Title: Integrin α4-dependent Interaction with the Niche During Development Programs Hematopoietic Stem and Progenitor Cells
Abstract: Hematopoietic stem and progenitor cells (HSPCs) reside in a microenvironment that regulates their behavior by interactions with niche support cells. In the mammalian embryo fetal liver HSPCs proliferate and when transplanted have a greater capacity for reconstituting the blood system compared to quiescent adult HSPCs. However, the genetic networks regulating fetal liver HSPCs are poorly understood. To dissect these networks, we are characterizing a viable integrin α4 (itga4) mutant zebrafish model with perturbed interaction between HSPCs and the caudal hematopoietic tissue (CHT), the equivalent niche to mammalian fetal liver. To define niche reprogramming mechanisms, we sorted WT and itga4 mutant Runx:mCherry+ HSPCs via FACS after CHT stage at 5 days post fertilization (5 dpf). Bulk RNA-seq detected 286 up- and 49 down-regulated transcripts in itga4 mutant versus WT (q<0.05) and GSEA revealed upregulated inflammatory pathways in itga4 mutant HSPCs. Bulk ATAC-seq discovered 84,236 peaks unique to WT HSPCs and 16,875 peaks unique to itga4 mutant (q<0.05) with motif analysis identifying potential regulatory factors of HSPC reprogramming, such as ETS and AP-1 factors. To understand the loss of itga4 function through development and into adulthood, we compared whole kidney marrow (WKM) from young (6-month-old) itga4 mutant and WT adults, and aged (24-month-old) WT adults via split-pool barcoding scRNA-seq. Factors increased in aged WT WKM, such as cebpa, il1b, and nfkbiab, were also increased in young itga4 mutant WKM. Separation of HSPC lineages by forward and side scatter via flow cytometry revealed enriched myeloid bias of young itga4 mutant Runx:mCherry+ HSPCs versus young WT (4.46% vs 1.63%, p=0.0432). The percentage of mitotic HSPCs was increased in young itga4 mutant versus young WT WKM (47.42% vs 3.83%, p<0.001). We will utilize this system and multiomic approaches to further dissect the developmental networks of HSPC reprogramming.
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