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Mus musculus

ABSTRACT: Next Generation Sequencing analysis using splenic DCs freshly isolated from Wild Type and TBK1-DKO mice

PROVIDER: PRJNA371452 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR5230940.fastq.gz	Fastqsanger.gz
	SRR5230940_1.fastq.gz	Fastqsanger.gz
	SRR5230940_2.fastq.gz	Fastqsanger.gz
	SRR5230941.fastq.gz	Fastqsanger.gz
	SRR5230941_1.fastq.gz	Fastqsanger.gz

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Next Generation Sequencing analysis using splenic DCs freshly isolated from Wild Type and TBK1-DKO mice

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Transcriptome analysis of conventional dendritic cells (cDCs) introduced into the thymic microenvironment of re-aggregated thymic organ cultures (RTOCs)

Project description:cDCs home from the periphery into the thymus. In order to determine the phenotypic and functional changes introduced by the thymic microenvironment to these thymus-homing cDCs, RNA-based next generation sequencing (RNAseq) was performed with splenic (sp-DC) and thymic conventional DCs (t-DCs) re-isolated from RTOCs after two days of culture. Methods: RTOCs were generated from single-cell suspensions of thymi isolated from E14.5 to E16.5 fetuses of Foxp3hCD2 reporter mice (BALB/c background), and sp‑DCs or t‑DCs sorted from 4-8 weeks old female CD45.1xBALB/c mice were introduced. After two days of culture, 1‑2x103 cDCs were re-isolated from RTOCs as CD45.1+Lin-CD11chi by FACS, and total RNA for transcriptional profiling by low-input RNAseq was isolated. Total RNA from sp-DC and t-DCs isolated ex vivo from 4-8 weeks old female CD45.1xBALB/c mice was used as control. RNAseq was performed on an Illumina HiSeq2500 system. Differential gene expression between the four different conditions was computed from the read counts. Results: The number of differentially expressed genes between ex vivo isolated sp-DC and t-DC was drastically reduced comparing sp-DCs and t-DCs re-isolated from RTOCs. This finding implies that the thymic microenvironment within an RTOC modulates the gene expression in sp‑DCs, conferring a transcriptomic profile that more closely resembled the one from t‑DCs. Conclusion: The thymic microenvironment modulates the transcriptome of thymus-homing peripheral cDCs.

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