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| Status |
Public on Mar 29, 2021 |
| Title |
Regulation of B lymphocyte development by histone H2A deubiquitinase BAP1 (Bap1KO_RNA-Seq) |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
BAP1 is a deubiquitinase (DUB) of the Ubiquitin C-terminal Hydrolase (UCH) family that regulates gene expression and other cellular processes, via deubiquitination of histone H2AK119ub and other substrates. BAP1 is an important tumour suppressor in human, expressed and functional across many cell-types and tissues, including those of the immune system. B lymphocytes are the mediators of humoral immune response, however the role of BAP1 in B cell development and physiology remains poorly understood. Here we characterize a mouse line with a selective deletion of BAP1 within the B cell lineage (Bap1fl/fl mb1-Cre) and establish a cell intrinsic role of BAP1 in the regulation of B cell development. We demonstrate a depletion of large pre-B cells, transitional B cells, and mature B cells in Bap1fl/fl mb1-Cre mice. We characterize broad transcriptional changes in BAP1-deficient pre-B cells, map BAP1 binding across the genome, and analyze the effects of BAP1-loss on histone H2AK119ub levels and distribution. Overall, our work establishes a cell intrinsic role of BAP1 in B lymphocyte development, and suggests its contribution to the regulation of the transcriptional programs of cell cycle progression, via the deubiquitination of histone H2AK119ub.
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| Overall design |
We performed RNA-Seq transcriptional profiling of FACS-sorted pre-B and immature B cells. Briefly, bone marrow cells were harvested from Bap1 fl/fl mb1-Cre, Bap1 fl/+ mb1-Cre, and Bap1fl/+ mice. The cells were sorted using FACS into pre-B and immature B cells, gating on B220+CD3-CD11b-NK1.1-CD11c-TER119-Gr1-, and IgM-IgD-CD19+CD43- for pre-B cells, and CD19+IgM+IgD- for immature B cells. RNA was isolated from the FACS-sorted cells using the MagMAX total RNA kit (Ambion, Life Technologies), and quality assessed using Bioanalyzer RNA Pico chips (Agilent). rRNA depletion and library preparation were performed using the SMARTer Stranded RNA-Seq kit (Takara Clontech). The libraries were sequenced on an Illumina HiSeq 4000 sequencer in paired-end 50bp configuration. Cells from multiple mice were pooled to achieve yields of >1.7ng of RNA per sample, and 3-4 independent samples were analyzed per genotype for each cell-type.
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| Contributor(s) |
Wang H, Lin YH, Langlais D, Nijnik A |
| Citation(s) |
33912157 |
| |
| Submission date |
Nov 24, 2020 |
| Last update date |
Jun 28, 2021 |
| Contact name |
Anastasia Nijnik |
| E-mail(s) |
anastasia.nijnik@mcgill.ca
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| Phone |
514-398-5567
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| Organization name |
McGill University
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| Department |
Physiology
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| Lab |
Nijnik Lab
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| Street address |
Rm 368 - 3649 Promenade Sir William Osler
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| City |
Montreal |
| State/province |
Quebec |
| ZIP/Postal code |
H3G 0B1 |
| Country |
Canada |
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| Platforms (1) |
| GPL21103 |
Illumina HiSeq 4000 (Mus musculus) |
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| Samples (21)
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| This SubSeries is part of SuperSeries: |
| GSE162085 |
Regulation of B lymphocyte development by histone H2A deubiquitinase BAP1 |
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| Relations |
| BioProject |
PRJNA680533 |
| SRA |
SRP293910 |
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