pHuji, a pH-sensitive red fluorescent protein for imaging of exo- and endocytosis

Shen, Yi; Rosendale, Morgane; Campbell, Robert E.; Perrais, David

doi:10.1085/jgp.1446oia52

Cited by 20 publications

(39 citation statements)

References 27 publications

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“…First, a number of red fluorescent pH sensors appeared [35][36][37] (Figure 2a). With pHRed [35], quantitative imaging of pH changes is possible due to the ratiometric response.…”

Section: Genetically Encoded Fluorescent Biosensors Go Redmentioning

confidence: 99%

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Novel uses of fluorescent proteins

Mishin

Belousov

Solntsev

et al. 2015

Current Opinion in Chemical Biology

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The field of genetically encoded fluorescent probes is developing rapidly. New chromophore structures were characterized in proteins of green fluorescent protein (GFP) family. A number of red fluorescent sensors, for example, for pH, Ca(2+) and H2O2, were engineered for multiparameter imaging. Progress in development of microscopy hardware and software together with specially designed FPs pushed superresolution fluorescence microscopy towards fast live-cell imaging. Deeper understanding of FPs structure and photophysics led to further development of imaging techniques. In addition to commonly used GFP-like proteins, unrelated types of FPs on the base of flavin-binding domains, bilirubin-binding domains or biliverdin-binding domains were designed. Their distinct biochemical and photophysical properties opened previously unexplored niches of FP uses such as labeling under anaerobic conditions, deep tissue imaging and even patients' blood analysis.

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“…First, a number of red fluorescent pH sensors appeared [35][36][37] (Figure 2a). With pHRed [35], quantitative imaging of pH changes is possible due to the ratiometric response.…”

Section: Genetically Encoded Fluorescent Biosensors Go Redmentioning

confidence: 99%

“…But the sensor was used also to quantify mitochondrial pH changes. Later, two other red pH reporters appeared, pHTomato [36] and pHuji [37]. Both are intensiometric and used mostly to monitor endocytosis and exocytosis events.…”

Section: Genetically Encoded Fluorescent Biosensors Go Redmentioning

confidence: 99%

Novel uses of fluorescent proteins

Mishin

Belousov

Solntsev

et al. 2015

Current Opinion in Chemical Biology

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“…Next, we directly assessed membrane scission achieved by the dyn2 mutants using the ppH assay. This assay relies on the overexpression of TfR fused to pH sensitive superecliptic pHluorin (TfR-SEP) as an endocytosis marker to detect the formation of single CCVs in living cells with a temporal resolution of 2 s (Merrifield et al, 2005;Shen et al, 2014;Taylor et al, 2011). We co-transfected TKO cells with TfR-SEP and dyn2-mCherry constructs.…”

Section: Recruitment Kinetics Of Dynamin Prd Mutantsmentioning

confidence: 99%

Functional recruitment of dynamin requires multimeric interactions for efficient endocytosis

Rosendale

Van

Grillo-Bosch

et al. 2018

Preprint

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During clathrin mediated endocytosis (CME), membrane scission is achieved by the concerted action of dynamin and its interacting partners. Essential interactions occur between the proline/arginine-rich domain of dynamin (dynPRD) and the Src-homology domain 3 (SH3) of various proteins including amphiphysins. Here we show that multiple SH3 domains must bind simultaneously to dynPRD through three adjacent motifs for dynamin’s efficient recruitment and function. First, we show in dynamin triple knock-out cells that mutant dynamins modified in a single motif, including the central amphiphysin SH3 (amphSH3) binding motif, are partially capable of rescuing CME. However, mutating two motifs largely prevents that ability. To support this observation, we designed divalent dynPRD-derived peptides. These ligands bind multimers of amphSH3 with >100-fold higher affinity than monovalent ones in vitro. Accordingly, dialyzing living cells with these divalent peptides through a patch-clamp pipette blocks CME 2 to 3 times more effectively than with monovalent ones. Finally, the frequency of endocytic events decreases with competing peptides or hypomorphic rescue mutants but the kinetics of dynamin recruitment is unaffected. This suggests that PRD-SH3 interactions act upstream of dynamin accumulation at the neck of nascent vesicles. We conclude from these data that dynamin drives vesicle scission via multivalent interactions in vivo.

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“…Another level of regulation of neuronal communication and plasticity is correlated to the amount of post-synaptic receptors inserted into the plasma membrane through tight regulation between cycles of endocytosis and exocytosis. David Perrais developed tools based on TIRFM to visualise receptor endocytosis in live neuronal dendrites (Rosendale et al, 2017;Shen et al, 2014).…”

Section: Exocytosis and Membrane Trafficking In Neurons And Endocrinementioning

confidence: 99%

Meeting after meeting: 20 years of discoveries by the members of the Exocytosis–Endocytosis Club

Niedergang

Gasman

Vitale

et al. 2017

Biology of the Cell

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Twenty years ago, a group of French cell biologists merged two scientific clubs with the aim of bringing together researchers in the fields of Endocytosis and Exocytosis. Founded in 1997, the first annual meeting of the Exocytosis Club was held in 1998. The Endocytosis Club held quarterly meetings from its founding in 1999. The first joint annual meeting of the Exocytosis-Endocytosis Club took place in Paris in April, 2001. What started as a modest gathering of enthusiastic scientists working in the field of cell trafficking has gone from strength to strength, rapidly becoming an unmissable yearly meeting, vividly demonstrating the high quality of science performed in our community and beyond. On the occasion of the 20th meeting of our club, we want to provide historic insight into the fields of exocytosis and endocytosis, and by extension, to subcellular trafficking, highlighting how French scientists have contributed to major advances in these fields. Today, the Exocytosis-Endocytosis Club represents a vibrant and friendly community that will hold its 20th meeting at the Presqu'Ile de Giens, near Toulon in the South of France, on May 11-13, 2017.

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pHuji, a pH-sensitive red fluorescent protein for imaging of exo- and endocytosis

Cited by 20 publications

References 27 publications

Novel uses of fluorescent proteins

Novel uses of fluorescent proteins

Functional recruitment of dynamin requires multimeric interactions for efficient endocytosis

Meeting after meeting: 20 years of discoveries by the members of the Exocytosis–Endocytosis Club

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