Figure 1.
Mitochondrial DNA content and enzyme activity in muscle from PolG mice.
(A) Mitochondrial DNA (mtDNA) was measured by Real Time-PCR (qT-PCR) in muscle from young (3–6 mo) and older (8–15 mo) WT and PolG mice. Relative mtDNA content was normalized to β-actin and a graphical representation of the summary data is shown (n = 7–9). (B) Cytochrome c oxidase (COX) activity was expressed as unit per gram of tissue (n = 7–9). Significance was set at P<0.05 and all data are represented as mean ± SE. *P<0.05 vs. age-matched WT.
Figure 2.
PolG mice display higher levels of mitochondrial regulatory proteins and transcription factors.
AMPKα activation (A), PGC-1α (B), NRF-1 (C), and Tfam (D) were determined by Western Blotting in muscle from young (3–6 mo) and older (8–15 mo) WT and PolG animals. AMPKα activation is determined by phosphorylated AMPKα over total AMPKα. Representative blots are shown above with a graphical summary of the data below (n = 7–13). Significance was set at P<0.05 and all data are represented as mean ± SE. Data are expressed as arbitrary units (AU). #P<0.05 main effect of age. *P<0.05 main effect of genotype.
Figure 3.
Altered mitochondrial fusion proteins in mtDNA mutator mice.
Immunoblotting of mitochondrial fusion proteins Mfn1 (A), Mfn2 (B) and Opa1 (C) in skeletal muscle from young (3–6 mo) and older (8–15 mo) WT and PolG animals. Representative blots are shown above with a graphical summary of the data below (n = 7–13). Two bands were detected for Opa1 representing the long and short isoforms. Both bands were quantified and total Opa1 protein content displayed in the graph. Significance was set at P<0.05 and all data are represented as mean ± SE. Data are expressed as arbitrary units (AU). #P<0.05 main effect of age.
Figure 4.
Altered expression of mitochondrial fission proteins in mtDNA mutator mice.
Fis1 (A) and Drp1 (B) protein content was determined in muscle from young (3–6 mo) and older (8–15 mo) WT and PolG animals. A graphical summary along with representative blots are depicted (n = 7–13). Significance was set at P<0.05 and all data are represented as mean ± SE. Data are expressed as arbitrary units (AU). #P<0.05 main effect of age. *P<0.05 main effect of genotype.
Figure 5.
Autophagy proteins are upregulated in muscle from PolG animals.
The content of autophagy proteins Beclin-1 (A), Atg5 (B), ULK1 (C), p62 (D), and LC3-II (E) was measured with Western Blotting in skeletal muscle from young (3–6 mo) and older (8–15 mo) WT and PolG animals. Representative blots are shown above with a graphical summary of the data below (n = 7–13). Significance was set at P<0.05 and all data are represented as mean ± SE. Data are expressed as arbitrary units (AU). #P<0.05 main effect of age. *P<0.05 main effect of genotype.
Figure 6.
Effect of mtDNA mutations on mitochondrial protein import machinery.
Levels of mitochondrial protein import machinery cytosolic Hsp70 (A), mitochondrial Hsp70 (B), Tim23 (C), and Tom22 (D) were measured in muscle from young (3–6 mo) and older (8–15 mo) WT and PolG animals. Representative blots are shown above with a graphical summary of the data below (n = 7–13). Significance was set at P<0.05 and all data are represented as mean ± SE. Data are expressed as arbitrary units (AU). #P<0.05 main effect of age. *P<0.05 main effect of genotype.
Table 1.
Comparison of changes in mitochondrial quality control proteins in premature and normally-aging mice.