title: Alterations in the Ure2 αCap domain elicit different GATA factor responses to rapamycin treatment and nitrogen limitation.
abstract: Ure2 is a phosphoprotein and central negative regulator of nitrogen-responsive Gln3/Gat1 localization and their ability to activate transcription. This negative regulation is achieved by the formation of Ure2-Gln3 and -Gat1 complexes that are thought to sequester these GATA factors in the cytoplasm of cells cultured in excess nitrogen. Ure2 itself is a dimer the monomer of which consists of two core domains and a flexible protruding αcap. Here, we show that alterations in this αcap abolish rapamycin-elicited nuclear Gln3 and, to a more limited extent, Gat1 localization. In contrast, these alterations have little demonstrable effect on the Gln3 and Gat1 responses to nitrogen limitation. Using two-dimensional PAGE we resolved eight rather than the two previously reported Ure2 isoforms and demonstrated Ure2 dephosphorylation to be stimulus-specific, occurring after rapamycin treatment but only minimally if at all in nitrogen-limited cells. Alteration of the αcap significantly diminished the response of Ure2 dephosphorylation to the TorC1 inhibitor, rapamycin. Furthermore, in contrast to Gln3, rapamycin-elicited Ure2 dephosphorylation occurred independently of Sit4 and Pph21/22 (PP2A) as well as Siw14, Ptc1, and Ppz1. Together, our data suggest that distinct regions of Ure2 are associated with the receipt and/or implementation of signals calling for cessation of GATA factor sequestration in the cytoplasm. This in turn is more consistent with the existence of distinct pathways for TorC1- and nitrogen limitation-dependent control than it is with these stimuli representing sequential steps in a single regulatory pathway.
authors.affiliation: Institut de Recherches Microbiologiques J.-M. Wiame, Laboratoire de Microbiologie Université Libre de Bruxelles, B1070 Brussels, Belgium.
authors.forename: Andre
authors.initials: A
authors.lastname: Feller
authors.forename: Isabelle
authors.initials: I
authors.lastname: Georis
authors.forename: Jennifer J
authors.initials: JJ
authors.lastname: Tate
authors.forename: Terrance G
authors.initials: TG
authors.lastname: Cooper
authors.forename: Evelyne
authors.initials: E
authors.lastname: Dubois
mesh_terms: D000935:Antifungal Agents / Q000494:pharmacology*; D050980:GATA Transcription Factors / Q000235:genetics / Q000378:metabolism; D015966:Gene Expression Regulation, Fungal / Q000187:drug effects*; D005979:Glutathione Peroxidase / Q000235:genetics / Q000378:metabolism; D009154:Mutation; D009584:Nitrogen / Q000172:deficiency*; D011328:Prions / Q000235:genetics / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012441:Saccharomyces cerevisiae / Q000235:genetics* / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects; D020123:Sirolimus / Q000494:pharmacology*; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D000935:Antifungal Agents; C096100:GAT1 protein, S cerevisiae; D050980:GATA Transcription Factors; C071664:GLN3 protein, S cerevisiae; D011328:Prions; D029701:Saccharomyces cerevisiae Proteins; C561842:TORC1 protein complex, S cerevisiae; D014157:Transcription Factors; D005979:Glutathione Peroxidase; C067020:URE2 protein, S cerevisiae; D009584:Nitrogen; D020123:Sirolimus