RESUMEN
Microbes influence cancer initiation, progression and therapy responsiveness. IL-17 signaling contributes to gut barrier immunity by regulating microbes but also drives tumor growth. A knowledge gap remains regarding the influence of enteric IL-17-IL-17RA signaling and their microbial regulation on the behavior of distant tumors. We demonstrate that gut dysbiosis induced by systemic or gut epithelial deletion of IL-17RA induces growth of pancreatic and brain tumors due to excessive development of Th17, primary source of IL-17 in human and mouse pancreatic ductal adenocarcinoma, as well as B cells that circulate to distant tumors. Microbial dependent IL-17 signaling increases DUOX2 signaling in tumor cells. Inefficacy of pharmacological inhibition of IL-17RA is overcome with targeted microbial ablation that blocks the compensatory loop. These findings demonstrate the complexities of IL-17-IL-17RA signaling in different compartments and the relevance for accounting for its homeostatic host defense function during cancer therapy.
Asunto(s)
Interleucina-17 , Neoplasias Pancreáticas , Ratones , Animales , Humanos , Receptores de Interleucina-17/genética , Ratones Noqueados , Transducción de Señal , Neoplasias Pancreáticas/patologíaRESUMEN
Adenosine deaminase acting on RNA 1 (ADAR1) is an RNA-binding protein that deaminates adenosine (A) to inosine (I). A-to-I editing alters post-transcriptional RNA processing, making ADAR1 a crucial regulator of gene expression. Consequently, Adar1 has been implicated in organogenesis. To determine the role of Adar1 in pancreatic development and homeostasis, we conditionally deleted Adar1 from the murine pancreas (Ptf1aCre/+; Adar1Fl/Fl). The resulting mice had stunted growth, likely due to malabsorption associated with exocrine pancreatic insufficiency. Analyses of pancreata revealed ductal cell expansion, heightened interferon-stimulated gene expression and an increased influx of immune cells. Concurrent deletion of Adar1 and Mavs, a signaling protein implicated in the innate immune pathway, rescued the degenerative phenotype and resulted in normal pancreatic development. Taken together, our work suggests that the primary function of Adar1 in the pancreas is to prevent aberrant activation of the Mavs-mediated innate immune pathway, thereby maintaining pancreatic homeostasis.
Asunto(s)
Páncreas Exocrino , Animales , Ratones , Páncreas Exocrino/metabolismo , Interferones/genética , Interferones/metabolismo , Fenotipo , Adenosina Desaminasa/genética , Adenosina Desaminasa/metabolismoRESUMEN
Conventional genetically engineered mouse models (GEMM) are time-consuming, laborious, and offer limited spatiotemporal control. Here, we describe the development of a streamlined platform for in vivo gene activation using CRISPR activation (CRISPRa) technology. Unlike conventional GEMMs, this model system allows for flexible, sustained, and timed activation of one or more target genes using single or pooled lentiviral guides. Myc and Yap1 were used as model oncogenes to demonstrate gene activation in primary pancreatic organoid cultures in vitro and enhanced tumorigenic potential in Myc-activated organoids when transplanted orthotopically in vivo. Implementation of this model as an autochthonous lung cancer model showed that transduction-mediated activation of Myc led to accelerated tumor progression and significantly reduced overall survival relative to nontargeted tumor controls. Furthermore, Myc activation led to the acquisition of an immune suppressive, "cold" tumor microenvironment. Cross-species validation of these results using publicly available RNA/DNA-seq datasets linked MYC to a previously described immunosuppressive molecular subtype in patient tumors, thus identifying a patient cohort that may benefit from combined MYC- and immune-targeted therapies. Overall, this work demonstrates how CRISPRa can be used for rapid functional validation of putative oncogenes and may allow for the identification and evaluation of potential metastatic and oncogenic drivers through competitive screening. SIGNIFICANCE: A streamlined platform for programmable CRISPR gene activation enables rapid evaluation and functional validation of putative oncogenes in vivo.
Asunto(s)
Adenocarcinoma del Pulmón , Neoplasias Pulmonares , Proteínas Proto-Oncogénicas c-myc , Adenocarcinoma del Pulmón/genética , Animales , Carcinogénesis/genética , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Humanos , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patología , Ratones , Oncogenes/genética , Proteínas Proto-Oncogénicas c-myc/genética , Microambiente Tumoral/genéticaRESUMEN
Despite efforts in understanding its underlying mechanisms, the etiology of chromosomal instability (CIN) remains unclear for many tumor types. Here, we identify CIN initiation as a previously undescribed function for APOBEC3A (A3A), a cytidine deaminase upregulated across cancer types. Using genetic mouse models of pancreatic ductal adenocarcinoma (PDA) and genomics analyses in human tumor cells we show that A3A-induced CIN leads to aggressive tumors characterized by enhanced early dissemination and metastasis in a STING-dependent manner and independently of the canonical deaminase functions of A3A. We show that A3A upregulation recapitulates numerous copy number alterations commonly observed in patients with PDA, including co-deletions in DNA repair pathway genes, which in turn render these tumors susceptible to poly (ADP-ribose) polymerase inhibition. Overall, our results demonstrate that A3A plays an unexpected role in PDA as a specific driver of CIN, with significant effects on disease progression and treatment.
Asunto(s)
Citidina Desaminasa , Neoplasias Pancreáticas , Animales , Inestabilidad Cromosómica/genética , Citidina Desaminasa/genética , Humanos , Ratones , Neoplasias Pancreáticas/genética , Proteínas/genética , Neoplasias PancreáticasRESUMEN
The transferrin receptor is the primary means of iron importation for most mammalian cells and understanding its regulatory mechanisms is relevant to hematologic, oncologic, and other disorders in which iron homeostasis is perturbed. The 3' UTR of the transferrin receptor mRNA contains an instability element that is protected from degradation during iron depletion through interactions of iron regulatory proteins (IRPs) with five iron-responsive elements (IREs). The structural features required for degradation and the site of IRP binding required for in situ protection remain unclear. An RNA-CLIP strategy is described here that identifies the predominant site of IRP-1 interaction within a nontransformed primary cell line. This approach avoided complications associated with the use of elevated concentrations of protein and/or mRNA and detected interactions within the native environment of the mRNA. A compensatory mutagenesis strategy indicates that the instability element at minimum consists of three non-IRE stem-loops that can function additively, suggesting that they are not forming one highly interdependent structure. Although the IREs are not essential for instability, they enhance instability when IRP interactions are inhibited. These results are supportive of a mechanism for a graded response to the intracellular iron resulting from a progressive loss of IRP protection.