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Tracking immunoglobulin gene editing and transcriptomic changes in GC B cells by single-cell analysis

ABSTRACT: Tracking immunoglobulin gene editing and transcriptomic changes in GC B cells by single-cell analysis

PROVIDER: PRJNA779703 | ENA |

REPOSITORIES: ENA

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Tracking immunoglobulin gene editing and transcriptomic changes in GC B cells by single-cell analysis

Project description:In response to T cell-dependent antigens, mature B cells are stimulated to form germinal centers, which are the place of B cell affinity maturation and selection. Here, we dissected the heterogeneity of germinal center B cells and reconstructed their pathway of differentiation by integrating single-cell transcriptomic data with the information on immunoglobulin gene editing associated with somatic hypermutation and class switch recombination.

2022-02-02 | GSE188617 | GEO

RNA-seq on MHV68 infected GC B cells expressing distinct immunoglobulin light chains

Project description:RNA-seq on MHV68 infected GC B cells expressing distinct immunoglobulin light chains

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Clonal Tracking Uncovers Barriers and Validate New Strategies to Enhance Gene Editing in Human Hematopoietic Stem Cells [RNA-seq]

Project description:By applying a barcoding strategy to clonal tracking of edited cells (BAR-seq), we show that p53 activation triggered by HDR editing significantly shrink the HSPC clonal repertoire in hematochimeric mice, despite engrafted edited clones preserved multilineage and self-renewing capacity. Transient inhibition of p53 restored polyclonal graft composition. We then overcame HSC constraints to HDR by forcing cell cycle progression and upregulating components of the HDR machinery through transient expression of Adenovirus 5 E4orf6/7 protein, which operates the major cell cycle controller E2F. Combined E4orf6/7 expression and p53 inhibition resulted in high and stable HDR editing efficiencies in the human graft, without perturbing repopulation and self-renewing properties of edited HSCs.

2020-04-22 | GSE143995 | GEO

Clonal Tracking Uncovers Barriers and Validate New Strategies to Enhance Gene Editing in Human Hematopoietic Stem Cells [BAR-seq]

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Tracking epigenetic changes during human leukemogenesis

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Tracking genome-editing and associated molecular perturbations by SWATH mass spectrometry

Project description:Advances in gene editing now allow reverse genetics to be applied to a broad range of biological systems. Ultimately, any modification to coding sequences requires confirmation at the protein level, although immunoblotting is often hampered by antibody quality or availability especially in non-mammalian species. Here, we show that Sequential Window Acquisition of All Theoretical Spectra (SWATH) - Mass Spectrometry (MS) is a robust antibody-independent alternative for monitoring gene editing at the protein level and concomitantly defines proteome responses which may provide pertinent biological insights.

2019-10-24 | PXD013176 | Pride

RNA-seq on MHV68 infected GC B cells expressing distinct immunoglobulin light chains

Project description:We performed RNA-seq to examine the differences in GC B cells that were either infected with MHV68 or uninfected and between B cells that expressed IgL or IgK B cell receptor light chains.

2020-03-02 | GSE140056 | GEO

Evolutionary and ontogenetic changes in RNA editing in human, chimpanzee and macaque brains

Project description:we used RNA-Seq to quantify the RNA editing level at more than 8,000 previously annotated exonic A-to-I RNA editing sites in two brain regions - prefrontal cortex and cerebellum - of humans, chimpanzees and rhesus macaques. We observed substantial conservation of RNA editing levels between the brain regions, as well as among the three primate species. Evolutionary changes in RNA editing were nonetheless evident among the species. Across lifespan, we observed an increase of the RNA editing level with advanced age in both brain regions of all three primate species. poly(A) enriched RNAs extracted from pooled samples of two brain regions: CBC and PFC of chimpanzee and macaque, fragmented, revers transcribed to double-stranded cDNA using random hexamers. Sequencing libraries were prepared according to the paired-end non-strand-specific sample preparation protocol of Illumina. Each sample was sequenced in a separate lane in the Illumina Genome Analyzer II system, using the 75-bp paired-end sequencing protocol. human data was downloaded from SRA [SRP005169]

2014-01-07 | E-GEOD-45244 | biostudies-arrayexpress

Transcriptome analysis of mouse WT and CD22 KO GC B cells

Project description:Germinal centres (GC) are specialized sites where B cells expand and diversify their antibody genes through somatic hypermutation. GC B-cells are routinely identified through distinct changes on their surface carbohydrates, known as glycans. One striking modification relates to the monosaccharide sialic acid. In mice, this change is mediated through downregulation of an enzyme called CMAH, which results in a GC-specific loss of preferred ligand for CD22, a member of the sialic acid-binding immunoglobulin-type lectins (Siglecs) and an inhibitory co-receptor of the B-cell antigen receptor (BCR). Here, we identified that glycan remodeling, mediated by downregulation of CMAH, is crucial for the GC B-cell response, and production of memory B-cells, plasma cells, and high affinity antibodies. We also demonstrated that the function of these altered glycans is dependent on CD22, highlighting that coordinated loss of preferred ligands acts to modulates the CD22 activity in the GC B-cells. Overall, our study reveals that intrinsic glycan remodeling functions to optimize the B-cell responses in the GC by controlling CD22.

2022-02-09 | GSE196206 | GEO

PTIP promotes chromatin changes critical for immunoglobulin class switch recombination

Project description:Programmed genetic rearrangements in lymphocytes require transcription at antigen receptor genes to promote accessibility for initiating double-strand break (DSB) formation critical for DNA recombination and repair. Here we show that activated B cells deficient in the PTIP component of the MLL3 (mixed-lineage leukemia 3) /MLL4 complex display impaired histone methylation (H3K4me3) and transcription initiation of downstream switch regions at the immunoglobulin heavy-chain (Igh) locus leading to defective immunoglobulin class-switching. We also show that PTIP accumulation at DSBs contributes to class-switch recombination (CSR) and genome stability independently from Igh switch transcription. These results demonstrate that PTIP promotes specific chromatin changes that control the accessibility of the Igh locus to CSR, and suggest a non-redundant role for the MLL3/MLL4 complex in altering antibody effector function. Genome-wide analysis of histone modifications, PTIP, and Pol II in PTIP-WT and PTIP-KO mouse activated B cells.

2010-08-25 | E-GEOD-20852 | biostudies-arrayexpress

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