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Elongator's toxin-target (TOT) function is nuclear localization sequence dependent and suppressed by
Lars Fichtner1, Daniel Jablonowski, Angelika Schierhorn
1Biologicum, Institut für Genetik, Martin-Luther-Universität Halle-Wittenberg, Weinbergweg 10, D-06120 Halle (Saale), Germany.
Abstract:
The toxin target (TOT) function of the Saccharomyces cerevisiae Elongator complex enables Kluyveromyces lactis zymocin to induce a G1 cell cycle arrest. Loss of a ubiquitin-related system (URM1-UBA4 ) and KTI11 enhances post-translational modification/proteolysis of Elongator subunit Tot1p (Elp1p) and abrogates its TOT function. Using TAP tagging, Kti11p contacts Elongator and translational proteins (Rps7Ap, Rps19Ap Eft2p, Yil103wp, Dph2p). Loss of YIL103w and DPH2 (involved in diphtheria toxicity) suppresses zymocicity implying that both toxins overlap in a manner mediated by Kti11p. Among the pool that co-fractionates with RNA polymerase II (pol II) and nucleolin, Nop1p, unmodified Tot1p dominates. Thus, modification/proteolysis may affect association of Elongator with pol II or its localization. Consistently, an Elongator-nuclear localization sequence (NLS) targets green fluorescent protein (GFP) to the nucleus, and its truncation yields TOT deficiency. Similarly, KAP120 deletion rescues cells from zymocin, suggesting that Elongator's TOT function requires NLS- and karyopherin-dependent nuclear import.
Insights
The Elongator complex
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- The Elongator complex in Saccharomyces cerevisiae is crucial for the toxin target (TOT) function.
- This function is essential for Kluyveromyces lactis zymocin to induce cell cycle arrest.
- Post-translational modification and proteolysis of Elongator subunit Tot1p (Elp1p) impact its TOT function.
Purpose of the Study:
- To investigate the molecular mechanisms underlying the Elongator complex's toxin target (TOT) function.
- To identify proteins interacting with the Elongator complex and their role in zymocin-induced cell cycle arrest.
- To elucidate the role of nuclear import in Elongator's TOT function.
Main Methods:
- TAP tagging to identify protein interactions.
- Analysis of gene deletions affecting Elongator modification and function.
- Localization studies using green fluorescent protein (GFP) fusions and nuclear localization sequences (NLS).
Main Results:
- Loss of ubiquitin-related system (URM1-UBA4) and KTI11 leads to increased Tot1p modification/proteolysis and abrogates TOT function.
- Kti11p interacts with Elongator and translational proteins, suggesting a role in toxin response.
- Loss of YIL103w and DPH2 suppresses zymocin toxicity, indicating overlapping pathways mediated by Kti11p.
- Unmodified Tot1p is found in a complex with RNA polymerase II (pol II) and nucleolin.
- Elongator's nuclear import via NLS and karyopherin-dependent import is essential for its TOT function.
Conclusions:
- Post-translational modification and proteolysis of Elongator may regulate its association with pol II and cellular localization.
- The Elongator complex's toxin target (TOT) function is dependent on its nuclear import pathway.
- Kti11p acts as a scaffold, linking Elongator to translational machinery and potentially mediating overlapping toxin response pathways.