Final Report Summary - DNAMETHYLOOPMCL (Of DNA methylation and looping of distant regulatory elements in mantle cell lymphoma)
1. Introduction.
1.1.Epigenetics.
Each cell in the body of an individual contains the same DNA. Our DNA is packed into chromosomes, long molecules that contain the information necessary for each cell to execute their function. More specifically, through the information that is stored in our DNA each cell in our body knows how to build the proteins necessary for their cellular functions. Each protein in the cell is encoded by a gene in the DNA, however, only a very small part of the DNA contains genes. A major part of the DNA contains regulatory elements, also known as enhancers, that regulate which genes are active and which are inactive.
One intriguing question is how different cell types, like liver cells, blood cells or neurons are formed. How do these cells know which part of the DNA to use, i.e. which proteins they need, to become a liver cell, a blood cell or a neuron? Epigenetic processes can regulate which part of the DNA is active and which part of the DNA is inactive and hence can regulate which proteins are generated in each cell type. The epigenetic code consist of two major components, the methylation of the DNA and the modification of histones, the proteins around which the DNA is wrapped. With different techniques we can determine the epigenetic code in order to understand which regions in the DNA are active in specific cell types.
1.2.The three-dimensional (3D) chromatin structure.
Chromosomes are very long molecules and we tend to think of them in a linear, two-dimensional way. With this view, however, we omit that the genome is highly organized in 3D space and that genomic regions might be far in linear space, but can be close in 3D space by the formation of loops. With different technique we can determine the 3D genome structure in order to understand which regulatory regions, i.e. enhancers, loop to which genes. In this way, distant enhancers can loop to their target genes, resulting in their activation.
1.3.Mantle cell lymphoma.
Lymphomas are tumors that arise in the lymph nodes upon malignant transformation of normal white blood cells, i.e. lymphocytes. Before and during malignant transformation normal lymphocytes acquire changes in their genetic code as well as in their epigenetic processes. Mantle cell lymphoma (MCL) is a specific type of lymphoma with an aggressive clinical course and a poor clinical outcome, however few cases show an indolent behavior. From the genetic point of view, it is known that MCLs carry a fusion of a part of chromosome 11 and chromosome 14, that leads to the activation of the protein cyclin D1. Furthermore, the difference between aggressive and indolent MCLs is characterized by the activation of another protein, SOX11, in the aggressive MCL cases specifically. However, it is not clear how the SOX11 gene becomes activated in these cases.
2. Summary of project objectives.
During this project, we aimed to study the epigenetic and 3D genomic landscape of MCL in order to discover unknown regions within the DNA, whose deregulation contribute to the development and/or aggressiveness of this lymphoma.
3. Performed work and results
We have mapped the DNA methylation landscape of 82 MCL samples as well as of normal lymphocytes. Furthermore, for 2 representative MCL cases and 2 normal lymphocyte subtypes, we have mapped the histone modification landscape. The latter enabled us to determine the activity of enhancers and genes within these representative samples.
We have analysed the DNA methylation patterns of the 82 MCL cases in an unsupervised way and detected 2 MCL subgroups. One subgroup comprised all aggressive cases, the other comprised indolent and aggressive cases.
When comparing the 2 MCL subgroups with each other and with normal lymphocytes as controls, we observed that regions of hypomethylation in the MCL samples are enriched for enhancer regions. One of these regions was located within proximity of the SOX11 gene. The DNA hypomethylation and enhancer activation of this region was specific for the MCL sample that had high levels of the SOX11 protein. Next, we performed an analysis of the 3D genome structure at this region and we showed that in the SOX11 positive MCL case, this hypomethylated enhancer region loops in 3D space to the SOX11 gene, while it did not loop to other genes that were located between SOX11 and the enhancer region.
4. Conclusions
From our analyses, we can conclude that DNA hypomethylation in MCL can be used to detect new active enhancer regions that may regulate the activation of proteins that contribute to the development and aggressiveness of MCL. This was exemplified by the discovery of a new enhancer region that potentially regulates SOX11 expression.
In addition we can conclude that differentially methylated regions in MCL subgroups can coincide with differential looping of these regions towards their target genes. This was shown by the existence of a differentially methylated enhancer region in SOX11 postivive cases that only loops to the SOX11 gene in these respective cases.
Furthermore, we can conclude that analysis of the 3D genome structure in MCL is essential to determine target genes of enhancers, as enhancers may not necessary activate their most neighboring gene. This was seen for the newly detected SOX11 enhancer that did not loop to other genes within the same region.
5. Potential impact and use
This project has lead to the detection of new regulatory regions in MCL that may regulate the activation of proteins that contribute to the development and aggressiveness of MCL. This was exemplified by a newly identified enhancer region that potentially regulate SOX11 expression. In line with this finding, we can now discover other regions that show a similar behaviour to better understand the biology of this aggressive disease. In the future this may aid in developing targeted therapies. In addition, the differential DNA methylation patterns may be useful to develop biomarkers that are useful for the detection and stratification of MCLs.
1.1.Epigenetics.
Each cell in the body of an individual contains the same DNA. Our DNA is packed into chromosomes, long molecules that contain the information necessary for each cell to execute their function. More specifically, through the information that is stored in our DNA each cell in our body knows how to build the proteins necessary for their cellular functions. Each protein in the cell is encoded by a gene in the DNA, however, only a very small part of the DNA contains genes. A major part of the DNA contains regulatory elements, also known as enhancers, that regulate which genes are active and which are inactive.
One intriguing question is how different cell types, like liver cells, blood cells or neurons are formed. How do these cells know which part of the DNA to use, i.e. which proteins they need, to become a liver cell, a blood cell or a neuron? Epigenetic processes can regulate which part of the DNA is active and which part of the DNA is inactive and hence can regulate which proteins are generated in each cell type. The epigenetic code consist of two major components, the methylation of the DNA and the modification of histones, the proteins around which the DNA is wrapped. With different techniques we can determine the epigenetic code in order to understand which regions in the DNA are active in specific cell types.
1.2.The three-dimensional (3D) chromatin structure.
Chromosomes are very long molecules and we tend to think of them in a linear, two-dimensional way. With this view, however, we omit that the genome is highly organized in 3D space and that genomic regions might be far in linear space, but can be close in 3D space by the formation of loops. With different technique we can determine the 3D genome structure in order to understand which regulatory regions, i.e. enhancers, loop to which genes. In this way, distant enhancers can loop to their target genes, resulting in their activation.
1.3.Mantle cell lymphoma.
Lymphomas are tumors that arise in the lymph nodes upon malignant transformation of normal white blood cells, i.e. lymphocytes. Before and during malignant transformation normal lymphocytes acquire changes in their genetic code as well as in their epigenetic processes. Mantle cell lymphoma (MCL) is a specific type of lymphoma with an aggressive clinical course and a poor clinical outcome, however few cases show an indolent behavior. From the genetic point of view, it is known that MCLs carry a fusion of a part of chromosome 11 and chromosome 14, that leads to the activation of the protein cyclin D1. Furthermore, the difference between aggressive and indolent MCLs is characterized by the activation of another protein, SOX11, in the aggressive MCL cases specifically. However, it is not clear how the SOX11 gene becomes activated in these cases.
2. Summary of project objectives.
During this project, we aimed to study the epigenetic and 3D genomic landscape of MCL in order to discover unknown regions within the DNA, whose deregulation contribute to the development and/or aggressiveness of this lymphoma.
3. Performed work and results
We have mapped the DNA methylation landscape of 82 MCL samples as well as of normal lymphocytes. Furthermore, for 2 representative MCL cases and 2 normal lymphocyte subtypes, we have mapped the histone modification landscape. The latter enabled us to determine the activity of enhancers and genes within these representative samples.
We have analysed the DNA methylation patterns of the 82 MCL cases in an unsupervised way and detected 2 MCL subgroups. One subgroup comprised all aggressive cases, the other comprised indolent and aggressive cases.
When comparing the 2 MCL subgroups with each other and with normal lymphocytes as controls, we observed that regions of hypomethylation in the MCL samples are enriched for enhancer regions. One of these regions was located within proximity of the SOX11 gene. The DNA hypomethylation and enhancer activation of this region was specific for the MCL sample that had high levels of the SOX11 protein. Next, we performed an analysis of the 3D genome structure at this region and we showed that in the SOX11 positive MCL case, this hypomethylated enhancer region loops in 3D space to the SOX11 gene, while it did not loop to other genes that were located between SOX11 and the enhancer region.
4. Conclusions
From our analyses, we can conclude that DNA hypomethylation in MCL can be used to detect new active enhancer regions that may regulate the activation of proteins that contribute to the development and aggressiveness of MCL. This was exemplified by the discovery of a new enhancer region that potentially regulates SOX11 expression.
In addition we can conclude that differentially methylated regions in MCL subgroups can coincide with differential looping of these regions towards their target genes. This was shown by the existence of a differentially methylated enhancer region in SOX11 postivive cases that only loops to the SOX11 gene in these respective cases.
Furthermore, we can conclude that analysis of the 3D genome structure in MCL is essential to determine target genes of enhancers, as enhancers may not necessary activate their most neighboring gene. This was seen for the newly detected SOX11 enhancer that did not loop to other genes within the same region.
5. Potential impact and use
This project has lead to the detection of new regulatory regions in MCL that may regulate the activation of proteins that contribute to the development and aggressiveness of MCL. This was exemplified by a newly identified enhancer region that potentially regulate SOX11 expression. In line with this finding, we can now discover other regions that show a similar behaviour to better understand the biology of this aggressive disease. In the future this may aid in developing targeted therapies. In addition, the differential DNA methylation patterns may be useful to develop biomarkers that are useful for the detection and stratification of MCLs.