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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Jun 01, 2023 |
| Title |
Comparative analysis of Tet2 catalytic deficient and knockout bone marrow over time |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
Methylation profiling by high throughput sequencing
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| Summary |
In this study, we investigated the role of Tet2 in regulation of hematopoietic genes by performing transcriptomic analysis of Tet2 WT, Mut, and KO LSK and Lin– cells at 3, 6, 9, and 12 months by RNA-seq, and mapping the DNA methylation landscape of 3-month-old Tet2 WT, Mut, and KO LSK and Lin– cells. (1) We find that loss of Tet2 (Tet2 KO) vs. a loss of Tet2 catalytic function (Tet2 Mut) at different time points and cell types causes distinct gene expression changes when compared to WT samples as assessed by RNA-seq. (2) Additionally, we find that many genes implicated in lymphoma and leukemia are deregulated in Tet2 Mut and KO LSK and Lin– cells, consistent with the phenotypes observed in mice transplanted with Tet2 Mut and KO bone marrow. (3) We also mapped genome-wide DNA methylation levels of WT, Tet2 Mut, and KO LSK and Lin– cells at 3 months by WGBS and establish a role for Tet2 in demethylating genes implicated in lymphoma and leukemia initiation and progression.
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| Overall design |
We studied the role of Tet2 in regulation of hematopoietic genes by performing transcriptomic analysis of Tet2 WT, Mut, and KO LSK and Lin– cells at 3, 6, 9, and 12 months by RNA-seq. Bone marrow from three mice was pooled and LSK and Lin– fractions were sorted by FACS. Total RNA was extracted (Qaigen RNeasy Total RNA kit). Library preparation and RNA-seq were performed at Novogene (https://en.novogene.com/) on an Illumina Novoseq 6000 platform. About 20-30 million reads were generated per sample. Details of RNA-seq and data analysis are described in the methods sections of the manuscript. We also performed WGBS to assess changes in DNA methylation in the absence of Tet2 or its catalytic function alone. We isolated LSK and Lin– cells from Tet2 WT, Mut, and KO 3-month-old mouse bone marrow and subjected them to WGBS. High-quality DNA was extracted (Quick-DNA miniprep kit (Zymo, D3024) and sent for WGBS analysis at BGI genomics company (https://en.genomics.cn/). Lamda DNA spike-in was added to confirm the bisulfite conversion efficiency. Details of WGBS and data analysis are described in the methods sections of the manuscript.
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| Contributor(s) |
Dawlaty MM, Ito K, Flores JC |
| Citation(s) |
37225048 |
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| Submission date |
Mar 22, 2023 |
| Last update date |
Aug 31, 2023 |
| Contact name |
Meelad M Dawlaty |
| E-mail(s) |
meelad.dawlaty@einsteinmed.org
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| Phone |
718-678-1224
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| Organization name |
Albert Einstein College of Medicine
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| Department |
Genetics
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| Lab |
Dawlaty Lab
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| Street address |
1301 Morris Park Ave, Price 419, Bronx
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| City |
New York |
| State/province |
New York |
| ZIP/Postal code |
10461 |
| Country |
USA |
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| Platforms (2) |
| GPL21103 |
Illumina HiSeq 4000 (Mus musculus) |
| GPL24247 |
Illumina NovaSeq 6000 (Mus musculus) |
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| Samples (30)
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| Relations |
| BioProject |
PRJNA947607 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE227977_RAW.tar |
1.8 Gb |
(http)(custom) |
TAR (of BW) |
| GSE227977_normalizedCounts.csv.gz |
3.0 Mb |
(ftp)(http) |
CSV |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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