Overexpression of the EVI1 oncogene is associated typically with aggressive myeloid leukemia, but is also detectable in breast carcinoma where its contributions are unexplored. Analyzing a tissue ...microarray of 608 breast carcinoma patient specimens, we documented EVI1 overexpression in both estrogen receptor-positive (ER
) and estrogen receptor-negative (ER
) breast carcinomas. Here, we report prognostic relevance of EVI1 overexpression in triple-negative breast carcinoma but not in the HER2-positive breast carcinoma subset. In human breast cancer cells, EVI1 silencing reduced proliferation, apoptosis resistance, and tumorigenicity, effects rescued by estrogen supplementation in ER
breast carcinoma cells. Estrogen addition restored ERK phosphorylation in EVI1-silenced cells, suggesting that EVI1 and estradiol signaling merge in MAPK activation. Conversely, EVI1 silencing had no effect on constitutive ERK activity in HER2
breast carcinoma cells. Microarray analyses revealed G-protein-coupled receptor (GPR) signaling as a prominent EVI1 effector mechanism in breast carcinoma. Among others, the GPR54-ligand KISS1 was identified as a direct transcriptional target of EVI1, which together with other EVI1-dependent cell motility factors such as RHOJ regulated breast carcinoma cell migration. Overall, our results establish the oncogenic contributions of EVI1 in ER- and HER2-negative subsets of breast cancer.
.
The zebrafish has become a powerful vertebrate model for genetic studies of embryonic development and organogenesis and increasingly for studies in cancer biology. Zebrafish facilitate the ...performance of reverse and forward genetic approaches, including mutagenesis and small molecule screens. Moreover, several studies report the feasibility of xenotransplanting human cells into zebrafish embryos and adult fish. This model provides a unique opportunity to monitor tumor‐induced angiogenesis, invasiveness, and response to a range of treatments in vivo and in real time. Despite the high conservation of gene function between fish and humans, concern remains that potential differences in zebrafish tissue niches and/or missing microenvironmental cues could limit the relevance and translational utility of data obtained from zebrafish human cancer cell xenograft models. Here, we summarize current data on xenotransplantation of human cells into zebrafish, highlighting the advantages and limitations of this model in comparison to classical murine models of xenotransplantation.
The Nck-associated protein 1-like (NCKAP1L) gene, alternatively called hematopoietic protein 1 (HEM-1), encodes a hematopoietic lineage-specific regulator of the actin cytoskeleton. Nckap1l-deficient ...mice have anomalies in lymphocyte development, phagocytosis, and neutrophil migration. Here we report, for the first time, NCKAP1L deficiency cases in humans. In two unrelated patients of Middle Eastern origin, recessive mutations in NCKAP1L abolishing protein expression led to immunodeficiency, lymphoproliferation, and hyperinflammation with features of hemophagocytic lymphohistiocytosis. Immunophenotyping showed an inverted CD4/CD8 ratio with a major shift of both CD4+ and CD8+ cells toward memory compartments, in line with combined RNA-seq/proteomics analyses revealing a T cell exhaustion signature. Consistent with the core function of NCKAP1L in the reorganization of the actin cytoskeleton, patients' T cells displayed impaired early activation, immune synapse morphology, and leading edge formation. Moreover, knockdown of nckap1l in zebrafish led to defects in neutrophil migration. Hence, NCKAP1L mutations lead to broad immune dysregulation in humans, which could be classified within actinopathies.
The reversible transitions of cancer cells between epithelial and mesenchymal states comprise cellular and molecular processes essential for local tumor growth and respective dissemination. We report ...here that globoside glycosphingolipid (GSL) glycosyltransferase-encoding genes are elevated in epithelial cells and correlate with characteristic EMT signatures predictive of disease outcome. Depletion of globosides through CRISPR-
-mediated deletion of the key enzyme A4GALT induces EMT, enhances chemoresistance, and increased CD24
/CD44
cells. The cholera toxin-induced mesenchymal-to-epithelial transition occurred only in cells with functional A4GALT. Cells undergoing EMT lost E-cadherin expression through epigenetic silencing at the promoter region of
However, in
cells, demethylation was able to rescue E-cadherin-mediated cell-cell adhesion only in the presence of exogenous A4GALT. Overall, our data suggest another class of biomolecules vital for epithelial cancer cells and for maintaining cell integrity and function.
This study highlights the essential role of glycosphingolipids in the maintenance of epithelial cancer cell properties.
.
The extracellular matrix (ECM) plays critical roles in tumor progression and metastasis. However, the contribution of ECM proteins to early metastatic onset in the peritoneal cavity remains ...unexplored. Here, we suggest a new route of metastasis through the interaction of integrin alpha 2 (ITGA2) with collagens enriched in the tumor coinciding with poor outcome in patients with ovarian cancer. Using multiple gene-edited cell lines and patient-derived samples, we demonstrate that ITGA2 triggers cancer cell adhesion to collagen, promotes cell migration, anoikis resistance, mesothelial clearance, and peritoneal metastasis in vitro and in vivo. Mechanistically, phosphoproteomics identify an ITGA2-dependent phosphorylation of focal adhesion kinase and mitogen-activated protein kinase pathway leading to enhanced oncogenic properties. Consequently, specific inhibition of ITGA2-mediated cancer cell-collagen interaction or targeting focal adhesion signaling may present an opportunity for therapeutic intervention of metastatic spread in ovarian cancer.
Transcriptome analysis of adult hematopoietic stem cells (HSCs) and their progeny has revealed mechanisms of blood differentiation and leukemogenesis, but a similar analysis of HSC development is ...lacking. Here, we acquired the transcriptomes of developing HSCs purified from >2,500 murine embryos and adult mice. We found that embryonic hematopoietic elements clustered into three distinct transcriptional states characteristic of the definitive yolk sac, HSCs undergoing specification, and definitive HSCs. We applied a network-biology-based analysis to reconstruct the gene regulatory networks of sequential stages of HSC development and functionally validated candidate transcriptional regulators of HSC ontogeny by morpholino-mediated knockdown in zebrafish embryos. Moreover, we found that HSCs from in vitro differentiated embryonic stem cells closely resemble definitive HSCs, yet lack a Notch-signaling signature, likely accounting for their defective lymphopoiesis. Our analysis and web resource will enhance efforts to identify regulators of HSC ontogeny and facilitate the engineering of hematopoietic specification.
Display omitted
► Gene expression profiling of hematopoietic stem cells during embryonic development ► Expression profiles used to construct gene regulatory networks ► Multiple regulators of HSC development implicated ► ESC-derived HSCs resemble definitive HSCs but lack a Notch-signaling signature
Network-based analysis of HSC development using transcriptome profiling reveals stage-specific developmental regulators and differences relative to HSCs derived from ESCs.
During development, hematopoietic stem cells (HSCs) emerge from aortic endothelial cells (ECs) through an intermediate stage called hemogenic endothelium by a process known as ...endothelial‐to‐hematopoietic transition (EHT). While Notch signaling, including its upstream regulator Vegf, is known to regulate this process, the precise molecular control and temporal specificity of Notch activity remain unclear. Here, we identify the zebrafish transcriptional regulator evi1 as critically required for Notch‐mediated EHT. In vivo live imaging studies indicate that evi1 suppression impairs EC progression to hematopoietic fate and therefore HSC emergence. evi1 is expressed in ECs and induces these effects cell autonomously by activating Notch via pAKT. Global or endothelial‐specific induction of notch, vegf, or pAKT can restore endothelial Notch and HSC formations in evi1 morphants. Significantly, evi1 overexpression induces Notch independently of Vegf and rescues HSC numbers in embryos treated with a Vegf inhibitor. In sum, our results unravel evi1–pAKT as a novel molecular pathway that, in conjunction with the shh–vegf axis, is essential for activation of Notch signaling in VDA endothelial cells and their subsequent conversion to HSCs.
Synopsis
During embryonic development, definitive hematopoietic stem cells (HSCs) arise from aortic endothelial cells by a process known as endothelial‐to‐hematopoietic transition (EHT). While shh–vegf signaling is known to regulate EHT via Notch, here, a novel evi1–pAKT pathway is shown to be essential for activation of Notch signaling in ventral dorsal aorta endothelial cells and their subsequent conversion to HSCs.
The zebrafish transcriptional regulator evi1 activates pAKT and is critically required for Notch‐mediated EHT.
In vivo live imaging indicates that evi1 suppression impairs endothelial cell progression to hematopoietic fate cell autonomously.
Global or endothelial‐specific induction of notch, vegf, or pAKT can restore HSC formation in evi1 morphants.
evi1 overexpression induces Notch independently of Vegf and rescues HSC numbers in embryos treated with Vegf inhibitor.
A new VEGF‐independent pathway triggers Notch‐mediated conversion of dorsal aorta endothelial cells into definitive hematopoietic stem cells during embryonic endothelial‐to‐hematopoietic transition.
Zebrafish offer a powerful vertebrate model for studies of development and disease. The major advantages of this model include the possibilities of conducting reverse and forward genetic screens and ...of observing cellular processes by
imaging of single cells. Moreover, pathways regulating blood development are highly conserved between zebrafish and mammals, and several discoveries made in fish were later translated to murine and human models. This review and accompanying poster provide an overview of zebrafish hematopoiesis and discuss the existing zebrafish models of blood disorders, such as myeloid and lymphoid malignancies, bone marrow failure syndromes and immunodeficiencies, with a focus on how these models were generated and how they can be applied for translational research.
Abstract
Oncogenic
Ras
mutations occur in various leukemias. It was unclear if, besides the direct transforming effect via constant RAS/MEK/ERK signaling, an inflammation-related effect of KRAS ...contributes to the disease. Here, we identify a functional link between oncogenic
Kras
G12D
and NLRP3 inflammasome activation in murine and human cells. Mice expressing active
Kras
G12D
in the hematopoietic system developed myeloproliferation and cytopenia, which is reversed in
Kras
G12D
mice lacking NLRP3 in the hematopoietic system. Therapeutic IL-1-receptor blockade or NLRP3-inhibition reduces myeloproliferation and improves hematopoiesis. Mechanistically, Kras
G12D
-RAC1 activation induces reactive oxygen species (ROS) production causing NLRP3 inflammasome-activation. In agreement with our observations in mice, patient-derived myeloid leukemia cells exhibit KRAS/RAC1/ROS/NLRP3/IL-1β axis activity. Our findings indicate that oncogenic KRAS not only act via its canonical oncogenic driver function, but also enhances the activation of the pro-inflammatory RAC1/ROS/NLRP3/IL-1β axis. This paves the way for a therapeutic approach based on immune modulation via NLRP3 blockade in KRAS-mutant myeloid malignancies.
Patients suffering from acute myeloid leukemia (AML) show highly heterogeneous clinical outcomes. Next to variabilities in patient-specific parameters influencing treatment decisions and outcome, ...this is due to differences in AML biology. In fact, different genetic drivers may transform variable cells of origin and co-exist with additional genetic lesions (e.g., as observed in clonal hematopoiesis) in a variety of leukemic (sub)clones. Moreover, AML cells are hierarchically organized and contain subpopulations of more immature cells called leukemic stem cells (LSC), which on the cellular level constitute the driver of the disease and may evolve during therapy. This genetic and hierarchical complexity results in a pronounced phenotypic variability, which is observed among AML cells of different patients as well as among the leukemic blasts of individual patients, at diagnosis and during the course of the disease. Here, we review the current knowledge on the heterogeneous landscape of AML surface markers with particular focus on those identifying LSC, and discuss why identification and targeting of this important cellular subpopulation in AML remains challenging.
PDF
