3069 – GRANULIN, REVEALED BY SINGLE-CELL RNA-SEQUENCING OF DEVELOPING HEMATOPOIETIC CELLS, DRIVES MACROPHAGE AND NEUTROPHIL DIFFERENTIATION

Abstract

Myeloid cell differentiation leads to the formation of neutrophils and macrophages, which are critical cells for host defenses against infections. In addition, a multitude of other functions beyond their classical inflammatory roles have been assigned to these cells, including wound healing, neurodegenerative diseases and tumorigenesis. Thus, the identification of novel genes that regulate myeloid cell development has undoubtedly the potential to impact diverse areas of human health. Here, we have used deep single-cell RNA-sequencing during the active phase of myelopoiesis in the zebrafish embryo to create a platform for the identification of novel genes required for myeloid cell formation. Using this platform, we have uncovered a role for granulin (GRN) during neutrophil and macrophage differentiation. Leveraging the restricted expression of the zebrafish orthologue grna to the myeloid cell lineage, enabled us to perform loss of function experiments without altering its function in non-hematopoietic cells. With the generation of a zebrafish model of grna deficiency, we reveal that myeloid progenitors are unable to differentiate into neutrophils and macrophages. These grna deficient progenitors are also incapable of triggering a myelopoiesis emergency response, resulting in abnormal wound healing with aberrant collagen depositions. Utilizing CUT&RUN, we identified that Pu.1 directly binds grna enhancers, triggering its expression. Similar to the zebrafish grna, the mammalian granulin is also upregulated in myeloid cells, and controlled by the myeloid transcription factors PU.1 and IRF8. Altogether, our findings uncover the role of Granulin during myeloid cell differentiation, opening a new field of study that will help elucidate the pleiotropic role of this enigmatic protein in inflammation, wound healing, tumor progression and neurodegenerative disease.