Stem cells are highly resistant to viral infection compared to their differentiated progeny; however, the mechanism is mysterious. Here, we analyzed gene expression in mammalian stem cells and cells at various stages of differentiation. We find that, conserved across species, stem cells express a subset of genes previously classified as interferon (IFN) stimulated genes (ISGs) but that expression is intrinsic, as stem cells are refractory to interferon. This intrinsic ISG expression varies in a cell-type-specific manner, and many ISGs decrease upon differentiation, at which time cells become IFN responsive, allowing induction of a broad spectrum of ISGs by IFN signaling. Importantly, we show that intrinsically expressed ISGs protect stem cells against viral infection. We demonstrate the in vivo importance of intrinsic ISG expression for protecting stem cells and their differentiation potential during viral infection. These findings have intriguing implications for understanding stem cell biology and the evolution of pathogen resistance.

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The erythroblastic island (EBI), composed of a central macrophage and surrounding erythroid cells, was the first hematopoietic niche discovered. The identity of EBI macrophages has thus far remained elusive. Given that Epo is essential for erythropoiesis and that Epor is expressed in numerous nonerythroid cells, we hypothesized that EBI macrophages express Epor so that Epo can act on both erythroid cells and EBI macrophages simultaneously to ensure efficient erythropoiesis. To test this notion, we used Epor-eGFPcre knockin mouse model. We show that in bone marrow (BM) and fetal liver, a subset of macrophages express Epor-eGFP. Imaging flow cytometry analyses revealed that >90% of native EBIs comprised F4/80+Epor-eGFP+ macrophages. Human fetal liver EBIs also comprised EPOR+ macrophages. Gene expression profiles of BM F4/80+Epor-eGFP+ macrophages suggest a specialized function in supporting erythropoiesis. Molecules known to be important for EBI macrophage function such as Vcam1, CD169, Mertk, and Dnase2α were highly expressed in F4/80+Epor-eGFP+ macrophages compared with F4/80+Epor-eGFP− macrophages. Key molecules involved in iron recycling were also highly expressed in BM F4/80+Epor-eGFP+ macrophages, suggesting that EBI macrophages may provide an iron source for erythropoiesis within this niche. Thus, we have characterized EBI macrophages in mouse and man. Our findings provide important resources for future studies of EBI macrophage function during normal as well as disordered erythropoiesis in hematologic diseases such as thalassemia, polycythemia vera, and myelodysplastic syndromes.

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The ten-eleven translocation (TET) family of proteins plays important roles in a wide range of biological processes by oxidizing 5-methylcytosine (5mC) to 5-hydroxy-methylcytosine. However, their function in erythropoiesis has remained unclear. We show here that TET2 and TET3 but not TET1 are expressed in human erythroid cells, and we explore the role of these proteins in erythropoiesis. Knockdown experiments revealed that TET2 and TET3 have different functions. Suppression of TET3 expression in human CD34 cells markedly impaired terminal erythroid differentiation, as reflected by increased apoptosis, the generation of bi/multinucleated polychromatic/orthochromatic erythroblasts, and impaired enucleation, although without effect on erythroid progenitors. In marked contrast, TET2 knockdown led to hyper-proliferation and impaired differentiation of erythroid progenitors. Surprisingly, knockdown of neither TET2 nor TET3 affected global levels of 5mC. Thus, our findings have identified distinct roles for TET2 and TET3 in human erythropoiesis, and provide new insights into their role in regulating human erythroid differentiation at distinct stages of development. Moreover, because knockdown of TET2 recapitulates certain features of erythroid development defects characteristic of myelodysplastic syndromes (MDSs), and the TET2 gene mutation is one of the most common mutations in MDS, our findings may be relevant for improved understanding of dyserythropoiesis of MDS.

Increased production of fetal hemoglobin (HbF) can ameliorate the severity of sickle cell disease and β-thalassemia 1. BCL11A represses the genes encoding HbF and regulates human hemoglobin switching through variation in its expression during development 2-7. However, the mechanisms underlying the developmental expression of BCL11A remain mysterious. Here we show that BCL11A is regulated at the level of mRNA translation during human hematopoietic development. Despite decreased BCL11A protein synthesis earlier in development, BCL11A mRNA continues to be associated with ribosomes. Through unbiased genomic and proteomic analyses, we demonstrate that the RNA-binding protein LIN28B, which is developmentally expressed in a reciprocal pattern to BCL11A, directly interacts with ribosomes and BCL11A mRNA. Furthermore, we show that BCL11A mRNA translation is suppressed by LIN28B through direct interactions, independent of its role in regulating let-7 microRNAs, and BCL11A is the major target of LIN28B-mediated HbF induction. Our results reveal a previously unappreciated mechanism underlying human hemoglobin switching that illuminates new therapeutic opportunities. The developmental switch from fetal to adult hemoglobin in humans has been extensively studied and is of substantial interest for developing approaches to induce fetal hemoglobin (HbF) to treat sickle cell disease and β-thalassemia 1,8. Through functional and genetic follow up of genome-wide association studies for HbF levels 9,10 , BCL11A has been identified as a key regulator of both developmental hemoglobin switching and silencing of HbF in the adult 1-7. BCL11A protein levels are developmentally regulated in humans such that, at the earlier developmental stages when HbF is highly expressed in erythroid cells, there is little or no BCL11A protein 2,3. In contrast, BCL11A protein is robustly expressed in adult erythroid cells that have low levels of HbF expression. Despite extensive studies, the basis of this developmental regulation of BCL11A protein expression and thereby the upstream regulators of human hemoglobin switching remain undefined. Consistent with earlier studies 2,3 , we found that BCL11A protein showed a gradient in expression across fetal, newborn, and adult erythroid cells at all stages of maturation (Fig. 1a and Extended Data Fig. 1a,b,d,h,i). Surprisingly, there was no substantial change in BCL11A mRNA expression between fetal, newborn, or adult erythroid cells (Fig. 1b and Extended Data Fig. 1c,e-g), suggesting a post-transcriptional mechanism underlying the observed variation in protein expression. This difference in protein levels between the developmental stages could not be attributed to variation in the maturation state of cells or to differences in BCL11A mRNA splicing (Extended Data Fig. 2). One possible post-transcriptional mechanism underlying the observed variation in protein expression could involve BCL11A protein being more readily degraded at the earlier developmental stages, as compared with adult erythroid cells. Since newborn...

Multiple sclerosis is associated with an imbalance between tumour necrosis factor-alpha (TNF-a )-and IL-10-secreting blood cells that is corrected by interferon-beta (IFN-b ) treatment SUMMARYThe up-regulated B cell responses detectable in cerebrospinal fluid (CSF) and the augmented myelin antigen-specific T cell responses observed in the CSF as well as systematically in patients with multiple sclerosis (MS) suggest the involvement of cytokines in disease development and perpetuation. Here we report on the parallel involvement of TNF-a , IL-6, IFN-g and IL-10 in MS and controls, using enzymelinked immunospot (ELISPOT) assays to detect and enumerate cytokine-secreting mononuclear cells (MNC) prepared from blood and, for IL-6 and IL-10, from CSF without in vitro stimulation. MS is associated with elevated levels of TNF-a -secreting blood MNC when compared with levels in groups of control patients with myasthenia gravis (MG) and other neurological diseases (OND) or healthy subjects. This elevation was confined to patients with untreated MS and not present in those examined during ongoing treatment with IFN-b . Untreated patients with MS had lower numbers of IL-10-secreting blood MNC compared with the three control groups. In patients undergoing treatment with IFN-b , numbers of IL-10-secreting cells were in the same range as in controls. Normalization of TNF-a from elevated, and of IL-10 from decreased levels could be one reason for the beneficial effects of IFN-b in MS, although it remains to be shown whether these changes reflect phenomena primarily involved in MS pathogenesis or secondary changes. In CSF, levels of IL-10-secreting cells were higher than in blood in both MS and OND, with no difference between these groups. Systemic aberrations of IL-6 and IFN-g and of IL-6 in CSF in MS versus controls were only minor, irrespective of treatment with IFN-b .