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A dual molecular analogue tuner for dissecting protein function in mammalian cells.

ABSTRACT: 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.

SUBMITTER: Brosh R

PROVIDER: S-EPMC4895048 | biostudies-literature | 2016 May

REPOSITORIES: biostudies-literature

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A dual molecular analogue tuner for dissecting protein function in mammalian cells.

Brosh Ran R Hrynyk Iryna I Shen Jessalyn J Waghray Avinash A Zheng Ning N Lemischka Ihor R IR

Nature communications 20160527

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 endoge ...[more]

PMID: 27230261

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A Dual Molecular Analog Tuner for Dissecting Mammalian Protein Function

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