Saccharomyces cerevisiae lives in boom and bust nutritional environments. Sophisticated
regulatory systems have evolved to rapidly cope with these changes while preserving
intracellular homeostasis. Target of Rapamycin Complex 1 (TorC1), is a serine/threonine
kinase complex and a principle nitrogen-responsive regulator. TorC1 is activated by
excess nitrogen and downregulated by limiting nitrogen. Two of TorC1’s many downstream
targets are Gln3 and Gat1 —GATA-family transcription activators—whose localization
and function are Nitrogen Catabolite Repression- (NCR-) sensitive. In nitrogen replete
environments, TorC1 is activated, thereby inhibiting the P Tap42-Sit4 and P Tap42-PP2A
( Pph21 / Pph22 - Tpd3 , Pph21 ,22- Rts1 / Cdc55 ) phosphatase complexes. Gln3
is phosphorylated, sequestered in the cytoplasm and NCR-sensitive transcription repressed.
In nitrogen-limiting conditions, TorC1 is downregulated and P Tap42-Sit4 and P Tap42-PP2A
are active. They dephosphorylate Gln3 , which dissociates from Ure2 , relocates
to the nucleus, and activates transcription. A paradoxical observation, however, led
us to suspect that Gln3 control was more complex than appreciated, i.e. , Sit4
dephosphorylates Gln3 more in excess than in limiting nitrogen conditions. This
paradox motivated us to reinvestigate the roles of these phosphatases in Gln3 regulation.
We discovered that: (i) Sit4 and PP2A actively function both in conditions where
TorC1 is activated as well as down-regulated; (ii) nuclear Gln3 is more highly phosphorylated
than when it is sequestered in the cytoplasm; (iii) in nitrogen-replete conditions,
Gln3 relocates from the nucleus to the cytoplasm, where it is dephosphorylated by
Sit4 and PP2A; and (iv) in nitrogen excess and limiting conditions, Sit4 , PP2A,
and Ure2 are all required to maintain cytoplasmic Gln3 in its dephosphorylated
form.