et al. (1992). Mrs2p was not identified as a putative Mg2+ transporter until Bui et al. (1999). Gregan et al. (2001a) identified LPE10 by homology to MRS2 and showed that both LPE10 and MRS2 mutants altered the Mg2+ content of yeast mitochondria and affected RNA splicing activity in the organelle. Mg2+ transport has been shown to be directly mediated by Mrs2p, but not for Lpe10p. The Mrs2p and Lpe10p proteins are 470 and 413 amino acid residues in size, respectively, and a 250–300 amino acid region in the middle of the proteins shows a weak similarity to the full CorA protein. The TM topologies of the Mrs2p and Lpe10p proteins have been assessed using a protease protection assay and are shown in the figure. TM 1 and 2 correspond to TM 2 and 3 in the CorA protein. The conserved GMN motif is at the outside end of the first TM domain, and when the glycine (G) in this motif was mutated to a cysteine (C) in Mrs2p, Mg2+ transport was strongly reduced. The figure shows the experimentally determined topology of Mrs2p and Lpe10p as adapted from Bui et al. (1999) and Gregan et al. (2001a). The GMN motif location is indicated in red and the TM domains in light blue. The orientation in the membrane and the positions of the N- and C-termini are indicated. The various sizes of the soluble domains are given in amino acids (AA), TM domains are numbered, and the figure is not drawn to scale. Mrs2p has been localised to the mitochondrial inner membrane by subcellular fractionation and immunodetection and Lpe10p to the mitochondria. Mitochondria lacking Mrs2p do not show a fast Mg2+ uptake, only a slow ‘leak’, and overaccumulation of Mrs2p leads to an increase in the initial rate of uptake. Additionally, CorA, when fused to the mitochondrial leader sequence of Mrs2p, can partially complement the mitochondrial defect conferred by the loss of either Mrs2p or Lpe10p. Hence, Mrs2p and/or Lpe10p may be the major Mg2+ uptake system for mitochondria. A possibility is that the proteins form heterodimers, as neither protein (when overexpressed) can fully complement the loss of the other. The characteristics of Mg2+ uptake in isolated mitochondria by Mrs2p were quantified using mag-fura 2. The uptake of Mg2+ by Mrs2p shared a number of attributes with CorA. First, Mg2+ uptake was directly dependent on the electric potential (ΔΨ) across the boundary