scholarly journals Analysis of the Conditions That Affect the Selective Processing of Endogenous Notch1 by ADAM10 and ADAM17

2021 ◽  
Vol 22 (4) ◽  
pp. 1846
Author(s):  
Rolake O. Alabi ◽  
Jose Lora ◽  
Arda B. Celen ◽  
Thorsten Maretzky ◽  
Carl P. Blobel
Keyword(s):  
Cell Fate ◽  
Cleavage Site ◽  
Cell Contact ◽  
Substrate Selectivity ◽  
Mouse Embryonic Fibroblasts ◽  
Embryonic Fibroblasts ◽  
Cell Fate Decisions ◽  
Selective Processing ◽  
A Disintegrin And Metalloproteinase ◽  
Notch Activation

Notch signaling is critical for controlling a variety of cell fate decisions during metazoan development and homeostasis. This unique, highly conserved signaling pathway relies on cell-to-cell contact, which triggers the proteolytic release of the cytoplasmic domain of the membrane-anchored transcription factor Notch from the membrane. A disintegrin and metalloproteinase (ADAM) proteins are crucial for Notch activation by processing its S2 site. While ADAM10 cleaves Notch1 under physiological, ligand-dependent conditions, ADAM17 mainly cleaves Notch1 under ligand-independent conditions. However, the mechanism(s) that regulate the distinct contributions of these ADAMs in Notch processing remain unclear. Using cell-based assays in mouse embryonic fibroblasts (mEFs) lacking ADAM10 and/or ADAM17, we aimed to clarify what determines the relative contributions of ADAM10 and ADAM17 to ligand-dependent or ligand-independent Notch processing. We found that EDTA-stimulated ADAM17-dependent Notch1 processing is rapid and requires the ADAM17-regulators iRhom1 and iRhom2, whereas the Delta-like 4-induced ligand-dependent Notch1 processing is slower and requires ADAM10. The selectivity of ADAM17 for EDTA-induced Notch1 processing can most likely be explained by a preference for ADAM17 over ADAM10 for the Notch1 cleavage site and by the stronger inhibition of ADAM10 by EDTA. The physiological ADAM10-dependent processing of Notch1 cannot be compensated for by ADAM17 in Adam10-/- mEFs, or by other ADAMs shown here to be able to cleave the Notch1 cleavage site, such as ADAMs9, 12, and 19. Collectively, these results provide new insights into the mechanisms underlying the substrate selectivity of ADAM10 and ADAM17 towards Notch1.

Immobilization of Notch ligand, Delta-1, is required for induction of notch signaling

2000 ◽  
Vol 113 (23) ◽  
pp. 4313-4318 ◽  
Author(s):  
B. Varnum-Finney ◽  
L. Wu ◽  
M. Yu ◽  
C. Brashem-Stein ◽  
S. Staats ◽  
...  
Keyword(s):  
Cell Fate ◽  
Extracellular Domain ◽  
Notch Ligand ◽  
Cell Fate Decisions ◽  
Plastic Surface ◽  
Ligand Stabilization ◽  
Potential Activity ◽  
Inhibition Of Differentiation ◽  
Notch Ligands ◽  
Notch Activation

Cell-cell interactions mediated by Notch and its ligands are known to effect many cell fate decisions in both invertebrates and vertebrates. However, the mechanisms involved in ligand induced Notch activation are unknown. Recently it was shown that, in at least some cases, endocytosis of the extracellular domain of Notch and ligand by the signaling cell is required for signal induction in the receptive cell. These results imply that soluble ligands (ligand extracellular domains) although capable of binding Notch would be unlikely to activate it. To test the potential activity of soluble Notch ligands, we generated monomeric and dimeric forms of the Notch ligand Delta-1 by fusing the extracellular domain to either a series of myc epitopes (Delta-1(ext-myc)) or to the Fc portion of human IgG-1 (Delta-1(ext-IgG)), respectively. Notch activation, assayed by inhibition of differentiation in C2 myoblasts and by HES1 transactivation in U20S cells, occurred when either Delta-1(ext-myc) or Delta-1(ext-IgG) were first immobilized on the plastic surface. However, Notch was not activated by either monomeric or dimeric ligand in solution (non-immobilized). Furthermore, both non-immobilized Delta-1(ext-myc) and Delta-1(ext-IgG) blocked the effect of immobilized Delta. These results indicate that Delta-1 extracellular domain must be immobilized to induce Notch activation in C2 or U20S cells and that non-immobilized Delta-1 extracellular domain is inhibitory to Notch function. These results imply that ligand stabilization may be essential for Notch activation.

scholarly journals SyNPL: Synthetic Notch pluripotent cell lines to monitor and manipulate cell interactions in vitro and in vivo

2021 ◽  
Author(s):  
Mattias Malaguti ◽  
Rosa Portero Migueles ◽  
Jennifer Annoh ◽  
Daina Sadurska ◽  
Guillaume Blin ◽  
...  
Keyword(s):  
Cell Fate ◽  
Cell Lines ◽  
Modular Design ◽  
Cell Interactions ◽  
Cell Populations ◽  
Cell Contact ◽  
Pluripotent Cells ◽  
Cell Fate Decisions ◽  
Cell Cell

ABSTRACTCell-cell interactions govern differentiation and cell competition in pluripotent cells during early development, but the investigation of such processes is hindered by a lack of efficient analysis tools. Here we introduce SyNPL: clonal pluripotent stem cell lines which employ optimised Synthetic Notch (SynNotch) technology to report cell-cell interactions between engineered “sender” and “receiver” cells in cultured pluripotent cells and chimaeric mouse embryos. A modular design makes it straightforward to adapt the system for programming differentiation decisions non-cell-autonomously in receiver cells in response to direct contact with sender cells. We demonstrate the utility of this system by enforcing neuronal differentiation at the boundary between two cell populations. In summary, we provide a new tool which could be used to identify cell interactions and to profile changes in gene or protein expression that result from direct cell-cell contact with defined cell populations in culture and in early embryos, and which can be adapted to generate synthetic patterning of cell fate decisions.

scholarly journals ADAM10 is essential for Notch2-dependent marginal zone B cell development and CD23 cleavage in vivo

2010 ◽  
Vol 207 (3) ◽  
pp. 623-635 ◽  
Author(s):  
David R. Gibb ◽  
Mohey El Shikh ◽  
Dae-Joong Kang ◽  
Warren J. Rowe ◽  
Rania El Sayed ◽  
...  
Keyword(s):  
B Cell ◽  
Cell Fate ◽  
Marginal Zone ◽  
Cell Development ◽  
B Cell Development ◽  
Mouse Embryos ◽  
Cell Fate Decisions ◽  
Important Regulator ◽  
A Disintegrin And Metalloproteinase

The proteolytic activity of a disintegrin and metalloproteinase 10 (ADAM10) regulates cell-fate decisions in Drosophila and mouse embryos. However, in utero lethality of ADAM10−/− mice has prevented examination of ADAM10 cleavage events in lymphocytes. To investigate their role in B cell development, we generated B cell–specific ADAM10 knockout mice. Intriguingly, deletion of ADAM10 prevented development of the entire marginal zone B cell (MZB) lineage. Additionally, cleavage of the low affinity IgE receptor, CD23, was profoundly impaired, but subsequent experiments demonstrated that ADAM10 regulates CD23 cleavage and MZB development by independent mechanisms. Development of MZBs is dependent on Notch2 signaling, which requires proteolysis of the Notch2 receptor by a previously unidentified proteinase. Further experiments revealed that Notch2 signaling is severely impaired in ADAM10-null B cells. Thus, ADAM10 critically regulates MZB development by initiating Notch2 signaling. This study identifies ADAM10 as the in vivo CD23 sheddase and an important regulator of B cell development. Moreover, it has important implications for the treatment of numerous CD23- and Notch-mediated pathologies, ranging from allergy to cancer.

Position dependent control of cell fate in the Fucus embryo: role of intercellular communication

Development ◽  
1998 ◽  
Vol 125 (11) ◽  
pp. 1999-2008 ◽  
Author(s):  
F.Y. Bouget ◽  
F. Berger ◽  
C. Brownlee
Keyword(s):  
Cell Wall ◽  
Pattern Formation ◽  
Cell Fate ◽  
Intercellular Communication ◽  
Cell Types ◽  
Early Embryo ◽  
Cell Contact ◽  
Dependent Manner ◽  
Intact Cells ◽  
Cell Fate Decisions

The early embryo of the brown alga Fucus comprises two cell types, i. e. rhizoid and thallus which are morphogically and cytologically distinguishable. Previous work has pointed to the cell wall as a source of position-dependent information required for polarisation and fate determination in the zygote and 2-celled embryo. In this study we have analysed the mechanism(s) of cell fate control and pattern formation at later embryonic stages using a combination of laser microsurgery and microinjection. The results indicate that the cell wall is required for maintenance of pre-existing polarity in isolated intact cells. However, all cell types ultimately have the capacity to re-differentiate or regenerate rhizoid cells in response to ablation of neighbouring cells. This regeneration is regulated in a position-dependent manner and is strongly influenced by intercellular communication, probably involving transport or diffusion of inhibitory signals which appear to be essential for regulation of cell fate decisions. This type of cell-to-cell communication does not involve symplastic transport or direct cell-cell contact inhibition. Apoplastic diffusible gradients appear to be involved in pattern formation in the multicellular embryo.

scholarly journals Getting to know your neighbor: Cell polarization in early embryos

2014 ◽  
Vol 206 (7) ◽  
pp. 823-832 ◽  
Author(s):  
Jeremy Nance
Keyword(s):  
Cell Fate ◽  
Cell Polarization ◽  
Cell Contact ◽  
Cell Fates ◽  
Par Proteins ◽  
Morphogenetic Movements ◽  
Cell Fate Decisions ◽  
Contact Patterns ◽  
Early Embryos ◽  
Contact Free

Polarization of early embryos along cell contact patterns—referred to in this paper as radial polarization—provides a foundation for the initial cell fate decisions and morphogenetic movements of embryogenesis. Although polarity can be established through distinct upstream mechanisms in Caenorhabditis elegans, Xenopus laevis, and mouse embryos, in each species, it results in the restriction of PAR polarity proteins to contact-free surfaces of blastomeres. In turn, PAR proteins influence cell fates by affecting signaling pathways, such as Hippo and Wnt, and regulate morphogenetic movements by directing cytoskeletal asymmetries.

scholarly journals Positive autoregulation of lag-1 in response to LIN-12 activation in cell fate decisions during C. elegans reproductive system development

Development ◽  
2020 ◽  
Vol 147 (18) ◽  
pp. dev193482
Author(s):  
Katherine Leisan Luo ◽  
Ryan S. Underwood ◽  
Iva Greenwald
Keyword(s):  
Cell Fate ◽  
Reproductive System ◽  
Target Gene ◽  
System Development ◽  
Animal Development ◽  
Cell Fate Decisions ◽  
C Elegans ◽  
Spatiotemporal Regulation ◽  
Target Gene Transcription ◽  
Notch Activation

ABSTRACTDuring animal development, ligand binding releases the intracellular domain of LIN-12/Notch by proteolytic cleavage to translocate to the nucleus, where it associates with the DNA-binding protein LAG-1/CSL to activate target gene transcription. We investigated the spatiotemporal regulation of LAG-1/CSL expression in Caenorhabditis elegans and observed that an increase in endogenous LAG-1 levels correlates with LIN-12/Notch activation in different cell contexts during reproductive system development. We show that this increase is via transcriptional upregulation by creating a synthetic endogenous operon, and identified an enhancer region that contains multiple LAG-1 binding sites (LBSs) embedded in a more extensively conserved high occupancy target (HOT) region. We show that these LBSs are necessary for upregulation in response to LIN-12/Notch activity, indicating that lag-1 engages in direct positive autoregulation. Deletion of the HOT region from endogenous lag-1 reduced LAG-1 levels and abrogated positive autoregulation, but did not cause hallmark cell fate transformations associated with loss of lin-12/Notch or lag-1 activity. Instead, later somatic reproductive system defects suggest that proper transcriptional regulation of lag-1 confers robustness to somatic reproductive system development.

scholarly journals The C. elegans Notch proteins LIN-12 and GLP-1 are tuned to lower force thresholds for activation than Drosophila Notch

2021 ◽  
Author(s):  
Paul D. Langridge ◽  
Jessica Yu Chan ◽  
Alejandro Garcia-Diaz ◽  
Iva Greenwald ◽  
Gary Struhl
Keyword(s):  
Signal Transduction ◽  
Cell Fate ◽  
Cleavage Site ◽  
Regulatory Region ◽  
Force Level ◽  
Cell Fate Decisions ◽  
C Elegans ◽  
Protease Cleavage ◽  
Cytosolic Domains ◽  
Pulling Force

AbstractThe conserved transmembrane receptor Notch mediates cell fate decisions in all animals. In the absence of ligand, a Negative Regulatory Region (NRR) in the Notch ectodomain adopts an autoinhibited confirmation, masking an ADAM protease cleavage site [1, 2]; ligand binding makes the cleavage site accessible, leading to shedding of the Notch ectodomain as the first step of signal transduction [3, 4]. In Drosophila and vertebrates, the ligands are all single-pass transmembrane Delta/Serrate/LAG-2 (DSL) proteins; the endocytic adaptor Epsin binds to the ubiquitinated intracellular domain, and the resulting Clathrin-mediated endocytosis exerts a “pulling force” that exposes the cleavage site in the NRR [4–6]. However, in C. elegans, the presence of natural secreted DSL proteins [7] and other observations suggested that Epsin-mediated endocytosis may not be required to activate the Notch proteins LIN-12 and GLP-1. Here, we confirm that neither Epsin nor the cytosolic domains of DSL proteins are required for Notch signaling in C. elegans. Furthermore, we provide evidence that the NRRs of LIN-12 and GLP-1 are tuned to a lower force level than the NRR of Drosophila Notch. Finally, we show that adding a Leucine “plug” that occludes the cleavage site in vertebrate and Drosophila Notch proteins but is absent in the C. elegans Notch proteins [1, 2] renders the LIN-12 and GLP-1 NRRs dependent on Epsin-mediated ligand endocytosis, indicating that greater force is now required to expose the cleavage site. Thus, the NRRs of LIN-12 and GLP-1 appear to be tuned to a lower force threshold, accounting for the different requirements for signaling in C. elegans.

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