Diversity in Notch ligand-receptor signaling interactions
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1.
California Institute of Technology
Abstract
The Notch signaling pathway uses families of ligands and receptors to transmit signals to nearby cells. These components are expressed in diverse combinations in different cell types, interact in a many-to-many fashion, both within the same cell (in cis) and between cells (in trans), and their interactions are modulated by Fringe glycosyltransferases. A fundamental question is how the strength of Notch signaling depends on which pathway components are expressed, at what levels, and in which cells. Here, we used a quantitative, bottom-up, cell-based approach to systematically characterize trans-activation, cis-inhibition, and cis-activation signaling efficiencies across a range of ligand and Fringe expression levels in Chinese hamster and mouse cell lines. Each ligand (Dll1, Dll4, Jag1, and Jag2) and receptor variant (Notch1 and Notch2) analyzed here exhibited a unique profile of interactions, Fringe dependence, and signaling outcomes. All four ligands were able to bind receptors in cis and in trans, and all ligands trans-activated both receptors, although Jag1-Notch1 signaling was substantially weaker than other ligand-receptor combinations. Cis-interactions were predominantly inhibitory, with the exception of the Dll1- and Dll4-Notch2 pairs, which exhibited cis-activation stronger than trans-activation. Lfng strengthened Delta-mediated trans-activation and weakened Jagged-mediated trans-activation for both receptors. Finally, cis-ligands showed diverse cis-inhibition strengths, which depended on the identity of the trans-ligand as well as the receptor. The map of receptor-ligand-Fringe interaction outcomes revealed here should help guide rational perturbation and control of the Notch pathway.
Copyright and License
© 2023, Kuintzle et al. This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.
Acknowledgement
RK, LS, and MBE conceived and designed the experiments RK and LS generated the cell lines and performed the experiments. RK and LS analyzed the experimental data. RK, LS, and MBE wrote the paper. We thank Irwin Bernstein for generously sharing the recombinant Dll1-ext-Fc ligand, and Igor Antoshechkin in the Millard and Muriel Jacobs Genetics and Genomics Laboratory for assistance with RNA-sequencing. We are grateful to Xun Wang and the Rothenberg Lab for providing the NGFR construct. We would also like to thank Ellen Rothenberg, David Sprinzak, Stephen Blacklow, Sandy Nandagopal, James Linton, Martin Tran, Jan Gregrowicz, Ronghui Zhu, Felix Horns, and Jacob Parres-Gold for discussions about this work and for critical feedback on this manuscript. This work was supported by the National Institutes of Health (NIH) (grant R01 HD7335C). RK was supported by an NIH Ruth L Kirschstein NRSA predoctoral fellowship (F31 HD100185). MBE is a Howard Hughes Medical Institute Investigator. This article is subject to HHMI’s Open Access to Publications policy. HHMI lab heads have previously granted a nonexclusive CC BY 4.0 license to the public and a sublicensable license to HHMI in their research articles. Pursuant to those licenses, the author-accepted manuscript of this article can be made freely available under a CC BY 4.0 license immediately upon publication.
Funding
- National Institutes of Health (R01 HD7335C)
- National Institutes of Health (F31 HD100185)
- Howard Hughes Medical Institute
Data Availability
RNA sequencing data are available at NCBI via the GEO accession GSE233573. All raw and processed datasets (RNA-seq, qRT-PCR, and flow cytometry) and the custom Python scripts used to make figures and support conclusions in this paper are available at data.caltech.edu: https://doi.org/10.22002/gjjkn-wrj28.
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Additional details
- National Institutes of Health
- R01 HD7335C
- National Institutes of Health
- F31 HD100185
- Howard Hughes Medical Institute
- Caltech groups
- Division of Biology and Biological Engineering (BBE)
- Publication Status
- Published