Mechanisms of granulocyte homeostasis

PROJECT SUMMARY/ABSTRACT A fundamental challenge in developmental biology is delineating hierarchical cellular states including rare intermediates and their underlying gene regulatory networks that mediate cell-type specification. Previously, we utilized scRNA-Seq, developed novel bioinformatics approaches and clonogenic assays to delineate hierarchical genomic and regulatory states culminating in neutrophil or macrophage specification. Our analysis captured prevalent mixed-lineage intermediates (MultiLin*) that manifested coincident expression of hematopoietic stem cell/progenitor (HSPC) and myeloid progenitor genes. Here, we used primary mouse bone marrow progenitors to: 1) rigorously identify stable developmental cell states and rare intermediates, 2) overcome major informatics hurdles to integrate multiomic data sets with flow cytometry to validate hypotheses, and 3) link neural-network-identified TF activity on chromatin to dynamic gene expression; predicting hematopoietic TF cistromes. Our Preliminary Data reveals nascent transcription factor programming within the MultiLin that restricts lineage potentials, yet precurses bipotential progenitors and lineage specification and commitment. We hypothesize that within the homeostatic hematopoietic system, successive myeloid lineage restriction events begin within MultiLin. The proposed work will isolate cluster-defined cell populations and define their developmental potentials, establish a validated network to establish the combinatorial regulatory TFs that program hemopoietic progenitors, then exploit this network to define the genomic and cellular impact of emergency stress adaptations which stimulate erythroid and neutrophil granulocyte production.