Identification of an E3 ligase that targets the catalytic subunit of RNA polymerase I upon transcription stress

Stephanie Pitts; Hester Liu; Adel Ibrahim; Amit Garg; Catarina Mendes Felgueira; Asma Begum; Wenjun Fan; Selina Teh; Jin-Yih Low; Brittany Ford; David A. Schneider; Ronald Hay; Marikki Laiho

doi:10.1016/j.jbc.2022.102690

Identification of an E3 ligase that targets the catalytic subunit of RNA polymerase I upon transcription stress

Stephanie Pitts, Hester Liu, Adel Ibrahim, Amit Garg, Catarina Mendes Felgueira, Asma Begum, Wenjun Fan, Selina Teh, Jin-Yih Low, Brittany Ford, David A. Schneider, Ronald Hay, Marikki Laiho (Lead / Corresponding author)

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Abstract

RNA Polymerase I (Pol I) synthesizes rRNA, which is the first and rate-limiting step in ribosome biogenesis. Factors governing the stability of the polymerase complex are not known. Previous studies characterizing Pol I inhibitor BMH-21 revealed a transcriptional stress-dependent pathway for degradation of the largest subunit of Pol I, RPA194. To identify the E3 ligase(s) involved, we conducted a cell-based RNAi screen for ubiquitin pathway genes. We establish Skp–Cullin–F-box protein complex F-box protein FBXL14 as an E3 ligase for RPA194. We show that FBXL14 binds to RPA194 and mediates RPA194 ubiquitination and degradation in cancer cells treated with BMH-21. Mutation analysis in yeast identified lysines 1150, 1153, and 1156 on Rpa190 relevant for the protein degradation. These results reveal the regulated turnover of Pol I, showing that the stability of the catalytic subunit is controlled by the F-box protein FBXL14 in response to transcription stress.

Original language	English
Article number	102690
Number of pages	18
Journal	Journal of Biological Chemistry
Volume	298
Issue number	12
Early online date	10 Nov 2022
DOIs	https://doi.org/10.1016/j.jbc.2022.102690
Publication status	Published - Dec 2022

Keywords

transcription
ubiquitin
proteasome
small molecule
cancer

ASJC Scopus subject areas

Molecular Biology
Biochemistry
Cell Biology

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1016/j.jbc.2022.102690Licence: CC BY

Final published versionFinal published version, 3.53 MBLicence: CC BY

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Pitts, S., Liu, H., Ibrahim, A., Garg, A., Felgueira, C. M., Begum, A., Fan, W., Teh, S., Low, J.-Y., Ford, B., Schneider, D. A., Hay, R., & Laiho, M. (2022). Identification of an E3 ligase that targets the catalytic subunit of RNA polymerase I upon transcription stress. Journal of Biological Chemistry, 298(12), Article 102690. https://doi.org/10.1016/j.jbc.2022.102690

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title = "Identification of an E3 ligase that targets the catalytic subunit of RNA polymerase I upon transcription stress",

abstract = "RNA Polymerase I (Pol I) synthesizes rRNA, which is the first and rate-limiting step in ribosome biogenesis. Factors governing the stability of the polymerase complex are not known. Previous studies characterizing Pol I inhibitor BMH-21 revealed a transcriptional stress-dependent pathway for degradation of the largest subunit of Pol I, RPA194. To identify the E3 ligase(s) involved, we conducted a cell-based RNAi screen for ubiquitin pathway genes. We establish Skp–Cullin–F-box protein complex F-box protein FBXL14 as an E3 ligase for RPA194. We show that FBXL14 binds to RPA194 and mediates RPA194 ubiquitination and degradation in cancer cells treated with BMH-21. Mutation analysis in yeast identified lysines 1150, 1153, and 1156 on Rpa190 relevant for the protein degradation. These results reveal the regulated turnover of Pol I, showing that the stability of the catalytic subunit is controlled by the F-box protein FBXL14 in response to transcription stress.",

keywords = "transcription, ubiquitin, proteasome, small molecule, cancer",

author = "Stephanie Pitts and Hester Liu and Adel Ibrahim and Amit Garg and Felgueira, {Catarina Mendes} and Asma Begum and Wenjun Fan and Selina Teh and Jin-Yih Low and Brittany Ford and Schneider, {David A.} and Ronald Hay and Marikki Laiho",

note = "Funding Information: This work was supported by grants from the National Cancer Institute, NIH (F31CA247077 to S.P.) and National Institute of General Medical Sciences, NIH (R01GM121404 to M.L. and D.A.S. and NIH P30CA006973 to M.L.). Copyright: {\textcopyright} 2022 The Authors.",

year = "2022",

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doi = "10.1016/j.jbc.2022.102690",

language = "English",

volume = "298",

journal = "Journal of Biological Chemistry",

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T1 - Identification of an E3 ligase that targets the catalytic subunit of RNA polymerase I upon transcription stress

AU - Pitts, Stephanie

AU - Liu, Hester

AU - Ibrahim, Adel

AU - Garg, Amit

AU - Felgueira, Catarina Mendes

AU - Begum, Asma

AU - Fan, Wenjun

AU - Teh, Selina

AU - Low, Jin-Yih

AU - Ford, Brittany

AU - Schneider, David A.

AU - Hay, Ronald

AU - Laiho, Marikki

N1 - Funding Information: This work was supported by grants from the National Cancer Institute, NIH (F31CA247077 to S.P.) and National Institute of General Medical Sciences, NIH (R01GM121404 to M.L. and D.A.S. and NIH P30CA006973 to M.L.). Copyright: © 2022 The Authors.

PY - 2022/12

Y1 - 2022/12

N2 - RNA Polymerase I (Pol I) synthesizes rRNA, which is the first and rate-limiting step in ribosome biogenesis. Factors governing the stability of the polymerase complex are not known. Previous studies characterizing Pol I inhibitor BMH-21 revealed a transcriptional stress-dependent pathway for degradation of the largest subunit of Pol I, RPA194. To identify the E3 ligase(s) involved, we conducted a cell-based RNAi screen for ubiquitin pathway genes. We establish Skp–Cullin–F-box protein complex F-box protein FBXL14 as an E3 ligase for RPA194. We show that FBXL14 binds to RPA194 and mediates RPA194 ubiquitination and degradation in cancer cells treated with BMH-21. Mutation analysis in yeast identified lysines 1150, 1153, and 1156 on Rpa190 relevant for the protein degradation. These results reveal the regulated turnover of Pol I, showing that the stability of the catalytic subunit is controlled by the F-box protein FBXL14 in response to transcription stress.

AB - RNA Polymerase I (Pol I) synthesizes rRNA, which is the first and rate-limiting step in ribosome biogenesis. Factors governing the stability of the polymerase complex are not known. Previous studies characterizing Pol I inhibitor BMH-21 revealed a transcriptional stress-dependent pathway for degradation of the largest subunit of Pol I, RPA194. To identify the E3 ligase(s) involved, we conducted a cell-based RNAi screen for ubiquitin pathway genes. We establish Skp–Cullin–F-box protein complex F-box protein FBXL14 as an E3 ligase for RPA194. We show that FBXL14 binds to RPA194 and mediates RPA194 ubiquitination and degradation in cancer cells treated with BMH-21. Mutation analysis in yeast identified lysines 1150, 1153, and 1156 on Rpa190 relevant for the protein degradation. These results reveal the regulated turnover of Pol I, showing that the stability of the catalytic subunit is controlled by the F-box protein FBXL14 in response to transcription stress.

KW - transcription

KW - ubiquitin

KW - proteasome

KW - small molecule

KW - cancer

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DO - 10.1016/j.jbc.2022.102690

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JO - Journal of Biological Chemistry

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ER -

Identification of an E3 ligase that targets the catalytic subunit of RNA polymerase I upon transcription stress

Abstract

Keywords

ASJC Scopus subject areas

UN SDGs

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