参考文献

Despite the clinical success of cancer immunotherapies,the majority of patients fail to respond or develop resistance through disruption of pathways that promote neo-antigen presentation on MHC I molecules. Here,we conducted a series of unbiased,genome-wide CRISPR/Cas9 screens to identify genes that limit natural killer (NK) cell anti-tumor activity. We identified that genes associated with antigen presentation and/or interferon-$\gamma$ (IFN-$\gamma$) signaling protect tumor cells from NK cell killing. Indeed,Jak1-deficient melanoma cells were sensitized to NK cell killing through attenuated NK cell-derived IFN-$\gamma$-driven transcriptional events that regulate MHC I expression. Importantly,tumor cells that became resistant to T cell killing through enrichment of MHC I-deficient clones were highly sensitive to NK cell killing. Taken together,we reveal the genes targeted by tumor cells to drive checkpoint blockade resistance but simultaneously increase their vulnerability to NK cells,unveiling NK cell-based immunotherapies as a strategy to antagonize tumor immune escape. View Publication

Humans and other mammalian hosts have evolved mechanisms to control the bacteria colonizing their mucosal barriers to prevent invasion. While the breach of barriers by bacteria typically leads to overt infection,increasing evidence supports a role for translocation of commensal bacteria across an impaired gut barrier to extraintestinal sites in the pathogenesis of autoimmune and other chronic,non-infectious diseases. Whether gut commensal translocation is a cause or consequence of the disease is incompletely defined. Here we discuss factors that lead to translocation of live bacteria across the gut barrier. We expand upon our recently published demonstration that translocation of the gut pathobiont Enterococcus gallinarum can induce autoimmunity in susceptible hosts and postulate on the role of Enterococcus species as instigators of chronic,non-infectious diseases. View Publication

Filoviruses of the genus Ebolavirus include five species with marked differences in their ability to cause disease in humans. From the highly virulent Ebola virus to the seemingly nonpathogenic Reston virus,case-fatality rates can range between 0-90{\%}. In order to understand the molecular basis of these differences it is imperative to establish disease models that recapitulate human disease as faithfully as possible. Non-human primates are the gold-standard models for filovirus pathogenesis,but comparative studies are skewed by the fact that Reston virus infection can be lethal for NHP. Here we have used HLA-A2 transgenic,NOD-scid-interleukin 2$\gamma$ receptor knockout (NSG-A2) mice reconstituted with human hematopoiesis to compare Ebola virus and Reston virus pathogenesis in a human-like environment. While significantly less pathogenic than Ebola virus,Reston virus killed 20{\%} of infected mice,a finding that was linked to exacerbated inflammation and viral replication in the liver. In addition,'humanized' mice recapitulated the case-fatality ratios of different Ebolavirus species in humans. Our findings point out at humanized mice as a putative model to test the pathogenicity of newly discovered filoviruses,and warrants further investigations on Reston virus pathogenesis in humans. View Publication

In patients with inactivating mutations in myosin Vb (Myo5B),enterocytes show large inclusions lined by microvilli. The origin of inclusions in small-intestinal enterocytes in microvillus inclusion disease is currently unclear. We postulated that inclusions in Myo5b KO mouse enterocytes form through invagination of the apical brush border membrane. 70-kD FITC-dextran added apically to Myo5b KO intestinal explants accumulated in intracellular inclusions. Live imaging of Myo5b KO-derived enteroids confirmed the formation of inclusions from the apical membrane. Treatment of intestinal explants and enteroids with Dyngo resulted in accumulation of inclusions at the apical membrane. Inclusions in Myo5b KO enterocytes contained VAMP4 and Pacsin 2 (Syndapin 2). Myo5b;Pacsin 2 double-KO mice showed a significant decrease in inclusion formation. Our results suggest that apical bulk endocytosis in Myo5b KO enterocytes resembles activity-dependent bulk endocytosis,the primary mechanism for synaptic vesicle uptake during intense neuronal stimulation. Thus,apical bulk endocytosis mediates the formation of inclusions in neonatal Myo5b KO enterocytes. View Publication

We introduce machine learning (ML) to perform classification and quantitation of images of nuclei from human blood neutrophils. Here we assessed the use of convolutional neural networks (CNNs) using free,open source software to accurately quantitate neutrophil NETosis,a recently discovered process involved in multiple human diseases. CNNs achieved {\textgreater}94{\%} in performance accuracy in differentiating NETotic from non-NETotic cells and vastly facilitated dose-response analysis and screening of the NETotic response in neutrophils from patients. Using only features learned from nuclear morphology,CNNs can distinguish between NETosis and necrosis and between distinct NETosis signaling pathways,making them a precise tool for NETosis detection. Furthermore,by using CNNs and tools to determine object dispersion,we uncovered differences in NETotic nuclei clustering between major NETosis pathways that is useful in understanding NETosis signaling events. Our study also shows that neutrophils from patients with sickle cell disease were unresponsive to one of two major NETosis pathways. Thus,we demonstrate the design,performance,and implementation of ML tools for rapid quantitative and qualitative cell analysis in basic science. View Publication

The contribution of mast cells in the microenvironment of solid malignancies remains controversial. Here we functionally assess the impact of tumor-adjacent,submucosal mast cell accumulation in murine and human intestinal-type gastric cancer. We find that genetic ablation or therapeutic inactivation of mast cells suppresses accumulation of tumor-associated macrophages,reduces tumor cell proliferation and angiogenesis,and diminishes tumor burden. Mast cells are activated by interleukin (IL)-33,an alarmin produced by the tumor epithelium in response to the inflammatory cytokine IL-11,which is required for the growth of gastric cancers in mice. Accordingly,ablation of the cognate IL-33 receptor St2 limits tumor growth,and reduces mast cell-dependent production and release of the macrophage-attracting factors Csf2,Ccl3,and Il6. Conversely,genetic or therapeutic macrophage depletion reduces tumor burden without affecting mast cell abundance. Therefore,tumor-derived IL-33 sustains a mast cell and macrophage-dependent signaling cascade that is amenable for the treatment of gastric cancer. View Publication

The objective of the study was to identify the mechanism of action for a radiation mitigator of the gastrointestinal (GI) acute radiation syndrome (ARS),identified in an unbiased high-throughput screen. We used mice irradiated with a lethal dose of radiation and treated with daily injections of the radiation mitigator 1-[(4-nitrophenyl)sulfonyl]-4-phenylpiperazine to study its effects on key pathways involved in intestinal stem cell (ISC) maintenance. RNASeq,quantitative reverse transcriptase-polymerase chain reaction,and immunohistochemistry were performed to identify pathways engaged after drug treatment. Target validation was performed with competition assays,reporter cells,and in silico docking. 1-[(4-Nitrophenyl)sulfonyl]-4-phenylpiperazine activates Hedgehog signaling by binding to the transmembrane domain of Smoothened,thereby expanding the ISC pool,increasing the number of regenerating crypts and preventing the GI-ARS. We conclude that Smoothened is a target for radiation mitigation in the small intestine that could be explored for use in radiation accidents as well as to mitigate normal tissue toxicity during and after radiotherapy of the abdomen. View Publication

Inflammatory bowel diseases (IBDs) are chronic and relapsing immune disorders that result,or possibly originate,from epithelial barrier defects. Intestinal organoids are a new reliable tool to investigate epithelial response in models of chronic inflammation. We produced organoids from the ulcerative colitis murine model Winnie to explore if the chronic inflammatory features observed in the parental intestine were preserved by the organoids. Furthermore,we investigated if quercetin administration to in vitro cultured organoids could suppress LPS-induced inflammation in wild-type organoids (WT-organoids) and spontaneous inflammation in ulcerative colitis organoids (UC-organoids). Our data demonstrate that small intestinal organoids obtained from Winnie mice retain the chronic intestinal inflammatory features characteristic of the parental tissue. Quercetin administration was able to suppress inflammation both in UC-organoids and in LPS-treated WT-organoids. Altogether,our data demonstrate that UC-organoids are a reliable experimental system for investigating chronic intestinal inflammation and pharmacological responses. View Publication

Post-transcriptional mechanisms have the potential to influence complex changes in gene expression,yet their role in cell fate transitions remains largely unexplored. Here,we show that suppression of the RNA helicase DDX6 endows human and mouse primed embryonic stem cells (ESCs) with a differentiation-resistant,hyper-pluripotent" state which readily reprograms to a naive state resembling the preimplantation embryo. We further demonstrate that DDX6 plays a key role in adult progenitors where it controls the balance between self-renewal and differentiation in a context-dependent manner. Mechanistically DDX6 mediates the translational suppression of target mRNAs in P-bodies. Upon loss of DDX6 activity P-bodies dissolve and release mRNAs encoding fate-instructive transcription and chromatin factors that re-enter the ribosome pool. Increased translation of these targets impacts cell fate by rewiring the enhancer heterochromatin and DNA methylation landscapes of undifferentiated cell types. Collectively our data establish a link between P-body homeostasis chromatin organization and stem cell potency." View Publication

mTORC1 regulates cellular growth and is activated by growth factors and by essential amino acids such as Leu. Leu enters cells via the Leu transporter LAT1-4F2hc (LAT1). Here we show that the Na+/K+/2Cl- cotransporter NKCC1 (SLC12A2),a known regulator of cell volume,is present in complex with LAT1. We further show that NKCC1 depletion or deletion enhances LAT1 activity,as well as activation of Akt and Erk,leading to activation of mTORC1 in cells,colonic organoids,and mouse colon. Moreover,NKCC1 depletion reduces intracellular Na+ concentration and cell volume (size) and mass and stimulates cell proliferation. NKCC1,therefore,suppresses mTORC1 by inhibiting its key activating signaling pathways. Importantly,by linking ion transport and cell volume regulation to mTORC1 function,NKCC1 provides a long-sought link connecting cell volume (size) to cell mass regulation. View Publication