CRISPR/Cas9-mediated gene disruption determines the roles of MITF and CITED2 in human mast cell differentiation

https://watermark.silverchair.com/bloodadvances.2023012279.pdf?token=AQECAHi208BE49Ooan9kkhW_Ercy7Dm3ZL_9Cf3qfKAc485ysgAABKUwggShBgkqhkiG9w0BBwagggSSMIIEjgIBADCCBIcGCSqGSIb3DQEHATAeBglghkgBZQMEAS4wEQQM4IVhdTD2wOJnv5CUAgEQgIIEWEgJaRIfU-L3SPO1RHttD3RI8vWvhu4edFN-AQG75rFWBRsVrkNOTy2vVQfn5ussNhMTVC-Mk8cwTuBo77rMZEhuSeelcV1LoLTF01gsYBwpw50rCzjzY9Upt8JvvUmkRTyoN_1KvqclG7E3Shd3I5DvLYgpcj059d0ETgx7v7syXUXW8MuZG_6Havx0CO2G3wfJc87M6o7cHyfYHs3sYG0giltwjz1nN2NVG-1w9KQzjJmTFiHf3LBdiJwH8_80BjJLYRDqqkyiP09g58k9UJNeEEpXLMtycuk9vbGG31KHueLOnuJA4VYNbvXpgbtMFQQGxqIrz7SBCfldsKOtAPBdn6KogT0TAJsBfiEp90CJhouAAaNbLyynf4yfjOQX3PtZZdyGQ1kjOISdPJYPrmMFDHvERkaP865rs03iZb839Sq7OXq0mkmo4YfHoM1fKXqya6bnw0oTqduUJ1-I7re7IsfOvTM5BVPmqqFi3KBdmiUnFaIZqeVLnRr8QCrp17Ed2pRf9B1t3xeKZ0LakI0Ed5aHQHfKBmEZPzK1AdZBhmxEG4Rt2jl_T3XuXDYsuvumGF2rWv3Fy0kJ4rLaAihQ_hqqdzFUOX_QE8UXmuT0ytNw222lliHEDFWNpXAThFmq8P603LToZWWUN3S0mmwDTZ86gAHnjOCIMmSonoJxHZ6N-HQTgDZanIM_kawc080HtL7FcrbZpRitpvmlDDUYLwI2KUKKID0C8_FRjFpePi-BmIjnqgY6i_ksC6DtOA5z4F7doAI4rz6i7dr4pKzK8ZcZsfvCb11B3nvBPi01M7WGehDm4v37a-YcAF_pmP7BLM1MyZ8qKNs9yoIuwzLc-XnVCQFFZUhdh-bl9GwXgocWuWPS5s9SZRxM0V5VWISMqdIhUGV-0PN_h28Lq2DRRo2XmNeH6M6OSF0BLHVH_k2mQ7PcRi4DktcO1AsoD_xSfgGvVfe09OaWqYe5rM1hQ9Ec7CpbK0WE8IBTz9MSVn25Ex5Lpe6N2Wc1r1eGT7Om4EpLDFlEaoGJm9Knd_eb4OcbTgqPHtyGy7c4JOMfGlSI4EeOthDSfnBXtcCWRdqssN-LDhs63i9hiRhiZYaaCSRNLdZLXL5cyNO3z4PFsvPj8hZmDeii2jtFhz5nIpmI47mXwL2LjMVm036ZXMrTHl5GrJyzrLQGDOMW8AR_7Defkci_rhnRm9lEa75FB_zOlzG0XQUya03adIAb3WJm2pXUNAfG8cDrlCTlrlwzMZEkL8kHd5dbTG9BLzqFisFghrhqdR3L7GtB_4uXEow1vSJJBdJRQZUiCT5qYSn1Pi5wNAEi9O6LV56nkWnU9OEJWnVv-aXVP2XGD7JCDhw1S0R_ySft7de2N0Uq2k-iPKK741cJ4Zot4pEp4TicvhXqmrCnDkzcysnCuXInvKvg5GRCjdDzSj9QGxALklkIMGGYKTzhgCk9bqJqZ3BzUrKEjBPHMpzZ

American Society of Hematology

Jiezhen Mo (Karolinska Institutet and Karolinska University Hospital, Sweden) Fredrik Wermeling (Karolinska Institutet and Karolinska University Hospital, Stockholm, Sweden, Sweden) Gunnar Nilsson (Department of Medical Sciences, Uppsala University, Uppsala, Sweden, Sweden) Joakim Dahlin (Karolinska Institutet and Karolinska University Hospital, Sweden)

Abstract:

The differentiation of hematopoietic stem and progenitor cells into the various cell lineages is regulated by cell-intrinsic factors and the progenitors’ microenvironment. Experiments in mice have allowed the identification of transcriptional modulators that control mast cell differentiation. However, a comprehensive approach that allows efficient disruption of individual transcriptional modulators in primary human hematopoietic progenitors coupled with a mast cell formation readout has not been described. Here, we report a simple electroporation- and ribonucleoprotein-based knockout system that allows the identification of genes that are required and dispensable for human mast cell differentiation. We show that the transcription factor MITF is upregulated in human mast cell progenitors and reveal that the loss of MITF results in the suppressed formation of mast cells. By contrast, CITED2, another transcriptional modulator that is upregulated along the mast cell differentiation trajectory, was dispensable for human mast cell differentiation. Taken together, we report a CRISPR/Cas9-based framework that serves to identify genes involved in regulating the formation of human mast cells, and the results uncover the role of two transcriptional modulators in controlling human mast cell differentiation.

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