The Saccharomyces cerevisiae LEP1/SAC3 gene is associated with leucine transport.
Leucine uptake by Saccharomyces cerevisiae is mediated by three transport systems, the general amino acid transport system (GAP), encoded by GAP1, and two group-specific systems (S1 and S2), which also transport isoleucine and valine. A new mutant defective in both group-specific transport activities was isolated by employing a gap1 leu4 strain and selecting for trifluoroleucine-resistant mutants which also showed greatly reduced ability to utilize L-leucine as sole nitrogen source and very low levels of [14C]L-leucine uptake. A multicopy plasmid containing a DNA fragment which complemented the leucine transport defect was isolated by selecting for transformants that grew normally on minimal medium containing leucine as nitrogen source and subsequently assaying [14C]L-leucine uptake. Transformation of one such mutant, lep1, restored sensitivity to trifluoroleucine. The complementing gene, designated LEP1, was subcloned and sequenced. The LEP1 ORF encodes a large protein that lacks characteristics of a transporter or permease (i.e., lacks hydrophobic domains necessary for membrane association). Instead, Lep1p is a very basic protein (pI of 9.2) that contains a putative bipartite signal sequence for targeting to the nucleus, suggesting that it might be a DNA-binding protein. A database search revealed that LEP1 encodes a polypeptide that is identical to Sac3p except for an N-terminal truncation. The original identification of SAC3 was based on the isolation of a mutant allele, sac3-1, that suppresses the temperature-sensitive growth defect of an actin mutant containing the allele act1-1. Sac3p has been previously shown to be localized in the nucleus. When a lep1 mutant was crossed with a sac3 deletion mutant, no complementation was observed, indicating that the two mutations are functionally allelic.
Stella
C A
CA
Departamento de Bioquimica, Facultad de Medicina, Universidad de Buenos Aires, Argentina. cstella@mail.fmed.uba.ar
Korch
C
C
Ramos
E H
EH
Bauer
A
A
Kölling
R
R
Mattoon
J R
JR
D000483:Alleles; D026905:Amino Acid Transport Systems; D001483:Base Sequence; D001692:Biological Transport; D003001:Cloning, Molecular; D004271:DNA, Fungal; D004352:Drug Resistance, Microbial; D005656:Fungal Proteins / Q000235:genetics / Q000378:metabolism*; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D007930:Leucine / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D026901:Membrane Transport Proteins; D008969:Molecular Sequence Data; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000378:metabolism*; D029543:Nucleocytoplasmic Transport Proteins; D018272:Porins; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins
D026905:Amino Acid Transport Systems; D004271:DNA, Fungal; D005656:Fungal Proteins; D026901:Membrane Transport Proteins; D009687:Nuclear Proteins; D029543:Nucleocytoplasmic Transport Proteins; D018272:Porins; C103382:SAC3 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; C035177:5',5',5'-trifluoroleucine; D007930:Leucine