Severe COVID-19 is characterized by persistent lung inflammation, inflammatory cytokine production, viral RNA and a sustained interferon (IFN) response, all of which are recapitulated and required for pathology in the SARS-CoV-2-infected MISTRG6-hACE2 humanized mouse model of COVID-19, which has a human immune system [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17][18][19][20] . Blocking either viral replication with remdesivir 21-23 or the downstream IFN-stimulated cascade with anti-IFNAR2 antibodies in vivo in the chronic stages of disease attenuates the overactive immune inflammatory response, especially inflammatory macrophages. Here we show that SARS-CoV-2 infection and replication in lung-resident human macrophages is a critical driver of disease. In response to infection mediated by CD16 and ACE2 receptors, human macrophages activate inflammasomes, release interleukin 1 (IL-1) and IL-18, and undergo pyroptosis, thereby contributing to the hyperinflammatory state of the lungs. Inflammasome activation and the accompanying inflammatory response are necessary for lung inflammation, as inhibition of the NLRP3 inflammasome pathway reverses chronic lung pathology. Notably, this blockade of inflammasome activation leads to the release of infectious virus by the infected macrophages. Thus, inflammasomes oppose host infection by SARS-CoV-2 through the production of inflammatory cytokines and suicide by pyroptosis to prevent a productive viral cycle.Acute SARS-CoV-2 infection resolves in most patients but becomes chronic and sometimes deadly in about 10-20% of patients [1][2][3][4][5][6][7][14][15][16]20,[24][25][26][27] . Two hallmarks of severe COVID-19 are a sustained IFN response and viral RNA persisting for months [1][2][3][4][5][6][7][8][9][10][11][12][13][14][15][16][17]20,[24][25][26][27][28] . This chronicity is recapitulated in SARS-CoV-2-infected MISTRG6-hACE2 humanized mice 19 . High levels of IL-1β, IL-18 and lactate dehydrogenase (LDH) are correlated with COVID-19 severity in patients, suggesting a role for inflammasome activation and pyroptosis in pathology [5][6][7][14][15][16][17][18]29 . Here we show that human lung macrophages are infected by SARS-CoV-2. Replicating SARS-CoV-2 in these human macrophages activates inflammasomes and initiates an inflammatory cascade with a unique transcriptome, results in pyroptosis, and contributes to the downstream type-I IFN response. Blocking viral replication, the downstream IFN response or inflammasome activation in vivo during the chronic phase of the disease attenuates many aspects of the overactive immune inflammatory response (especially the inflammatory macrophage response) and disease. Viral replication and the IFN responseChronic interferon is associated with disease severity and impaired recovery in influenza infection 30 . To test whether a viral-RNAdependent type-I IFN response was a driver of chronic disease, we treated SARS-CoV-2-infected MISTRG6-hACE2 mice with remdesivir [21][22][23] and/or anti-IFNAR2 antibodies (Fig. 1a) to inhibit vi...

SUMMARY Activation of the Fgf signaling pathway during preimplantation development of the mouse embryo is known to be essential for differentiation of the inner cell mass and the formation of the primitive endoderm (PrE). We now show using fluorescent reporter knock-in lines that Fgfr1 is expressed in all cell populations of the blastocyst, while Fgfr2 expression becomes restricted to extraembryonic lineages, including the PrE. We further show that loss of both receptors prevents the development of the PrE and demonstrate that Fgfr1 plays a more prominent role in this process than Fgfr2. Last, we document an essential role for Fgfrs in ES cell differentiation, with again Fgfr1 having a larger influence than Fgfr2 in ES cell exit from the pluripotent state. Collectively, these results identify mechanisms through which Fgf signaling regulates inner cell mass (ICM) lineage restriction and cell commitment during preimplantation development.

The initiation of an intestinal tumour is a probabilistic process that depends on the competition between mutant and normal epithelial stem cells in crypts 1 . Intestinal stem cells are closely associated with a diverse but poorly characterized network of mesenchymal cell types 2 , 3 . However, whether the physiological mesenchymal microenvironment of mutant stem cells affects tumour initiation remains unknown. Here we provide in vivo evidence that the mesenchymal niche controls tumour initiation in trans . By characterizing the heterogeneity of the intestinal mesenchyme using single-cell RNA-sequencing analysis, we identified a population of rare pericryptal Ptgs2 -expressing fibroblasts that constitutively process arachidonic acid into highly labile prostaglandin E 2 (PGE 2 ). Specific ablation of Ptgs2 in fibroblasts was sufficient to prevent tumour initiation in two different models of sporadic, autochthonous tumorigenesis. Mechanistically, single-cell RNA-sequencing analyses of a mesenchymal niche model showed that fibroblast-derived PGE 2 drives the expansion οf a population of Sca-1 + reserve-like stem cells. These express a strong regenerative/tumorigenic program, driven by the Hippo pathway effector Yap. In vivo, Yap is indispensable for Sca-1 + cell expansion and early tumour initiation and displays a nuclear localization in both mouse and human adenomas. Using organoid experiments, we identified a molecular mechanism whereby PGE 2 promotes Yap dephosphorylation, nuclear translocation and transcriptional activity by signalling through the receptor Ptger4. Epithelial-specific ablation of Ptger4 misdirected the regenerative reprogramming of stem cells and prevented Sca-1 + cell expansion and sporadic tumour initiation in mutant mice, thereby demonstrating the robust paracrine control of tumour-initiating stem cells by PGE 2 –Ptger4. Analyses of patient-derived organoids established that PGE 2 –PTGER4 also regulates stem cell function in humans. Our study demonstrates that initiation of colorectal cancer is orchestrated by the mesenchymal niche and reveals a mechanism by which rare pericryptal Ptgs2 -expressing fibroblasts exert paracrine control over tumour-initiating stem cells via the druggable PGE 2 –Ptger4–Yap signalling axis.

The fibroblast growth factor (Fgf) family of ligands and receptor tyrosine kinases is required throughout embryonic and postnatal development and also regulates multiple homeostatic functions in the adult. Aberrant Fgf signaling causes many congenital disorders and underlies multiple forms of cancer. Understanding the mechanisms that govern Fgf signaling is therefore important to appreciate many aspects of Fgf biology and disease. Here we review the mechanisms of Fgf signaling by focusing on genetic strategies that enable in vivo analysis. These studies support an important role for Erk1/2 as a mediator of Fgf signaling in many biological processes but have also provided strong evidence for additional signaling pathways in transmitting Fgf signaling in vivo.

The annotation of the mammalian protein coding genome is incomplete. Arbitrary open reading frame (ORF) size restriction and the absolute requirement for a methionine codon as the sole initiator of translation, have constrained identification of potentially important transcripts with non-canonical protein coding potential1,2. Using unbiased transcriptomic approaches in macrophages responding to bacterial infection, we show widespread ribosome association with a large number of RNAs that were previously annotated as “non-protein coding”. Although the ability of such non-canonical ORFs to encode functional protein is controversial3,4, we identify a plethora of novel short and non-ATG initiated ORFs with the ability to generate stable and spatially distinct proteins. Importantly, we show that the translation of a novel ORF ‘hidden’ within the long non-coding RNA Aw112010 is essential for the orchestration of mucosal immunity during both bacterial infection and colitis. Together this work expands our interpretation of the protein coding genome and demonstrates the critical nature of proteinaceous products generated from non-canonical ORFs to the immune response in vivo. We therefore propose that the misannotation of non-canonical ORF-containing genes as non-coding RNAs may obscure the essential role of a multitude of previously undiscovered protein coding genes in immunity and disease.