Benzoate X receptor zinc-finger gene switches for drug-inducible regulation of transcription

Schwimmer, Lauren J.; González, Beatríz; Barbas, Carlos F.

doi:10.1038/gt.2011.112

Cited by 7 publications

(5 citation statements)

References 30 publications

(46 reference statements)

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The ability to easily manipulate gene expression with spatial and temporal control in a tunable and reversible manner is critical to many potential applications in gene therapy, regenerative medicine, tissue engineering, metabolic engineering, synthetic biology, and basic research. The most common gene regulation systems are controlled by the presence of a small molecule such as an antibiotic or steroid analogue [1–9]; however, because small molecules quickly diffuse throughout tissues and cell culture, these systems cannot be used to achieve spatial control over gene regulation. Furthermore, the small molecule must be removed from the system to reverse its effect, making it difficult to dynamically control gene expression for some applications.…”

Section: Introductionmentioning

confidence: 99%

Light-Inducible Gene Regulation with Engineered Zinc Finger Proteins

Polstein

Gersbach

2014

Methods in Molecular Biology

View full text Add to dashboard Cite

The coupling of light-inducible protein-protein interactions with gene regulation systems has enabled the control of gene expression with light. In particular, heterodimer protein pairs from plants can be used to engineer a gene regulation system in mammalian cells that is reversible, repeatable, tunable, controllable in a spatiotemporal manner, and targetable to any DNA sequence. This system, Light-Inducible Transcription using Engineered Zinc finger proteins (LITEZ), is based on the blue light-induced interaction of GIGANTEA and the LOV domain of FKF1 that drives the localization of a transcriptional activator to the DNA-binding site of a highly customizable engineered zinc finger protein. This chapter provides methods for modifying LITEZ to target new DNA sequences, engineering a programmable LED array to illuminate cell cultures, and using the modified LITEZ system to achieve spatiotemporal control of transgene expression in mammalian cells.

show abstract

Section: Introductionmentioning

confidence: 99%

Light-Inducible Gene Regulation with Engineered Zinc Finger Proteins

Polstein

Gersbach

2014

Methods in Molecular Biology

View full text Add to dashboard Cite

show abstract

“…This can be due to the natural occurrence of hormones in the mammalian system or the presence of an ectopic overexpression of an activation domain. In a recent study, Schwimmer et al (2012) have partially addressed the first problem through the development of a benzoate X receptor gene switch. However, this has yet to be demonstrated in an endogenous context.…”

Section: Introductionmentioning

confidence: 99%

Orthogonal control of endogenous gene expression in mammalian cells using synthetic ligands

Liang

McLachlan

Zhao

2013

Biotech & Bioengineering

View full text Add to dashboard Cite

Gene switches have wide utility in synthetic biology, gene therapy, and developmental biology, and multiple orthogonal gene switches are needed to construct advanced circuitry or to control complex phenotypes. Endogenous vascular endothelial growth factor (VEGF-A) is crucial to angiogenesis, and it has been shown that multiple alternately spliced VEGF-A isoforms are necessary for proper blood vessel formation. Such a necessity limits the utility of direct transgene delivery, which can provide only one splice variant. To overcome this limitation, we constructed a gene switch that can regulate the (VEGF-A) locus in mammalian cells by combining an engineered estrogen receptor (ER) ligand-binding domain (LBD), a p65 activation domain, and an artificial zinc-finger DNA binding domain (DBD). Our gene switch is specifically and reversibly controlled by 4,4'-dyhydroxybenzil (DHB), a small molecule, non-steroid synthetic ligand, which acts orthogonally in a mammalian system. After optimization of the gene switch architecture, an endogenous VEGF-A induction ratio of >100-fold can be achieved in HEK293 cells at 1 µM DHB, which is the highest endogenous induction reported to date. In addition, induction has been shown to be reversible, repeatable, and sustainable. Another advantage is that the ligand response is tunable by varying the clonal composition of a stably integrated cell line. The integration of our findings with the technology to change ligand specificity and DNA binding specificity will provide the framework for generating a wide array of orthogonal gene switches that can control multiple genes with multiple orthogonal ligands.

show abstract

“…Upon ligand binding, however, these hormone receptors are released by the inactivating complex, undergo intermolecular homodimerization, and bind their target DNA to activate transcription (Figure 1a). We 41−43 and others 44,45 have previously shown that zinc-finger transcription factors 46 fused to these LBDs modulate gene expression in response to specific chemical stimuli. As the natural ligands of these mammalian receptors mediate significant biological activity and thus may stimulate induction of unwanted and potentially confounding gene networks, we employed mutants of the ER 47 and PR 48 proteins that no longer bind their natural substrates but instead recognize the synthetic agonists 4-hydroxytamoxifen (4-OHT) and RU486, respectively.…”

mentioning

confidence: 97%

Regulation of Endogenous Human Gene Expression by Ligand-Inducible TALE Transcription Factors

et al. 2013

View full text Add to dashboard Cite

The construction of increasingly sophisticated synthetic biological circuits is dependent on the development of extensible tools capable of providing specific control of gene expression in eukaryotic cells. Here, we describe a new class of synthetic transcription factors that activate gene expression in response to extracellular chemical stimuli. These inducible activators consist of customizable transcription activator-like effector (TALE) proteins combined with steroid hormone receptor ligand-binding domains. We demonstrate that these ligand-responsive TALE transcription factors allow for tunable and conditional control of gene activation and can be used to regulate the expression of endogenous genes in human cells. Since TALEs can be designed to recognize any contiguous DNA sequence, the conditional gene regulatory system described herein will enable the design of advanced synthetic gene networks.

show abstract

Benzoate X receptor zinc-finger gene switches for drug-inducible regulation of transcription

Cited by 7 publications

References 30 publications

Light-Inducible Gene Regulation with Engineered Zinc Finger Proteins

Light-Inducible Gene Regulation with Engineered Zinc Finger Proteins

Orthogonal control of endogenous gene expression in mammalian cells using synthetic ligands

Regulation of Endogenous Human Gene Expression by Ligand-Inducible TALE Transcription Factors

Contact Info

Product

Resources

About