Saccharomyces cerevisiae GATA Sequences Function as TATA Elements during Nitrogen Catabolite Repression and When Gln3p Is Excluded from the Nucleus by Overproduction of Ure2p

Abstract

Saccharomyces cerevisiae selectively uses good nitrogen sources (glutamine) in preference to poor ones (proline) by repressing GATA factor-dependent transcription of the genes needed to transport and catabolize poor nitrogen sources, a physiological process designated nitrogen catabolite repression (NCR). We show that some NCR-sensitive genes (CAN1, DAL5, DUR1,2, and DUR3) produce two transcripts of slightly different sizes. Synthesis of the shorter transcript is NCR-sensitive and that of the longer transcript is not. The longer transcript also predominates in gln3⌬ mutants irrespective of the nitrogen source provided. We demonstrate that the longer mRNA species arises through the use of an alternative transcription start site generated by Gln3p-binding sites (GATAAs) being able to act as surrogate TATA elements. The ability of GATAAs to serve as surrogate TATAs, i.e. when synthesis of the shorter, NCR-sensitive transcripts are inhibited, correlates with sequestration of enhanced green fluorescent protein (EGFP)-Gln3p in the cytoplasm in a way that is indistinguishable from that seen with EGFP-Ure2p. However, when the shorter, NCR-sensitive DAL5 transcript predominates, EGFP-Gln3p is nuclear. These data suggest that the mechanism underlying NCR involves the cytoplasmic association of Ure2p with Gln3p, an interaction that prevents Gln3p from reaching it is binding sites upstream of NCR-sensitive genes.

Methionine Sulfoximine Treatment and Carbon Starvation Elicit Snf1-independent Phosphorylation of the Transcription Activator Gln3 in Saccharomyces cerevisiae

“…24 -27). The correlations that Gln3 is nuclear with a poor nitrogen source (28) or in rich medium containing rapamycin (9,16) were consistent with the suggestion that Tor1/2 transmit the signal of nitrogen excess to Gln3. The observations that rapamycin induces Gln3 dephosphorylation and that this dephosphorylation depends on Sit4, a type-2A serine/threonine phosphatase regulated by Tor1/2, Tap42, and Tip41, further connected Gln3 intracellular localization to the upper portion of the Tor1/2 pathway (see Refs.…”

Mks1p Is Required for Negative Regulation of Retrograde Gene Expression in Saccharomyces cerevisiae but Does Not Affect Nitrogen Catabolite Repression-sensitive Gene Expression

“…When cells are grown in excess nitrogen, Gln3p and Gat1p are not bound to their target promoter sequences thus making these GATAs available, in some instances, to serve as surrogate TATA elements (10). This behavior correlates with green fluorescent protein-Gln3p and green fluorescent protein-Gat1p being nuclear when NCR-sensitive expression is high and cytoplasmic when low (10).…”

The crystal structure of the nitrogen regulation fragment of the yeast prion protein Ure2p

“…Ure2p directly interacts with the DNA binding protein Gln3p, preventing its entry into the nucleus to promote transcription of an array of genes involved in assimilation of poor nitrogen sources (12)(13)(14)(15). Ure2p is itself regulated by Mks1p, mediating nitrogen catabolite repression (16), and by the TOR system, a global regulator that transmits nutritional information to many cellular components (14,17,18).…”