Post-translational control of T cell development by the ESCRT protein CHMP5.

The acquisition of a protective vertebrate immune system hinges on the efficient generation of a diverse but self-tolerant repertoire of T cells by the thymus through mechanisms that remain incompletely resolved. Here we identified the endosomal-sorting-complex-required-for-transport (ESCRT) protein CHMP5, known to be required for the formation of multivesicular bodies, as a key sensor of thresholds for signaling via the T cell antigen receptor (TCR) that was essential for T cell development. CHMP5 enabled positive selection by promoting post-selection thymocyte survival in part through stabilization of the pro-survival protein Bcl-2. Accordingly, loss of CHMP5 in thymocyte precursor cells abolished T cell development, a phenotype that was 'rescued' by genetic deletion of the pro-apoptotic protein Bim or transgenic expression of Bcl-2. Mechanistically, positive selection resulted in the stabilization of CHMP5 by inducing its interaction with the deubiquitinase USP8. Our results thus identify CHMP5 as an essential component of the post-translational machinery required for T cell development.

Efficient hemogenic endothelial cell specification by RUNX1 is dependent on baseline chromatin accessibility of RUNX1-regulated TGFß target genes.

Hematopoietic stem and progenitor cells (HSPCs) are generated de novo in the embryo from hemogenic endothelial cells (HECs) via an endothelial-to-hematopoietic transition (EHT) that requires the transcription factor RUNX1. Ectopic expression of RUNX1 alone can efficiently promote EHT and HSPC formation from embryonic endothelial cells (ECs), but less efficiently from fetal or adult ECs. Efficiency correlated with baseline accessibility of TGFß-related genes associated with endothelial-to-mesenchymal transition (EndoMT) and participation of AP-1 and SMAD2/3 to initiate further chromatin remodeling along with RUNX1 at these sites. Activation of TGFß signaling improved the efficiency with which RUNX1 specified fetal ECs as HECs. Thus, the ability of RUNX1 to promote EHT depends on its ability to recruit the TGFß signaling effectors AP-1 and SMAD2/3, which in turn is determined by the changing chromatin landscape in embryonic versus fetal ECs. This work provides insight into regulation of EndoMT and EHT that will guide reprogramming efforts for clinical applications.

The transcription factor XBP1 is selectively required for eosinophil differentiation.

The transcription factor XBP1 has been linked to the development of highly secretory tissues such as plasma cells and Paneth cells, yet its function in granulocyte maturation has remained unknown. Here we discovered an unexpectedly selective and absolute requirement for XBP1 in eosinophil differentiation without an effect on the survival of basophils or neutrophils. Progenitors of myeloid cells and eosinophils selectively activated the endoribonuclease IRE1α and spliced Xbp1 mRNA without inducing parallel endoplasmic reticulum (ER) stress signaling pathways. Without XBP1, nascent eosinophils exhibited massive defects in the post-translational maturation of key granule proteins required for survival, and these unresolvable structural defects fed back to suppress critical aspects of the transcriptional developmental program. Hence, we present evidence that granulocyte subsets can be distinguished by their differential reliance on secretory-pathway homeostasis.

Pluripotent stem cell-derived epithelium misidentified as brain microvascular endothelium requires ETS factors to acquire vascular fate.

Cells derived from pluripotent sources in vitro must resemble those found in vivo as closely as possible at both transcriptional and functional levels in order to be a useful tool for studying diseases and developing therapeutics. Recently, differentiation of human pluripotent stem cells (hPSCs) into brain microvascular endothelial cells (ECs) with blood-brain barrier (BBB)-like properties has been reported. These cells have since been used as a robust in vitro BBB model for drug delivery and mechanistic understanding of neurological diseases. However, the precise cellular identity of these induced brain microvascular endothelial cells (iBMECs) has not been well described. Employing a comprehensive transcriptomic metaanalysis of previously published hPSC-derived cells validated by physiological assays, we demonstrate that iBMECs lack functional attributes of ECs since they are deficient in vascular lineage genes while expressing clusters of genes related to the neuroectodermal epithelial lineage (Epi-iBMEC). Overexpression of key endothelial ETS transcription factors (ETV2, ERG, and FLI1) reprograms Epi-iBMECs into authentic endothelial cells that are congruent with bona fide endothelium at both transcriptomic as well as some functional levels. This approach could eventually be used to develop a robust human BBB model in vitro that resembles the human brain EC in vivo for functional studies and drug discovery.

Conversion of adult endothelium to immunocompetent haematopoietic stem cells.

Developmental pathways that orchestrate the fleeting transition of endothelial cells into haematopoietic stem cells remain undefined. Here we demonstrate a tractable approach for fully reprogramming adult mouse endothelial cells to haematopoietic stem cells (rEC-HSCs) through transient expression of the transcription-factor-encoding genes Fosb, Gfi1, Runx1, and Spi1 (collectively denoted hereafter as FGRS) and vascular-niche-derived angiocrine factors. The induction phase (days 0-8) of conversion is initiated by expression of FGRS in mature endothelial cells, which results in endogenous Runx1 expression. During the specification phase (days 8-20), RUNX1+ FGRS-transduced endothelial cells commit to a haematopoietic fate, yielding rEC-HSCs that no longer require FGRS expression. The vascular niche drives a robust self-renewal and expansion phase of rEC-HSCs (days 20-28). rEC-HSCs have a transcriptome and long-term self-renewal capacity similar to those of adult haematopoietic stem cells, and can be used for clonal engraftment and serial primary and secondary multi-lineage reconstitution, including antigen-dependent adaptive immune function. Inhibition of TGFß and CXCR7 or activation of BMP and CXCR4 signalling enhanced generation of rEC-HSCs. Pluripotency-independent conversion of endothelial cells into autologous authentic engraftable haematopoietic stem cells could aid treatment of haematological disorders.

Reprogramming human endothelial cells to haematopoietic cells requires vascular induction.

Generating engraftable human haematopoietic cells from autologous tissues is a potential route to new therapies for blood diseases. However, directed differentiation of pluripotent stem cells yields haematopoietic cells that engraft poorly. Here, we have devised a method to phenocopy the vascular-niche microenvironment of haemogenic cells, thereby enabling reprogramming of human endothelial cells into engraftable haematopoietic cells without transition through a pluripotent intermediate. Highly purified non-haemogenic human umbilical vein endothelial cells or adult dermal microvascular endothelial cells were transduced with the transcription factors FOSB, GFI1, RUNX1 and SPI1 (hereafter referred to as FGRS), and then propagated on serum-free instructive vascular niche monolayers to induce outgrowth of haematopoietic colonies containing cells with functional and immunophenotypic features of multipotent progenitor cells (MPPs). These endothelial cells that have been reprogrammed into human MPPs (rEC-hMPPs) acquire colony-forming-cell potential and durably engraft into immune-deficient mice after primary and secondary transplantation, producing long-term rEC-hMPP-derived myeloid (granulocytic/monocytic, erythroid, megakaryocytic) and lymphoid (natural killer and B cell) progenies. Conditional expression of FGRS transgenes, combined with vascular induction, activates endogenous FGRS genes, endowing rEC-hMPPs with a transcriptional and functional profile similar to that of self-renewing MPPs. Our approach underscores the role of inductive cues from the vascular niche in coordinating and sustaining haematopoietic specification and may prove useful for engineering autologous haematopoietic grafts to treat inherited and acquired blood disorders.

Divergent angiocrine signals from vascular niche balance liver regeneration and fibrosis.

Chemical or traumatic damage to the liver is frequently associated with aberrant healing (fibrosis) that overrides liver regeneration. The mechanism by which hepatic niche cells differentially modulate regeneration and fibrosis during liver repair remains to be defined. Hepatic vascular niche predominantly represented by liver sinusoidal endothelial cells deploys paracrine trophogens, known as angiocrine factors, to stimulate regeneration. Nevertheless, it is not known how pro-regenerative angiocrine signals from liver sinusoidal endothelial cells is subverted to promote fibrosis. Here, by combining an inducible endothelial-cell-specific mouse gene deletion strategy and complementary models of acute and chronic liver injury, we show that divergent angiocrine signals from liver sinusoidal endothelial cells stimulate regeneration after immediate injury and provoke fibrosis after chronic insult. The pro-fibrotic transition of vascular niche results from differential expression of stromal-derived factor-1 receptors, CXCR7 and CXCR4 (refs 18, 19, 20, 21), in liver sinusoidal endothelial cells. After acute injury, CXCR7 upregulation in liver sinusoidal endothelial cells acts with CXCR4 to induce transcription factor Id1, deploying pro-regenerative angiocrine factors and triggering regeneration. Inducible deletion of Cxcr7 in sinusoidal endothelial cells (Cxcr7(iΔEC/iΔEC)) from the adult mouse liver impaired liver regeneration by diminishing Id1-mediated production of angiocrine factors. By contrast, after chronic injury inflicted by iterative hepatotoxin (carbon tetrachloride) injection and bile duct ligation, constitutive FGFR1 signalling in liver sinusoidal endothelial cells counterbalanced CXCR7-dependent pro-regenerative response and augmented CXCR4 expression. This predominance of CXCR4 over CXCR7 expression shifted angiocrine response of liver sinusoidal endothelial cells, stimulating proliferation of desmin(+) hepatic stellate-like cells and enforcing a pro-fibrotic vascular niche. Endothelial-cell-specific ablation of either Fgfr1 (Fgfr1(iΔEC/iΔEC)) or Cxcr4 (Cxcr4(iΔEC/iΔEC)) in mice restored the pro-regenerative pathway and prevented FGFR1-mediated maladaptive subversion of angiocrine factors. Similarly, selective CXCR7 activation in liver sinusoidal endothelial cells abrogated fibrogenesis. Thus, we demonstrate that in response to liver injury, differential recruitment of pro-regenerative CXCR7-Id1 versus pro-fibrotic FGFR1-CXCR4 angiocrine pathways in vascular niche balances regeneration and fibrosis. These results provide a therapeutic roadmap to achieve hepatic regeneration without provoking fibrosis.

Akt-activated endothelium promotes ovarian cancer proliferation through notch activation.

BACKGROUND: One main challenge in ovarian cancer rests on the presence of a relapse and an important metastatic disease, despite extensive surgical debulking and chemotherapy. The difficulty in containing metastatic cancer is partly due to the heterotypic interaction of tumor and its microenvironment. In this context, evidence suggests that endothelial cells (EC) play an important role in ovarian tumor growth and chemoresistance. Here, we studied the role of tumor endothelium on ovarian cancer cells (OCCs). METHODS: We evaluated the effect of activated endothelial cells on ovarian cancer cell proliferation and resistance to chemotherapy and investigated the survival pathways activated by endothelial co-culture. RESULTS: The co-culture between OCCs and E4+ECs, induced an increase of OCCs proliferation both in vitro and in vivo. This co-culture induced an increase of Notch receptors expression on OCC surface and an increase of Jagged 1 expression on E4+ECs surface and activation of survival pathways leading to chemoresistance by E4+ECs. CONCLUSION: The targeting of aberrant NOTCH signaling could constitute a strategy to disrupt the pro-tumoral endothelial niche.

CCL2/CCL5 secreted by the stroma induce IL-6/PYK2 dependent chemoresistance in ovarian cancer.

BACKGROUND: Minimal residual disease is the main issue of advanced ovarian cancer treatment. According to the literature and previous results, we hypothesized that Mesenchymal Stromal Cells (MSC) could support this minimal residual disease by protecting ovarian cancer cells (OCC) from chemotherapy. In vitro study confirmed that MSC could induce OCC chemoresistance without contact using transwell setting. Further experiments showed that this induced chemoresistance was dependent on IL-6 OCC stimulation. METHODS: We combined meticulous in vitro profiling and tumor xenograft models to study the role of IL-6 in MSC/OCC intereactions. RESULTS: We demonstrated that Tocilizumab® (anti-IL-6R therapy) in association with chemotherapy significantly reduced the peritoneal carcinosis index (PCI) than chemotherapy alone in mice xenografted with OCCs+MSCs. Further experiments showed that CCL2 and CCL5 are released by MSC in transwell co-culture and induce OCCs IL-6 secretion and chemoresistance. Finally, we found that IL-6 induced chemoresistance was dependent on PYK2 phosphorylation. CONCLUSIONS: These findings highlight the potential key role of the stroma in protecting minimal residual disease from chemotherapy, thus favoring recurrences. Future clinical trials targeting stroma could use anti-IL-6 therapy in association with chemotherapy.

Human ESC-derived hemogenic endothelial cells undergo distinct waves of endothelial to hematopoietic transition.

UNLABELLED: Several studies have demonstrated that hematopoietic cells originate from endotheliumin early development; however, the phenotypic progression of progenitor cells during human embryonic hemogenesis is not well described. Here, we define the developmental hierarchy among intermediate populations of hematopoietic progenitor cells (HPCs) derived from human embryonic stem cells (hESCs). We genetically modified hESCs to specifically demarcate acquisition of vascular (VE-cadherin) and hematopoietic (CD41a) cell fate and used this dual-reporting transgenic hESC line to observe endothelial to hematopoietic transition by real-time confocal microscopy. Live imaging and clonal analyses revealed a temporal bias in commitment of HPCs that recapitulates discrete waves of lineage differentiation noted during mammalian hemogenesis. Specifically, HPCs isolated at later time points showed reduced capacity to form erythroid/ megakaryocytic cells and exhibited a tendency toward myeloid fate that was enabled by expression of the Notch ligand Dll4 on hESC-derived vascular feeder cells. These data provide a framework for defining HPC lineage potential, elucidate a molecular contribution from the vascular niche in promoting hematopoietic lineage progression, and distinguish unique subpopulations of hemogenic endothelium during hESC differentiation. KEY POINTS: Live imaging of endothelial to hematopoietic conversion identifies distinct subpopulations of hESC-derived hemogenic endothelium. Expression of the Notch ligand DII4 on vascular ECs drives induction of myeloid fate from hESC-derived hematopoietic progenitors.

Predictive markers of chemoresistance in advanced stages epithelial ovarian carcinoma.

OBJECTIVE: DNA repair mechanisms, environment-mediated drug resistance and cancer initiating cells (CIC) are three major research concepts that can explain the chemoresistance of epithelial ovarian cancer (EOC). The objective was to test if changes in the expression of potential markers associated with drug resistance before and after chemotherapy would correlate with platinum resistance, defined as a recurrence within the first year after chemotherapy cessation, and with survival, in advanced EOC. METHODS: We included 32 patients with stage IIIC-IV EOC who underwent laparoscopy to evaluate the extent of carcinomatosis, neoadjuvant chemotherapy (carboplatin/taxol) and interval surgery. Biopsies taken during the initial laparoscopies and interval surgeries were evaluated using immunohistochemistry for the expression of 7 proteins: CD117, CD44 and ALDH1 to evaluate CIC; IL-6, IL-8 and BMP2 to evaluate environment-mediated drug resistance; and ERCC1 to evaluate DNA repair. Expression measurements were correlated with platin resistance and survival. The markers' relevance was confirmed in vitro using chemoresistance tests and flow cytometric measurements of the proportion of CD44+ cells. RESULTS: 17 patients were chemoresistant and 15 patients were chemosensitive. We observed increases in CD44, IL-6 and ERCC1 expression and stable ALDH1, CD117, IL-8, and BMP2 expression. Reduced expression of cancer initiating cell markers and increased expression of environment-mediated drug resistance markers were associated with poor prognosis. We also demonstrated that CD44+ cells had survival advantages in vitro. CONCLUSIONS: Changes in CD44 and IL-8 expression on tumor cells appeared to correlate with overall survival and should be further tested as predictors of chemoresistance using larger cohort.

Role of mesenchymal cells in the natural history of ovarian cancer: a review.

BACKGROUND: Ovarian cancer is the deadliest gynaecologic malignancy. Despite progresses in chemotherapy and ultra-radical surgeries, this locally metastatic disease presents a high rate of local recurrence advocating for the role of a peritoneal niche. For several years, it was believed that tumor initiation, progression and metastasis were merely due to the changes in the neoplastic cell population and the adjacent non-neoplastic tissues were regarded as bystanders. The importance of the tumor microenvironment and its cellular component emerged from studies on the histopathological sequence of changes at the interface between putative tumor cells and the surrounding non-neoplastic tissues during carcinogenesis. METHOD: In this review we aimed to describe the pro-tumoral crosstalk between ovarian cancer and mesenchymal stem cells. A PubMed search was performed for articles published pertaining to mesenchymal stem cells and specific to ovarian cancer. RESULTS: Mesenchymal stem cells participate to an elaborate crosstalk through direct and paracrine interaction with ovarian cancer cells. They play a role at different stages of the disease: survival and peritoneal infiltration at early stage, proliferation in distant sites, chemoresistance and recurrence at later stage. CONCLUSION: The dialogue between ovarian and mesenchymal stem cells induces the constitution of a pro-tumoral mesencrine niche. Understanding the dynamics of such interaction in a clinical setting might propose new therapeutic strategies.

Mesenchymal cell interaction with ovarian cancer cells induces a background dependent pro-metastatic transcriptomic profile.

BACKGROUND: The cross talk between the stroma and cancer cells plays a major role in phenotypic modulation. During peritoneal carcinomatosis ovarian cancer cells interact with mesenchymal stem cells (MSC) resulting in increased metastatic ability. Understanding the transcriptomic changes underlying the phenotypic modulation will allow identification of key genes to target. However in the context of personalized medicine we must consider inter and intra tumoral heterogeneity. In this study we used a pathway-based approach to illustrate the role of cell line background in transcriptomic modification during a cross talk with MSC. METHODS: We used two ovarian cancer cell lines as a surrogate for different ovarian cancer subtypes: OVCAR3 for an epithelial and SKOV3 for a mesenchymal subtype. We co-cultured them with MSCs. Genome wide gene expression was determined after cell sorting. Ingenuity pathway analysis was used to decipher the cell specific transcriptomic changes related to different pro-metastatic traits (Adherence, migration, invasion, proliferation and chemoresistance). RESULTS: We demonstrate that co-culture of ovarian cancer cells in direct cellular contact with MSCs induces broad transcriptomic changes related to enhance metastatic ability. Genes related to cellular adhesion, invasion, migration, proliferation and chemoresistance were enriched under these experimental conditions. Network analysis of differentially expressed genes clearly shows a cell type specific pattern. CONCLUSION: The contact with the mesenchymal niche increase metastatic initiation and expansion through cancer cells' transcriptome modification dependent of the cellular subtype. Personalized medicine strategy might benefit from network analysis revealing the subtype specific nodes to target to disrupt acquired pro-metastatic profile.

Schizophrenia endothelial cells exhibit higher permeability and altered angiogenesis patterns in patient-derived organoids.

Schizophrenia (SCZ) is a complex neurodevelopmental disorder characterized by the manifestation of psychiatric symptoms in early adulthood. While many research avenues into the origins of SCZ during brain development have been explored, the contribution of endothelial/vascular dysfunction to the disease remains largely elusive. To model the neuropathology of SCZ during early critical periods of brain development, we utilized patient-derived induced pluripotent stem cells (iPSCs) to generate 3D cerebral organoids and define cell-specific signatures of disease. Single-cell RNA sequencing revealed that while SCZ organoids were similar in their macromolecular diversity to organoids generated from healthy controls (CTRL), SCZ organoids exhibited a higher percentage of endothelial cells when normalized to total cell numbers. Additionally, when compared to CTRL, differential gene expression analysis revealed a significant enrichment in genes that function in vessel formation, vascular regulation, and inflammatory response in SCZ endothelial cells. In line with these findings, data from 23 donors demonstrated that PECAM1+ microvascular vessel-like structures were increased in length and number in SCZ organoids in comparison to CTRL organoids. Furthermore, we report that patient-derived endothelial cells displayed higher paracellular permeability, implicating elevated vascular activity. Collectively, our data identified altered gene expression patterns, vessel-like structural changes, and enhanced permeability of endothelial cells in patient-derived models of SCZ. Hence, brain microvascular cells could play a role in the etiology of SCZ by modulating the permeability of the developing blood brain barrier (BBB).

Lung lymphatic endothelial cells undergo inflammatory and prothrombotic changes in a model of chronic obstructive pulmonary disease.

The lymphatic vasculature regulates lung homeostasis through drainage of fluid and trafficking of immune cells and plays a key role in the response to lung injury in several disease states. We have previously shown that lymphatic dysfunction occurs early in the pathogenesis of chronic obstructive pulmonary disease (COPD) caused by cigarette smoke (CS) and that this is associated with increased thrombin and fibrin clots in lung lymph. However, the direct effects of CS and thrombin on lymphatic endothelial cells (LECs) in COPD are not entirely clear. Studies of the blood vasculature have shown that COPD is associated with increased thrombin after CS exposure that causes endothelial dysfunction characterized by changes in the expression of coagulation factors and leukocyte adhesion proteins. Here, we determined whether similar changes occur in LECs. We used an in vitro cell culture system and treated human lung microvascular lymphatic endothelial cells with cigarette smoke extract (CSE) and/or thrombin. We found that CSE treatment led to decreased fibrinolytic activity in LECs, which was associated with increased expression of plasminogen activator inhibitor 1 (PAI-1). LECs treated with both CSE and thrombin together had a decreased expression of tissue factor pathway inhibitor (TFPI) and increased expression of adhesion molecules. RNA sequencing of lung LECs isolated from mice exposed to CS also showed upregulation of prothrombotic and inflammatory pathways at both acute and chronic exposure time points. Analysis of publicly available single-cell RNA sequencing of LECs as well as immunohistochemical staining of lung tissue from COPD patients supported these data and showed increased expression of inflammatory markers in LECs from COPD patients compared to those from controls. These studies suggest that in parallel with blood vessels, the lymphatic endothelium undergoes inflammatory changes associated with CS exposure and increased thrombin in COPD. Further research is needed to unravel the mechanisms by which these changes affect lymphatic function and drive tissue injury in COPD.

Mesenchymal stem cells enhance ovarian cancer cell infiltration through IL6 secretion in an amniochorionic membrane based 3D model.

BACKGROUND: The early peritoneal invasion of epithelial ovarian cancer (EOC) by tumoral aggregates presents in ascites is a major concern. The role of the microenvironment seems to be important in this process but the lack of adequate models to study cellular interactions between cancer cells and stromal cells does not allow to uncover the molecular pathways involved. Our goal was to study the interactions between ovarian cancer cells (OCC) and mesenchymal stem cells (MSC) using a 3D model. METHODS: We used millimetric pieces of amniochorionic membrane - referred to as amniotic membrane scaffold (AMS) - to create 3D peritoneal nodules mimicking EOC early invasion. We were able to measure the distribution and the depth of infiltration using confocal microsopy. We extracted MSC from the amniochorionic membrane using the markers CD34-, CD45-, CD73+, CD90+, CD105+ and CD29+ at the Fluorescence Activated Cell Sorting (FACS) analysis. We used transwell and wound healing tests to test OCC migration and invasion in vitro. RESULTS: Here we show that OCC tumors were located in regions rich in MSC (70%). The tumors infiltrated deeper within AMS in regions rich in MSC (p<0.001). In vitro tests revealed that higher IL6 secretion in a context of MSC-OCC co-culture could enhance migration and invasion of OCC. After IL6 receptor antagonism, OCC infiltration was significantly decreased, mostly in regions rich in MSCs, indicating that recruitment and tridimensional invasion of OCC was dependent of IL6 secretion. CONCLUSIONS: The use of tridimensional models using AMS could be a useful tool to decipher early molecular events in ovarian cancer metastasis. Cytokine inhibitors interrupting the cross-talk between OCCs and MSCs such as IL6 should be investigated as a new therapeutic approach in ovarian cancer.

Lymphatic Dysfunction Models an Autoimmune Emphysema Phenotype of Chronic Obstructive Pulmonary Disease.

Chronic Obstructive Pulmonary Disease (COPD) is a heterogeneous disease that is characterized by many clinical phenotypes. One such phenotype of COPD is defined by emphysema, pathogenic lung tertiary lymphoid organs (TLOs), and autoantibody production. We have previously shown that lymphatic dysfunction can cause lung TLO formation and lung injury in mice. We now sought to uncover whether underlying lymphatic dysfunction may be a driver of lung injury in cigarette smoke (CS)-induced COPD. We found that lung TLOs in mice with lymphatic dysfunction produce autoantibodies and are associated with a lymphatic endothelial cell subtype that expresses antigen presentation genes. Mice with underlying lymphatic dysfunction develop increased emphysema after CS exposure, with increased size and activation of TLOs. CS further increased autoantibody production in mice with lymphatic dysfunction. B-cell blockade prevented TLO formation and decreased lung injury after CS in mice with lymphatic dysfunction. Using tissue from human COPD patients, we also found evidence of a lymphatic gene signature that was specific to patients with emphysema and prominent TLOs compared to COPD patients without emphysema. Taken together, these data suggest that lymphatic dysfunction may underlie lung injury in a subset of COPD patients with an autoimmune emphysema phenotype.

Histone variant H3.3 maintains adult haematopoietic stem cell homeostasis by enforcing chromatin adaptability.

Histone variants and the associated post-translational modifications that govern the stemness of haematopoietic stem cells (HSCs) and differentiation thereof into progenitors (HSPCs) have not been well defined. H3.3 is a replication-independent H3 histone variant in mammalian systems that is enriched at both H3K4me3- and H3K27me3-marked bivalent genes as well as H3K9me3-marked endogenous retroviral repeats. Here we show that H3.3, but not its chaperone Hira, prevents premature HSC exhaustion and differentiation into granulocyte-macrophage progenitors. H3.3-null HSPCs display reduced expression of stemness and lineage-specific genes with a predominant gain of H3K27me3 marks at their promoter regions. Concomitantly, loss of H3.3 leads to a reduction of H3K9me3 marks at endogenous retroviral repeats, opening up binding sites for the interferon regulatory factor family of transcription factors, allowing the survival of rare, persisting H3.3-null HSCs. We propose a model whereby H3.3 maintains adult HSC stemness by safeguarding the delicate interplay between H3K27me3 and H3K9me3 marks, enforcing chromatin adaptability.

Specification of fetal liver endothelial progenitors to functional zonated adult sinusoids requires c-Maf induction.

The liver vascular network is patterned by sinusoidal and hepatocyte co-zonation. How intra-liver vessels acquire their hierarchical specialized functions is unknown. We study heterogeneity of hepatic vascular cells during mouse development through functional and single-cell RNA-sequencing. The acquisition of sinusoidal endothelial cell identity is initiated during early development and completed postnatally, originating from a pool of undifferentiated vascular progenitors at E12. The peri-natal induction of the transcription factor c-Maf is a critical switch for the sinusoidal identity determination. Endothelium-restricted deletion of c-Maf disrupts liver sinusoidal development, aberrantly expands postnatal liver hematopoiesis, promotes excessive postnatal sinusoidal proliferation, and aggravates liver pro-fibrotic sensitivity to chemical insult. Enforced c-Maf overexpression in generic human endothelial cells switches on a liver sinusoidal transcriptional program that maintains hepatocyte function. c-Maf represents an inducible intra-organotypic and niche-responsive molecular determinant of hepatic sinusoidal cell identity and lays the foundation for the strategies for vasculature-driven liver repair.