Molecular Integration of HoxB4 and STAT3 for Self‐Renewal of Hematopoietic Stem Cells: A Model of Molecular Convergence for Stemness

Abstract

The upregulation of HoxB4 promotes self-renewal of hematopoietic stem cells (HSCs) without overriding the normal stem cell pool size. A similar enhancement of HSC self-renewal occurs when signal transducer and activator of transcription 3 (STAT3) is activated in HSCs. In this study, to gain insight into the functional organization of individual transcription factors (TFs) that have similar effects on HSCs, we investigated the molecular interplay between HoxB4 and STAT3 in the regulation of HSC self-renewal. We found that while STAT3-C or HoxB4 similarly enhanced the in vitro self-renewal and in vivo repopulating activities of HSCs, simultaneous transduction of both TFs did not have additive effects, indicating their functional redundancy in HSCs. In addition, activation of STAT3 did not cause changes in the expression levels of HoxB4. In contrast, the inhibition of STAT3 activity in HoxB4-overexpressing hematopoietic cells significantly abrogated the enhancing effects of HoxB4, and the upregulation of HoxB4 caused a ligand-independent Tyr-phosphorylation of STAT3. Microarray analysis revealed a significant overlap of the transcriptomes regulated by STAT3 and HoxB4 in undifferentiated hematopoietic cells. Moreover, a gene set enrichment analysis showed significant overlap in the candidate TFs that can recapitulate the transcriptional changes induced by HoxB4 or STAT3. Interestingly, among these common TFs were the pluripotency-related genes Oct-4 and Nanog. These results indicate that tissue-specific TFs regulating HSC self-renewal are functionally organized to play an equivalent role in transcription and provide insights into the functional convergence of multiple entries of TFs toward a conserved transcription program for the stem cell state. STEM CELLS 2014;32:1313-1322

Functional compensation between hematopoietic stem cell clones in vivo

“…Genes that are known to promote HSC self-renewal, including Hoxb3 [42,43], Hoxb6 [44], and Gfi-1 [41], were downregulated ( Fig 5B). Genes that are known to promote HSC self-renewal, including Hoxb3 [42,43], Hoxb6 [44], and Gfi-1 [41], were downregulated ( Fig 5B).…”

“…We also found molecular evidence of reduced HSC self-renewal in the NSG co-transplantation group. Genes that are known to promote HSC self-renewal, including Hoxb3 [42,43], Hoxb6 [44], and Gfi-1 [41], were downregulated ( Fig 5B). Genes involved in differentiation such as Pax5 [33] and Ebf1 [45], and in development such as Vpreb2 [46] and Zap70 [47], were upregulated ( Fig 5B).…”

Microenvironmental Remodeling as a Parameter and Prognostic Factor of Heterogeneous Leukemogenesis in Acute Myelogenous Leukemia

“…MSC gene expression was analyzed by Illumina BeadChip (Illumina) array hybridization analysis, using the Linear Models for Microarray Data (LIMMA) method and the R-package for Statistics as described previously (37). For expression profiles, the median absolute deviation was calculated, and highly variable genes were selected for hierarchical clustering with average linkage.…”

Regulation of mesenchymal stromal cells through fine tuning of canonical Wnt signaling

“…MSCs were transduced with each retroviral vector with multiplicity of infection (MOI) of 30, then sorted for transduced (GFP + ) cells as described (Hong et al, 2014).…”

“…A gene expression study using microarrays was performed as previously described (Hong et al, 2014). Briefly, biotinlabeled cRNA samples were prepared using the Illumina TotalPrep RNA Amplification kit (Ambion, Austin, TX) and hybridized to a version 2 of the Illumina Mouse-6 BeadChip (48 K) according to the manufacturer's instructions (Illumina, San Diego, CA).…”