A single PXY motif located within the carboxyl terminus of Spt23p and Mga2p mediates a physical and functional interaction with ubiquitin ligase Rsp5p

Natalia Shcherbik, Younghoon Kee, Nancy Lyon, Jon M. Huibregtse, Dale S. Haines

Research output: Contribution to journalArticlepeer-review

40 Scopus citations

Abstract

Proteasome-dependent processing of the endoplasmic reticulum localized transcription factor Spt23p of Saccharomyces cerevisiae generates its transcriptionally competent form and requires the WW domain containing Rsp5p ubiquitin ligase. Although previous studies documented an Rsp5p-Spt23p association in cells, very little is known about the nature of this interaction. We report here the identification of an imperfect type I WW domain-binding site (LPKY) within the carbosyl-terminal region of Spt23p that is required for Rsp5p binding In vitro and in vivo. Deletion of this motif abrogates Rsp5p-induced ubiquitination of Spt23p in vitro and reduces ubiquitination of the Spt23p precursor in yeast. In addition, the Spt23pΔLPKY mutant is inefficiently processed and is defective at up-regulating target gene (OLE1) expression in cells. Deletion of the corresponding LPKY site within Mga2p, an Spt23p homologue, also abrogates Rsp5p binding and Rsp5p-dependent ubiquitination in vitro as well as Rsp5p binding and Mga2p polyubiquitination in cells. However, the Mga2pΔLPKY mutant undergoes efficient proteasome-dependent processing. These experiments indicate that the LPKY motif of Spt23p is required for Rsp5p binding, Rsp5-induced ubiquitination, proteasome-dependent processing, and its OLE1 inducing function. They also suggest that the LPKY motif of Mga2p is required for Rsp5p binding and ubiquitination, and Rsp5p regulates Mga2p function by a mechanism that is independent of providing the partial degradation signal.

Original languageEnglish (US)
Pages (from-to)53892-53898
Number of pages7
JournalJournal of Biological Chemistry
Volume279
Issue number51
DOIs
StatePublished - Dec 17 2004

All Science Journal Classification (ASJC) codes

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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