4-Hydroxytamoxifen probes for light-dependent spatiotemporal control of Cre-ER mediated reporter gene expression
| Autor: | Yumay Chen, Seok-Ki Choi, Pamela T. Wong, James R. Baker, Alexa Coulter, Shengzhuang Tang, Tannaz Faal, Christina H. Tu, Matthew A. Inlay |
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| Rok vydání: | 2014 |
| Předmět: |
Biochemistry & Molecular Biology
Light 1.1 Normal biological development and functioning Estrogen receptor lac operon Gene Expression Biology In Vitro Techniques Genetic recombination Green fluorescent protein Cell membrane In vivo Underpinning research Genes Reporter Receptors medicine Genetics Homologous Recombination Molecular Biology Reporter Reporter gene Molecular biology Estrogen Tamoxifen medicine.anatomical_structure Genes Gene Expression Regulation Receptors Estrogen Biochemistry and Cell Biology medicine.drug Biotechnology |
| Zdroj: | Molecular bioSystems, vol 11, iss 3 Faal, T; Wong, PT; Tang, S; Coulter, A; Chen, Y; Tu, CH; et al.(2015). 4-Hydroxytamoxifen probes for light-dependent spatiotemporal control of Cre-ER mediated reporter gene expression. Molecular BioSystems, 11(3), 783-790. doi: 10.1039/c4mb00581c. UC Irvine: Retrieved from: http://www.escholarship.org/uc/item/8781924k |
| ISSN: | 1742-2051 |
| DOI: | 10.1039/c4mb00581c. |
| Popis: | © 2015 The Royal Society of Chemistry. The tamoxifen inducible Cre-ER/loxP system provides tissue specific temporal control of gene recombination events, and can be used to induce expression of reporter genes (e.g. GFP, LacZ) for lineage tracing studies. Cre enzyme fused with estrogen receptor (Cre-ER) is released upon tamoxifen binding, resulting in permanent activation of reporter genes within cells and their progeny. Tamoxifen and its active metabolite, hydroxytamoxifen (4OHT) diffuses rapidly in vivo, making it difficult to restrict labeling to specific locations. In this study, we developed a photocaged 4OHT molecule by covalently attaching 4OHT to an ortho-nitrobenzyl (ONB1) group, rendering 4OHT inactive. Exposure to UV radiation cleaves the bond between ONB1and 4OHT, freeing the 4OHT to bind Cre-ER to result in downstream genetic recombination and reporter activation. We show that caged ONB1-4OHT crosses the cell membrane and uncages after short UV exposure, resulting in Cre-driven genetic recombination that can be localized to specific regions or tissues. ONB1-4OHT can provide spatial control of reporter activation and be adapted with any existing Cre-ER/loxP based system. |
| Databáze: | OpenAIRE |
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