5 years ago

More Than One Way In - Three Gln3 Sequences Required To Relieve Negative Ure2 Regulation and Support Nuclear Gln3 Import in Saccharomyces cerevisiae.

Terrance G Cooper, Rajendra Rai, Jennifer J Tate
Gln3 is responsible for Nitrogen Catabolite Repression-sensitive transcriptional activation in the yeast Saccharomyces cerevisiae In nitrogen-replete medium, Gln3 is cytoplasmic and NCR-sensitive transcription is repressed. In nitrogen-limiting medium, in cells treated with TorC1 inhibitor, rapamycin or glutamine synthetase inhibitor, methionine sulfoximine (Msx), Gln3 becomes highly nuclear and NCR-sensitive transcription derepressed. Previously, nuclear Gln3 localization was concluded to be mediated by a single nuclear localization sequence, NLS1. Here we show that nuclear Gln3-Myc(13) localization is significantly more complex than previously appreciated. We identify three Gln3 sequences, other than NLS1 that are highly required for nuclear Gln3-Myc(13) localization. Two of these sequences exhibit characteristics of monopartite (K/R-Rich NLS) and bipartite (S/R NLS) NLSs, respectively. Mutations altering these sequences are partially epistatic to a ure2Δ The third sequence, the Ure2 relief sequence, exhibits no predicted NLS homology and is only necessary when Ure2 is present. Substitution of the basic amino acid repeats in the Ure2 relief sequence or phosphomimetic aspartate substitutions for the serine residues between them abolishes nuclear Gln3-Myc(13) localization in response to both limiting nitrogen and rapamycin treatment. In contrast, Gln3-Myc(13) responses are normal in parallel serine to alanine substitution mutants. These observations suggest that Gln3 responses to specific nitrogen environments likely occur in multiple steps that can be genetically separated. At least one general step, that is associated with the Ure2 relief sequence, may be prerequisite for responses to the specific stimuli of growth in poor nitrogen sources and rapamycin inhibition of TorC1.

Publisher URL: http://doi.org/10.1534/genetics.117.300457

DOI: 10.1534/genetics.117.300457

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