Genetic targeting of chemical indicators in vivo

Yang, Guo‐Ping; Reis, Fernanda de Castro; Sundukova, Mayya; Pimpinella, Sofia; Asaro, Antonino; Castaldi, Laura; Batti, Laura; Bilbao, Daniel; Reymond, Luc; Johnsson, Kai; Heppenstall, Paul A.

doi:10.1038/nmeth.3207

Cited by 63 publications

(66 citation statements)

References 23 publications

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“…The generation and genotyping of all mouse lines used has been described previously (Kalebic et al, 2013b; Yang et al, 2015; Zurborg et al, 2011). …”

Section: Methodsmentioning

confidence: 99%

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Acetylated tubulin is essential for touch sensation in mice

Morley

Qi²,

Iovino

et al. 2016

eLife

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At its most fundamental level, touch sensation requires the translation of mechanical energy into mechanosensitive ion channel opening, thereby generating electro-chemical signals. Our understanding of this process, especially how the cytoskeleton influences it, remains unknown. Here we demonstrate that mice lacking the α-tubulin acetyltransferase Atat1 in sensory neurons display profound deficits in their ability to detect mechanical stimuli. We show that all cutaneous afferent subtypes, including nociceptors have strongly reduced mechanosensitivity upon Atat1 deletion, and that consequently, mice are largely insensitive to mechanical touch and pain. We establish that this broad loss of mechanosensitivity is dependent upon the acetyltransferase activity of Atat1, which when absent leads to a decrease in cellular elasticity. By mimicking α-tubulin acetylation genetically, we show both cellular rigidity and mechanosensitivity can be restored in Atat1 deficient sensory neurons. Hence, our results indicate that by influencing cellular stiffness, α-tubulin acetylation sets the force required for touch.DOI: http://dx.doi.org/10.7554/eLife.20813.001

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“…The generation and genotyping of all mouse lines used has been described previously (Kalebic et al, 2013b; Yang et al, 2015; Zurborg et al, 2011). …”

Section: Methodsmentioning

confidence: 99%

“…SNAP-tag labelling was carried out by intradermal injection of the finger in anaesthetized mice of 2 μM BG TMRstar substrate as described previously (Yang et al, 2015). After five hours the animals were sacrificed and the samples were mounted in 80% glycerol for imaging.…”

Section: Methodsmentioning

confidence: 99%

Acetylated tubulin is essential for touch sensation in mice

Morley

Qi²,

Iovino

et al. 2016

eLife

Self Cite

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“…Extension to living model organisms has lagged behind the applications in cells, but remains a very exciting future direction[41]. For these in-vivo applications, the optimal fluorogens will need to be coupled with highly active, highly robust fluorogen activating proteins that are functional in various intracellular compartments.…”

Section: Discussionmentioning

confidence: 99%

Dark dyes–bright complexes: fluorogenic protein labeling

Bruchez

2015

Current Opinion in Chemical Biology

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Complexes formed between organic dyes and genetically encoded proteins combine the advantages of stable and tunable fluorescent molecules and targetable, biologically integrated labels. To overcome the challenges imposed by labeling with bright fluorescent dyes, a number of approaches now exploit chemical or environmental changes to control the properties of a bound dye, converting dyes from a weakly fluorescent state to a bright, easily detectable complex. Optimized, such approaches avoid the need for removal of unbound dyes, facilitate rapid and simple assays in cultured cells and enable hybrid labeling to function more robustly in living model organisms.

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“…The fusion was found to react with a benzyl guanine partner both in vivo and ex vivo. 12 Another popular tag is the HaloTag (34 kD), which recognizes an alkyl chloride substrate and transfers the alkyl group to the active-site aspartate. 13 Recently, Crews and coworkers used the HaloTag in combination with an unusual substrate that destabilizes the tertiary structure of the tag.…”

Section: Enzyme-protein Fusionsmentioning

confidence: 99%

Recent Advances in Chemoenzymatic Bioconjugation Methods

Rabuka¹

2015

Organic Chemistry Insights

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Genetic fusions of either full enzymes or peptide tags with a protein of interest have enabled the synthesis of protein conjugates with precise control over the site of attachment and number of payloads incorporated. Engineering of protein glycans, depending on the application, can lead to similar control for nonengineered glycoproteins. Recent advances in the field of chemoenzymatic modifications of proteins for site-specific protein conjugation will be reviewed. These techniques have been used in an array of fields for the execution of innovative and valuable experiments. Specific industrial applications of these technologies will be highlighted.KEY WORDS: bioconjugation, antibody drug conjugate, protein tag, transferase, ligase, glycoconjugate, chemoenzymatic CITATION: mcFarland and rabuka. recent advances in chemoenzymatic Bioconjugation methods.

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Genetic targeting of chemical indicators in vivo

Cited by 63 publications

References 23 publications

Acetylated tubulin is essential for touch sensation in mice

Acetylated tubulin is essential for touch sensation in mice

Dark dyes–bright complexes: fluorogenic protein labeling

Recent Advances in Chemoenzymatic Bioconjugation Methods

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