Hematopoietic homeostasis is tightly regulated by controlling the balance between quiescence, self-renewal, and lineage-commitment of hematopoietic stem cells (HSCs). Deregulation of these processes and aberrant acquisition of stem cell-like properties is believed to be central to the pathogenesis of hematologic malignancies, such as myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML). However, little is known about the molecular networks maintaining the stem cell state and the epigenetic and post-transcriptional regulation of determinants that control these programs. MicroRNAs (miRNAs) represent a large class of post-transcriptional regulators that mediate repression of multiple target mRNAs. We have previously shown that miR-126 and miR-125a are differentially expressed across the human hematopoietic hierarchy and function to control self-renewal and cell fate decisions by reinforcing gene expression programs in a developmental stage-specific manner (Lechman et al. Cell Stem Cell, 2012; Wojtowitz et al. Cell Stem Cell, 2016).
To identify additional miRNA(s) that play a functional role in hematopoiesis, we performed an in vivo competitive repopulation screen in which candidate miRNAs were overexpressed (OE) in human CD34+CD38- umbilical cord blood (CB) cells and subsequently transplanted into immune-deficient mice for 24 weeks. miR-130a was shown to enhance long-term hematopoietic reconstitution and chosen for further investigation. At 12 and 24 weeks after transplantation, enforced miR-130a expression (including an mOrange-mO+ indicator) conferred a competitive advantage over untransduced CB cells demonstrated by increased CD45+ human chimerism in the injected femur (IF), bone marrow (BM), and spleen of recipient mice. miR-130 enforced expression (miR-130a OE) increased the proportion of mO+/hCD45+ cells by approximately 2- and 5-fold after 12 and 24 weeks of repopulation, respectively. miR-130a OE xenografts showed multilineage engraftment with increased myeloid lineage output and significantly enhanced erythroid development at the expense of B-lymphoid lineage output in BM and spleen of recipient mice. Detailed flow cytometry analysis of xenografts revealed accumulation of immature GlyA+/CD71+/CD36+ erythroid progenitors, suggesting a differentiation block at the polychromic erythroblast stage. Notably, miR-130a OE induced the expansion of CD34+CD38- Lin- compartment and increased proportion of CD34+CD38-CD90+CD45RA- immuno-phenotypic HSC. Secondary transplantation involving limiting dilution analysis revealed approximately a 10-fold increase in HSC frequency, consistent with a role of miR-130a in HSC self-renewal. The lineage potential of miR-130OE primitive cells was assessed in vitro using single-cell stromal-based myelo-erythroid differentiation assay. Enforced expression of miR-130a in human HSC and multipotent progenitors (MPP) resulted in the decreased frequency of unipotent myeloid output (M colonies) and increased multipotent output (M/E/Meg, E/Meg colonies), supporting a role of miR-130a in erythroid-megakaryocytic fate specification. Label-free semiquantitative proteomics and subsequent gene set enrichment pathway analysis (GSEA) were performed on miR-130a OE and control transduced CD34+ CB cells to elucidate molecular mechanism(s) of miR-130a function. We identified that miR-130a modulated pathways centered on translational regulation and chromatin modification. Together, our data suggest that miR-130a plays a role in the regulation of the HSC self-renewal and erythroid differentiation. Given that several studies showed aberrant expression of miR-130a in MDS and some AML subtypes, it is important to delineate the role of miR-130a in normal hematopoiesis to comprehend its potential contribution to the development of hematologic malignancies.
This abstract is also being presented as Poster 40.
Citation Format: Gabriela Krivdova, Eric R. Lechman, Erwin M. Schoof, Veronique Voisin, Olga I. Gan, Aaron Trotman-Grant, Karin G. Hermans, Gary D. Bader, John E. Dick. MicroRNA-130a regulates hematopoietic stem cell self-renewal and erythroid differentiation [abstract]. In: Proceedings of the Second AACR Conference on Hematologic Malignancies: Translating Discoveries to Novel Therapies; May 6-9, 2017; Boston, MA. Philadelphia (PA): AACR; Clin Cancer Res 2017;23(24_Suppl):Abstract nr PR07.