scholarly journals The GTPase-Activating Protein FgGyp1 Is Important for Vegetative Growth, Conidiation, and Virulence and Negatively Regulates DON Biosynthesis in Fusarium graminearium

2021 ◽  
Vol 12 ◽  
Author(s):  
Qiaojia Zheng ◽  
Zhi Yu ◽  
Yanping Yuan ◽  
Danli Sun ◽  
Yakubu Saddeeq Abubakar ◽  
...  
Keyword(s):  
Vegetative Growth ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Negative Regulator ◽  
Rab Gtpase ◽  
Wild Type ◽  
Gtpase Activating Protein ◽  
Replacement Method

Ypt1 is a small Rab GTPase in yeast, Gyp1 functions at the Golgi as a negative regulator of Ypt1. Gyp1 homologs are conserved in filamentous fungi. However, the roles of Gyp1 in phytopathogenic fungi are still unclear. Herein, we investigated the functions of FgGyp1 in the wheat pathogen Fusarium graminearum by live-cell imaging, genetic, and pathological analyses. Targeted gene replacement method was used to delete FgGYP1 in F. graminearum. Phenotypic analyses showed that FgGyp1 is critically important not only for the vegetative growth of F. graminearum but also its conidiation. The mutant’s vegetative growth was significantly reduced by 70% compared to the wild type PH-1. The virulence of FgGYP1 deletion mutant was significantly decreased when compared with the wild type PH-1. We further found that FgGyp1 negatively regulates DON production of the fungus. Live-cell imaging clearly demonstrated that FgGyp1 mainly localizes to the Golgi apparatus. Moreover, the TBC domain, C-terminal, and N-terminal regions of FgGyp1 are found to be indispensable for its biological functions and normal localization. The Arg357 residue of FgGyp1 is essential for its functions but dispensable for the normal localization of the protein, while the Arg284 residue is not required for both the functions and normal localization of the protein. Furthermore, we showed that FgGyp1 essentially hydrolyzes the GTP-bound FgRab1 (activated form) to its corresponding GDP-bound (inactive) form in vitro, suggesting that FgGyp1 is a GTPase-activating protein (GAP) for FgRab1. Finally, FgGyp1 was found to be important for FgSnc1-mediated fusion of secretory vesicles from the Golgi with the plasma membrane in F. graminearum. Put together, these data demonstrate that FgGyp1 functions as a GAP for FgRab1 and is important for vegetative growth, conidiation and virulence, and negatively regulates DON biosynthesis in F. graminearum.

A new visible-light-excitable ICT-CHEF-mediated fluorescence ‘turn-on’ probe for the selective detection of Cd2+ in a mixed aqueous system with live-cell imaging

2015 ◽  
Vol 44 (12) ◽  
pp. 5763-5770 ◽  
Author(s):  
Shyamaprosad Goswami ◽  
Krishnendu Aich ◽  
Sangita Das ◽  
Chitrangada Das Mukhopadhyay ◽  
Deblina Sarkar ◽  
...  
Keyword(s):  
Visible Light ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Aqueous System ◽  
Live Cell ◽  
Selective Detection ◽  
Turn On ◽  
Fluorescence Turn On

A new quinoline based sensor was developed and applied for the selective detection of Cd2+ both in vitro and in vivo.

scholarly journals tdLanYFP, a yellow, bright, photostable and pH insensitive fluorescent protein for live cell imaging and FRET-based sensing strategies

2021 ◽  
Author(s):  
Y. Bousmah ◽  
H. Valenta ◽  
G. Bertolin ◽  
U. Singh ◽  
V. Nicolas ◽  
...  
Keyword(s):  
Single Molecule ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Fluorescent Protein ◽  
Fluorescent Proteins ◽  
Yellow Fluorescent Protein ◽  
Resonance Energy ◽  
Live Cell ◽  
Live Cells

AbstractYellow fluorescent proteins (YFP) are widely used as optical reporters in Förster Resonance Energy Transfer (FRET) based biosensors. Although great improvements have been done, the sensitivity of the biosensors is still limited by the low photostability and the poor fluorescence performances of YFPs at acidic pHs. In fact, today, there is no yellow variant derived from the EYFP with a pK1/2 below ∼5.5. Here, we characterize a new yellow fluorescent protein, tdLanYFP, derived from the tetrameric protein from the cephalochordate B. lanceolatum, LanYFP. With a quantum yield of 0.92 and an extinction coefficient of 133 000 mol−1.L.cm−1, it is, to our knowledge, the brightest dimeric fluorescent protein available, and brighter than most of the monomeric YFPs. Contrasting with EYFP and its derivatives, tdLanYFP has a very high photostability in vitro and preserves this property in live cells. As a consequence, tdLanYFP allows the imaging of cellular structures with sub-diffraction resolution with STED nanoscopy. We also demonstrate that the combination of high brightness and strong photostability is compatible with the use of spectro-microscopies in single molecule regimes. Its very low pK1/2 of 3.9 makes tdLanYFP an excellent tag even at acidic pHs. Finally, we show that tdLanYFP can be a FRET partner either as donor or acceptor in different biosensing modalities. Altogether, these assets make tdLanYFPa very attractive yellow fluorescent protein for long-term or single-molecule live-cell imaging that is also suitable for FRET experiment including at acidic pH.

scholarly journals The Use of Live Cell Imaging and Automated Image Analysis to Assist With Determining Optimal Parameters for Angiogenic Assay in vitro

2019 ◽  
Vol 7 ◽  
Author(s):  
Brooke M. Huuskes ◽  
Ryan J. DeBuque ◽  
Peter G. Kerr ◽  
Chrishan S. Samuel ◽  
Sharon D. Ricardo
Keyword(s):  
Image Analysis ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Automated Image Analysis ◽  
Optimal Parameters

Naked-eye detection of Pd2+ ion using a highly selective fluorescent heterocyclic probe by “turn-off” response and in-vitro live cell imaging

2020 ◽  
Vol 394 ◽  
pp. 112441
Author(s):  
Satyajit Mahata ◽  
Araghni Bhattacharya ◽  
Jadi Praveen Kumar ◽  
Biman B. Mandal ◽  
Vadivelu Manivannan
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Eye Detection ◽  
Naked Eye ◽  
Naked Eye Detection

scholarly journals Live-cell imaging of nuclear–chromosomal dynamics in bovine in vitro fertilised embryos

2018 ◽  
Vol 8 (1) ◽  
Author(s):  
Tatsuma Yao ◽  
Rie Suzuki ◽  
Natsuki Furuta ◽  
Yuka Suzuki ◽  
Kyoko Kabe ◽  
...  
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell

scholarly journals Live Cell Imaging of In Vitro Human Trophoblast Syncytialization1

2014 ◽  
Vol 90 (6) ◽  
Author(s):  
Rui Wang ◽  
Yan-Li Dang ◽  
Ru Zheng ◽  
Yue Li ◽  
Weiwei Li ◽  
...  
Keyword(s):  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Human Trophoblast

scholarly journals Genotypes and phenotypes of resistance in Ecuadorian Plasmodium falciparum

2019 ◽  
Vol 18 (1) ◽  
Author(s):  
Gabriela Valenzuela ◽  
L. Enrique Castro ◽  
Julio Valencia-Zamora ◽  
Claudia A. Vera-Arias ◽  
Petra Rohrbach ◽  
...  
Keyword(s):  
Plasmodium Falciparum ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Chloroquine Resistance ◽  
In Vitro Assays ◽  
Wild Type ◽  
The Status ◽  
Transport Assay ◽  
Resistance Markers

Abstract Background Malaria continues to be endemic in the coast and Amazon regions of Ecuador. Clarifying current Plasmodium falciparum resistance in the country will support malaria elimination efforts. In this study, Ecuadorian P. falciparum parasites were analysed to determine their drug resistance genotypes and phenotypes. Methods Molecular analyses were performed to search for mutations in known resistance markers (Pfcrt, Pfdhfr, Pfdhps, Pfmdr1, k13). Pfmdr1 copy number was determined by qPCR. PFMDR1 transporter activity was characterized in live parasites using live cell imaging in combination with the Fluo-4 transport assay. Chloroquine, quinine, lumefantrine, mefloquine, dihydroartemisinin, and artemether sensitivities were measured by in vitro assays. Results The majority of samples from this study presented the CVMNT genotype for Pfcrt (72–26), NEDF SDFD mutations in Pfmdr1 and wild type genotypes for Pfdhfr, Pfdhps and k13. The Ecuadorian P. falciparum strain ESM-2013 showed in vitro resistance to chloroquine, but sensitivity to quinine, lumefantrine, mefloquine, dihydroartemisinin and artemether. In addition, transport of the fluorochrome Fluo-4 from the cytosol into the digestive vacuole (DV) of the ESM-2013 strain was minimally detected in the DV. All analysed samples revealed one copy of Pfmdr1. Conclusion This study indicates that Ecuadorian parasites presented the genotype and phenotype for chloroquine resistance and were found to be sensitive to SP, artemether-lumefantrine, quinine, mefloquine, and dihydroartemisinin. The results suggest that the current malaria treatment employed in the country remains effective. This study clarifies the status of anti-malarial resistance in Ecuador and informs the P. falciparum elimination campaigns in the country.

Human and equine endothelial cells in a live cell imaging scratch assay in vitro

2019 ◽  
Vol 70 (4) ◽  
pp. 495-509 ◽  
Author(s):  
Juliane Rieger ◽  
Carsten Hopperdietzel ◽  
Sabine Kaessmeyer ◽  
Ilka Slosarek ◽  
Sebastian Diecke ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Live Cell Imaging ◽  
Cell Imaging ◽  
Live Cell ◽  
Scratch Assay

Fluorogenic Protein Probes with Red or Near-Infrared Emission for Genetically Targeted Live-Cell Imaging

2020 ◽  
Author(s):  
Sylvestre P. J. T. Bachollet ◽  
Cyril Addi ◽  
Jean-Maurice Mallet ◽  
Blaise Dumat
Keyword(s):  
Live Cell Imaging ◽  
Near Infrared ◽  
Cell Imaging ◽  
Infrared Emission ◽  
Live Cell ◽  
Molecular Rotor ◽  
Model Protein ◽  
Near Infrared Emission

A series of red-emitting and near-infrared fluorogenic protein probes based on push-pull molecular rotor structures was developed. After characterization of their optical properties using Bovine Serum Albumin as a model protein, they were conjugated to a halogenoalkane ligand in order to target the protein self-labeling tag HaloTag. The interaction with HaloTag was investigated in vitro and then the most promising probes were applied to live-cell imaging in wash-free conditions using fluorogenic and chemogenetic targeting of HaloTag fusion proteins.<br>

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