Stem-cell ageing is thought to contribute to altered tissue maintenance and repair. Older humans experience increased bone marrow failure and poorer haematologic tolerance of cytotoxic injury. Haematopoietic stem cells (HSCs) in older mice have decreased per-cell repopulating activity, self-renewal and homing abilities, myeloid skewing of differentiation, and increased apoptosis with stress. Here we report that the cyclin-dependent kinase inhibitor p16INK4a, the level of which was previously noted to increase in other cell types with age, accumulates and modulates specific age-associated HSC functions. Notably, in the absence of p16INK4a, HSC repopulating defects and apoptosis were mitigated, improving the stress tolerance of cells and the survival of animals in successive transplants, a stem-cell-autonomous tissue regeneration model. Inhibition of p16INK4a may ameliorate the physiological impact of ageing on stem cells and thereby improve injury repair in aged tissue.

Wingless (Wnt) is a potent morphogen demonstrated in multiple cell lineages to promote the expansion and maintenance of stem and progenitor cell populations. Wnt effects are highly context dependent, and varying effects of Wnt signaling on hematopoietic stem cells (HSCs) have been reported. We explored the impact of Wnt signaling in vivo, specifically in the context of the HSC niche by using an osteoblast-specific promoter driving expression of the paninhibitor of canonical Wnt signaling, Dickkopf1 (Dkk1). Here we report that Wnt signaling was markedly inhibited in HSCs and, unexpectedly given prior reports, reduction in HSC Wnt signaling resulted in reduced p21Cip1 expression, increased cell cycling, and a progressive decline in regenerative function after transplantation. This effect was microenvironment determined, but irreversible if the cells were transferred to a normal host. Wnt pathway activation in the niche is required to limit HSC proliferation and preserve the reconstituting function of endogenous hematopoietic stem cells.

The zinc finger transcription factor GATA-2 plays a fundamental role in generating hematopoietic stem-cells in mammalian development. Less well defined is whether GATA-2 participates in adult stem-cell regulation, an issue we addressed using GATA-2 heterozygote mice that express reduced levels of GATA-2 in hematopoietic cells. While GATA

Inhibitors of the receptor tyrosine kinase c-MET are currently used in the clinic to target oncogenic signaling in tumor cells. We found that concomitant c-MET inhibition promoted adoptive T cell transfer and checkpoint immunotherapies in murine cancer models by increasing effector T cell infiltration in tumors. This therapeutic effect was independent of tumor cell-intrinsic c-MET dependence. Mechanistically, c-MET inhibition impaired the reactive mobilization and recruitment of neutrophils into tumors and draining lymph nodes in response to cytotoxic immunotherapies. In the absence of c-MET inhibition, neutrophils recruited to T cell-inflamed microenvironments rapidly acquired immunosuppressive properties, restraining T cell expansion and effector functions. In cancer patients, high serum levels of the c-MET ligand HGF correlated with increasing neutrophil counts and poor responses to checkpoint blockade therapies. Our findings reveal a role for the HGF/c-MET pathway in neutrophil recruitment and function and suggest that c-MET inhibitor co-treatment may improve responses to cancer immunotherapy in settings beyond c-MET-dependent tumors.

Throughout development, cell fate decisions are converted into epigenetic information that determines cellular identity. Covalent histone modifications are heritable epigenetic marks and are hypothesized to play a central role in this process. In this report, we assess the concordance of histone H3 lysine 4 dimethylation (H3K4me2) and trimethylation (H3K4me3) on a genome-wide scale in erythroid development by analyzing pluripotent, multipotent, and unipotent cell types. Although H3K4me2 and H3K4me3 are concordant at most genes, multipotential hematopoietic cells have a subset of genes that are differentially methylated (H3K4me2+/me3-). These genes are transcriptionally silent, highly enriched in lineage-specific hematopoietic genes, and uniquely susceptible to differentiation-induced H3K4 demethylation. Self-renewing embryonic stem cells, which restrict H3K4 methylation to genes that contain CpG islands (CGIs), lack H3K4me2+/me3- genes. These data reveal distinct epigenetic regulation of CGI and non-CGI genes during development and indicate an interactive relationship between DNA sequence and differential H3K4 methylation in lineage-specific differentiation.

Limited responsiveness to inflammatory cytokines is a feature of adult hematopoietic stem cells and contributes to the relative quiescence and durability of the stem cell population in vivo. Here we report that the executioner Caspase, Caspase-3, unexpectedly participates in that process. Mice deficient in Caspase-3 had increased numbers of immunophenotypic long-term repopulating stem cells in association with multiple functional changes, most prominently cell cycling. Though these changes were cell autonomous, they reflected altered activation by exogenous signals. Caspase-3(-/-) cells exhibited cell type-specific changes in phosphorylated members of the Ras-Raf-MEK-ERK pathway in response to specific cytokines, while notably, members of other pathways, such as pSTAT3, pSTAT5, pAKT, pp38 MAPK, pSmad2, and pSmad3, were unaffected. Caspase-3 contributes to stem cell quiescence, dampening specific signaling events and thereby cell responsiveness to microenvironmental stimuli.

Early T-cell precursor leukaemia (ETP-ALL) is a high-risk subtype of human leukaemia that is poorly understood at the molecular level. Here we report translocations targeting the zinc finger E-box-binding transcription factor ZEB2 as a recurrent genetic lesion in immature/ETP-ALL. Using a conditional gain-of-function mouse model, we demonstrate that sustained Zeb2 expression initiates T-cell leukaemia. Moreover, Zeb2-driven mouse leukaemia exhibit some features of the human immature/ETP-ALL gene expression signature, as well as an enhanced leukaemia-initiation potential and activated Janus kinase (JAK)/signal transducers and activators of transcription (STAT) signalling through transcriptional activation of IL7R. This study reveals ZEB2 as an oncogene in the biology of immature/ETP-ALL and paves the way towards pre-clinical studies of novel compounds for the treatment of this aggressive subtype of human T-ALL using our Zeb2-driven mouse model.

IntroductionZeb2 (also known as Sip1 and Zfhx1b) is a DNA-binding transcriptional regulator of the family of zinc-finger E-box-binding (ZEB) proteins. 1,2 Its expression and functioning during development have been associated with epithelial-to-mesenchymal transitions (EMTs). 3 EMTs encompass a series of events in which polarized epithelial cells become round in shape, lose their cell-cell contacts, and acquire the motile, migratory properties of mesenchymal cells. 4 This physiologic process is essential for many developmental processes, including mesoderm formation during gastrulation and neural crest delamination and migration. Similar EMT-like changes in cellular morphology can be observed during tumor progression, allowing tumor cells to acquire the capacity to invade the surrounding tissue and ultimately metastasize to a distant site. Subsequent tissue colonization occurs via a reverse transitional mechanism called the mesenchymal-to-epithelial transition. Given the importance of EMT and the mesenchymal-to-epithelial transition in developmental processes and disease, numerous studies have identified several EMT-inducing or EMT-regulating transcription factors, including Zeb2. 5 Recent studies of neoplastic tissues have demonstrated the existence of cancer stem cells (CSCs), tumor-initiating cells with a self-renewal capacity that exhibit an ability to induce new tumors when transplanted into nude and/or syngeneic mouse strains. 6 The existence of CSCs was initially discovered in leukemia samples, but they have also been identified in various solid tumor types. The origin of CSCs was until now unclear, but compelling results from Mani et al 7 now link EMT processes with the formation of CSCs. EMT induction in an immortalized human mammary epithelial cell line resulted in the acquisition of mesenchymal traits, the expression of stem cell markers, and an enhanced capacity to form mammospheres, a property previously and exclusively associated with mammary epithelial stem cells. 7 Suppression of the miR-200 family members, which together with miR-205 have previously been shown to negatively regulate Zeb family members, 8 not only elevates Zeb1 and/or Zeb2 expression, but also several stem cell factors (including Bmi), resulting in increased stemness and metastasisinitiating capacity. 9 These findings illustrate a potential direct link between EMT induction and the acquisition of stem cell properties.It is in this context that we analyzed the role of the EMT inducer Zeb2 in the formation of tissue-specific stem cells in vivo, specifically within the hematopoietic system. Mature blood cells arise from HSCs that are capable of generating every hematopoietic cell type, including the various lymphoid and myeloid lineages. Each HSC has the capacity to generate large numbers of mature hematopoietic cells throughout its life, and the HSC pool size in adults is regulated by finely tuned self-renewal and differentiation The online version of this article contains a data supplement.The publication costs of this article wer...