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Rapamycin‐Dependent Gln3 Transcription Factor Dephosphorylation and Nuclear Localization Depend On Distinct Regions of the Gln3 Protein
Author(s) -
Tate Jennifer J.,
Cooper Terrance G
Publication year - 2020
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.2020.34.s1.06551
Subject(s) - dephosphorylation , phosphorylation , nuclear export signal , phosphatase , serine , transcription factor , microbiology and biotechnology , protein phosphatase 2 , kinase , protein kinase a , intracellular , protein phosphatase 1 , biology , chemistry , cytoplasm , biochemistry , cell nucleus , gene
Yeast cell survival in nitrogen‐depleted environments depends on Gln3 entering the nucleus and activating: (i) Nitrogen Catabolite Repression‐sensitive genes whose products scavenge poor nitrogen sources from its extra‐cellular surroundings and (ii) the ATG14 gene required for autophagic recycling nitrogen from intra‐cellular constituents. The TorC1 global serine/threonine kinase complex is one of multiple regulators that controls intracellular Gln3 localization and function. When TorC1 is specifically inhibited by rapamycin, Gln3 is dephosphorylated in a Sit4 phosphatase‐dependent manner, enters the nucleus and activates transcription. However, rapamycin‐elicited Gln3 dephosphorylation itself is insufficient to achieve this outcome, as the hypo‐phosphorylated Gln3 fails to enter the nucleus in the absence of a second phosphatase, PP2A. These observations led to the hypothesis that rapamycin‐elicited outcomes involve distinct target sequences within the Gln3 protein. Supporting this hypothesis, our experiments demonstrate that abolishing the Gln3‐Tor1 interaction, rapamycin‐response or Gln3 cytoplasmic sequestration targets abrogates nuclear Gln3 localization but without effect on rapamycin‐elicited Gln3 dephosphorylation. Truncated Gln3 proteins further show that summary loss of all three regulatory targets also has no effect on Gln3 dephosphorylation. Finally, we have identified multiple specific serine/threonine residues in the N‐terminal region of Gln3 that are dephosphorylated in rapamycin treated cells and when substituted with phosphomimetic aspartate residues abolish Gln3 localization in an otherwise full‐length wild type protein. Support or Funding Information This work supported by NIH GM35642‐27 and LTAUSA18162.