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Identification of druggable regulators of cell secretion via a kinome-wide screen and high-throughput immunomagnetic cell sorting

Nature Biomedical Engineering volume 8, pages 263–277 (2024)Cite this article

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Abstract

The identification of genetic regulators of cell secretions is challenging because it requires the sorting of a large number of cells according to their secretion patterns. Here we report the development and applicability of a high-throughput microfluidic method for the analysis of the secretion levels of large populations of immune cells. The method is linked with a kinome-wide loss-of-function CRISPR screen, immunomagnetically sorting the cells according to their secretion levels, and the sequencing of their genomes to identify key genetic modifiers of cell secretion. We used the method, which we validated against flow cytometry for cytokines secreted from primary mouse CD4+ (cluster of differentiation 4-positive) T cells, to discover a subgroup of highly co-expressed kinase-coding genes that regulate interferon-gamma secretion by these cells. We validated the function of the kinases identified using RNA interference, CRISPR knockouts and kinase inhibitors and confirmed the druggability of selected kinases via the administration of a kinase inhibitor in an animal model of colitis. The technique may facilitate the discovery of regulatory mechanisms for immune-cell activation and of therapeutic targets for autoimmune diseases.

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Fig. 1: Identification of the druggable modulators of cell secretion via a high-throughput single-cell secretion assay.
Fig. 2: Validation of SECRE.
Fig. 3: SECRE combined with a CRISPR screen identifies regulators of IFNγ secretion.
Fig. 4: Validation of the identified hits.
Fig. 5: Validation of IFNγ inhibition in a DSS-induced colitis mouse model.

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Data availability

The sgRNA-sequencing data are available from figshare via the identifier https://doi.org/10.6084/m9.figshare.24151401. The raw and analysed datasets generated during the study are available for research purposes from the corresponding author on reasonable request. Source data are provided with this paper.

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Acknowledgements

The research reported in this publication was supported by the Canadian Institutes of Health Research (grant number FDN-148415), the Natural Sciences and Engineering Research Council of Canada (grant number 2016-06090), the Province of Ontario through the Ministry of Research, Innovation and Science (grant number RE05-009) and the National Cancer Institute of the National Institutes of Health (grant number 1R33CA204574). The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health or the other funding agencies.

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Author notes

  1. These authors contributed equally: Mahmoud Labib, Zongjie Wang.

Authors and Affiliations

  1. Peninsula Medical School, Faculty of Health, University of Plymouth, Plymouth, UK

    Mahmoud Labib

  2. Department of Chemistry, Northwestern University, Evanston, IL, USA

    Mahmoud Labib, Hanie Yousefi & Shana O. Kelley

  3. Department of Pharmaceutical Sciences, University of Toronto, Toronto, Ontario, Canada

    Mahmoud Labib, Yunhye Kim, Sichun Lin, Stéphane Angers & Shana O. Kelley

  4. Department of Biomedical Engineering, McCormick School of Engineering, Northwestern University, Evanston, IL, USA

    Zongjie Wang, Pei-Ying Lo, Edward H. Sargent & Shana O. Kelley

  5. Robert H. Laurie Comprehensive Cancer Center, Northwestern University, Chicago, IL, USA

    Abdalla Abdrabou & Shana O. Kelley

  6. Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada

    Stéphane Angers & Shana O. Kelley

  7. Donnelly Centre for Cellular and Biomolecular Research, Toronto, Ontario, Canada

    Stéphane Angers & Shana O. Kelley

  8. Department of Electrical & Computer Engineering, University of Toronto, Toronto, Ontario, Canada

    Edward H. Sargent

  9. Institute for Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada

    Shana O. Kelley

  10. Chan Zuckerberg Biohub Chicago, Chicago, IL, USA

    Shana O. Kelley

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Contributions

M.L., S.A., E.H.S. and S.O.K. conceived and designed the experiments; M.L., Z.W., Y.K., S.L. and H.Y. performed the experiments and analysed the data. A.A. performed the CRISPR KO experiments. P-Y.L. cultured and maintained the cells. M.L., Z.W., S.A., E.H.S. and S.O.K. contributed to the preparation and editing of the manuscript.

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Correspondence to Shana O. Kelley.

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Labib, M., Wang, Z., Kim, Y. et al. Identification of druggable regulators of cell secretion via a kinome-wide screen and high-throughput immunomagnetic cell sorting. Nat. Biomed. Eng 8, 263–277 (2024). https://doi.org/10.1038/s41551-023-01135-w

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