Project description:We report the application of RNA-seq technology for high-throughput profiling of gene transcription in RAW264.7 infected with Mycobacterium tuberculosis H37Ra. By obtaining over six billion bases of sequence from mRNA, we generated genome-wide gene transcription maps of RAW264.7 infected with CdhM-related Mycobacterium tuberculosis H37Ra. We find that numerous genes involved in ER stress are significantly affected by CdhM. This finding indicates that CdhM may induce ER stress during Mtb infection of host cells.
Project description:Transcriptional profiling of Mycobacterium tuberculosis H37Ra::pTetR-yidC (Test) compared with Mycobacterium tuberculosis H37Ra::pTetR (Control) bacteria after 4 days of treatment with 50ng/ml ATc with shaking at 200rpm at 37°C.
Project description:Purpose: The goals of this study are to obtain the NGS-derived transcriptome profiling (RNA-seq) for THP-1 macrophages response to Mycobacterium tuberculosis (H37Rv and H37Ra) Methods: mRNA and long noncoding RNA profiles of THP-1 macrophages infected with H37Rv and H37Ra for 1, 4, 12, 24, 48 hours were generated by deep sequencing, using Illumina Hiseq3000. The sequence reads that passed quality filters were first mapped to the latest UCSC transcript set using Bowtie2 (version 2.1.0). Then the gene expression level was estimated using RSEM (RNA-Seq by Expectation Maximization, v1.2.15), for lncRNA analysis, reads were mapped to lncRNA transcript set from LNCipedia.org. The sequence reads were normalized with TMM (trimmed mean of M-values) to identify differentially expressed genes (DEGs) using the edgeR package edgeR. qRT–PCR validation was performed using SYBR Green assays. Results: Using an optimized data analysis workflow, we mapped about 20 million sequence reads per sample to the human genome (GRCh38/hg38) and identified 25,343 mRNA and 47877 long non-coding RNA transcripts. Conclusions: Our study represents the detailed analysis of transcriptomes for THP-1 macrophages response to H37Rv and H37Ra, generated by RNA-seq technology.
Project description:Synchronised cultures of Mycobacterium tuberculosis (M. tb H37Ra DnaAcos115) were used to determine the cell cycle dependent gene expression. Mycobacterium tuberculosis H37Ra DnaAcos115 strain was grown in Middlebrook 7H9 broth at 30C for 30h which arrests replication and later cultures were allowed to grow at 37oC for 30h. Samples were collected at 0, 2, 6, 12,18, 24 and 30 hours. Samples at 30oC for 30h were considered as '0' time point. A total of 18 samples, consisting of triplicates for 6 time-points were analyzed using microarray.
Project description:This experiment is to compare the transcription patterns of mouse macrophages (J774A.1) infected with BCG, H37Ra and M. smegmatis under high multiplicity of infection (MOI). Through the global transcriptome profiling study, we define a pathogen specific host gene expression pattern and indicate that SRC likely plays a central role in regulating multiple unique signaling pathways activated by MTB infection. Mycobacterium tuberculosis (MTB) infects an estimated one-third of the global population and is one of the main causes of mortality due to an infectious agent.
Project description:Synchronised cultures of Mycobacterium tuberculosis (M. tb H37Ra DnaAcos115) were used to determine the cell cycle dependent gene expression.
Project description:Histone modifications plays vital role in the regulation of immune response following Mycobacterium tuberculosis infection. Here, mouse macrophages (RAW264.7) were infected with M. tuberculosis (H37Rv) or left uninfected, followed by Chromatin immunoprecipitation with anti-H3K4me3 or anti-H3K27ac antibody and high throughput sequencing.
Project description:Host macrophage transcriptional responses to intracellular pathogens remain poorly characterized. We screened transcriptional enhancers engaged in response to M. tuberculosis (Mtb) infection by ChIPseq analysis of histone H3 lysine 4 monomethylation (H3K4me1). De novo monomethylation during infection was associated with genes implicated in host defense and apoptosis. These regions were enriched for binding sites for ETS transcription factor family members and response elements for nuclear receptors, including liver X receptors (LXRs) and peroxisomal proliferator activated receptors (PPARs), many of which were encompassed by transposable elements. LXRa expression was strongly induced by infection, whereas that of PPARs was unaffected. LXR DNA binding and NCoR corepressor recruitment increased proportionately in infected cells but coactivator association was unchanged, consistent with a lack of induction of endogenous agonists. However, treatment of infected cells with LXR agonist T0901317 strongly increased coactivator recruitment and induced a gene expression program characterized by enhanced innate immune signaling and lipid metabolism. Remarkably, T0901317 treatment selectively induced apoptosis in infected macrophages, and was accompanied by Mtb death, reducing mycobacterial burden 18-fold relative to vehicle 5d after infection. These studies define macrophage transcriptional responses to Mtb infection, and suggest that tissue-specific LXRa agonists may be efficacious in clinical management of tuberculosis.
Project description:We report the application of RNA-seq technology for high-throughput profiling of gene transcription in BMDMs. By obtaining over six billion bases of sequence from mRNA, we generated genome-wide gene transcription maps of BMDMs infected with recombinant Mycobacterium tuberculosis H37Ra strains. We find that a group of genes is significantly affected by MpbR. This finding indicates that MpbR regulates the host immune.
