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Eukaryotic Cell, November 2009, p. 1637-1647, Vol. 8, No. 11
1535-9778/09/$08.00+0     doi:10.1128/EC.00205-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Distinct Subsets of Sit4 Holophosphatases Are Required for Inhibition of Saccharomyces cerevisiae Growth by Rapamycin and Zymocin {triangledown} ,{dagger}

Daniel Jablonowski,1,§ Jens-Eike Täubert,1,{ddagger} Christian Bär,1,§ Michael J. R. Stark,2 and Raffael Schaffrath1*

Institut für Biologie, Bereich Genetik, Martin-Luther-Universität, Halle-Wittenberg, Weinbergweg 10, D-06120 Halle (Saale), Germany,1 Wellcome Trust Centre for Gene Regulation and Expression, College of Life Sciences, MSI/WTB Complex, University of Dundee, Dundee DD1 5EH, Scotland2

Received 14 July 2009/ Accepted 7 September 2009

Protein phosphatase Sit4 is required for growth inhibition of Saccharomyces cerevisiae by the antifungals rapamycin and zymocin. Here, we show that the rapamycin effector Tap42, which interacts with Sit4, is dispensable for zymocin action. Although Tap42 binding-deficient sit4 mutants are resistant to zymocin, these mutations also block interaction between Sit4 and the Sit4-associating proteins Sap185 and Sap190, previously shown to mediate zymocin toxicity. Among the four different SAP genes, we found that SAP190 deletions specifically induce rapamycin resistance but that this phenotype is reversed in the additional absence of SAP155. Similarly, the rapamycin resistance of an rrd1{Delta} mutant lacking the Sit4 interactor Rrd1 specifically requires the Sit4/Sap190 complex. Thus, Sit4/Sap190 and Sit4/Sap155 holophosphatases apparently play opposing roles following rapamycin treatment, although rapamycin inhibition is operational in the absence of all Sap family members or Sit4. We further identified a Sit4-interacting region on Sap185 in sap190{Delta} cells that mediates Sit4/Sap185 complex formation and is essential for dephosphorylation of Elp1, a subunit of the Elongator complex. This suggests that Sit4/Sap185 and Sit4/Sap190 holophosphatases promote Elongator functions, a notion supported by data showing that their inactivation eliminates Elongator-dependent processes, including tRNA suppression by SUP4 and tRNA cleavage by zymocin.

* Corresponding author. Present address: Department of Genetics, University of Leicester, University Road, Leicester LE1 7RH, United Kingdom. Phone: 44-116-2525233. Fax: 44-116-2523378. E-mail: rs240{at}le.uk.ac

{triangledown} Published ahead of print on 1 September 2009.

{dagger} Supplemental material for this article may be found at http://ec.asm.org/.

§ Present address: Department of Genetics, University of Leicester, University Road, Leicester LE1 7RH, United Kingdom.

{ddagger} Present address: Department für Tierwissenschaften, Technische Universität München-Weihenstephan, Mühlenweg 22, D-85350 Freising, Germany.

Eukaryotic Cell, November 2009, p. 1637-1647, Vol. 8, No. 11
1535-9778/09/$08.00+0     doi:10.1128/EC.00205-09
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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