regulated expression, and gain of a mt targeting sequence. Nucleic acids could escape from the mitochondrion by several mechanisms, as thoroughly discussed by Thorsness and Weber (1996). The rate of mtDNA escape and uptake by the nucleus has been estimated to be relatively high (Thorsness and Fox, 1990; Thorsness and Weber, 1996). Once inside the nucleus, nucleic acids can integrate into the nuclear genome by double-strand break repair, as shown recently for yeast (Ricchetti et al., 1999), and perhaps by other mechanisms. Clues have been revealed as to the mechanisms of gene activation and targeting sequence acquisition, including gain of a targeting sequence (and perhaps upstream promoter and other regulatory elements) from a preexisting gene for a mt protein by a shuffling process (Kadowaki et al., 1996; our unpublished data) and by integration into a preexisting gene for a mt protein (Figueroa et al., 1999; Kubo et al., 1999).
After the nuclear copy of a transferred gene is activated and gains a mt targeting sequence, both genes must be expressed at least transiently, as described above for cox2 in legumes (Fig. 2; Adams et al., 1999). It is possible that the genes in both genomes could become fixed. Both nuclear and mt atp9 genes have been retained in Neurospora crassa (van den Boogaart et al., 1982) and Aspergillus nidulans (Brown et al., 1984; Ward and Turner, 1986), and both are functional at certain times during the life cycle of Neurospora (Bittner-Eddy et al., 1994). However, the most commonly observed outcome is that one gene (usually the mt gene, although this is influenced by sampling biases) becomes silenced and lost. Inactivation of the nuclear copy of a transferred gene results in a failed transfer, but the opportunity for repeated “attempts” at transfer can create a gene transfer ratchet (Doolittle, 1998). Both selection and chance factors may play a role in determining which gene is retained and which gene is inactivated. Our finding of approximately equal numbers of cox2 silencings in legume mt and nuclear genomes raises the possibility that it is largely random as to which gene became silenced in a given species, with disabling mutations inactivating either cox2 gene at comparable frequencies. Alternatively, if the rate of production of disabling mutations is higher in one genome or the other, then the equal numbers of silencings would reflect selection favoring the gene's retention in the high-rate genome. This is difficult to assess because, although we know that substitution rates are much higher in legume nuclear than mt genes (Wolfe et al., 1987), we do not know what the overall rate of disabling mutations is in either genome.
Several hypotheses have been proposed for selection favoring retention of the nuclear copy of a transferred and activated organelle gene and loss of the organellar copy. Presence of a gene in the nucleus allows for crossing over during meiosis, perhaps enabling beneficial mutations to be fixed more rapidly than in asexual organelle genomes (Blanchard and