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CRISPR/Cas9 and active genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity
The knirps (kni) locus encodes transcription factors required for induction of the L2 wing vein in Drosophila. Here, we employ diverse CRISPR/Cas9 genome editing tools to generate a series of targeted lesions within the endogenous cis-regulatory module (CRM) required for kni expression in the L2 vei...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
eLife Sciences Publications, Ltd
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5800851/ https://www.ncbi.nlm.nih.gov/pubmed/29274230 http://dx.doi.org/10.7554/eLife.30281 |
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author | Xu, Xiang-Ru Shannon Gantz, Valentino Matteo Siomava, Natalia Bier, Ethan |
author_facet | Xu, Xiang-Ru Shannon Gantz, Valentino Matteo Siomava, Natalia Bier, Ethan |
author_sort | Xu, Xiang-Ru Shannon |
collection | PubMed |
description | The knirps (kni) locus encodes transcription factors required for induction of the L2 wing vein in Drosophila. Here, we employ diverse CRISPR/Cas9 genome editing tools to generate a series of targeted lesions within the endogenous cis-regulatory module (CRM) required for kni expression in the L2 vein primordium. Phenotypic analysis of these ‘in locus’ mutations based on both expression of Kni protein and adult wing phenotypes, reveals novel unexpected features of L2-CRM function including evidence for a chromosome pairing-dependent process that promotes transcription. We also demonstrate that self-propagating active genetic elements (CopyCat elements) can efficiently delete and replace the L2-CRM with orthologous sequences from other divergent fly species. Wing vein phenotypes resulting from these trans-species enhancer replacements parallel features of the respective donor fly species. This highly sensitive phenotypic readout of enhancer function in a native genomic context reveals novel features of CRM function undetected by traditional reporter gene analysis. |
format | Online Article Text |
id | pubmed-5800851 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | eLife Sciences Publications, Ltd |
record_format | MEDLINE/PubMed |
spelling | pubmed-58008512018-02-08 CRISPR/Cas9 and active genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity Xu, Xiang-Ru Shannon Gantz, Valentino Matteo Siomava, Natalia Bier, Ethan eLife Developmental Biology The knirps (kni) locus encodes transcription factors required for induction of the L2 wing vein in Drosophila. Here, we employ diverse CRISPR/Cas9 genome editing tools to generate a series of targeted lesions within the endogenous cis-regulatory module (CRM) required for kni expression in the L2 vein primordium. Phenotypic analysis of these ‘in locus’ mutations based on both expression of Kni protein and adult wing phenotypes, reveals novel unexpected features of L2-CRM function including evidence for a chromosome pairing-dependent process that promotes transcription. We also demonstrate that self-propagating active genetic elements (CopyCat elements) can efficiently delete and replace the L2-CRM with orthologous sequences from other divergent fly species. Wing vein phenotypes resulting from these trans-species enhancer replacements parallel features of the respective donor fly species. This highly sensitive phenotypic readout of enhancer function in a native genomic context reveals novel features of CRM function undetected by traditional reporter gene analysis. eLife Sciences Publications, Ltd 2017-12-23 /pmc/articles/PMC5800851/ /pubmed/29274230 http://dx.doi.org/10.7554/eLife.30281 Text en © 2017, Xu et al http://creativecommons.org/licenses/by/4.0/ http://creativecommons.org/licenses/by/4.0/This article is distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/) , which permits unrestricted use and redistribution provided that the original author and source are credited. |
spellingShingle | Developmental Biology Xu, Xiang-Ru Shannon Gantz, Valentino Matteo Siomava, Natalia Bier, Ethan CRISPR/Cas9 and active genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity |
title | CRISPR/Cas9 and active genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity |
title_full | CRISPR/Cas9 and active genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity |
title_fullStr | CRISPR/Cas9 and active genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity |
title_full_unstemmed | CRISPR/Cas9 and active genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity |
title_short | CRISPR/Cas9 and active genetics-based trans-species replacement of the endogenous Drosophila kni-L2 CRM reveals unexpected complexity |
title_sort | crispr/cas9 and active genetics-based trans-species replacement of the endogenous drosophila kni-l2 crm reveals unexpected complexity |
topic | Developmental Biology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5800851/ https://www.ncbi.nlm.nih.gov/pubmed/29274230 http://dx.doi.org/10.7554/eLife.30281 |
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