Patterned CpG methylation of silenced B cell gene promoters in classical Hodgkin lymphoma-derived and primary effusion lymphoma cell lines
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Published on: January 20, 2019
Jeanette R Doerr1, Cindy S Malone, Francesca M Fike
1Department of Microbiology, David Geffen School of Medicine at the University of California, Los Angeles, CA 90095, USA.
Insights
Epigenetic mechanisms, specifically DNA methylation, silence B cell genes in Hodgkin lymphoma (HL) and primary effusion lymphoma (PEL) cells. Reactivation of these genes by 5-aza-2'-deoxycytidine confirms methylation
Area of Science:
- Hematology
- Epigenetics
- Molecular Biology
Background:
- Hodgkin and Reed-Sternberg (HRS) cells in classical Hodgkin lymphoma (cHL) and primary effusion lymphoma (PEL) cells originate from germinal center (GC) B cells.
- These malignant B cells exhibit a significant loss of typical B cell gene and surface marker expression.
- This gene silencing is not caused by a deficiency in necessary transcription factors.
Purpose of the Study:
- To investigate the epigenetic mechanisms responsible for the loss of B cell gene expression in HRS and PEL cells.
- To determine if DNA methylation plays a role in silencing key B cell genes.
- To characterize the patterns of DNA methylation at silenced gene promoters.
Main Methods:
- Treatment of HRS and PEL cells with 5-aza-2 -deoxycytidine to assess gene reactivation.
- Genomic bisulfite sequencing to analyze DNA methylation patterns.
- Analysis of promoter methylation in silenced B cell genes including B29, TCL1, and Bob1.
Main Results:
- 5-aza-2 -deoxycytidine treatment reactivated silenced B cell genes (B29, TCL1, Bob1), indicating DNA methylation as the silencing mechanism.
- Genomic bisulfite sequencing revealed distinct DNA methylation patterns at the promoters of silenced B29 and TCL1 genes.
- Three specific methylation patterns were identified for B29 and TCL1 promoters: 5 promoter CpG methylation only, 5 and 3 promoter CpG methylation with central sparing, and complete CpG methylation.
- The Bob1 promoter exhibited dense CpG methylation across nearly all sites.
Conclusions:
- DNA methylation is the primary epigenetic mechanism responsible for the silencing of B cell genes in classical Hodgkin lymphoma and primary effusion lymphoma.
- The patterns of DNA methylation at silenced gene promoters are specific and consistent.
- While some methylation patterns mimic normal gene silencing, complete CpG methylation throughout promoters suggests abnormal activation and targeting of methylation in certain cases.
Abstract:
Hodgkin and Reed-Sternberg (HRS) cells of classical Hodgkin lymphoma (cHL) and primary effusion lymphoma (PEL) are derived from germinal center (GC) and post-GC B cells, respectively. Neither express many of the B cell genes or surface markers typically expressed by other GC-derived B cell lymphomas or normal B cells. This loss of B cell gene expression is not due to a lack of essential transcription factors, as studies have shown that the ectopic expression of missing transcription factors failed to reactivate endogenous target genes. These results implicate epigenetic mechanisms extinguishing B cell gene expression. Silenced endogenous B cell genes representing a surface receptor, B29 (Igbeta, CD79b), a signaling molecule, TCL1, and a transcription factor, Bob1 (OCA-B, OBF-1), were reactivated by 5-aza-2'-deoxycytidine, indicating that gene silencing in HRS and PEL cells is due to DNA methylation. Genomic bisulfite sequencing corroborated this prediction and revealed three distinct patterns of methylation for the silenced B29 and TCL1 promoters. These distinct patterns consisted of 5' promoter CpG methylation alone, 5' and 3' promoter CpG methylation sparing sites in the central cores, and complete CpG methylation throughout the promoter regions. The silenced Bob1 promoter showed one pattern of dense CpG methylation at essentially all sites. These consistent patterns predict that, although gene silencing in many HRS and PEL cells mimics appropriate gene silencing, in some cases of complete CpG methylation throughout entire promoters both the activation and targeting of methylation is abnormal.
