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A remodeled RNA polymerase II complex catalyzing viroid RNA-templated transcription

Viroids, a fascinating group of plant pathogens, are subviral agents composed of single-stranded circular noncoding RNAs. It is well-known that nuclear-replicating viroids exploit host DNA-dependent RNA polymerase II (Pol II) activity for transcription from circular RNA genome to minus-strand interm...

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Autores principales: Dissanayaka Mudiyanselage, Shachinthaka D., Ma, Junfei, Pechan, Tibor, Pechanova, Olga, Liu, Bin, Wang, Ying
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Public Library of Science 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9521916/
https://www.ncbi.nlm.nih.gov/pubmed/36121876
http://dx.doi.org/10.1371/journal.ppat.1010850
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author Dissanayaka Mudiyanselage, Shachinthaka D.
Ma, Junfei
Pechan, Tibor
Pechanova, Olga
Liu, Bin
Wang, Ying
author_facet Dissanayaka Mudiyanselage, Shachinthaka D.
Ma, Junfei
Pechan, Tibor
Pechanova, Olga
Liu, Bin
Wang, Ying
author_sort Dissanayaka Mudiyanselage, Shachinthaka D.
collection PubMed
description Viroids, a fascinating group of plant pathogens, are subviral agents composed of single-stranded circular noncoding RNAs. It is well-known that nuclear-replicating viroids exploit host DNA-dependent RNA polymerase II (Pol II) activity for transcription from circular RNA genome to minus-strand intermediates, a classic example illustrating the intrinsic RNA-dependent RNA polymerase activity of Pol II. The mechanism for Pol II to accept single-stranded RNAs as templates remains poorly understood. Here, we reconstituted a robust in vitro transcription system and demonstrated that Pol II also accepts minus-strand viroid RNA template to generate plus-strand RNAs. Further, we purified the Pol II complex on RNA templates for nano-liquid chromatography-tandem mass spectrometry analysis and identified a remodeled Pol II missing Rpb4, Rpb5, Rpb6, Rpb7, and Rpb9, contrasting to the canonical 12-subunit Pol II or the 10-subunit Pol II core on DNA templates. Interestingly, the absence of Rpb9, which is responsible for Pol II fidelity, explains the higher mutation rate of viroids in comparison to cellular transcripts. This remodeled Pol II is active for transcription with the aid of TFIIIA-7ZF and appears not to require other canonical general transcription factors (such as TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH, and TFIIS), suggesting a distinct mechanism/machinery for viroid RNA-templated transcription. Transcription elongation factors, such as FACT complex, PAF1 complex, and SPT6, were also absent in the reconstituted transcription complex. Further analyses of the critical zinc finger domains in TFIIIA-7ZF revealed the first three zinc finger domains pivotal for RNA template binding. Collectively, our data illustrated a distinct organization of Pol II complex on viroid RNA templates, providing new insights into viroid replication, the evolution of transcription machinery, as well as the mechanism of RNA-templated transcription.
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spelling pubmed-95219162022-09-30 A remodeled RNA polymerase II complex catalyzing viroid RNA-templated transcription Dissanayaka Mudiyanselage, Shachinthaka D. Ma, Junfei Pechan, Tibor Pechanova, Olga Liu, Bin Wang, Ying PLoS Pathog Research Article Viroids, a fascinating group of plant pathogens, are subviral agents composed of single-stranded circular noncoding RNAs. It is well-known that nuclear-replicating viroids exploit host DNA-dependent RNA polymerase II (Pol II) activity for transcription from circular RNA genome to minus-strand intermediates, a classic example illustrating the intrinsic RNA-dependent RNA polymerase activity of Pol II. The mechanism for Pol II to accept single-stranded RNAs as templates remains poorly understood. Here, we reconstituted a robust in vitro transcription system and demonstrated that Pol II also accepts minus-strand viroid RNA template to generate plus-strand RNAs. Further, we purified the Pol II complex on RNA templates for nano-liquid chromatography-tandem mass spectrometry analysis and identified a remodeled Pol II missing Rpb4, Rpb5, Rpb6, Rpb7, and Rpb9, contrasting to the canonical 12-subunit Pol II or the 10-subunit Pol II core on DNA templates. Interestingly, the absence of Rpb9, which is responsible for Pol II fidelity, explains the higher mutation rate of viroids in comparison to cellular transcripts. This remodeled Pol II is active for transcription with the aid of TFIIIA-7ZF and appears not to require other canonical general transcription factors (such as TFIIA, TFIIB, TFIID, TFIIE, TFIIF, TFIIH, and TFIIS), suggesting a distinct mechanism/machinery for viroid RNA-templated transcription. Transcription elongation factors, such as FACT complex, PAF1 complex, and SPT6, were also absent in the reconstituted transcription complex. Further analyses of the critical zinc finger domains in TFIIIA-7ZF revealed the first three zinc finger domains pivotal for RNA template binding. Collectively, our data illustrated a distinct organization of Pol II complex on viroid RNA templates, providing new insights into viroid replication, the evolution of transcription machinery, as well as the mechanism of RNA-templated transcription. Public Library of Science 2022-09-19 /pmc/articles/PMC9521916/ /pubmed/36121876 http://dx.doi.org/10.1371/journal.ppat.1010850 Text en © 2022 Dissanayaka Mudiyanselage et al https://creativecommons.org/licenses/by/4.0/This is an open access article distributed under the terms of the Creative Commons Attribution License (https://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
spellingShingle Research Article
Dissanayaka Mudiyanselage, Shachinthaka D.
Ma, Junfei
Pechan, Tibor
Pechanova, Olga
Liu, Bin
Wang, Ying
A remodeled RNA polymerase II complex catalyzing viroid RNA-templated transcription
title A remodeled RNA polymerase II complex catalyzing viroid RNA-templated transcription
title_full A remodeled RNA polymerase II complex catalyzing viroid RNA-templated transcription
title_fullStr A remodeled RNA polymerase II complex catalyzing viroid RNA-templated transcription
title_full_unstemmed A remodeled RNA polymerase II complex catalyzing viroid RNA-templated transcription
title_short A remodeled RNA polymerase II complex catalyzing viroid RNA-templated transcription
title_sort remodeled rna polymerase ii complex catalyzing viroid rna-templated transcription
topic Research Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9521916/
https://www.ncbi.nlm.nih.gov/pubmed/36121876
http://dx.doi.org/10.1371/journal.ppat.1010850
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