Dissecting the effects of mtDNA variations on complex traits using mouse conplastic strains

Abstract

Previous reports have demonstrated that the mtDNA of mouse common inbred strains (CIS) originated from a single female ancestor and that mtDNA mutations occurred during CIS establishment. This situation provides a unique opportunity to investigate the impact of individual mtDNA variations on complex traits in mammals. In this study, we compiled the complete mtDNA sequences of 52 mouse CIS. Phylogenetic analysis demonstrated that 50 of the 52 CIS descended from a single female Mus musculus domesticus mouse, and mtDNA mutations have accumulated in 26 of the CIS. We then generated conplastic strains on the C57BL/6J background for 12 mtDNA variants with one to three functional mtDNA mutations. We also generated conplastic strains for mtDNA variants of the four M. musculus subspecies, each of which contains hundreds of mtDNA variations. In total, a panel of conplastic strains was generated for 16 mtDNA variants. Phenotypic analysis of the conplastic strains demonstrated that mtDNA variations affect susceptibility to experimental autoimmune encephalomyelitis and anxiety-related behavior, which confirms that mtDNA variations affect complex traits. Thus, we have developed a unique genetic resource that will facilitate exploration of the biochemical and physiological roles of mitochondria in complex traits.

Figure 1.

Phylogenetic tree of mtDNA from mouse inbred strains. The tree was generated with ClustalW software on the basis of the mtDNA sequences of 56 mouse inbred strains, including 52 classic inbred strains (CIS) and four wild-derived inbred strains (WIS). For the four WIS, the taxonomic classification is indicated. Numbers correspond to bootstrap values (percentage of 1000 total bootstrap replicates).

Figure 2.

The most parsimonious tree for all mouse mtDNA variations found in 50 common inbred strains. The mtDNA sequence of the AKR/J strain (AB042432) was taken as the reference sequence, with the positions of the mtDNA variations indicated. nt9821, an A-repeat variation, and some other heteroplasmic variations were not included.

Figure 3.

mtDNA variations affect susceptibility to EAE. (A) Development of EAE in five conplastic strains. The x-axis presents the days post-immunization, and the y-axis indicates the EAE score. (B) Comparison of EAE clinical phenotypes, including disease incidence, onset day (only the disease mice were included), maximal disease score, and area under the curve (AUC). The C57BL/6J-mt^AKR/J strain was taken as the reference strain, and the statistical significance was calculated for each strain compared with the C57BL/6J-mt^AKR/J strain. Asterisks indicate significant differences: *P < 0.05; **P < 0.01; ***P < 0.001.

Figure 4.

Behavior of C57BL/6J (B6), C57BL/6J-mt^AKR/J (B6_AKR), and C57BL/6J-mt^FVB/N (B6_FVB) mice on the elevated plus maze. (A) Anxiety was measured by the percentage of entries into open arms, the percentage of time spent in open arms, and the distance walked in open arms. (B) Locomotor activity was recorded as total entries into open and closed arms, the number of times mice entered closed arms, and the distance walked in closed arms. Data represent means and SEM of nine to 12 mice. Asterisks indicate significant differences between strains; : *P < 0.05; **P < 0.01; Mann-Whitney test.