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PMID:19521502

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Citation

Neklesa, TK and Davis, RW (2009) A genome-wide screen for regulators of TORC1 in response to amino acid starvation reveals a conserved Npr2/3 complex. PLoS Genet. 5:e1000515

Abstract

TORC1 is a central regulator of cell growth in response to amino acid availability, yet little is known about how it is regulated. Here, we performed a reverse genetic screen in yeast for genes necessary to inactivate TORC1. The screen consisted of monitoring the expression of a TORC1 sensitive GFP-based transcriptional reporter in all yeast deletion strains using flow cytometry. We find that in response to amino acid starvation, but not to carbon starvation or rapamycin treatment, cells lacking NPR2 and NPR3 fail to fully (1) activate transcription factors Gln3/Gat1, (2) dephosphorylate TORC1 effector Npr1, and (3) repress ribosomal protein gene expression. Both mutants show proliferation defects only in media containing a low quality nitrogen source, such as proline or ammonia, whereas no defects are evident when cells are grown in the presence of glutamine or peptone mixture. Proliferation defects in npr2Delta and npr3Delta cells can be completely rescued by artificially inhibiting TORC1 by rapamycin, demonstrating that overactive TORC1 in both strains prevents their ability to adapt to an environment containing a low quality nitrogen source. A biochemical purification of each demonstrates that Npr2 and Npr3 form a heterodimer, and this interaction is evolutionarily conserved since the human homologs of NPR2 and NPR3 (NPRL2 and NPRL3, respectively) also co-immunoprecipitate. We conclude that, in yeast, the Npr2/3 complex mediates an amino acid starvation signal to TORC1.

Links

PubMed PMC2686269 Online version:10.1371/journal.pgen.1000515

Keywords

Amino Acids/metabolism; Dimerization; Evolution, Molecular; Gene Expression Regulation, Fungal; HeLa Cells; Humans; Intracellular Signaling Peptides and Proteins/genetics; Intracellular Signaling Peptides and Proteins/metabolism; Protein Binding; Saccharomyces cerevisiae/genetics; Saccharomyces cerevisiae/metabolism; Saccharomyces cerevisiae Proteins/genetics; Saccharomyces cerevisiae Proteins/metabolism

Significance

Annotations

Gene product Qualifier GO Term Evidence Code with/from Aspect Extension Notes Status

YEAST:NPR3

involved_in

GO:1903833: positive regulation of cellular response to amino acid starvation

ECO:0000315: mutant phenotype evidence used in manual assertion

P

Seeded From UniProt

complete

YEAST:NPR3

involved_in

GO:1904262: negative regulation of TORC1 signaling

ECO:0000315: mutant phenotype evidence used in manual assertion

P

Seeded From UniProt

complete

YEAST:NPR3

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:P39923

F

Seeded From UniProt

complete

YEAST:NPR2

involved_in

GO:1903833: positive regulation of cellular response to amino acid starvation

ECO:0000315: mutant phenotype evidence used in manual assertion

P

Seeded From UniProt

complete

YEAST:NPR2

involved_in

GO:1904262: negative regulation of TORC1 signaling

ECO:0000315: mutant phenotype evidence used in manual assertion

P

Seeded From UniProt

complete

YEAST:NPR2

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:P38742

F

Seeded From UniProt

complete

HUMAN:NPRL3

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:Q8WTW4

F

Seeded From UniProt

complete

HUMAN:NPRL2

enables

GO:0005515: protein binding

ECO:0000353: physical interaction evidence used in manual assertion

UniProtKB:Q12980

F

Seeded From UniProt

complete


See also

References

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