strong red signal indicates markedly increased membrane potential at regions 2B and 3 in merged image. The ratio of red to green fluorescence intensity is shown as the pseudo-color ratiometric image. The developing regions of germarium germ cells are outlined. Scale bar, 10 µm. (D) Mitochondrial respiratory activity in ovary using the colorimetric assay. Ovaries from control and cox5A knockdown flies driven by nanos-gal4 were stained for dual succinate dehydrogenase (complex II) and cytochrome c oxidase (complex IV) activity. Representative images for each group are shown. Intense brown color, indicating that both complex II and complex IV are active, was prominent in region 2B but mostly absent in regions 1 and 2A. The blue color indicates that complex IV activity is greatly reduced in cox5A knockdown ovary. The developing regions of germarium germ cells are outlined. Scale bar, 10 µm. (E) Selection against the deleterious mtDNA mutation (ts) in heteroplasmic flies was compromised by knocking down cox5A in the germline using nanos-gal4 driver (n = 10 for each genotype). Data presented are means ± SD. P < 0.001.

Mitochondrial respiration is activated in region 2B and is essential for mtDNA selective inheritance. (A) Expression of mtDNA- and nuclear-encoded ETCs genes in the germarium. The spatial patterns of nuclear and mitochondrial encoded mRNAs were revealed by smFISH assay. Fluorescently labeled probes targeted mtDNA-encoded ND4 and CoI transcripts and the nuclear-encoded ND-SDGH and cox5A transcripts. Scale bar, 10 µm. (B) Quantification of total immunofluorescence intensity per volume in regions 2A and 2B (n = 6). The expression of all RNAs was markedly increased at region 2B. Data presented are means ± SD. **, P < 0.005; ***, P < 0.001. (C) Mitochondria membrane potential staining demonstrated by the mitochondrial membrane potential indicator TMRM (red) and mitochondrial fluorescent dye MitoTracker (green). The strong red signal indicates markedly increased membrane potential at regions 2B and 3 in merged image. The ratio of red to green fluorescence intensity is shown as the pseudo-color ratiometric image. The developing regions of germarium germ cells are outlined. Scale bar, 10 µm. (D) Mitochondrial respiratory activity in ovary using the colorimetric assay. Ovaries from control and cox5A knockdown flies driven by nanos-gal4 were stained for dual succinate dehydrogenase (complex II) and cytochrome c oxidase (complex IV) activity. Representative images for each group are shown. Intense brown color, indicating that both complex II and complex IV are active, was prominent in region 2B but mostly absent in regions 1 and 2A. The blue color indicates that complex IV activity is greatly reduced in cox5A knockdown ovary. The developing regions of germarium germ cells are outlined. Scale bar, 10 µm. (E) Selection against the deleterious mtDNA mutation (ts) in heteroplasmic flies was compromised by knocking down cox5A in the germline using nanos-gal4 driver (n = 10 for each genotype). Data presented are means ± SD. P < 0.001.

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