Cited 0 time in webofscience Cited 0 time in scopus

Single-cell mechanogenetics using monovalent magnetoplasmonic nanoparticles

Title
Single-cell mechanogenetics using monovalent magnetoplasmonic nanoparticles
Authors
Kim, Ji-wookSeo, DaehaLee, Jung-ukSouthard, Kaden M.Lim, YongjunKim, DaehyunGartner, Zev J.Jun, Young-wookCheon, Jinwoo
DGIST Authors
Seo, Daeha
Issue Date
2017-09
Citation
Nature Protocols, 12(9), 1871-1889
Type
Article
Article Type
Article
Keywords
Endosomal EscapeFe NanoparticlesIntracellular DeliveryLiving CellsMagnetic Nano Particles (MNPs)Molecule Force SpectroscopyQuantum DotsRemote ControlSignal TransductionSpatio Temporal Control
ISSN
1754-2189
Abstract
Spatiotemporal interrogation of signal transduction at the single-cell level is necessary to answer a host of important biological questions. This protocol describes a nanotechnology-based single-cell and single-molecule perturbation tool, termed mechanogenetics, that enables precise spatial and mechanical control over genetically encoded cell-surface receptors in live cells. The key components of this tool are a magnetoplasmonic nanoparticle (MPN) actuator that delivers defined spatial and mechanical cues to receptors through target-specific one-to-one engagement and a micromagnetic tweezers (μMT) that remotely controls the magnitude of force exerted on a single MPN. In our approach, a SNAP-tagged cell-surface receptor of interest is conjugated with a single-stranded DNA oligonucleotide, which hybridizes to its complementary oligonucleotide on the MPN. This protocol consists of four major stages: (i) chemical synthesis of MPNs, (ii) conjugation with DNA and purification of monovalent MPNs, (iii) modular targeting of MPNs to cell-surface receptors, and (iv) control of spatial and mechanical properties of targeted mechanosensitive receptors in live cells by adjusting the μMT-to-MPN distance. Using benzylguanine (BG)-functionalized MPNs and model cell lines expressing either SNAP-tagged Notch or vascular endothelial cadherin (VE-cadherin), we provide stepwise instructions for mechanogenetic control of receptor clustering and for mechanical receptor activation. The ability of this method to differentially control spatial and mechanical inputs to targeted receptors makes it particularly useful for interrogating the differential contributions of each individual cue to cell signaling. The entire procedure takes up to 1 week. © 2017 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.
URI
http://hdl.handle.net/20.500.11750/4502
DOI
10.1038/nprot.2017.071
Publisher
Nature Publishing Group
Files:
There are no files associated with this item.
Collection:
Emerging Materials ScienceETC1. Journal Articles


qrcode mendeley

Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.

BROWSE