The viral defense gene RNaseL acts as a regeneration repressor
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ABSTRACT: Normal Human Epidermal Keratinocytes treated with either Q-VD-OPh or vehicle,were prepared for protein analysis using proteomics approach.Protein expression changes were discussed.
Project description:Mammalian injury responses are predominantly characterized by fibrosis and scarring rathe than functional regeneration.This limited regenerative capacity in mammals could reflect loss of pro-regeneration programs or active suppression by genes functioning akin to tumor suppressors.
Project description:RNA sequencing of wild-type or Interferon Alpha receptor 1 Knockout MEF cells treated with DMSO or the Broad-spectrum Caspase Inhibitor Q-VD-OPh The mechanism by which cells undergo death determines whether dying cells trigger inflammatory responses or remain immunologically silent. Mitochondria play a central role in the induction of cell death, as well as in immune signaling pathways. Here, we identify of a mechanism by which mitochondria and downstream pro-apoptotic caspases regulate the activation of antiviral immunity. In the absence of active caspases, mitochondrial outer membrane permeabilization by Bax and Bak results in the expression of type I interferons (IFNs). This induction is mediated by mitochondrial DNA-dependent activation of the cGAS/STING pathway and results in the establishment of a potent state of viral resistance. Our results show that mitochondria have the capacity to simultaneously expose a cell-intrinsic inducer of the IFN response, and to inactivate this response in a caspase-dependent manner. This mechanism provides a dual control, which determines whether mitochondria initiate an immunologically silent or a pro-inflammatory type of cell death. To determine whether the pharmacological inhibition of caspases could recapitulate the phenotype caused by genetic deficiencies, we treated WT MEFs with broad-spectrum inhibitors of caspases (Q-VD-OPH). The inhibitor induced an increased expression of ISGs, similar to the effect of caspase or Apaf-1 deficiency. Next, we compared the transcriptional changes induced by caspase inhibition in WT and IFNAR1 KO cells, which lack a critical subunit of the receptor for IFNα/β (Muller et al., 1994). We observed that the absence of the IFNAR receptor abrogated the transcriptional response of the cells to caspase inhibition by Q-VD-OPH, demonstrating the role of type I IFNs in this response. RNA was extracted from duplicate samples and libraries generated for sequencing using the directional RNA-Seq library prep at the Yale Center for Genome Analysis. Libraries were sequenced using a Hiseq2500 sequencer to generate 76bp single-end reads. Duplicate samples were analyzed for each condition.
Project description:Congenital infection of Trypanosoma cruzi allows transmission of this parasite through generations. Despite the problematic that this entails, little is known about the placenta environment genetic response produced against infection. We performed functional genomics by microarray analysis in C57Bl/6J mice comparing placentas from uninfected animals and from animals infected with two different T. cruzi strains: K98, a clone of the non-lethal myotropic CA-I strain (TcI), and VD (TcVI), isolated from a human case of congenital infection. Analysis of networks by GeneMANIA of differentially expressed genes showed that âSecretory Granuleâ was a pathway down-regulated in both infected groups, whereas âInnate Immune Responseâ and âResponse to Interferon-gammaâ were pathways up-regulated in VD infection but not in K98. Applying another approach, the GSEA algorithm that detects small changes in predetermined gene sets, we found that metabolic processes, transcription and macromolecular transport were down-regulated in infected placentas environment and some pathways related to cascade signaling had opposite regulation: over-represented in VD and down-regulated in K98 group. We also have found a stronger placental tropism by VD strain, by detection of parasite DNA and RNA, suggesting living parasites in that tissue. Our study is the first one to describe in a murine model the genetic response of placental environment to T. cruzi infection and suggests the development of a strong immune response, parasite genotype-dependent, to the detriment of cellular metabolism. Total RNA obtained from isolated placentas from mice with chronic T. cruzi infection (K98 or VD strains) at 18.5 days of gestation compared to uninfected control mice.
Project description:Objective: The aim of this study was to identify novel diagnostic biomarkers of diabetic vascular dementia (DVD) and unraveling the underlying mechanisms using mass spectrometry (MS). Methods: Label-Free LC-MS/MS proteomics analysis was applied to urine samples from four groups including 14 vascular dementia CC(VD), 22 type 2 diabetes mellitus (T2DM), 12 DVD, 21 Normal Control (NC). Searching the MS data by Proteome Discoverer software, protein abundances were analyzed qualitatively and quantitatively and compared between these groups. Combining bioinformatics analysis using GO, pathway crosstalk analysis using KEGG, PPI network analysis using STRING and literature searching, the differentially expressed proteins (DEPs) of DVD can be comprehensively judged and were further quantified by receiver operating characteristic (ROC) curve methods. Results: The proteomic findings showed quantitative changes in DVD was compared to NC, T2DM, and VD groups; among 7,527 identified urine proteins, 1222, 1152, and 1180 of the proteins displayed quantitative changes unique to DVD vs NC, T2DM, and VD, respectively, including 481 overlapped common DEPs. Then 9 unique proteins and 2 composite markers (CM) were confirmed by a ROC curve method. Conclusion: This study provided a novel insight into the potential pathogenesis of DVD and elucidated a method for the earlier detection.
Project description:Comparison of the hepatic transcriptomes for two half-sib-families of European sea bass fed on vegetable and fish diet. These two half-sib-families exhibit similar growth on fish diet while significantly different on vegetable diet. The aim of the study is to point out the large panel of metabolic and physiological effects induced by total substitution of both fish meal and fish oil in the diets of European sea bass and to reveal physiological characteristics associated to the two half-sib-families. Fish from both two half-sib-families (G and g) were fed a a fish diet (FD) or a vegetable diet (VD) diet for 9 months. Five to eight independent experiments were performed for each experimental groups (G-FD; G-VD; g-FD; g-VD) using different fishes for each experiment.
Project description:To clarify the relationship between VD deficiency and airway epithelial disorders, we investigated the effects of VD on 2.5-D airway epithelial organoids cultured at the air–liquid interface (ALI) and performed an in-silico analysis. We performed gene expression profiling analysis using data obtained from RNA-seq of 14 mature airway organoids with 1,25(OH)2D3 treatment followed by HI1N1 infection.
Project description:Our goal is to find new genes regulated by p21 in human primary cells . To get it we carried out a gene expression profiling in two different models, human myeloid leukemia K562 cells and human keratinocytes both of them with conditional expression of p21. In order to identify genes specifically modulated by p21 we compared with the cell line with overexpression of p27, because p21 and p27 belong to the same gene family and regulated the same genes specially in cell cycle. So, our intention is to identify only genes regulated by p21 and not p27. In order to confirm these results we studied the p21-dependent repression of mitotic genes in a different cellular system. We chose human primary keratinocytes because they are non-tumorigenic, non-immortalized and epithelial cells, in contrast to human myeloid leukemia K562 cells. Human primary keratinocytes were infected with recombinant adenoviruses expressing the full-length p21 protein. A dramatic increase in p21 in infected keratinocytes was demonstrated by RT-qPCR (as we show in the manuscript). As controls, we also infected the keratinocytes with adenovirus carrying the genes for p27 which overexpression was also confirmed by RT-qPCR (as we show in the manuscript). We prepared RNA 24 h after infection and performed large-scale expression assay using the Afftymetrix platform. The clustering analysis revealed that p21 provoked the down-regulation of a number genes involved in cell cycle control not shared by cells expressing p27 (as we show in the manuscript). Our goal, has been getting genes regulated more strongly by p21 and not by p27 in cell cycle and mitosis. Our result are supported because we have found the same genes in two different models and also we have validated (by RT-qPCR) more than 20 cell cycle and mitotic genes, found in our affymetrix arrays. Also we have found the region of p21 that is sufficient for gene regulation and for one gene we have described as p21 bind to the promoter. Finally, we have discussed in our manuscript how p21 can do this regulation by bioinformatic analysis of p21-target genes. The success of this study is to describe a new role of p21 as a transcriptional co-repressor in some systems.
Project description:The skin serves as a barrier to the environment and is constantly exposed to viral pathogens. Whether viral pathogens contribute to the oncogenesis of rare skin cancers has not been systematically explored. Here, we analyzed 18 skin cancer types by exploiting off-target reads from commonly available next-generation sequencing data. We identified human papillomavirus 42 (HPV42) in all digital papillary adenocarcinoma (DPA) cases but not in any of 11,091 tumors from common cancer types. HPV42 was previously not described as an oncogenic driver, yet we show that HPV42 recapitulates the molecular hallmarks of oncogenic HPVs. Using a machine learning approach, we identified a conserved transcriptomic fingerprint of HPV transformation common to oncogenic HPVs, including HPV42. Collectively, our results establish HPV42 as an oncogenic HPV, which has implications for the diagnosis and treatment for DPA patients.
Project description:Differentiating agents have been proposed to overcome the impaired cellular differentiation in acute myeloid leukemia (AML). However, only the combinations of all-trans retinoic acid or arsenic trioxide with chemotherapy have been successful, and only in treating acute promyelocytic leukemia (also called AML3). Here we showed that iron homeostasis is an effective target in the treatment of AML. Iron chelating therapy induces the differentiation of leukemia blasts and normal bone marrow precursors into monocytes/macrophages in a manner involving modulation of reactive oxygen species expression and the activation of mitogen-activated protein kinases (MAPK). Thirty percent of the genes most strongly induced by iron deprivation are also targeted by vitamin D3 (VD), a well-known differentiating agent. Iron chelating agents induce expression and phosphorylation of the VD receptor, and iron deprivation and VD act synergistically. VD magnifies activation of MAPK JNK and the induction of VD receptor target genes. When used to treat one AML patient refractory to chemotherapy, the combination of iron chelating agents and VD resulted in reversal of pancytopenia and blast differentiation. We propose that iron availability modulates myeloid cell commitment, and that targeting this cellular differentiation pathway together with conventional differentiating agents provides new therapeutic modalities for AML. This study presents 8 arrays 4x44K Agilent.