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Multi-modal single cell analysis reveals gut microbiota reshapes brain immune landscape in aged mouse model

ABSTRACT: Multi-modal single cell analysis reveals gut microbiota reshapes brain immune landscape in aged mouse model

PROVIDER: PRJNA623317 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR11486406.fastq.gz	Fastqsanger.gz
	SRR11486407.fastq.gz	Fastqsanger.gz
	SRR11486408.fastq.gz	Fastqsanger.gz
	SRR11486409.fastq.gz	Fastqsanger.gz
	SRR11486410.fastq.gz	Fastqsanger.gz

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Multi-modal single cell analysis reveals gut microbiota reshapes brain immune landscape in aged mouse model.

Project description:Multi-modal single cell analysis reveals gut microbiota reshapes brain immune landscape in aged mouse model.

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Multi-modal single cell analysis reveals gut microbiota reshapes brain immune landscape in aged mouse model

Project description:We used CITE-seq (10x Gemoics-based) to profile and compare the transcriptomes and cell surface expression of a set of immune markers of naïve brain myeloid cells from young-adult (12 weeks old) wild type C57Bl/6 mice, aged (72 weeks old) wild type C57Bl/6 mice and young-adult and aged C57Bl/6 mice with antibiotics induced gut microbiota depletion. In one sequencing round, we sequenced a total of 12 different mouse brains (3 young control, 3 aged control, 3 young depleted gut microbiota (ABX) and 3 aged depleted gut microbiota (ABX)). We created an antibody pool consisting of 31 different antibodies (CCR2/CD192, C117/c-kit, CD11b, CD11c, CD172a/SIRPα, CD25, CD3, CD38, CD4, CD44, CD45, CD45R/B220, CD86, CD8a, CD90.1, Cx3cr1, F4/80, I-A/I-E, Ly6C, Ly6G, NK1.1, PD-1, PD-L1, CD169/Siglec-1, Siglec-H, TMEM119, XCR1, CD24, CD103, CD64, CD83) and stained each brain individually with this antibody pool. Then, we stained each brain with it's own unique hashing antibody so that we could subsequently pool the samples for loading onto the 10x Chromium. Young Control and Young ABX brains were tagged with HTOs 1-6 (HTO 1-3 for young control and HTO 4-6 for young ABX). Aged control and Aged ABX brains were tagged with HTOs 1-6 (HTO 1-3 for aged control and HTO 4-6 for aged ABX). Young and aged were loaded in separate wells on the 10X Chromium. The samples were tagged with unique nextera index primers to allow for preparation of one sequencing library consisting of all 12 samples and were separated in silico by it's unique hashing antibody and nextera index barcode combinations.

2020-11-05 | GSE148127 | GEO

Multi-modal single cell analysis reveals gut microbiota reshapes brain immune landscape in aged mouse model.

Project description:We used 16S V3/V4 region amplification to evaluate the composition of bacteria species in mouse fecal pellets. Fecel pellets were collected from young-adult (12 weeks old) wild type C57Bl/6 mice and aged (72 weeks old) wild type C57Bl/6 mice after 21 days of vehicle or antibiotics treatment (to induce gut microbiota depletion). In one sequencing round, we sequenced a total of 12 different fecal samples (3 young control, 3 aged control, 3 young depleted gut microbiota (ABX) and 3 aged depleted gut microbiota (ABX)). Amplicons were indexed using the Nextera XT Index Kit and pooled into a library for Illumina sequencing.

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Project description:The expression of inhibitory immune checkpoint molecules such as PD-L1 is frequently observed in human cancers and can lead to the suppression of T-cell mediated immune responses. Here we apply ECCITE-seq, a technology which combines pooled CRISPR screens with single-cell mRNA and surface protein measurements, to explore the molecular networks that regulate PD-L1 expression. We also develop a computational framework, mixscape, that substantially improves the signal-to-noise ratio in single-cell perturbation screens by identifying and removing confounding sources of variation. Applying these tools, we identify and validate regulators of PD-L1, and leverage our multi-modal data to identify both transcriptional and post-transcriptional modes of regulation. In particular, we discover that the kelch-like protein KEAP1 and the transcriptional activator NRF2, mediate levels of PD-L1 upregulation after IFNγ stimulation. Our results identify a novel mechanism for the regulation of immune checkpoints and present a powerful analytical framework for the analysis of multi-modal single-cell perturbation screens.

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