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A mating-type mutagenesis screen identifies a zinc-finger protein required for specific DNA excision events in Paramecium

In the ciliate Paramecium tetraurelia, functional genes are reconstituted during development of the somatic macronucleus through the precise excision of ∼45 000 single-copy Internal Eliminated Sequences (IESs), thought to be the degenerate remnants of ancient transposon insertions. Like introns, IES...

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Autores principales: Bhullar, Simran, Denby Wilkes, Cyril, Arnaiz, Olivier, Nowacki, Mariusz, Sperling, Linda, Meyer, Eric
Formato: Online Artículo Texto
Lenguaje:English
Publicado: Oxford University Press 2018
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6182129/
https://www.ncbi.nlm.nih.gov/pubmed/30165457
http://dx.doi.org/10.1093/nar/gky772
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author Bhullar, Simran
Denby Wilkes, Cyril
Arnaiz, Olivier
Nowacki, Mariusz
Sperling, Linda
Meyer, Eric
author_facet Bhullar, Simran
Denby Wilkes, Cyril
Arnaiz, Olivier
Nowacki, Mariusz
Sperling, Linda
Meyer, Eric
author_sort Bhullar, Simran
collection PubMed
description In the ciliate Paramecium tetraurelia, functional genes are reconstituted during development of the somatic macronucleus through the precise excision of ∼45 000 single-copy Internal Eliminated Sequences (IESs), thought to be the degenerate remnants of ancient transposon insertions. Like introns, IESs are marked only by a weak consensus at their ends. How such a diverse set of sequences is faithfully recognized and precisely excised remains unclear: specialized small RNAs have been implicated, but in their absence up to ∼60% of IESs are still correctly excised. To get further insight, we designed a mutagenesis screen based on the hypersensitivity of a specific excision event in the mtA gene, which determines mating types. Unlike most IES-containing genes, the active form of mtA is the unexcised one, allowing the recovery of hypomorphic alleles of essential IES recognition/excision factors. Such is the case of one mutation recovered in the Piwi gene PTIWI09, a key player in small RNA-mediated IES recognition. Another mutation identified a novel protein with a C2H2 zinc finger, mtGa, which is required for excision of a small subset of IESs characterized by enrichment in a 5-bp motif. The unexpected implication of a sequence-specific factor establishes a new paradigm for IES recognition and/or excision.
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spelling pubmed-61821292018-10-18 A mating-type mutagenesis screen identifies a zinc-finger protein required for specific DNA excision events in Paramecium Bhullar, Simran Denby Wilkes, Cyril Arnaiz, Olivier Nowacki, Mariusz Sperling, Linda Meyer, Eric Nucleic Acids Res Genome Integrity, Repair and Replication In the ciliate Paramecium tetraurelia, functional genes are reconstituted during development of the somatic macronucleus through the precise excision of ∼45 000 single-copy Internal Eliminated Sequences (IESs), thought to be the degenerate remnants of ancient transposon insertions. Like introns, IESs are marked only by a weak consensus at their ends. How such a diverse set of sequences is faithfully recognized and precisely excised remains unclear: specialized small RNAs have been implicated, but in their absence up to ∼60% of IESs are still correctly excised. To get further insight, we designed a mutagenesis screen based on the hypersensitivity of a specific excision event in the mtA gene, which determines mating types. Unlike most IES-containing genes, the active form of mtA is the unexcised one, allowing the recovery of hypomorphic alleles of essential IES recognition/excision factors. Such is the case of one mutation recovered in the Piwi gene PTIWI09, a key player in small RNA-mediated IES recognition. Another mutation identified a novel protein with a C2H2 zinc finger, mtGa, which is required for excision of a small subset of IESs characterized by enrichment in a 5-bp motif. The unexpected implication of a sequence-specific factor establishes a new paradigm for IES recognition and/or excision. Oxford University Press 2018-10-12 2018-08-27 /pmc/articles/PMC6182129/ /pubmed/30165457 http://dx.doi.org/10.1093/nar/gky772 Text en © The Author(s) 2018. Published by Oxford University Press on behalf of Nucleic Acids Research. http://creativecommons.org/licenses/by-nc/4.0/ This is an Open Access article distributed under the terms of the Creative Commons Attribution Non-Commercial License (http://creativecommons.org/licenses/by-nc/4.0/), which permits non-commercial re-use, distribution, and reproduction in any medium, provided the original work is properly cited. For commercial re-use, please contact journals.permissions@oup.com
spellingShingle Genome Integrity, Repair and Replication
Bhullar, Simran
Denby Wilkes, Cyril
Arnaiz, Olivier
Nowacki, Mariusz
Sperling, Linda
Meyer, Eric
A mating-type mutagenesis screen identifies a zinc-finger protein required for specific DNA excision events in Paramecium
title A mating-type mutagenesis screen identifies a zinc-finger protein required for specific DNA excision events in Paramecium
title_full A mating-type mutagenesis screen identifies a zinc-finger protein required for specific DNA excision events in Paramecium
title_fullStr A mating-type mutagenesis screen identifies a zinc-finger protein required for specific DNA excision events in Paramecium
title_full_unstemmed A mating-type mutagenesis screen identifies a zinc-finger protein required for specific DNA excision events in Paramecium
title_short A mating-type mutagenesis screen identifies a zinc-finger protein required for specific DNA excision events in Paramecium
title_sort mating-type mutagenesis screen identifies a zinc-finger protein required for specific dna excision events in paramecium
topic Genome Integrity, Repair and Replication
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6182129/
https://www.ncbi.nlm.nih.gov/pubmed/30165457
http://dx.doi.org/10.1093/nar/gky772
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