In vivo genetic screen in xenotransplanted cord blood-derived macrophages to identify regulators of human border-associated macrophage identities

Border-associated macrophages (BAMs) are macrophages that reside in specific anatomical interfaces between the brain and the blood circulation. Due to limited access to fresh human tissues from those anatomical sites, our understanding of human BAM development and function remains poor. It was previously observed that human hematopoietic stem cells from umbilical cord blood take up residence and differentiate into BAM-like macrophages when injected into the brains of neonatal mice expressing human colony-stimulating factor 1 (hCSF1). Single-cell RNA-sequencing confirmed strikingly similar transcriptomic signatures of those human macrophages to BAMs. The resulting chimeric mouse model represents an innovative platform in which the development and function of human BAMs could be investigated. In the current study, brain-xenotransplanted human cord blood-derived macrophages will be further characterized to assess whether they show location-specific molecular signatures, as do human BAMs. Then, an in vivo CRISPR knockout screen will be performed on those cells in order to decipher the genetic regulators of human BAM identities. I, the applicant, have extensive experience in brain immunology and in working with macrophages. I will work on developing and applying an in vivo CRISPR screen workflow in the chimeric mouse model developed by the laboratory of Dr. Movahedi (VUB, Belgium) who has established himself as a leader in brain macrophage research. This project will offer me the invaluable opportunity to learn genome editing techniques, design and execution of mouse experiments, as well as single-cell -omic experiments using state-of-the-art instrument. By bridging my existing knowledge with the cutting-edge techniques offered at the host laboratory, I aim to lay the foundations for an innovative system to study human BAMs.