Thiolutin has complex effects in vivo but is a direct inhibitor of RNA polymerase II in vitro.

Autor:	Qiu C; Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843, USA., Arora P; Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843, USA.; Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA., Malik I; Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843, USA., Laperuta AJ; Stevenson University, Stevenson, MD 21153, USA., Pavlovic EM; Earlham College, Richmond, IN 47374, USA., Ugochukwu S; Texas State University, San Marcos, TX 78666, USA., Naik M; Department of Biochemistry and Biophysics, Texas A&M University, College Station, TX 77843, USA., Kaplan CD; Department of Biological Sciences, University of Pittsburgh, Pittsburgh, PA 15260, USA.
Jazyk:	angličtina
Zdroj:	Nucleic acids research [Nucleic Acids Res] 2024 Mar 21; Vol. 52 (5), pp. 2546-2564.
DOI:	10.1093/nar/gkad1258
Abstrakt:	Thiolutin is a natural product transcription inhibitor with an unresolved mode of action. Thiolutin and the related dithiolopyrrolone holomycin chelate Zn2+ and previous studies have concluded that RNA Polymerase II (Pol II) inhibition in vivo is indirect. Here, we present chemicogenetic and biochemical approaches to investigate thiolutin's mode of action in Saccharomyces cerevisiae. We identify mutants that alter sensitivity to thiolutin. We provide genetic evidence that thiolutin causes oxidation of thioredoxins in vivo and that thiolutin both induces oxidative stress and interacts functionally with multiple metals including Mn2+ and Cu2+, and not just Zn2+. Finally, we show direct inhibition of RNA polymerase II (Pol II) transcription initiation by thiolutin in vitro in support of classical studies that thiolutin can directly inhibit transcription in vitro. Inhibition requires both Mn2+ and appropriate reduction of thiolutin as excess DTT abrogates its effects. Pause prone, defective elongation can be observed in vitro if inhibition is bypassed. Thiolutin effects on Pol II occupancy in vivo are widespread but major effects are consistent with prior observations for Tor pathway inhibition and stress induction, suggesting that thiolutin use in vivo should be restricted to studies on its modes of action and not as an experimental tool. (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)
Databáze:	MEDLINE
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