Abstract
BackgroundAdditional sex combs-like 1 (ASXL1) is frequently mutated in myeloid malignancies. Recent studies showed that hematopoietic-specific deletion of Asxl1 or overexpression of mutant ASXL1 resulted in myelodysplasia-like disease in mice. However, actual effects of a “physiological” dose of mutant ASXL1 remain unexplored.MethodsWe established a knock-in mouse model bearing the most frequent Asxl1 mutation and studied its pathophysiological effects on mouse hematopoietic system.ResultsHeterozygotes (Asxl1tm/+) marrow cells had higher in vitro proliferation capacities as shown by more colonies in cobblestone-area forming assays and by serial re-plating assays. On the other hand, donor hematopoietic cells from Asxl1tm/+ mice declined faster in recipients during transplantation assays, suggesting compromised long-term in vivo repopulation abilities. There were no obvious blood diseases in mutant mice throughout their life-span, indicating Asxl1 mutation alone was not sufficient for leukemogenesis. However, this mutation facilitated engraftment of bone marrow cell overexpressing MN1. Analyses of global gene expression profiles of ASXL1-mutated versus wild-type human leukemia cells as well as heterozygote versus wild-type mouse marrow precursor cells, with or without MN1 overexpression, highlighted the association of in vivo Asxl1 mutation to the expression of hypoxia, multipotent progenitors, hematopoietic stem cells, KRAS, and MEK gene sets. ChIP-Seq analysis revealed global patterns of Asxl1 mutation-modulated H3K27 tri-methylation in hematopoietic precursors.ConclusionsWe proposed the first Asxl1 mutation knock-in mouse model and showed mutated Asxl1 lowered the threshold of MN1-driven engraftment and exhibited distinct biological functions on physiological and malignant hematopoiesis, although it was insufficient to lead to blood malignancies.
Highlights
Additional sex combs-like 1 (ASXL1) is frequently mutated in myeloid malignancies
An additional guanine was inserted into this 8-G cassette located at mouse Asxl1 exon 13 to mimic this frequent human ASXL1 mutation (Fig. 1a)
This insertion causes frame shift in the reading frame and introduces a premature stop codon so that the mutant Asxl1 would be shorter than wild-type form and lack the c-terminal region which contains a plant homeodomain (PHD)
Summary
Additional sex combs-like 1 (ASXL1) is frequently mutated in myeloid malignancies. Since the discovery of ASXL1 mutation in myeloid malignancies in 2009 [15], many studies about its pathophysiology have been reported. In vivo deletion of Asxl was shown to result in subtle phenotypes including defects in the frequencies of myeloid and lymphoid cells in blood, marrow or other hematopoietic organs in mice but not myelodysplastic syndrome (MDS) or leukemia [16]. In other studies, knockout of Asxl led to systemic developmental defects including MDS-like presentation, with alteration of the self-renewal and repopulation capacities of the mutant hematopoietic stem/progenitor cells and global reduction of H3K27 tri-methylation (H3K27me3) [17, 18]
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