Project description:The closely related protozoan parasites Toxoplasma gondii and Neospora caninum display similar life cycles, subcellular ultrastructure, invasion mechanisms, metabolic pathways, and genome organization, but differ in their host range and disease pathogenesis. Type II (γ) interferon has long been known to be the major mediator of innate and adaptive immunity to Toxoplasma infection, but genome-wide expression profiling of infected host cells indicates that Neospora is a potent activator of the type I (α/β) interferon pathways typically associated with antiviral responses. Infection of macrophages from mice with targeted deletions in various innate sensing genes demonstrates that host responses to Neospora are dependent on the toll-like receptor Tlr3 and the adapter protein Trif. Consistent with this observation, RNA from Neospora elicits type I interferon responses when targeted to the host endo-lysosomal system. While live Toxoplasma fails to induce type I interferon, heat-killed parasites do trigger this response, and co-infection studies reveal that T. gondii actively suppresses the production of type I interferon. These findings reveal that eukaryotic pathogens can be potent inducers of type I interferon and that some parasite species, like Toxoplasma gondii, have evolved mechanisms to suppress this response. Human foreskin fibroblasts (HFF; line BJ-5ta) were cultured to confluency in T25 flasks, infected with one representative of each of the three architypial strains of Toxoplasma gondii: GT1 (type I), Prugniaud (type II) and VEG (type III), or the closely related parasite species, Neospora caninum (strain Nc-Liv). RNA was collected from biological replicates for expression profiling by microarray. Uninfected HFF cells were used as a reference.
Project description:The closely related protozoan parasites Toxoplasma gondii and Neospora caninum display similar life cycles, subcellular ultrastructure, invasion mechanisms, metabolic pathways, and genome organization, but differ in their host range and disease pathogenesis. Type II (?) interferon has long been known to be the major mediator of innate and adaptive immunity to Toxoplasma infection, but genome-wide expression profiling of infected host cells indicates that Neospora is a potent activator of the type I (?/?) interferon pathways typically associated with antiviral responses. Infection of macrophages from mice with targeted deletions in various innate sensing genes demonstrates that host responses to Neospora are dependent on the toll-like receptor Tlr3 and the adapter protein Trif. Consistent with this observation, RNA from Neospora elicits type I interferon responses when targeted to the host endo-lysosomal system. While live Toxoplasma fails to induce type I interferon, heat-killed parasites do trigger this response, and co-infection studies reveal that T. gondii actively suppresses the production of type I interferon. These findings reveal that eukaryotic pathogens can be potent inducers of type I interferon and that some parasite species, like Toxoplasma gondii, have evolved mechanisms to suppress this response. In vitro cultures of bone marrow-derived macrophages from WT or IFNAR2-/- mice were infected with either Toxoplasma gondii (VEG strain) or Neospora caninum (Nc2 strain) for 17 hours. RNA was collected from biological replicates for expression profiling by microarray. Uninfected controls for both WT and IFNAR2-/- were used as a reference.
Project description:miRNA expression profiles of WI38 primary human fibroblasts with an active or inactive p53. Cells were compared under normal untreated conditions (young and proliferating cells), after DNA damage with Doxorubicin, and upon entry into replicative senescence. Keywords: miRNA, WI-38, p53, GSE56, Senescence, Doxorubicin, Cancer, DNA-damage, fibroblasts. 6 samples of WI38 cells were analyzed on 12 Exiqon miRcurry LNA arrays in biological duplicates (2 different cell culture plates for each experimental condition). The six conditions included: 1. [Con_Young] - Primary Young WI38 cells (passage 20) with a control retroviral vector (pLXSN-NEO). Untreated. 2. [GSE_Young] -Primary Young WI38 cells (passage 20) with a retroviral vector encoding for the p53-inactivating peptide GSE56 (pLXSN-NEO-GSE56).Untreated. 3. [Con_Dox] -Primary Young WI38 cells (passage 20) with a control retroviral vector (pLXSN-NEO). Treated with Doxorubicin (0.2 micrograms/ml) for 24 hours). 4. [GSE_Dox] Primary Young WI38 cells (passage 20) with a retroviral vector encoding for the p53-inactivating peptide GSE56 (pLXSN-NEO-GSE56). Treated with Doxorubicin (0.2 micrograms/ml) for 24 hours). 5. [Con_Old] - Sesescent WI38 cells (passage 30) with a control retroviral vector (pLXSN-NEO). Untreated. 6. [GSE_Old] - Senescent WI38 cells (passage 26) with a retroviral vector encoding for the p53-inactivating peptide GSE56 (pLXSN-NEO-GSE56).Untreated. RNA was extracted with TRI-Reagent and sent for labeling and hybridization in Exiqon laboratories (In Denamark). Samples were labeled with Cy5. Reference sample (Cy3) was an RNA mix of all samples. Log2 for Ratio(Cy5/Cy3) was used for further analysis.
Project description:This dataset consists of two individual sample-multiplexing (MULTI-seq) single-cell RNA sequencing experiments, MB10x01 and MB10x02. Single-cell RNA sequencing (10X Genomics) analyses were performed on a microfluidic 3D in vitro blood-brain-barrier model (containing primary human brain microvascular endothelial cells, brain vascular pericytes, and astrocytes) perfused with P. falciparum egress product (MB10x01) or P. falciparum-infected red blood cells (RBC) (MB10x02). Dataset MB10x01 included two samples multiplexed by MULTI-seq sample barcoding (TCCTCGAA for control RBC lysate, ATGCGATG for P. falciparum egress product). P. falciparum egress product was obtained by letting tightly synchronized P. falciparum-infected RBC egress in media used for perfusions (5x10^7 infected RBC/ml). 3D blood-brain-barrier models perfused with P. falciparum egress products were incubated for 24 hours and compared to a control perfused with uninfected red blood cell lysate. MULTI-seq barcoding (McGinnis et al.Ê2019) was used for sample-barcoding of these two conditions, and the dataset contains cDNA (transcriptome) and sample barcode read files. Dataset MB10x02 included three samples multiplexed by MULTI-seq sample barcoding (GCTATGCA for control RBC, CGATACTG for Trophozoite stage, TACGCAGT for Schizont stage). 3D blood-brain-barrier models were perfused for 30 minutes with P. falciparum-infected RBC in the Trophozoite stage (26-34 hours post invasion) or Schizont stage (42-48 hours post invasion) (5x10^7 infected RBC/ml). After a 20-minute wash, the 3D blood-brain-barrier models were incubated with the bound P. falciparum-infected RBC for 6 hours and compared to uninfected RBC perfused controls. MULTI-seq barcoding was used for sample-barcoding of the three conditions, and the dataset contains cDNA (transcriptome) and sample barcode read files.
Project description:The poly(A)+ and poly(A)− fractions of interacting and non-interacting cells were used for distinct library preparation of interacting and non-interacting prokaryotic pathogen and eukaryotic host cells by deepSuperSAGE. Sequencing was performed with the Illumina HiSeq 2000 platform, and one point of time post infection (early interaction) was additionally prepared by Massive Analysis of cDNA Ends (MACE) as alternative tag-based library preparation method. 10 deepSuperSAGE and 2 MACE libraries. Please consult the publication mentioned in the following for more details.
Project description:Determine the role of interferons in the transcriptional profile of Ad-F7 transduced primary human macrophages using neutralizing antibody for the type I IFN receptor (IFNAR2). Primary human macrophage preparations were transduced with Ad-GFP or Ad-F7 and treated with control isotype (IgG) or neutralizing antibody for the type I IFN receptor (IFNAR2). RNA was collected 24 hours later and subjected to microarray analysis. Data represents the average of 5 donors.
Project description:The Human T-cell Leukemia Virus (HTLV)-type-I non-structural protein p30 plays an important role in virus transmission and gene regulation. p30 has been documented to inhibit the export of certain viral mRNA transcripts from the nucleus to the cytoplasm. This nuclear retainment of RNA molecules essentially results in gene silencing, where protein products are not produced. Considering this unique function of p30, we used microarray analysis to assess the ability of p30 to inhibit not only the regulation of transcription of cellular genes, but also the ability of p30 to regulate the export of cellular transcripts to the cytoplasm. Experiment Overall Design: Total or cytoplamsic RNA from peripheral blood mononuclear cells expressing HTLV-I p30 was isolated and analyzed by microarray analysis, in comparison with mock-transcuced cells.
Project description:This SuperSeries is composed of the following subset Series: GSE11926: Transcriptional profiles of Ad-F3 and Ad-F7 transduced macrophages. GSE12002: Transcriptional profiles of Ad-F7 transduced macrophages in the presence of neutralizing antibody for the type I interferon receptor (IFNAR2) or control isotype (IgG). Refer to individual Series
Project description:We combined metabolic pulse labeling and quantitative shotgun proteomics to globally monitor protein synthesis upon infection of human cells with a human- and a bird-adapted IAV strain. While production of host proteins was remarkably similar, we observed striking differences in the kinetics of viral protein synthesis over the course of infection. Most importantly, the matrix protein M1 was inefficiently produced by the bird-adapted strain at later stages.
Project description:While infection of chickens with highly pathogenic avian influenza (HPAI) H5N1 subtypes often leads to complete mortality within 24 to 48 h, infection of ducks in contrast causes mild or no clinical signs. Rapid onsets of fatal disease in chickens, but with no evidence of severe clinical symptoms in ducks, suggest underlying differences in their innate immune mechanisms. To understand the molecular basis for such difference, chicken and duck primary lung cells, infected with a low-pathogenicity avian influenza (LPAI) and two HPAI H5N1 viruses, were subjected to RNA expression profiling using Affymetrix Chicken GeneChip arrays. We used microarrays to analyze the gene expression profiles of primary chicken and duck lung cells infected with H2N3 LPAI and two H5N1 influenza virus subtypes to understand the molecular basis of host susceptibility and resistance. We have identified a set of key genes and pathways that could play an important role in mediating innate host resistance to avian influenza in chickens and ducks. 24 hours following infection, total RNA from cells was extracted. Replicate RNA samples from each of the virus-infected (H2N3, H5N1 50-92, or H5N1 ty-Ty) or mock-infected chicken and duck cells (4 treatment groups for each species) were used for microarray analysis. Each of the RNA samples was hybridized to one GeneChipM-BM-. Chicken Genome Array (Affymetrix), and a total of 16 array chips were used.