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Efficient CRISPR-Cas9-mediated mutagenesis in primary human B cells for identifying plasma cell regulators

Van Duc (57608893100) | Thomas (7005054399); Dang | Dania (57219669445); Rajewsky Faculty of Biology, VNU University of Science, Vietnam National University, Hanoi, Viet Nam| Eva (55370246500); Kressler Berlin Institute of Health (BIH), Berlin, Germany|

Molecular Therapy - Nucleic Acids Số , năm 2022 (Tập 30, trang 621-632)

ISSN: 21622531

ISSN: 21622531

DOI: 10.1016/j.omtn.2022.11.016

Tài liệu thuộc danh mục:

Article

English

Tóm tắt tiếng anh
Human B lymphocytes are attractive targets for immunotherapies in autoantibody-mediated diseases. Gene editing technologies could provide a powerful tool to determine gene regulatory networks regulating B cell differentiation into plasma cells, and identify novel therapeutic targets for prevention and treatment of autoimmune disorders. Here, we describe a new approach that uses CRISPR-Cas9 technology to target genes in primary human B cells in vitro for identifying plasma cell regulators. We found that sgRNA and Cas9 components can be efficiently delivered into primary human B cells through RD114-pseudotyped retroviral vectors. Using this system, we achieved approximately 80% of gene knockout efficiency. We disrupted expression of a triad of transcription factors, IRF4, PRDM1, and XBP1, and showed that human B cell survival and plasma cell differentiation are severely impaired. Specifically, that IRF4, PRDM1, and XBP1 were expressed at different stages during plasma cell differentiation, IRF4, PRDM1, and XBP1-targeted B cells failed to progress to the pre-plasmablast, plasma cell state, and plasma cell survival, respectively. Our method opens a new avenue to study gene functions in primary human B cells and identify novel plasma cell regulators for therapeutic applications. � 2022 The Author(s)

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