Isolation of the NPR1 gene responsible for the reactivation of ammonia-sensitive amino-acid permeases in Saccharomyces cerevisiae. RNA analysis and gene dosage effects.
The NPR1 gene codes for a protein, called the nitrogen permease reactivator protein or Npr1, which appears to promote the activity of several permeases for nitrogenous substances under conditions of nitrogen catabolite derepression, but fails to do so in the presence of ammonium ions. This gene has been cloned. Its transcription seems unaffected by growth on ammonia, so any ammonia regulation of Npr1 function most likely occurs at another level. In order to elucidate further the mechanism of permease inactivation, which requires an intact NPI1 gene product (NPI1 for nitrogen permease inactivator gene, formerly termed MUT2) and the role of Npr1 in counteracting this process, we have studied the effects of NPR1 and NPI1 gene dosage on general amino-acid permease activity. On nitrogen-derepressing media, NPR1 gene dose can be increased from 1 copy in a diploid to 16 plasmid-borne copies in a haploid strain without altering general amino-acid permease activity. On minimal ammonia medium, the plasmid-bearing haploid cells exhibit low but increased general amino-acid permease activity with respect to non-transformed cells. The adverse effect of the NPI1 gene product on general amino-acid permease activity is reduced when NPI1 gene dose is decreased to 1 gene copy in a diploid strain, regardless of the nitrogen source. We hypothesize that this product inactivates the permease by stoichiometric binding and that the Npr1 protein or a product of its catalytic action opposes this binding under conditions of nitrogen derepression.
D026905:Amino Acid Transport Systems; D000641:Ammonia / Q000494:pharmacology*; D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000032:analysis; D004789:Enzyme Activation; D005786:Gene Expression Regulation; D005800:Genes, Fungal; D026901:Membrane Transport Proteins / Q000235:genetics*; D009154:Mutation; D012331:RNA, Fungal / Q000032:analysis*; D012441:Saccharomyces cerevisiae / Q000201:enzymology* / Q000235:genetics
D026905:Amino Acid Transport Systems; D004271:DNA, Fungal; D026901:Membrane Transport Proteins; D012331:RNA, Fungal; D000641:Ammonia