Taken together, the results suggest that different transporters from the same superfamily do transport the same substances, but use different transport mechanisms. This may not be too surprising, since different functional relationships are found in diverse classes of membrane transporters, such as glucose transporters of the major facilitator superfamily (for example, the yeast protein ScHXT1 compared with the Arabidopsis protein AtSTP1) and amino acid transporters of the amino acid/polyamine/choline (APC) family (for example, the human protein HsCAT1 compared with the Arabidopsis protein AtCAT1). In both cases, the plant protein probably mediates proton co-transport, whereas the other mediates uniport.
One might speculate that Rh-like proteins are NH3
uniporters or NH4+
/cation antiporters (Figure ). In contrast, Mep and AMT ammonium transporters may either function as NH3
co-transporters or as NH4+
uniporters (Figure ). A chemical interaction of ammonium or ammonia with the proteins might explain why ammonium transporters are so selective for ammonium (compared with K+
, for example). It might be possible that members of different clades of the superfamily recognize either uncharged or charged forms of the substrate. Does this fit the evolutionary pattern? Phylogenetic analyses suggest that Rh proteins and Mep/AMT have an ancestral relationship (Figure ). Interestingly, the nematode worm and fruitfly contain both Mep/AMT and Rh homologs in their genomes. One higher plant sequence from Arabidopsis,
that of AtAMT2, groups closely to bacterial sequences, which are also closely related to yeast sequences [23
]. The archaebacterial sequences are closely related to some plant and animal sequences. Within a single species, co-assembly of different subunits into a heteromeric complex may lead to fine tuning of ammonium transport by Mep, AMT and Rh proteins.
Figure 2 A phylogenetic tree of Mep, AMT and Rh-like proteins. Rh and AMT/Mep proteins are found in bacteria, archaebacteria and eukaryotes. The amino acid sequence alignment was created using the program ClustalW, and phylogenetic analysis was done using the (more ...)
The absence of Rh-like proteins in higher plants suggests that either some of the AMT proteins also use a different coupling mechanism or that other, sequence-unrelated proteins are responsible for ammonium/ammonia export. In contrast to urea-excreting mammals, fish excrete mainly ammonium, through the gills. We would therefore predict that Rh-like proteins (such as AAF63256, also known as DrRh1) may be highly expressed in the gills and may be responsible for ammonium/ammonia excretion in fish. Knock-out mutants in plants, mice and fish will be excellent tools for dissecting the individual roles of AMT, Mep and Rh-like proteins in the different genomes and their relative contribution to net ammonium flux compared with other transporters that can mediate ammonium transport (such as potassium transporters, Na+/H+ antiporters or Na+/K+/Cl- cotransporters). Genome-wide analysis will also help to unravel the evolution of the mechanism of ammonium/ammonia transport.