Best part- mouse Notch 1 core

Our favorite part for this year’s iGEM competition is mouse Notch 1 receptor’s core domain.

Notch core consists of the negatively regulated region and transmembrane region of the mouse Notch 1 receptor. It is the joint that empowers SynNotch the high modularity and proper activation. From the N terminal to C terminal, it is composed of three lin12-repeats (LNR) domains LNR-A, LNR-B, LNR-C, the heterodimerization domain (HD), and the transmembrane domain. The LNR-AB linker between LNR-A and B is like a plug that occludes ADAM from cleavts wild type extracellular domain can be substituted by antiGFP scfvs like LaG16 and anti-CD19, while its intracellular domain can be replaced by specialized transcription factors. Notch core is also the key to the activation of both wild type Notch receptors and SynNotch.

Figure: Structure of the negative regulatory region of Notch 1 receptor, surface delivery, and optimization. a.-c. Shows structure of the Notch 1 core, with more detail on the NRR. (With structural information adapted from Gordon et al., 2009) d. Shows that mouse SynNotch with LaG16-2 scfv (single chain fragment variants) as ectodomain is expressed on the 293T cell membrane as an example via anti-myc AF488 immunofluorescence staining. Shown here the light parts. e. A conclusive diagram of our Notch optimization. We have 5 mutations that exhibit better activation performance to SynNotch with LaG17 as scfv.

It is suggested by recent research that Notch activation begin as the receptor is first "opened up" at its negatively regulatory region (NRR) by the mechanical force exerted by Notch-bound ligand endocytosis on signal-sender cell, exposing its cleavage sites to proteases. Intracellular proteolytic action releases the Notch intracellular domain (NICD). In the majority of cellular interactions, the free Notch intracellular domain then translocate into the cell nucleus via nuclear localization sequence to regulate downstream signaling. For wild type Notch, the Notch intracellular domain would interacted with its major downstream effector CBF-1/Suppressor of Hairless/Lag-1)(CSL) on their target DNA. Together they recruit co-factors to activate endogenous downstream transcription. For SynNotch, specialized transcription factors will exclusively participate in regulation of genetic circuits that allow user-defined cellular responses.