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Development of a CRISPR-Cas9 KO Screening Platform to Study MHC-II Antigen Presentation in Dendritic Cells 2282

Year: 2025
Type: article
Abstract: Description Major histocompatibility complex class II (MHC-II) molecules are crucial for immune response initiation and regulation through peptide presentation to CD4+ T cells. While MHC-II expression is predominantly restricted to professional antigen-presenting cells, particularly dendritic cells (DCs), the molecular machinery governing antigen processing remains partially undefined. Understanding the molecular components involved in antigen uptake, trafficking, processing, and presentation is essential for therapeutic manipulation of MHC-II presentation in cancer and autoimmune disorders. We developed an in vitro CRISPR KO screening platform using the murine MutuDC1940 cells and the soluble Ea antigen to identify novel regulators of this pathway. Cells were transduced with a custom CRISPR KO library targeting genes involved in phagocytosis, autophagy and endocytosis. Antigen presentation was quantified by flow cytometry using the Y-Ae antibody, recognizing the pMHC-II/Ea complex. The inclusion of established MHC-II regulators as positive controls validated our screening approach. This systematic analysis has the potential to provide new insights into the machinery required for MHC-II antigen processing in DCs. This approach will be complemented using diverse exogenous antigenic sources and a whole-genome CRISPR KO library for unbiased screening. These preliminary findings warrant further validation and could reveal promising therapeutic targets for optimizing DC-based immunotherapies. Funding Sources Supported by the American Lebanese Syrian Associated Charities (ALSAC), St. Jude Children’s Research Hospital. Topic Categories Antigen and Dendritic Cell Processing, Presentation, and Biology (AGDC)
DOI: 10.1093/jimmun/vkaf283.217
Language: English
Subfield: Immunology
Open Access status: bronze
OpenAlex ID: W4416449816
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