WT naïve vs. M. avium infected scRNA-seq
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ABSTRACT:
PROVIDER: PRJNA810168 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR18134100.fastq.gz | Fastqsanger.gz | |||
| SRR18134101.fastq.gz | Fastqsanger.gz | |||
| SRR18134086.fastq.gz | Fastqsanger.gz | |||
| SRR18134087.fastq.gz | Fastqsanger.gz | |||
| SRR18134088.fastq.gz | Fastqsanger.gz |
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Project description:purpose: Chronic inflammation induced by Mycobacterium avium infection (timepoint= 1 month) results in transcriptomic changes within the HSC population subset upon bulk RNA-seq, particularly in pathways involved in the immune response, interferon gamma signaling (Th1 immune response) pathway, stat signaling, and antigen processing and presentation. We have also shown that chronic inflammation results in a myeloid differentiation bias of HSPCs. We hypothesize that these transcriptomic changes induced by chronic inflammation may be specific to a certain subset of HSPCs and could be passed on to specific terminally differentiated subsets. To investigate this hypothesis, we performed scRNA-seq using the 10x genomics platform to test the transcriptomic differences induced upon infection in HSC, progenitor, myeloid, and lymphoid cell populations along with heterogeneity observed within the HSC population. methods: CD45.2 mice were infected with 2 x 10^6 CFU M. avium for one month prior to bone marrow harvesting. One month post infection, experimental mice were euthanized and infection was confirmed by measuring splenic weight and size. WBM from the hips, tibias, and femurs of each mouse was extracted through crushing with a mortar and pestle. The suspension was then RBC lysed and CD117 (c-kit)+ and cKit- cells were separated using a manual magnetic separation system (Miltenyi). Experimental samples were stained with desired antibodies (listed in key resources table) and sorted using a BD FACS Aria cell sorter. The following cell types were isolated according to the following phenotypic definitions (please see file phenotypic_definitions.txt). The cellular purity and count of each sample was confirmed prior to pooling for submission to the BCM Single Cell Core. Depending on the rarity of the isolated population, 20,000-50,000 sorted cells were combined to a concentration of ~7200 cells/uL. Samples were loaded into the Chromium controller system and generated 3’ 10x scRNAseq barcoded libraries for each sample. Prior to sequencing, quality control of the libraries was completed using a Bioanalyzer (Agilent, Santa Clara, CA) and Nanodrop system. Samples were sequenced on a NovaSeq at a sequencing depth of 300M reads at the BCM Genomic and RNA Profiling (GARP) core. After sequencing, fastq files for each sample were generated (R1, R2, and I1 files) to demultiplex the data using the CellRanger pipeline results: scRNA-seq studies revealed that training led to a bimodal distribution of gene expression, with a subset showing activation of immunity-related genes. This gene signature was propagated to innate immune cells but was not conserved in B cells, indicating specific patterns of gene signature inheritance. M. avium training also led to the emergence of an “activated” HSC subpopulation, indicating subsets of HSCs are more responsive to training conclusions: Transcriptomic signatures induced by M. avium exposure, particularly in IFNg response genes, are heterogeneous and are propagated in a myeloid biased manner
2023-07-30 | GSE197407 | GEO
Project description:Mycobacterium avium infection in mice induces granuloma necrosis in the lung which is dependent on IFNg. IRF1 is a transcription factor activated by IFNg signaling. The effect of IFNg and IRF1 on immunopathology and transcriptional changes in the lung were analysed using gene-deficient mice. The data from the Ifng-/- experiment have been described (Aly et al. 2007. J Pathol 212:295). Keywords: Infection, Genotype comparison
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Project description:Once entering the cell, M. avium subsp.paratuberculosis is known to survive harsh microenvironments, especially those inside activated macrophages. To improve our understanding of M. avium subsp.paratuberculosis pathogenesis, we examined the phagosome maturation associated with transcriptional responses of M. avium subsp.paratuberculosis during macrophage infection. Monitoring cellular markers, only live M. avium subsp.paratuberculosis bacilli were able to prevent phagosome maturation and reduce its acidification. On the transcriptional level, over 300 of M. avium subsp.paratuberculosis genes were significantly, differentially regulated in both naïve and IFN-γ-activated macrophages. These genes include the sigma factor H (sigH) that was shown to be important during persistent infection in M. tuberculosis.
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Project description:We studied the membrane protein compositions of Streptococcus pneumoniae WT and scRNA mutant strains, with or without the CSP1 induction into competence state.
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Project description:Mycobacterium avium subspecies paratuberculosis (MAP) infection is established following the ingestion of bacteria, their penetration of the intestinal mucosa and subsequent events of the host-parasite interaction. These bacteria invade M cells, macrophages and are capable of resisting host defenses and multiply to reach very high numbers intracellular. Mycobactrium avium subsp. avium (MAA) is an antigenically and genetically similar organisms but is relatively nonpathogenic for cattle. MAA organisms appear to infect cattle, but unlike cattle infected with MAP, cattle infected with MAA typically mount an effective systemic immune response, form caseous granuloma, and eliminate the infection. This study was designed to understand the similarities and differences in the host responses during MAP and MAA infection. Neonatal calves were infected (ligated ileal loops) with PBS, MAP, or avirulent MAA following which samples were collected at 30 min, 1h, 2h, 4h, 8h, or 12h. Post-infection, RNA was collected, processed, and then hybridized to custom bovine gene expression arrays, each of which represented 13,258 transcripts spotted in duplicate. Arrays were normalized by scaling against the average reference intensity value (i.e., average across all arrays), normalized by global mean, and then log transformed before statistical analyses were performed. Pairwise comparisons of averaged signal values and Student’s t test were performed using GeneSifter software (VizX Labs, Seattle, WA). A fold-change of at least 1.5-fold and a p value of 0.05 or less was required for a difference in signal to be considered statistically significant. When comparing the transcriptional responses of calves infected with the MAA versus MAP, unique patterns of expression were clearly visible. In general, numbers of transcripts altered in response to infection were much greater for MAA-infected animals than for those infected with the MAP. No genes were significantly up-regulated in MAP-infected animals at the earliest time point tested (30 min), and only modest numbers of genes were increased in expression over the experimental time course. On the other hand, up-regulated transcripts were detected by 30 min in MAA-infected animals and peaked by 2 hr. Differences in the numbers of down-regulated genes between MAP-infected and MAA-infected animals (compared to PBS-infected controls) were less pronounced. At the earliest time points, MAP-infected calves had a greater number of down-regulated genes than did animals infected with MAA. This trend was reversed at 8 and 12 hr post-infection. Keywords: Gene expression profiling by microarray
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Project description:Deep sequencing was implemented to study the transcriptional landscape of Mycobacterium avium TMC724. High-resolution transcriptome analysis identified the transcription start points for 652 genes. One third of these coincided with the start codons and therefore belong to leaderless transcripts, whereas the rest of the transcripts had 5' UTRs with the mean length of 83 nt. In addition, the 5' UTRs of 6 genes contained SAM-IV and Ykok types of riboswitches. 87 antisense RNAs and 9 intergenic small RNAs were mapped. Four of the revealed intergenic small RNAs, including igMAV_1034-1035 expressed at a very high level, have no homologs in M. tuberculosis, whilst M. avium lacks several intergenic sRNAs present in M. tuberculosis. Among those, MTS479 and MTS1338 are of special interest due to their possible implication in pathogenesis. Elucidation of differences in the repertoire of intergenic sRNAs between the two mycobacterial species may improve our understanding of mycobacterial diseases pathogenesis. Transcriptional profile of Mycobacterium avium TMC724, grown at 37M-BM-0C in Dubos broth until mid-logarithmic growth phase
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