Summary Smoothened (SMO) inhibitors recently entered clinical trials for sonic-hedgehog-driven medulloblastoma (SHH-MB). Clinical response is highly variable. To understand the mechanism(s) of primary resistance and identify pathways cooperating with aberrant SHH signaling, we sequenced and profiled a large cohort of SHH-MBs (n = 133). SHH pathway mutations involved PTCH1 (across all age groups), SUFU (infants, including germline), and SMO (adults). Children >3 years old harbored an excess of downstream MYCN and GLI2 amplifications and frequent TP53 mutations, often in the germline, all of which were rare in infants and adults. Functional assays in different SHH-MB xenograft models demonstrated that SHH-MBs harboring a PTCH1 mutation were responsive to SMO inhibition, whereas tumors harboring an SUFU mutation or MYCN amplification were primarily resistant.
Liver fibrosis is orchestrated by a complex network of signaling pathways regulating the deposition of extracellular matrix proteins during fibrogenesis. MicroRNAs (miRNAs) represent a family of small noncoding RNAs controlling translation and transcription of many genes. Recently, miRNAs have been suggested to crucially modulate cellular processes in the liver such as hepatocarcinogenesis. However, their role in liver fibrosis is not well understood. We systematically analyzed the regulation of miRNAs in a mouse model of carbon tetrachloride-induced hepatic fibrogenesis (CCl 4) by gene array analysis, which revealed a panel of miRNA that were specifically regulated in livers of mice undergoing hepatic fibrosis. Within those, all three members of the miR-29-family were significantly down-regulated in livers of CCl 4-treated mice as well as in mice that underwent bile duct ligation. Specific regulation of miR-29 members in murine fibrosis models correlated with lower expression of miR-29 in livers from patients with advanced liver fibrosis. Moreover, patients with advanced liver cirrhosis showed significantly lower levels of miR-29a in their serum when compared with healthy controls or patients with early fibrosis. On a cellular level, down-regulation of miR-29 in murine hepatic stellate cells (HSCs) was mediated by transforming growth factor beta (TGF-b) as well as inflammatory signals, namely, lipopolysaccharide (LPS) and nuclear factor kappa B (NF-jB). Furthermore, overexpression of miR-29b in murine HSC resulted in down-regulation of collagen expression. Conclusion: Our data indicate that miR-29 mediates the regulation of liver fibrosis and is part of a signaling nexus involving TGF-band NF-jB-dependent down-regulation of miR-29 family members in HSC with subsequent up-regulation of extracellular matrix genes. Thus they may represent targets for novel therapeutic strategies against hepatic fibrogenesis and also might evolve as biomarkers in the diagnosis of liver fibrosis.
Pilocytic astrocytoma, the most common childhood brain tumor1, is typically associated with mitogen-activated protein kinase (MAPK) pathway alterations2. Surgically inaccessible midline tumors are therapeutically challenging, showing sustained tendency for progression3 and often becoming a chronic disease with substantial morbidities4. Here we describe whole-genome sequencing of 96 pilocytic astrocytomas, with matched RNA sequencing (n=73), conducted by the International Cancer Genome Consortium (ICGC) PedBrain Tumor Project. We identified recurrent activating mutations in FGFR1 and PTPN11 and novel NTRK2 fusion genes in non-cerebellar tumors. New BRAF activating changes were also observed. MAPK pathway alterations affected 100% of tumors analyzed, with no other significant mutations, indicating pilocytic astrocytoma as predominantly a single-pathway disease. Notably, we identified the same FGFR1 mutations in a subset of H3F3A-mutated pediatric glioblastoma with additional alterations in NF15. Our findings thus identify new potential therapeutic targets in distinct subsets of pilocytic astrocytoma and childhood glioblastoma.
Fibrosis is a common pathology in cardiovascular disease1. In the heart, fibrosis causes mechanical and electrical dysfunction1,2 and in the kidney, it predicts the onset of renal failure3. Transforming growth factor β1 (TGFβ1) is the principal pro-fibrotic factor4,5, but its inhibition is associated with side effects due to its pleiotropic roles6,7. We hypothesized that downstream effectors of TGFβ1 in fibroblasts could be attractive therapeutic targets and lack upstream toxicity. Here we show, using integrated imaging–genomics analyses of primary human fibroblasts, that upregulation of interleukin-11 (IL-11) is the dominant transcriptional response to TGFβ1 exposure and required for its pro-fibrotic effect. IL-11 and its receptor (IL11RA) are expressed specifically in fibroblasts, in which they drive non-canonical, ERK-dependent autocrine signalling that is required for fibrogenic protein synthesis. In mice, fibroblast-specific Il11 transgene expression or Il-11 injection causes heart and kidney fibrosis and organ failure, whereas genetic deletion of Il11ra1 protects against disease. Therefore, inhibition of IL-11 prevents fibroblast activation across organs and species in response to a range of important pro-fibrotic stimuli. These results reveal a central role of IL-11 in fibrosis and we propose that inhibition of IL-11 is a potential therapeutic strategy to treat fibrotic diseases.
Artemisinin and its derivatives (ARTs) are the frontline drugs against malaria, but resistance is jeopardizing their effectiveness. ART resistance is mediated by mutations in the parasite’s Kelch13 protein, but Kelch13 function and its role in resistance remain unclear. In this study, we identified proteins located at a Kelch13-defined compartment. Inactivation of eight of these proteins, including Kelch13, rendered parasites resistant to ART, revealing a pathway critical for resistance. Functional analysis showed that these proteins are required for endocytosis of hemoglobin from the host cell. Parasites with inactivated Kelch13 or a resistance-conferring Kelch13 mutation displayed reduced hemoglobin endocytosis. ARTs are activated by degradation products of hemoglobin. Hence, reduced activity of Kelch13 and its interactors diminishes hemoglobin endocytosis and thereby ART activation, resulting in parasite resistance.
Summary Paragraph Medulloblastoma is a highly malignant paediatric brain tumour currently treated with a combination of surgery, radiation, and chemotherapy, posing a considerable burden of toxicity to the developing child. Genomics has illuminated the extensive intertumoural heterogeneity of medulloblastoma, identifying four distinct molecular subgroups. Group 3 and Group 4 subgroup medulloblastomas account for the majority of paediatric cases; yet, oncogenic drivers for these subtypes remain largely unidentified. Here we describe a series of prevalent, highly disparate genomic structural variants, restricted to Groups 3 and 4, resulting in specific and mutually exclusive activation of the growth factor independent 1 family protooncogenes, GFI1 and GFI1B. Somatic structural variants juxtapose GFI1/GFI1B coding sequences proximal to active enhancer elements, including super-enhancers, instigating oncogenic activity. Our results, supported by evidence from mouse models, identify GFI1 and GFI1B as prominent medulloblastoma oncogenes and implicate ‘enhancer hijacking’ as an efficient mechanism driving oncogene activation in a childhood cancer.
Integration of genome-wide expression profiling with linkage analysis is a new approach to identifying genes underlying complex traits. We applied this approach to the regulation of gene expression in the BXH/HXB panel of rat recombinant inbred strains, one of the largest available rodent recombinant inbred panels and a leading resource for genetic analysis of the highly prevalent metabolic syndrome. In two tissues important to the pathogenesis of the metabolic syndrome, we mapped cis- and trans-regulatory control elements for expression of thousands of genes across the genome. Many of the most highly linked expression quantitative trait loci are regulated in cis, are inherited essentially as monogenic traits and are good candidate genes for previously mapped physiological quantitative trait loci in the rat. By comparative mapping we generated a data set of 73 candidate genes for hypertension that merit testing in human populations. Mining of this publicly available data set is expected to lead to new insights into the genes and regulatory pathways underlying the extensive range of metabolic and cardiovascular disease phenotypes that segregate in these recombinant inbred strains.
Nucleoid-associated proteins (NAPs) are global regulators of gene expression in Escherichia coli, which affect DNA conformation by bending, wrapping and bridging the DNA. Two of these—H-NS and Fis—bind to specific DNA sequences and structures. Because of their importance to global gene expression, the binding of these NAPs to the DNA was previously investigated on a genome-wide scale using ChIP-chip. However, variation in their binding profiles across the growth phase and the genome-scale nature of their impact on gene expression remain poorly understood. Here, we present a genome-scale investigation of H-NS and Fis binding to the E. coli chromosome using chromatin immunoprecipitation combined with high-throughput sequencing (ChIP-seq). By performing our experiments under multiple time-points during growth in rich media, we show that the binding regions of the two proteins are mutually exclusive under our experimental conditions. H-NS binds to significantly longer tracts of DNA than Fis, consistent with the linear spread of H-NS binding from high- to surrounding lower-affinity sites; the length of binding regions is associated with the degree of transcriptional repression imposed by H-NS. For Fis, a majority of binding events do not lead to differential expression of the proximal gene; however, it has a significant indirect effect on gene expression partly through its effects on the expression of other transcription factors. We propose that direct transcriptional regulation by Fis is associated with the interaction of tandem arrays of Fis molecules to the DNA and possible DNA bending, particularly at operon-upstream regions. Our study serves as a proof-of-principle for the use of ChIP-seq for global DNA-binding proteins in bacteria, which should become significantly more economical and feasible with the development of multiplexing techniques.
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