scholarly journals A novel GSK3-regulated APC:Axin interaction regulates Wnt signaling by driving a catalytic cycle of efficient βcatenin destruction

eLife ◽  
2015 ◽  
Vol 4 ◽  
Author(s):  
Mira I Pronobis ◽  
Nasser M Rusan ◽  
Mark Peifer
Keyword(s):  
Wnt Signaling ◽  
Dynamic Model ◽  
High Throughput ◽  
Mammalian Cells ◽  
Super Resolution ◽  
Negative Regulator ◽  
Functional Tests ◽  
Interaction Site ◽  
Signaling Complex ◽  
Super Resolution Microscopy

APC, a key negative regulator of Wnt signaling in development and oncogenesis, acts in the destruction complex with the scaffold Axin and the kinases GSK3 and CK1 to target βcatenin for destruction. Despite 20 years of research, APC's mechanistic function remains mysterious. We used FRAP, super-resolution microscopy, functional tests in mammalian cells and flies, and other approaches to define APC's mechanistic role in the active destruction complex when Wnt signaling is off. Our data suggest APC plays two roles: (1) APC promotes efficient Axin multimerization through one known and one novel APC:Axin interaction site, and (2) GSK3 acts through APC motifs R2 and B to regulate APC:Axin interactions, promoting high-throughput of βcatenin to destruction. We propose a new dynamic model of how the destruction complex regulates Wnt signaling and how this goes wrong in cancer, providing insights into how this multiprotein signaling complex is assembled and functions via multivalent interactions.

scholarly journals STED super-resolution microscopy in Drosophila tissue and in mammalian cells

2011 ◽  
Author(s):  
Lana Lau ◽  
Yin Loon Lee ◽  
Maja Matis ◽  
Jeff Axelrod ◽  
Tim Stearns ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Super Resolution ◽  
Super Resolution Microscopy

scholarly journals Photonic chip-based multimodal super-resolution microscopy for histopathological assessment of cryopreserved tissue sections

2021 ◽  
Author(s):  
Luis E. Villegas-Hernandez ◽  
Vishesh Kumar Dubey ◽  
Mona Nystad ◽  
Jean-Claude Tinguely ◽  
David A. Coucheron ◽  
...  
Keyword(s):  
High Throughput ◽  
Single Molecule ◽  
Operating Cost ◽  
Super Resolution ◽  
Anatomical Variations ◽  
Histopathological Analysis ◽  
Tissue Samples ◽  
Tissue Sections ◽  
Imaging Tool ◽  
Super Resolution Microscopy

Histopathological assessment involves the identification of anatomical variations in tissues that are associated with diseases. While diffraction-limited optical microscopes assist in the diagnosis of a wide variety of pathologies, their resolving capabilities are insufficient to visualize some anomalies at subcellular level. Although a novel set of super-resolution optical microscopy techniques can fulfill the resolution demands in such cases, the system complexity, high operating cost, lack of multimodality, and low-throughput imaging of these methods limit their wide adoption in clinical settings. In this study, we interrogate the photonic chip as an attractive high-throughput super-resolution microscopy platform for histopathology. Using cryopreserved ultrathin tissue sections of human placenta, mouse kidney, and zebrafish eye retina prepared by the Tokuyasu method, we validate the photonic chip as a multi-modal imaging tool for histo-anatomical analysis. We demonstrate that photonic-chip platform can deliver multi-modal imaging capabilities such as total internal reflection fluorescence microscopy, intensity fluctuation-based optical nanoscopy, single-molecule localization microscopy, and correlative light-electron microscopy. Our results demonstrate that the photonic chip-based super-resolution microscopy platform has the potential to deliver high-throughput multimodal histopathological analysis of cryopreserved tissue samples.

scholarly journals Distinctive nuclear zone for RAD51-mediated homologous recombinational DNA repair

2021 ◽  
Author(s):  
Yasunori Horikoshi ◽  
Hiroki Shima ◽  
Wataru Kobayashi ◽  
Jiying Sun ◽  
Volker J Schmid ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Nuclear Architecture ◽  
Super Resolution ◽  
Double Strand Breaks ◽  
Genome Integrity ◽  
Dna Double Strand Breaks ◽  
Strand Breaks ◽  
Sister Chromatids ◽  
Super Resolution Microscopy ◽  
Damaged Dna

Genome-based functions are inseparable from the dynamic higher-order architecture of the cell nucleus. In this context, the repair of DNA damage is coordinated by precise spatiotemporal controls that target and regulate the repair machinery required to maintain genome integrity. However, the mechanisms that pair damaged DNA with intact template for repair by homologous recombination (HR) without illegitimate recombination remain unclear. This report highlights the intimate relationship between nuclear architecture and HR in mammalian cells. RAD51, the key recombinase of HR, forms spherical foci in S/G2 phases spontaneously. Using super-resolution microscopy, we show that following induction of DNA double-strand breaks RAD51 foci at damaged sites elongate to bridge between intact and damaged sister chromatids; this assembly occurs within bundle-shaped distinctive nuclear zones, requires interactions of RAD51 with various factors, and precedes ATP-dependent events involved the recombination of intact and damaged DNA. We observed a time-dependent transfer of single-stranded DNA overhangs, generated during HR, into such zones. Our observations suggest that RAD51-mediated homologous pairing during HR takes place within the distinctive nuclear zones to execute appropriate recombination.

A Platinum(II) Based Correlative Probe for Bacteria

2019 ◽  
Author(s):  
Anna Maria Ranieri ◽  
Kathryn Leslie ◽  
Song Huang ◽  
Stefano Stagni ◽  
Denis Jacquemin ◽  
...  
Keyword(s):  
Mammalian Cells ◽  
Super Resolution ◽  
Molecular Probes ◽  
Live Cells ◽  
Structured Illumination ◽  
Luminescent Probe ◽  
Structured Illumination Microscopy ◽  
Cellular Localisation ◽  
Super Resolution Microscopy ◽  
Nanosims Analysis

There is a lack of molecular probes for imaging bacteria, in comparison to the array of such tools available for the imaging of mammalian cells. This is especially so for correlative probes, which are proving to be powerful tools for enhancing the imaging of live cells. In this work a platinum(II)-naphthalimide molecule has been developed to extend small molecule correlative probes to bacterial imaging. The probe was designed to exploit the naphthalimide moiety as a luminescent probe for super-resolution microscopy, with the platinum(II) centre enabling visualisation of the complex with ion nanoscopy. Photophysical characterisation and theoretical studies confirmed that the emission properties of the naphthalimide are not altered by the platinum(II) centre. Structured illumination microscopy (SIM) imaging on live <i>Bacillus cereus</i>revealed that the platinum(II) centre does not change the sub-cellular localisation of the naphthalimide, and confirmed the suitability of the probe for super-resolution microscopy. NanoSIMS analysis of the sample was used to monitor the uptake of the platinum(II) complex within the bacteria and proved the correlative action of the probe. The successful combination of these two probe moieties with no perturbation of their individual detection introduces a platform for a versatile range of new correlative probes for bacteria.

scholarly journals 3D Single-Molecule Super-Resolution Microscopy in Mammalian Cells Using a Tilted Light Sheet

2018 ◽  
Vol 114 (3) ◽  
pp. 14a
Author(s):  
Anna-Karin Gustavsson ◽  
Petar N. Petrov ◽  
Maurice Y. Lee ◽  
Yoav Shechtman W.E. Moerner
Keyword(s):  
Single Molecule ◽  
Mammalian Cells ◽  
Super Resolution ◽  
Light Sheet ◽  
Super Resolution Microscopy

scholarly journals Nradd Acts as a Negative Feedback Regulator of Wnt/β-Catenin Signaling and Promotes Apoptosis

Biomolecules ◽  
2021 ◽  
Vol 11 (1) ◽  
pp. 100
Author(s):  
Ozgun Ozalp ◽  
Ozge Cark ◽  
Yagmur Azbazdar ◽  
Betul Haykir ◽  
Gokhan Cucun ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Mammalian Cells ◽  
Genetic Disorders ◽  
Negative Regulator ◽  
Neurotrophin Receptor ◽  
P75 Neurotrophin Receptor ◽  
Death Domain ◽  
C Terminus ◽  
And Function ◽  
Functional Analyses

Wnt/β-catenin signaling controls many biological processes for the generation and sustainability of proper tissue size, organization and function during development and homeostasis. Consequently, mutations in the Wnt pathway components and modulators cause diseases, including genetic disorders and cancers. Targeted treatment of pathway-associated diseases entails detailed understanding of the regulatory mechanisms that fine-tune Wnt signaling. Here, we identify the neurotrophin receptor-associated death domain (Nradd), a homolog of p75 neurotrophin receptor (p75NTR), as a negative regulator of Wnt/β-catenin signaling in zebrafish embryos and in mammalian cells. Nradd significantly suppresses Wnt8-mediated patterning of the mesoderm and neuroectoderm during zebrafish gastrulation. Nradd is localized at the plasma membrane, physically interacts with the Wnt receptor complex and enhances apoptosis in cooperation with Wnt/β-catenin signaling. Our functional analyses indicate that the N-glycosylated N-terminus and the death domain-containing C-terminus regions are necessary for both the inhibition of Wnt signaling and apoptosis. Finally, Nradd can induce apoptosis in mammalian cells. Thus, Nradd regulates cell death as a modifier of Wnt/β-catenin signaling during development.

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