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A dual molecular analogue tuner for dissecting protein function in mammalian cells
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A dual molecular analogue tuner for dissecting protein function in mammalian cells
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A dual molecular analogue tuner for dissecting protein function in mammalian cells
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Journal Article
A dual molecular analogue tuner for dissecting protein function in mammalian cells
2016
Overview
Loss-of-function studies are fundamental for dissecting gene function. Yet, methods to rapidly and effectively perturb genes in mammalian cells, and particularly in stem cells, are scarce. Here we present a system for simultaneous conditional regulation of two different proteins in the same mammalian cell. This system harnesses the plant auxin and jasmonate hormone-induced degradation pathways, and is deliverable with only two lentiviral vectors. It combines RNAi-mediated silencing of two endogenous proteins with the expression of two exogenous proteins whose degradation is induced by external ligands in a rapid, reversible, titratable and independent manner. By engineering molecular tuners for NANOG, CHK1, p53 and NOTCH1 in mammalian stem cells, we have validated the applicability of the system and demonstrated its potential to unravel complex biological processes.
Loss-of-function approaches are fundamental for dissecting the roles played by genes but methods to simultaneously perturb several proteins in the same mammalian cell are scarce. Here the authors harness the plant auxin and jasmonate hormone-degradation pathways and RNAi technology, to control the levels of two proteins and validate its application in stem cells.
Publisher
Nature Publishing Group UK,Nature Publishing Group,Nature Portfolio
Subject
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/ 13/89
/ 14/63
/ 38/1
/ 38/35
/ 38/39
/ 38/90
/ 64/60
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/ Animals
/ Biology
/ Checkpoint Kinase 1 - genetics
/ Checkpoint Kinase 1 - metabolism
/ Embryonic Stem Cells - drug effects
/ Embryonic Stem Cells - metabolism
/ Enzymes
/ Gene Expression Profiling - methods
/ Gene Expression Regulation - drug effects
/ Gene Expression Regulation - genetics
/ Genes
/ Humanities and Social Sciences
/ Humans
/ Mammals
/ Medicine
/ Nanog Homeobox Protein - genetics
/ Nanog Homeobox Protein - metabolism
/ Plant Growth Regulators - pharmacology
/ Proteins
/ Receptor, Notch1 - metabolism
/ Science
