No abstract

Many human mitochondrial diseases are associated with defects in the mitochondrial DNA (mtDNA). Mutated and wild-type forms of mtDNA often coexist in the same cell in a state called heteroplasmy. Here, we report the isolation of a Caenorhabditis elegans strain bearing the 3.1-kb uaDf5 deletion that removes 11 genes from the mtDNA. The uaDf5 deletion is maternally transmitted and has been maintained for at least 100 generations in a stable heteroplasmic state in which it accounts for approximately 60% of the mtDNA content of each developmental stage. Heteroplasmy levels vary between individual animals (from approximately 20 to 80%), but no observable phenotype is detected. The total mtDNA copy number in the uaDf5 mutant is approximately twice that of the wild type. The maternal transmission of the uaDf5 mtDNA is controlled by at least two competing processes: one process promotes the increase in the average proportion of uaDf5 mtDNA in the offspring, while the second promotes a decrease. These two forces prevent the segregation of the mtDNAs to homoplasmy.

The growth and development of Caenorhabditis elegans are energy-dependent and rely on the mitochondrial respiratory chain (MRC) as the major source of ATP. The MRC is composed of ϳ70 nuclear and 12 mitochondrial gene products. Complexes I and V are multisubunit proteins of the MRC. The nuo-1 gene encodes the NADH-and FMN-binding subunit of complex I, the NADH-ubiquinone oxidoreductase. The atp-2 gene encodes the active-site subunit of complex V, the ATP synthase. The nuo-1(ua1) and atp-2(ua2) mutations are both lethal. They result in developmental arrest at the third larval stage (L3), arrest of gonad development at the second larval stage (L2), and impaired mobility, pharyngeal pumping, and defecation. Surprisingly, the nuo-1 and atp-2 mutations significantly lengthen the life spans of the arrested animals. When MRC biogenesis is blocked by chloramphenicol or doxycycline (inhibitors of mitochondrial translation), a quantitative and homogeneous developmental arrest as L3 larvae also results. The common phenotype induced by the mutations and drugs suggests that the L3-to-L4 transition may involve an energy-sensing developmental checkpoint. Since ϳ200 gene products are needed for MRC assembly and mtDNA replication, transcription, and translation, we predict that L3 arrest will be characteristic of mutations in these genes. The human mitochondrial respiratory chain (MRC)1 is composed of Ͼ80 subunits, but requires Ͼ100 additional genes for its assembly (1). These additional genes encode the import apparatus for transporting polypeptides into the organelle, chaperones, and other assembly factors, as well as the machinery needed to replicate, transcribe, and translate the 13 MRC subunits encoded by mtDNA. The MRC generates the majority of cellular ATP. Thus, the loss of one member of this large class of genes can compromise the entire pathway of energy production. The consequences of such events are diverse and debilitating (2-4). The metabolism and structure of the Caenorhabditis elegans MRC closely parallel its mammalian counterpart (5). Moreover, the nematode mtDNA is similar in size and gene content to the human mtDNA (6).The MRC is organized into five multisubunit proteins. Complexes I-IV form the electron transport chain, which couples the oxidation of NADH and succinate to the reduction of oxygen and the formation of a proton gradient. Complex I (NADHubiquinone oxidoreductase) is an elaborate enzyme that catalyzes the transfer of electrons from NADH to ubiquinone. Bovine complex I consists of ϳ43 subunits, an FMN cofactor, and up to eight iron-sulfur clusters (7, 8). The FMN-and NADHbinding sites, as well as one iron-sulfur cluster, are considered to reside in the 51-kDa subunit. One predicted C. elegans gene product (C09H10.3) bears strong sequence resemblance (75% identical amino acids) to the human gene. We have designated the gene encoding the C. elegans NADH-ubiquinone oxidoreductase 51-kDa subunit as nuo-1. Complex I deficiency is one of the most frequently encountered mitochondrial defects (9 -11). Mutat...