Personalized in vitro models for dysplasia and carcinogenesis in the pancreas have been constrained by insufficient differentiation of human pluripotent stem cells (hPSCs) into the exocrine ...pancreatic lineage. Here, we differentiate hPSCs into pancreatic duct-like organoids (PDLOs) with morphological, transcriptional, proteomic, and functional characteristics of human pancreatic ducts, further maturing upon transplantation into mice. PDLOs are generated from hPSCs inducibly expressing oncogenic GNAS, KRAS, or KRAS with genetic covariance of lost CDKN2A and from induced hPSCs derived from a McCune-Albright patient. Each oncogene causes a specific growth, structural, and molecular phenotype in vitro. While transplanted PDLOs with oncogenic KRAS alone form heterogenous dysplastic lesions or cancer, KRAS with CDKN2A loss develop dedifferentiated pancreatic ductal adenocarcinomas. In contrast, transplanted PDLOs with mutant GNAS lead to intraductal papillary mucinous neoplasia-like structures. Conclusively, PDLOs enable in vitro and in vivo studies of pancreatic plasticity, dysplasia, and cancer formation from a genetically defined background.
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•Robust differentiation of hPSCs into functional pancreatic duct-like organoids•RNA-seq and proteome measurements confirm ductal identity and maturity•Inducing KRASG12D in PDLOs causes EMT and growth arrest, GNASR201C/H causes cystic growth•PDAC- and IPMN-like tumor formation of oncogenic PDLO grafts
Kleger and colleagues developed a differentiation protocol guiding hPSCs into functional pancreatic duct-like organoids. Focusing on early pancreatic tumor formation, they show that the PDLO system is applicable for disease modeling in vitro and in vivo. The genetically defined background of PDLOs will allow the tracking of human pancreatic cancer development.
Since acute myeloid leukemia (AML) is characterized by the blockade of hematopoietic differentiation and cell death, we interrogated RIPK3 signaling in AML development. Genetic loss of Ripk3 ...converted murine FLT3-ITD-driven myeloproliferation into an overt AML by enhancing the accumulation of leukemia-initiating cells (LIC). Failed inflammasome activation and cell death mediated by tumor necrosis factor receptor caused this accumulation of LIC exemplified by accelerated leukemia onset in Il1r1−/−, Pycard–/–, and Tnfr1/2−/− mice. RIPK3 signaling was partly mediated by mixed lineage kinase domain-like. This link between suppression of RIPK3, failed interleukin-1β release, and blocked cell death was supported by significantly reduced RIPK3 in primary AML patient cohorts. Our data identify RIPK3 and the inflammasome as key tumor suppressors in AML.
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•RIPK3 suppresses HSPC transformation by promoting cell death and differentiation•Cell death in leukemia-initiating cells is driven by TNFR-driven RIPK3 activation•RIPK3 promotes differentiation of LIC by activating the inflammasome•RIPK3 expression is reduced in several subtypes of primary de novo AML
Höckendorf et al. demonstrate that RIPK3 restricts malignant myeloproliferation by activating the inflammasome, which promotes differentiation and cell death, and that loss of RIPK3 increases leukemic burden in mice. Reduced RIPK3 expression is observed across several human acute myeloid leukemia subtypes.
The immune system is in place to assist in ensuring tissue homeostasis, which can be easily perturbed by invading pathogens or nonpathogenic stressors causing tissue damage. Extracellular nucleotides ...are well known to contribute to innate immune signaling specificity and strength, but how their signaling is relayed downstream of cell surface receptors and how this translates into antiviral immunity is only partially understood. Here, we systematically investigated the responses of human macrophages to extracellular nucleotides, focusing on the nucleotide‐sensing GPRC receptors of the P2Y family. Time‐resolved transcriptomic analysis showed that adenine‐ and uridine‐based nucleotides induce a specific, immediate, and transient cytokine response through the MAPK signaling pathway that regulates transcriptional activation by AP‐1. Using receptor trans‐complementation, we identified a subset of P2Ys (P2Y1, P2Y2, P2Y6, and P2Y11) that govern inflammatory responses via cytokine induction, while others (P2Y4, P2Y11, P2Y12, P2Y13, and P2Y14) directly induce antiviral responses. Notably, P2Y11 combined both activities, and depletion or inhibition of this receptor in macrophages impaired both inflammatory and antiviral responses. Collectively, these results highlight the underappreciated functions of P2Y receptors in innate immune processes.
Synopsis
How signaling via extracellular nucleotides contributes to innate immunity and antiviral responses is incompletely understood. Here, a systematic survey of the eight P2Y family nucleotide receptors in human macrophages reveals two distinct subsets controlling either inflammation or viral replication.
P2Y1, P2Y2, P2Y6 and P2Y11 induces cytokines in response to nucleotide treatment.
P2Y4, P2Y11, P2Y12, P2Y13 and P2Y14 inhibit replication of reporter Semliki forest virus (SFV).
P2Y11 receptor combines both properties and induces a specific cytokine expression pattern in nucleotide‐stimulated macrophages.
P2Y11‐dependent transcriptional regulation involves downstream signaling via the p38 MAPK pathway driving activation of AP‐1 transcription factors.
Most human nucleotide‐sensing P2Y macrophage receptors are involved either in cytokine induction or antiviral responses, while P2Y11 uniquely combines these two roles.
Although glioblastoma multiforme (GBM) is not an invariably cold tumor, checkpoint inhibition has largely failed in GBM. In order to investigate T cell–intrinsic properties that contribute to the ...resistance of GBM to endogenous or therapeutically enhanced adaptive immune responses, we sorted CD4+ and CD8+ T cells from the peripheral blood, normal-appearing brain tissue, and tumor bed of nine treatment-naive patients with GBM. Bulk RNA sequencing of highly pure T cell populations from these different compartments was used to obtain deep transcriptomes of tumor-infiltrating T cells (TILs). While the transcriptome of CD8+ TILs suggested that they were partly locked in a dysfunctional state, CD4+ TILs showed a robust commitment to the type 17 T helper cell (TH17) lineage, which was corroborated by flow cytometry in four additional GBM cases. Therefore, our study illustrates that the brain tumor environment in GBM might instruct TH17 commitment of infiltrating T helper cells. Whether these properties of CD4+ TILs facilitate a tumor-promoting milieu and thus could be a target for adjuvant anti-TH17 cell interventions needs to be further investigated.
In certain instances, Th17 responses are associated with severe immunopathology. T cell–intrinsic mechanisms that restrict pathogenic effector functions have been described for type 1 and 2 responses ...but are less well studied for Th17 cells. Here, we report a cell-intrinsic feedback mechanism that controls the pathogenicity of Th17 cells. Th17 cells produce IL-24, which prompts them to secrete IL-10. The IL-10–inducing function of IL-24 is independent of the cell surface receptor of IL-24 on Th17 cells. Rather, IL-24 is recruited to the inner mitochondrial membrane, where it interacts with the NADH dehydrogenase (ubiquinone) 1 α subcomplex subunit 13 (also known as Grim19), a constituent of complex I of the respiratory chain. Together, Grim19 and IL-24 promote the accumulation of STAT3 in the mitochondrial compartment. We propose that IL-24–guided mitochondrial STAT3 constitutes a rheostat to blunt extensive STAT3 deflections in the nucleus, which might then contribute to a robust IL-10 response in Th17 cells and a restriction of immunopathology in experimental autoimmune encephalomyelitis.
Although long noncoding RNAs (lncRNAs) dominate the transcriptome, their functions are largely unexplored. The extensive overlap of lncRNAs with coding and regulatory sequences restricts their ...systematic interrogation by DNA-directed perturbation. Here we developed genome-scale lncRNA transcriptome screening using Cas13d/CasRx. We show that RNA targeting overcomes limitations inherent to other screening methods, thereby considerably expanding the explorable space of the lncRNAome. By evolving the screening system toward pan-cancer applicability, it supports molecular and phenotypic data integration to contextualize screening hits or infer lncRNA function. We thereby addressed challenges posed by the enormous transcriptome size and tissue specificity through a size-reduced multiplexed gRNA library termed Albarossa, targeting 24,171 lncRNA genes. Its rational design incorporates target prioritization based on expression, evolutionary conservation and tissue specificity, thereby reconciling high discovery power and pan-cancer representation with scalable experimental throughput. Applied across entities, the screening platform identified numerous context-specific and common essential lncRNAs. Our work sets the stage for systematic exploration of lncRNA biology in health and disease.
The identity of tumor-initiating cells in many cancer types is unknown. Tumors often express genes associated with embryonic development, although the contributions of zygotic programs to tumor ...initiation and formation are poorly understood. Here, we show that regeneration-induced loss of quiescence in p53-deficient muscle stem cells (MuSCs) results in rhabdomyosarcoma formation with 100% penetrance. Genomic analyses of purified tumor cells revealed spontaneous and discrete oncogenic amplifications in MuSCs that drive tumorigenesis, including, but not limited to, the amplification of the cleavage-stage Dux transcription factor (TF) Duxbl. We further found that Dux factors drive an early embryonic gene signature that defines a molecular subtype across a broad range of human cancers. Duxbl initiates tumorigenesis by enforcing a mesenchymal-to-epithelial transition, and targeted inactivation of Duxbl specifically in Duxbl-expressing tumor cells abolishes their expansion. These findings reveal how regeneration and genomic instability can interact to activate zygotic genes that drive tumor initiation and growth.
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•Regeneration elicits tumor formation from genomically instable muscle stem cells•Tumor cell purification eases sequencing-based identification of oncogenic drivers•Dux transcription factors define a molecular subtype of a broad range of cancers•DuxB-Duxbl initiates tumorigenesis by a mesenchymal-to-epithelial-like transition
Kim and colleagues show that tissue regeneration elicits formation of tumors originating from adult stem cells. Purification of tumor cells enabled identification of oncogenic drivers, including Duxbl, that are normally expressed in early zygotes. Dux factors initiate tumorigenesis through an MET-like transition and define a broad range of human cancers.
ABSTRACT
Regulatory T cells (Tregs) are essential for the inhibition of immunity and the maintenance of tissue homeostasis. Signals from the T‐cell antigen receptor (TCR) are critical for early Treg ...development, their expansion, and inhibitory activity. Although TCR‐engaged activation of the paracaspase MALT1 is important for these Treg activities, the MALT1 effector pathways in Tregs remain ill‐defined. Here, we demonstrate that MALT1 protease activity controls the TCR‐induced upregulation of the transcription factor MYC and the subsequent expression of MYC target genes in Tregs. These mechanisms are important for Treg‐intrinsic mitochondrial function, optimal respiratory capacity, and homeostatic Treg proliferation. Consistently, conditional deletion of Myc in Tregs results similar to MALT1 inactivation in a lethal autoimmune inflammatory syndrome. Together, these results identify a MALT1 protease‐mediated link between TCR signaling in Tregs and MYC control that coordinates metabolism and Treg expansion for the maintenance of immune homeostasis.
In murine CD4+Foxp3+ regulatory T cells (Tregs), T‐cell receptor (TCR)/CD28‐induced MALT1 protease activity promotes MYC and MYC‐controlled gene expression in a c‐Rel‐dependent manner. This MALT1‐MYC axis drives mitochondrial metabolism and the respiratory capacity of Tregs, which is critical for Treg proliferation and the maintenance of immune homeostasis.
Although glioblastoma multiforme (GBM) is not an invariably cold tumor, checkpoint inhibition has largely failed in GBM. In order to investigate T cell–intrinsic properties that contribute to the ...resistance of GBM to endogenous or therapeutically enhanced adaptive immune responses, we sorted CD4
+
and CD8
+
T cells from the peripheral blood, normal-appearing brain tissue, and tumor bed of nine treatment-naive patients with GBM. Bulk RNA sequencing of highly pure T cell populations from these different compartments was used to obtain deep transcriptomes of tumor-infiltrating T cells (TILs). While the transcriptome of CD8
+
TILs suggested that they were partly locked in a dysfunctional state, CD4
+
TILs showed a robust commitment to the type 17 T helper cell (T
H
17) lineage, which was corroborated by flow cytometry in four additional GBM cases. Therefore, our study illustrates that the brain tumor environment in GBM might instruct T
H
17 commitment of infiltrating T helper cells. Whether these properties of CD4
+
TILs facilitate a tumor-promoting milieu and thus could be a target for adjuvant anti-T
H
17 cell interventions needs to be further investigated.
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