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The prodomain of Ssy5 protease controls receptor-activated proteolysis of transcription factor Stp1
Stockholm University, Faculty of Science, The Wenner-Gren Institute, Cell Biology.
Stockholm University, Faculty of Science, The Wenner-Gren Institute, Cell Biology.
Stockholm University, Faculty of Science, The Wenner-Gren Institute, Cell Biology.
Stockholm University, Faculty of Science, The Wenner-Gren Institute, Cell Biology.
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2010 (English)In: Molecular and Cellular Biology, ISSN 0270-7306, E-ISSN 1098-5549, Vol. 30, no 13, 3299-309 p.Article in journal (Refereed) Published
Abstract [en]

Extracellular amino acids induce the yeast SPS sensor to endoproteolytically cleave transcription factors Stp1 and Stp2 in a process termed receptor-activated proteolysis (RAP). Ssy5, the activating endoprotease, is synthesized with a large N-terminal prodomain and a C-terminal chymotrypsin-like catalytic (Cat) domain. During biogenesis, Ssy5 cleaves itself and the prodomain and Cat domain remain associated, forming an inactive primed protease. Here we show that the prodomain is a potent inhibitor of Cat domain activity and that its inactivation is a requisite for RAP. Accordingly, amino acid-induced signals trigger proteasome-dependent degradation of the prodomain. A mutation that stabilizes the prodomain prevents Stp1 processing, whereas destabilizing mutations lead to constitutive RAP-independent Stp1 processing. We fused a conditional degron to the prodomain to synthetically reprogram the amino acid-responsive SPS signaling pathway, placing it under temperature control. Our results define a regulatory mechanism that is novel for eukaryotic proteases functioning within cells

Place, publisher, year, edition, pages
2010. Vol. 30, no 13, 3299-309 p.
National Category
Cell Biology
Research subject
Cell Biology
Identifiers
URN: urn:nbn:se:su:diva-44307DOI: 10.1128/MCB.00323-10ISI: 000278626100013PubMedID: 20421414OAI: oai:DiVA.org:su-44307DiVA: diva2:360941
Available from: 2010-11-05 Created: 2010-11-05 Last updated: 2013-12-09Bibliographically approved
In thesis
1. Regulatory mechanisms of amino acid-induced signaling in Saccharomyces cerevisiae
Open this publication in new window or tab >>Regulatory mechanisms of amino acid-induced signaling in Saccharomyces cerevisiae
2014 (English)Doctoral thesis, comprehensive summary (Other academic)
Abstract [en]

This thesis describes studies aimed at elucidating the molecular mechanisms that regulate the SPS (Ssy1-Ptr3-Ssy5) signal transduction pathway in the yeast Saccharomyces cerevisiae. This pathway is induced by extracellular amino acids and facilitates their uptake. The most downstream effectors of the SPS pathway, the homologous transcription factors Stp1 and Stp2 (Stp1/2), are synthesized as latent precursors with N-terminal regulatory domains that restrict their nuclear accumulation. Amino acid-induced signaling, initiated by the plasma membrane localized receptor Ssy1, leads via Ptr3 to the activation of the endoprotease Ssy5. Active Ssy5 cleaves the regulatory domains in Stp1/2. As a consequence, the processed transcription factors lacking their N-terminal domains accumulate in the nucleus and activate the transcription of amino acid permease genes to enhance the uptake capacity of cells.

Ssy5 is synthesized as a zymogen precursor that processes itself into a prodomain and catalytic (Cat) domain that remain non-covalently associated. We found that the prodomain functions as an inhibitor of the Cat domain. Signaling triggers the degradation of the prodomain by the proteasome, thereby releasing Cat domain activity (paper I). We identified a motif in the prodomain that functions as inducible phosphodegron. Upon signaling, this motif is phosphorylated which triggers prodomain polyubiquitylation, and as a consequence, its proteasomal degradation (paper II). Also, we found that Ptr3 functions to mediate prodomain phosphorylation upon signaling and that protein phosphatase 2A constitutively mutes phosphorylation-dependent activation of Ssy5 (paper III).

Finally, in addition to the regulation of the processing protease Ssy5, the control of transcriptional activity of Stp1 depends on a motif within its N-terminal regulatory domain, designated Region I. We found that Region I mediates latency by functioning as cytoplasmic retention determinant and nuclear degron (paper IV).

Place, publisher, year, edition, pages
Stockholm: Department of Molecular Biosciences, The Wenner-Gren Institute, Stockholm University, 2014. 70 p.
Keyword
Signal transduction, regulatory mechanisms, nutrient sensing, Saccharomyces cerevisiae
National Category
Cell Biology
Research subject
Cell Biology
Identifiers
urn:nbn:se:su:diva-97329 (URN)978-91-7447-830-3 (ISBN)
Public defence
2014-02-04, sal E306, Arrheniuslaboratorierna, Svante Arrhenius väg 20 C, Stockholm, 10:00 (English)
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Note

At the time of the doctoral defense, the following paper was unpublished and had a status as follows: Paper 4: Manuscript.

Available from: 2014-01-13 Created: 2013-12-08 Last updated: 2014-01-02Bibliographically approved

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