Oligomerization of the Mg²⁺‐transport proteins Alr1p and Alr2p in yeast plasma membrane

Abstract

Alr1p is an integral plasma membrane protein essential for uptake of Mg2+ into yeast cells. Homologs of Alr1p are restricted to fungi and some protozoa. Alr1‐type proteins are distant relatives of the mitochondrial and bacterial Mg2+‐transport proteins, Mrs2p and CorA, respectively, with which they have two adjacent TM domains and a short Mg2+ signature motif in common. The yeast genome encodes a close homolog of Alr1p, named Alr2p. Both proteins are shown here to be present in the plasma membrane. Alr2p contributes poorly to Mg2+ uptake. Substitution of a single arginine with a glutamic acid residue in the loop connecting the two TM domains at the cell surface greatly improves its function. Both proteins are shown to form homo‐oligomers as well as hetero‐oligomers. Wild‐type Alr2p and mutant Alr1 proteins can have dominant‐negative effects on wild‐type Alr1p activity, presumably through oligomerization of low‐function with full‐function proteins. Chemical cross‐linking indicates the presence of Alr1 oligomers, and split‐ubiquitin assays reveal Alr1p–Alr1p, Alr2p–Alr2p, and Alr1p–Alr2p interactions. These assays also show that both the N‐terminus and C‐terminus of Alr1p and Alr2p are exposed to the inner side of the plasma membrane.

Nonsense-mediated mRNA decay maintains translational fidelity by limiting magnesium uptake

“…S3; data not shown). Consistent with the notion that Alr2p plays a minor role in Mg 2+ uptake (MacDiarmid and Gardner 1998; Wachek et al 2006), an alr2D allele does not counteract the nonsense suppression phenotype of upf1D cells or prevent accumulation of Mg 2+ in the presence of elevated Alr1p levels (data not shown; see below). Collectively, these results suggest a model in which the readthrough phenotype of NMD-deficient cells is caused by elevated Mg 2+ levels, primarily through increased Alr1p expression, and their effects on translational fidelity.…”

MNR2 Regulates Intracellular Magnesium Storage in Saccharomyces cerevisiae

“…Several studies have suggested that ALR2 does not contribute to Mg homeostasis in S288C-derived strains (MacDiarmid and Gardner 1998; Wachek et al 2006;da Costa et al 2007). However, ALR1 inactivation had a less severe effect on viability in a W303-derived strain (Tran et al 2000), suggesting that ALR2 was functional in this genetic background.…”

“…ALR1 inactivation conferred Mg-dependent growth and blocked Mg uptake (MacDiarmid and Gardner 1998;Graschopf et al 2001). The closely related ortholog Alr2 was not essential for growth, but could compensate for the loss of Alr1 when overexpressed and was found to physically associate with Alr1 in vivo (MacDiarmid and Gardner 1998; Wachek et al 2006). Further studies identified two related proteins in the mitochondrial inner membrane (Mrs2 and Lpe10).…”

“…If the Alr proteins do play a role in divalent cation uptake, this phenotype suggests that their activity and/or expression might be increased in mnr2 strains. A mechanism for this effect is suggested by the observation that Alr1 and Alr2 expression may be regulated by Mg supply (Graschopf et al 2001;Wachek et al 2006), which in turn may be determined by Mnr2 activity. An upregulation of the Alr systems might also explain the divalent cation sensitivity of mnr2 strains (Table 2 and Figure 1A).…”

Molecular identification of ancient and modern mammalian magnesium transporters

“…Alr1 protein comprises two TMs and possesses the conserved GMN motif, which has been shown to be essential for yeast Mg 2ϩ transport (78). As with CorA and Mrs2, Alr1 functions as an oligomer (162). The Alr1 transcript is increased in low-Mg 2ϩ environments, and the Alr1 protein is diminished with high external Mg 2ϩ , indicating that Alr1 expression is regulated in yeast (46).…”