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The epithelial-mesenchymal transition (EMT) is the crucial step that cancer cells must pass before they can undergo metastasis. The transition requires the activity of complex functional networks that downregulate properties of the epithelial phenotype and upregulate characteristics of the mesenchymal phenotype. The networks frequently include reciprocal repressions between transcription factors (TFs) driving the EMT and microRNAs (miRs) inducing the reverse process, termed mesenchymal-epithelial transition (MET). In this work we develop four kinetic models that are based on experimental data and hypotheses describing how autocrine transforming growth factor-β (TGF-β) signal transduction induces and maintains an EMT by upregulating the TFs ZEB1 and ZEB2 which repress the expression of the miR-200b/c family members. After successful model calibration we validate our models by predicting requirements for the maintenance of the mesenchymal steady state which agree with experimental data. Finally, we apply our validated kinetic models for the design of experiments in cancer therapy. We demonstrate how steady state properties of the kinetic models, combined with data from tumor-derived cell lines of individual patients, can predict the minimal amount of an inhibitor to induce a MET. [ABSTRACT FROM AUTHOR]

Summary Hepatitis C virus (HCV) enters human hepatocytes through a multistep mechanism involving, among other host proteins, the virus receptor CD81. How CD81 governs HCV entry is poorly characterized, and CD81 protein interactions after virus binding remain elusive. We have developed a quantitative proteomics protocol to identify HCV-triggered CD81 interactions and found 26 dynamic binding partners. At least six of these proteins promote HCV infection, as indicated by RNAi. We further characterized serum response factor binding protein 1 (SRFBP1), which is recruited to CD81 during HCV uptake and supports HCV infection in hepatoma cells and primary human hepatocytes. SRFBP1 facilitates host cell penetration by all seven HCV genotypes, but not of vesicular stomatitis virus and human coronavirus. Thus, SRFBP1 is an HCV-specific, pan-genotypic host entry factor. These results demonstrate the use of quantitative proteomics to elucidate pathogen entry and underscore the importance of host protein-protein interactions during HCV invasion. [ABSTRACT FROM AUTHOR]

Dengue virus (DENV) is a human arboviral pathogen accounting for 390 million infections every year. The available vaccine has limited efficacy, and DENV-specific drugs have not been generated. To better understand DENV-host cell interaction, we employed RNA interference-based screening of the human kinome and identified fibroblast growth factor receptor 4 (FGFR4) to control the DENV replication cycle. Pharmacological inhibition of FGFR exerts a reciprocal effect by reducing DENV RNA replication and promoting the production of infectious virus particles. Addressing the latter effect, we found that the FGFR signaling pathway modulates intracellular distribution of DENV particles in a PI3K-dependent manner. Upon FGFR inhibition, virions accumulate in the trans -Golgi network compartment, where they undergo enhanced maturation cleavage of the envelope protein precursor membrane (prM), rendering virus particles more infectious. This study reveals an unexpected reciprocal role of a cellular receptor tyrosine kinase regulating DENV RNA replication and the production of infectious virions. • DENV host dependency and restriction kinases are identified through a kinome screen • FGFR4 reciprocally affects viral replication and infectivity • Inhibition of FGFR4 decreases replication and increases specific infectivity of virions • Increase in specific infectivity is linked to enhanced proteolytic cleavage of prM Cortese et al. conduct a human kinome RNAi-based screen and identify fibroblast growth factor receptor 4 (FGFR4) as a kinase that has a reciprocal effect on the DENV life cycle. Inhibition of the FGFR pathway reduces RNA replication while increasing production of infectious virus particles through enhanced proteolytic cleavage of prM. [ABSTRACT FROM AUTHOR]

Abstract: Neuroblastoma is the most common extracranial childhood tumor, comprising 15% of all childhood cancer deaths. In an initial study, we used Affymetrix oligonucleotide microarrays to analyse gene expression in 68 primary neuroblastomas and compared different data mining approaches for prediction of early relapse. Here, we performed re-analyses of the data including prolonged follow-up and applied support vector machine (SVM) algorithms and outer cross-validation strategies to improve reliability of expression profiling based predictors. Accuracy of outcome prediction was significantly improved by the use of innovative SVM algorithms on the updated data. In addition, CASPAR, a hierarchical Bayesian approach, was used to predict survival times for the individual patient based on expression profiling data. CASPAR reliably predicted event-free survival, given a cut-off time of three years. Differential expression of genes used by CASPAR to predict patient outcome was validated in an independent cohort of 117 neuroblastomas. In conclusion, we show here for the first time that reanalysis of microarray data using improved methodology, state-of-the-art performance tests and updated follow-up data improves prognosis prediction, and may further improve risk stratification of individual patients. [Copyright &y& Elsevier]

The incidence of breast cancer increases with age until menopause, and breast cancer is more aggressive in younger women. The existence of epidemiological links between breast cancer and aging indicates that both processes share some common mechanisms of development. Oxidative stress is associated with both cancer susceptibility and aging. Here we observed that ERBB2-positive breast cancer, which developed in genetically heterogeneous ERBB2-positive transgenic mice generated by a backcross, is more aggressive in chronologically younger than in older mice (differentiated by the median survival of the cohort that was 79 weeks), similar to what occurs in humans. In this cohort, we estimated the oxidative biological age using a mathematical model that integrated several subphenotypes directly or indirectly related to oxidative stress. The model selected the serum levels of HDL-cholesterol and magnesium and total AKT1 and glutathione concentrations in the liver. The grade of aging was calculated as the difference between the predicted biological age and the chronological age. This comparison permitted the identification of biologically younger and older mice compared with their chronological age. Interestingly, biologically older mice developed more aggressive breast cancer than the biologically younger mice. Genomic regions on chromosomes 2 and 15 linked to the grade of oxidative aging were identified. The levels of expression of Zbp1 located on chromosome 2, a gene related to necroptosis and inflammation, positively correlated with the grade of aging and tumour aggressiveness. Moreover, the pattern of gene expression of genes linked to the inflammation and the response to infection pathways was enriched in the livers of biologically old mice. This study shows part of the complex interactions between breast cancer and aging. [ABSTRACT FROM AUTHOR]

Cerebral cavernous malformations (CCM) are vascular lesions of the central nervous system that can cause headaches, seizures and hemorrhagic stroke. Disease-associated mutations have been identified in three genes: CCM1/KRIT1 , CCM2 and CCM3/PDCD10 . The precise proportion of deep-intronic variants in these genes and their clinical relevance is yet unknown. Here, a long-range PCR (LR-PCR) approach for target enrichment of the entire genomic regions of the three genes was combined with next generation sequencing (NGS) to screen for coding and non-coding variants. NGS detected all six CCM1/KRIT1 , two CCM2 and four CCM3/PDCD10 mutations that had previously been identified by Sanger sequencing. Two of the pathogenic variants presented here are novel. Additionally, 20 stringently selected CCM index cases that had remained mutation-negative after conventional sequencing and exclusion of copy number variations were screened for deep-intronic mutations. The combination of bioinformatics filtering and transcript analyses did not reveal any deep-intronic splice mutations in these cases. Our results demonstrate that target enrichment by LR-PCR combined with NGS can be used for a comprehensive analysis of the entire genomic regions of the CCM genes in a research context. However, its clinical utility is limited as deep-intronic splice mutations in CCM1/KRIT1 , CCM2 and CCM3/PDCD10 seem to be rather rare. [ABSTRACT FROM AUTHOR]

Malaria, caused by protozoan Plasmodium parasites, remains a prevalent infectious human disease due to the lack of an efficient and safe vaccine. This is directly related to the persisting gaps in our understanding of the parasite's interactions with the infected host, especially during the clinically silent yet essential liver stage of Plasmodium development. Previously, we and others showed that genetically attenuated parasites (GAP) that arrest in the liver induce sterile immunity, but only upon multiple administrations. Here, we comprehensively studied hepatic gene and miRNA expression in GAP-injected mice, and found both a broad activation of IFNγ-associated pathways and a significant increase of murine microRNA-155 (miR-155), that was especially pronounced in non-parenchymal cells including liver-resident macrophages (Kupffer cells). Remarkably, ectopic upregulation of this miRNA in the liver of mice using robust hepatotropic adeno-associated virus 8 (AAV8) vectors enhanced GAP's protective capacity substantially. In turn, this AAV8-mediated miR-155 expression permitted a reduction of GAP injections needed to achieve complete protection against infectious parasite challenge from previously three to only one. Our study highlights a crucial role of mammalian miRNAs in Plasmodium liver infection in vivo and concurrently implies their great potential as future immune-augmenting agents in improved vaccination regimes against malaria and other diseases. [ABSTRACT FROM AUTHOR]

RIG-I is a major innate immune sensor for viral infection, triggering an interferon (IFN)-mediated antiviral response upon cytosolic detection of viral RNA. Double-strandedness and 5′-terminal triphosphates were identified as motifs required to elicit optimal immunological signaling. However, very little is known about the response dynamics of the RIG-I pathway, which is crucial for the ability of the cell to react to diverse classes of viral RNA while maintaining self-tolerance. In the present study, we addressed the molecular mechanism of RIG-I signal detection and its translation into pathway activation. By employing highly quantitative methods, we could establish the length of the double-stranded RNA (dsRNA) to be the most critical determinant of response strength. Size exclusion chromatography and direct visualization in scanning force microscopy suggested that this was due to cooperative oligomerization of RIG-I along dsRNA. The initiation efficiency of this oligomerization process critically depended on the presence of high affinity motifs, like a 5′-triphosphate. It is noteworthy that for dsRNA longer than 200 bp, internal initiation could effectively compensate for a lack of terminal triphosphates. In summary, our data demonstrate a very flexible response behavior of the RIG-I pathway, in which sensing and integration of at least two distinct signals, initiation efficiency and double strand length, allow the host cell to mount an antiviral response that is tightly adjusted to the type of the detected signal, such as viral genomes, replication intermediates, or small by-products. [ABSTRACT FROM AUTHOR]

Standard cancer therapy targets tumor cells without considering possible damage on the tumor microenvironment that could impair therapy response. In rectal cancer patients we find that inflammatory cancer-associated fibroblasts (iCAFs) are associated with poor chemoradiotherapy response. Employing a murine rectal cancer model or patient-derived tumor organoids and primary stroma cells, we show that, upon irradiation, interleukin-1α (IL-1α) not only polarizes cancer-associated fibroblasts toward the inflammatory phenotype but also triggers oxidative DNA damage, thereby predisposing iCAFs to p53-mediated therapy-induced senescence, which in turn results in chemoradiotherapy resistance and disease progression. Consistently, IL-1 inhibition, prevention of iCAFs senescence, or senolytic therapy sensitizes mice to irradiation, while lower IL-1 receptor antagonist serum levels in rectal patients correlate with poor prognosis. Collectively, we unravel a critical role for iCAFs in rectal cancer therapy resistance and identify IL-1 signaling as an attractive target for stroma-repolarization and prevention of cancer-associated fibroblasts senescence. [Display omitted] • Inflammatory CAFs are associated with poor prognosis in rectal cancer • Oxidative DNA damage in iCAFs is a pre-requisite for irradiation-induced senescence • IL-1 inhibition prevents iCAF senescence upon irradiation, improving therapy response • IL-1 signaling is enhanced by low serum IL-1RA in patients with poor prognosis Nicolas et al. highlight the important role of inflammatory cancer-associated fibroblasts (iCAFs) for therapy response of rectal cancer patients. They demonstrate that IL-1-dependent signaling elevates oxidative DNA damage in iCAFs, which upon irradiation undergo senescence. This causes tissue remodeling and therapy resistance that can be overcome by inhibiting IL-1. [ABSTRACT FROM AUTHOR]