Exbio — About exbio — Blog — Notch at a glance – part two

Notch at a glance – part two

DLL4 (Delta-like 4) is one of five Notch receptor ligands (agonistic DLL1, 4, Jagged 1, 2, and antagonistic DLL3). DLL4 is up-regulated at sites of physiological and pathological angiogenesis, whereas its expression is low in most adult normal tissues. It is also highly expressed in human clear-cell renal carcinomas, bladder cancers, and breast cancers. In arterial endothelial cells, DLL4 is one of the earliest expressed antigens, and functions as a regulator to prevent angiogenic sprouting. Blocking of its interaction with Notch receptors results in forming defective vessels, which has, however, benefits in case of tumor therapy, as it reduces tumor growth. This blocking can be performed even by soluble extracellular domain of DLL4. In muscles, DLL4 is upregulated and released as a response to muscle atrophy. It is also an active player in insulin secretion by pancreatic β-cells. Concerning T-cell lymphopoiesis, Notch activation by DLL1 and DLL4 differs in effects on induction of T-lineage differentiation steps, and DLL4 plays the more important role here. DLL4 can be used also for culturing of T cells from pluripotent stem cells.

CD156c (ADAM10) is involved in the Notch system by cleavage Notch extracellular domain upon DLL or Jagged ligand binding, which is a necessary step to enable downstream signaling. This type I transmembrane glycoprotein with a zinc-dependent metalloprotease activity serves as an endopeptidase of broad specificity, which is expressed mainly in thymus, liver, and muscles. Its expression can be induced in inflamed central nervous system, and in arthritic tissues. CD156c is involved in multiple sclerosis-associated myelin degradation. It also solubilizes various membrane proteins, including CD23, CD44, CD126, CD171, ephrin-A2, and other. Based on calcium channel-type sensors CD156c reacts to frictional force from fluid flow and transduces this information to the Notch system. Inhibition of CD156c enzymatic activity is one of the ways how to target Notch signaling.

Fig. 1: Schematic drawing describing the signal transduction process of the Notch system.

The mouse monoclonal antibody MHD4-46 recognizes extracellular part of human DLL4, and can be used for flow cytometry, and functional applications (blocking). Formats: Purified (11-717-C100), purified low endotoxin (12-717-C100), PE (1P-717-C100), and APC (1A-717-C100).

Fig. 2: Separation of DLL4-transfected CHO cells stained using anti-DLL4 (MHD4-46) APC antibody (concentration in sample 5 μg/ml, red-filled) from DLL4-transfected CHO cells stained using mouse IgG1 isotype control (MOPC-21) APC antibody (concentration in sample 5 μg/ml, same as DLL4 APC antibody concentration, black-dashed) in flow cytometry analysis (surface staining).

The mouse monoclonal antibody 11G2 recognizes an extracellular/luminal epitope of human CD156c, and can be used for flow cytometry, Western blotting, immunoprecipitation, and immunohistochemistry. Formats: Purified (11-938-C100), PE (1P-938-T100), and APC (1A-938-T100).

Fig. 3: Separation of human monocytes (red-filled) from CD156c negative blood debris (black-dashed) in flow cytometry analysis (surface staining) of human peripheral whole blood stained using anti-human CD156c (11G2) PE antibody (10 μl reagent / 100 μl of peripheral whole blood).

Further reading:

Lobov IB et al.: Delta-like ligand 4 (DLL4) is induced by VEGF as a negative regulator of angiogenic sprouting. PNAS 2007, 104(9): 3219-3224.

Scehnet JS et al.: Inhibition of DLL4-mediated signaling induces proliferation of immature vessels and results in poor tissue perfusion. Blood 2007, 109(11): 4753-4760.

Mohtashami M et al.: Direct comparison of DLL1- and DLL4-mediated Notch activation levels shows differential lymphomyeloid lineage commitment outcomes. J. Immunol. 2010, 185: 867-876.

Brandstadter JD and Maillard I: Notch signaling in T cell homeostasis and differentiation. Open Biol. 9: 190187.

Rubey M et al.: DLL1- and DLL4-mediated Notch signaling is essential for adult pancreatic islet homeostasis. Diabetes 2020, 69: 915-926.

Fujimaki S et al.: The endothelial DLL4-muscular Notch2 axis regulates skeletal muscle mass. Nat. Metabol. 2022, 4:180-189.

Michaels YS et al.: DLL4 and VCAM1 enhance the emergence of T cell-competent hematopoietic progenitors from human pluripotent stem cells. Sci. Adv. 2022, 8:5522.

Tian L et al.: ADAM10 is essential for proteolytic activation of Notch during thymocyte development. Int. Immunol. 2008, 9:1181-1187.

van Tetering G et al.: Metalloprotease ADAM10 is required for Notch1 site 2 cleavage. J. Biol. Chem. 2009, 284(45): 31018-31027.

Caolo V et al.: Shear stress activates ADAM10 sheddase to regulate Notch1 via the Piezo1 force sensor in endothelial cells. eLife 2020, 9:e50684.