Gln3p nuclear localization and interaction with Ure2p in Saccharomyces cerevisiae

Gln3p is one of two well characterized GATA family transcriptional activati on factors whose function is regulated by the nitrogen supply of the cell. When nitrogen is limiting, Gln3p and Gat1p are concentrated in the nucleus where they bind GATA sequences upstream of nitrogen catabolite repression ( NCR)-sensitive genes and activate their transcription. Conversely, in exces s nitrogen, these GATA sequences are unoccupied by Gln3p and Gat1p because these transcription activators are excluded from the nucleus. Ure2p binds t o Gln3p and Gat1p and is required for NCR-sensitive transcription to be rep ressed and for nuclear exclusion of these transcription factors. Here we sh ow the following. (i) Gln3p residues 344-365 are required for nuclear local ization. (ii) Replacing Ser-344, Ser-347, and Ser-355 with alanines has min imal effects on GFP-Gln3p localization. However, replacing Gln3p Ser-344, S er-347, and Ser-355 with aspartates results in significant loss of its abil ity to be concentrated in the nucleus. (iii) N and C termini of the Gln3p r egion required for it to complex with Ure2p and be excluded from the nucleu s are between residues 1-103 and 301-365, respectively. (iv) N and C termin i of the Ure2p region required for it to interact with Gln3p are situated b etween residues 101-151 and 330-346, respectively. (v) Loss of Ure2p residu es participating in either dimer or prion formation diminishes its ability to carry out NCR-sensitive regulation of Gln3p activity.