Application of Chromatrap® Solid-State Chromatin Immunoprecipitation (ChIP) for epigenetic profiling of insects
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ABSTRACT:
PROVIDER: PRJNA498662 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR8113880_1.fastq.gz | Fastqsanger.gz | |||
| SRR8113880_2.fastq.gz | Fastqsanger.gz | |||
| SRR8113890_1.fastq.gz | Fastqsanger.gz | |||
| SRR8113890_2.fastq.gz | Fastqsanger.gz | |||
| SRR8113881_1.fastq.gz | Fastqsanger.gz |
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Project description:We report the application of Chromatraps® Solid-State Chromatin Immunoprecipitation technology for epigenetic profiling of histone modifications in insects. Here, we present the optimised protocol and conditions of Chromatrap® kits for successful ChIP and high-throughput sequencing of established model species, Drosophila melanogaster and the emerging model for behavioural plasticity Nicrophorus vespilloides. We highlight successful ChIP-seq of Drosophila melanogaster (Oregon-R) of comparable quality to modENCODE data and present successful enrichment of histone marks for Nicrophorus vespilloides. The addition of this insect-based ChIP-seq protocol provides a set of optimal guidelines to aid streamline end-users epigenetic research focus and reduce experimental time.
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Project description:Background: In addition to triggering endosomal escape, the Accum® platform was recently reported for its ability to instill antigen cross-presentation properties in mesenchymal stromal cells (MSCs). Despite the promising results obtained with the first-generation vaccine using the A1 Accum® derivative (ARM vaccine), large quantities of cancer antigens were required to achieve meaningful therapeutic effects. Given this limitation, additional Accum® variants were engineered and tested for their ability to lower the need for large antigen quantities. A leading variant, AccuTOX®, was selected for that purpose. Methods: Several functional studies, including a series of antigen cross-presentation assays, were conducted using the SIINFEKL-specific T-cell clone B3Z. Analysis of endosomal escape and the effect of various anti-oxidant compounds were used to decipher the AccuTOX® mode of action in MSCs. The potency of the AccuTOX®-reprogramed MSCs (ARM-X) cells was evaluated in the context of therapeutic vaccination using immunocompetent C57BL/6 mice with three different pre-established solid tumor models. Various depletion studies were also conducted in animals to identify effector cells involved in the therapeutic response mediated by the ARM-X cells. Finally, the effect observed on murine ARM-X cells was validated on human MSCs along with an immunopeptidome study reflecting the cross-presentation potency of these reprogrammed human cells. Results: AccuTOX® can indeed trigger MSCs to cross-present, even if pulsed with low dose tumor antigens while retaining most of the innate properties of A1, including increased antigen uptake and processing, production of reactive oxygen species, endosomal escape and induction of the unfolded protein response (UPR). When tested against melanoma, pancreatic and colon cancer, therapeutic administration of the ARM-X vaccine, in combination with anti-PD-1, impairs tumor growth. Mechanistically, the ARM-X vaccine relies on efferocytosis by endogenous phagocytes and requires both CD4 and CD8 T cells, as their depletion leads to a loss in therapeutic function. Conclusion: Altogether, this second-generation ARM-X vaccine represents a platform adaptable to multiple solid tumors. In addition, our data clearly allude to a direct link between AccuTOX®-mediated UPR activation and antigen cross-presentation by MSCs. The fact that these modulated MSCs become antigen-presenting cells via UPR stimulation opens-up a new line of investigation to search for additional agents capable of specifically activating this pathway to convert culture-adapted MSCs to a cellular vaccination tool adaptable to various cancer indications.
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