Cross-talk between ammonium transporters in yeast and interference by the soybean SAT1 protein.
Ammonium uptake in the yeast Saccharomyces cerevisiae involves three membrane transporters (Mep1, -2 and -3) belonging to an evolutionarily conserved protein family that also includes the rhesus (Rh) blood group polypeptides of erythrocytes. We show here that, in the 26972c mutant defective in NH4+ transport, the Mep1 protein carrying an amino acid substitution in its cytoplasmic C-terminus trans-inhibits the closely related Mep3 protein. The same mutation introduced into Mep3 leads to loss of transport activity and this inactive form also trans-inhibits native Mep3. Inhibition of Mep3 is post-translational and can be overcome by overexpression. These results are consistent with a direct interaction between Mep proteins, as is the case for the Rh polypeptides. The soybean GmSAT1 gene, recently cloned for its ability to complement the NH4+ transport defect of strain 26972c, has been described as an NH4+ channel protein involved in the transfer of fixed nitrogen from the bacteroid to the host plant. We show here that GmSAT1 contains a sequence homologous to the DNA-binding domain of basic helix-loop-helix (bHLH) transcription factors. We also show that GmSAT1 restores NH4+ uptake in the yeast mutant by interfering with the inhibition of Mep3. Our results are not consistent with a direct role of GmSAT1 in ammonium transport.
Marini
A M
AM
Laboratoire de Physiologie Cellulaire, Université Libre de Bruxelles, CP300, Institut de Biologie et de Médecine Moléculaires, Rue des Pr. Jeener et Brachet, 12, 6041 Gosselies, Belgium.
Springael
J Y
JY
Frommer
W B
WB
André
B
B
D000595:Amino Acid Sequence; D001692:Biological Transport; D002352:Carrier Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D027682:Cation Transport Proteins; D003470:Culture Media; D004591:Electrophoresis, Polyacrylamide Gel; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D015966:Gene Expression Regulation, Fungal; D018257:Helix-Loop-Helix Motifs; D006801:Humans; D015151:Immunoblotting; D008969:Molecular Sequence Data; D009154:Mutation; D010957:Plasmids / Q000235:genetics; D000644:Quaternary Ammonium Compounds / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D016415:Sequence Alignment; D030262:Soybean Proteins
D002352:Carrier Proteins; D027682:Cation Transport Proteins; D003470:Culture Media; D005656:Fungal Proteins; C088951:MEP1 protein, S cerevisiae; C107270:MEP2 protein, S cerevisiae; C107271:MEP3 protein, S cerevisiae; D000644:Quaternary Ammonium Compounds; C114002:SAT1 protein, glycine max; D029701:Saccharomyces cerevisiae Proteins; D030262:Soybean Proteins