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Genome-wide maps of chromatin states during B-cell to Macrophage lineage switching

ABSTRACT: Using homogenous populations of cells undergoing lineage switching we studied enhancer nucleation.

ORGANISM(S): Mus musculus

PROVIDER: GSE53173 | GEO | 2015/07/29

SECONDARY ACCESSION(S): PRJNA231058

REPOSITORIES: GEO

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Genome-wide maps of chromatin states during B-cell to Macrophage lineage switching

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Nucleosome density map during B-cell to Macrophage lineage switching

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Dynamic chromatin connectivity maps reveal lineage specific regulation

Project description:Using genome-wide Chromatin Interaction Analysis with Paired-End-Tag sequencing, we mapped long-range chromatin interactions associated with RNA polymerase II in three different mouse cell lines and uncovered widespread promoter-centered interactions. These interactions further aggregated into higher-order clusters, in which proximal and distant genes are engaged through enhancer-promoter interactions. Comparative analyses of different cell lines imply that cell specific enhancer interactions are dynamic among different cell specific transcription, and suggest significant enrichment of enhancer-promoter interactions for cell specific manner. Overall, our study provides novel insights into the three-dimensional basis of transcription activity in mouse cells. RNA polymerase II (RNAPII) guided chromatin interactions were discovered by Chromatin Interaction Analysis with Paired-End Tag (ChIA-PET) sequencing, in order to study genome-wise the enhancer-promoter interactions. Three cell lines, namely mouse embryonic stem cell E14, Neural stem cell NS5 and neuroshpere cells were grown under standard culture conditions and harvested at log phase. Harvested cells were cross-linked using 1% formaldehyde followed by neutralization with 0.2M glycine. Chromatin was isolated and subjected to ChIA-PET protocol as described in Fullwood et al, 2009. The ChIA-PET sequence reads were processed and analyzed using ChIA-PET Tool (Li et al, 2010)

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Next Generation Sequencing confirmed exon alternative splicing and lineage switching in one patient

Project description:Purpose: To analyze the expression of two different CD19 isoforms during BiTE treatment, and to further determine cell lineage specific molecules before and after BiTE treatment Methods: For transcriptome sequencing, we isolated patient's bone marrow samples, quality of total RNA was evaluated by Agilent 2100. RNA libraries were prepared for sequencing using NEBNext® UltraTM RNA Library Prep Kit for Illumina (NEB,USA). RNA sequencing process and the data acquisition were finished on Novogene Experimental Department according to the manufacturer’s protocol. Reads were aligned to hg38 reference genome. Relative abundance of CD19 and other gene transcripts were mapped and estimated at both gene and transcript level (FPKM) by feature Counts. Results: The expression of CD19 isoform with exon 2 deletion was found at diagnosis before BiTE therapy. The patient did not achieve remission after BiTE treatment, and the expression of CD19 antigen turned negative by flow cytometry detection. But the expression ratio of exon 2 deleted CD19 was not increased, and the flow cytometry phenotype and transcriptome sequencing confirmed that no linage switching occurred, which suggested the expression of CD19 isoform caused by exon alternative splicing and lineage switching was not the driving mechanism of CD19 epitope loss in this patient. Immune escape could not be prevented by targeting alternative exons. This patient achieved complete remission by sequential infusions of our own developed CD22 CAR-T and CD19 CAR-T after disease progression Conclusions: transcriptome sequencing confirmed that no linage switching occurred in this patient, which suggested the expression of CD19 isoform caused by exon alternative splicing and lineage switching was not the driving mechanism of CD19 epitope loss in this patient.

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