Engineering light-inducible nuclear localization signals for precise spatiotemporal control of protein dynamics in living cells

Niopek, Dominik; Benzinger, Dirk; Roensch, Julia; Draebing, Thomas; Wehler, Pierre; Eils, Roland; Ventura, Barbara Di

doi:10.1038/ncomms5404

Cited by 218 publications

(260 citation statements)

References 26 publications

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“…15 Here exposure to blue light provokes the uncaging of an added NLS that is otherwise buried within the protein structure in the dark. The system benefits from its small size and is useful for triggering cellular processes, yet requires optimization (e.g., reduce leakiness, increase dynamic range) for generalized applications.…”

Section: Acs Synthetic Biologymentioning

confidence: 99%

Red Light-Regulated Reversible Nuclear Localization of Proteins in Mammalian Cells and Zebrafish

et al. 2015

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Protein trafficking in and out of the nucleus represents a key step in controlling cell fate and function. Here we report the development of a red light-inducible and far-red light-reversible synthetic system for controlling nuclear localization of proteins in mammalian cells and zebrafish. First, we synthetically reconstructed and validated the red light-dependent Arabidopsis phytochrome B nuclear import mediated by phytochrome-interacting factor 3 in a nonplant environment and support current hypotheses on the import mechanism in planta. On the basis of this principle we next regulated nuclear import and activity of target proteins by the spatiotemporal projection of light patterns. A synthetic transcription factor was translocated into the nucleus of mammalian cells and zebrafish to drive transgene expression. These data demonstrate the first in vivo application of a plant phytochrome-based optogenetic tool in vertebrates and expand the repertoire of available light-regulated molecular devices.

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Section: Acs Synthetic Biologymentioning

confidence: 99%

Red Light-Regulated Reversible Nuclear Localization of Proteins in Mammalian Cells and Zebrafish

et al. 2015

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“…A LOV2 domain of phototropin 1 from Avena sativa ( As LOV2) and a modified PDZ domain (ePDZ) are combined into an optogenetic system based on heterodimerization 4 . As LOV2-based optogenetic tools enable light control of nuclear–cytoplasmic protein shuttling 5–7 . Cryptochrome 2 (CRY2) from Arabidopsis thaliana is another photoreceptor, which initially was applied to PPI heterodimerization approaches 8 .…”

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confidence: 99%

Near-infrared optogenetic pair for protein regulation and spectral multiplexing

et al. 2017

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Multifunctional optogenetic systems are in high demand for use in basic and biomedical research. Near-infrared-light-inducible binding of bacterial phytochrome BphP1 to its natural PpsR2 partner is beneficial for simultaneous use with blue-light-activatable tools. However, applications of the BphP1–PpsR2 pair are limited by the large size, multidomain structure and oligomeric behavior of PpsR2. Here, we engineered a single-domain BphP1 binding partner, Q-PAS1, which is three-fold smaller and lacks oligomerization. We exploited a helix–PAS fold of Q-PAS1 to develop several near-infrared-light-controllable transcription regulation systems, enabling either 40-fold activation or inhibition. The light-induced BphP1–Q-PAS1 interaction allowed modification of the chromatin epigenetic state. Multiplexing the BphP1–Q-PAS1 pair with a blue-light-activatable LOV-domain-based system demonstrated their negligible spectral crosstalk. By integrating the Q-PAS1 and LOV domains in a single optogenetic tool, we achieved tridirectional protein targeting, independently controlled by near-infrared and blue light, thus demonstrating the superiority of Q-PAS1 for spectral multiplexing and engineering of multicomponent systems.

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“…Proof-ofconcept was first realized with the small GTPase Rac1, for which light-dependent conformational changes controlled access of Rac1 to its effector protein kinase PAK and, thus, cell motility [59] (Figure 4A). Similar types of light-inducible protein switch have since been constructed to activate kinase inhibitors [60], control membrane and promoter localization through PDZ affinity-clamp interactions [61,62], control ion channel permeability [63], regulate gene expression and proteosomal degradation in E. coli through ipaA-vinculin and SsrA-SspB interactions [64] and control nuclear localization [65]. In the latter two cases, Ja of AsLov2 was engineered such that the molecular specificity feature-binding of the AsLov2 core overlapped with effector binding.…”

Section: Light-dependent Protein Switchesmentioning

confidence: 99%