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Analysis of DNA methylation landscape reveals the roles of DNA methylation in the regulation of drug metabolizing enzymes.

Habano W, Kawamura K, Iizuka N, Terashima J, Sugai T, Ozawa S - Clin Epigenetics (2015)

Bottom Line: Moreover, tissue-specific and age-dependent expression of UDP-glucuronosyltransferase 1A splicing variants was associated with DNA methylation status of individual first exons.Some DME genes were regulated by DNA methylation, potentially resulting in inter- and intra-individual differences in drug metabolism.Analysis of DNA methylation landscape facilitated elucidation of the role of DNA methylation in the regulation of DME genes, such as mediator of inter-individual variability, guide for correct alternative splicing, and potential tumor-suppressor or housekeeper.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacodynamics and Molecular Genetics, School of Pharmacy, Iwate Medical University, 2-1-1 Nishitokuta, Yahaba-Cho, Shiwa-Gun 028-3694 Japan.

ABSTRACT

Background: Drug metabolizing enzymes (DMEs) exhibit dramatic inter- and intra-individual variability in expression and activity. However, the mechanisms determining this variability have not been fully elucidated. The aim of this study was to evaluate the biological significance of DNA methylation in the regulation of DME genes by genome-wide integrative analysis.

Results: DNA methylation and mRNA expression profiles of human tissues and hepatoma cells were examined by microarrays. The data were combined with GEO datasets of liver tissues, and integrative analysis was performed on selected DME genes. Detailed DNA methylation statuses at individual CpG sites were evaluated by DNA methylation mapping. From analysis of 20 liver tissues, highly variable DNA methylation was observed in 37 DME genes, 7 of which showed significant inverse correlations between DNA methylation and mRNA expression. In hepatoma cells, treatment with a demethylating agent resulted in upregulation of 5 DME genes, which could be explained by DNA methylation status. Interestingly, some DMEs were suggested to act as tumor-suppressor or housekeeper based on their unique DNA methylation features. Moreover, tissue-specific and age-dependent expression of UDP-glucuronosyltransferase 1A splicing variants was associated with DNA methylation status of individual first exons.

Conclusions: Some DME genes were regulated by DNA methylation, potentially resulting in inter- and intra-individual differences in drug metabolism. Analysis of DNA methylation landscape facilitated elucidation of the role of DNA methylation in the regulation of DME genes, such as mediator of inter-individual variability, guide for correct alternative splicing, and potential tumor-suppressor or housekeeper.

No MeSH data available.


Related in: MedlinePlus

Representative results of DNA methylation mapping of UGT1A isoforms. The levels of the DNA methylation of hepatic-type (UGT1A1, UGT1A4, UGT1A6, and UGT1A9) and intestinal-type (UGT1A10 and UGT1A8) genes were examined in adult liver tissues (NLA and NL2), fetal liver tissue (NLF), and adult small intestinal tissue (NSI). In each panel, the open or closed circles located on the horizontal axis indicate the positions of CpG sites arranged in the 5′ to 3′ direction according to the relative distance of each CpG site. The closed circles indicate CpG sites in which tissue-specific expression could be explained by the DNA methylation status
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Fig6: Representative results of DNA methylation mapping of UGT1A isoforms. The levels of the DNA methylation of hepatic-type (UGT1A1, UGT1A4, UGT1A6, and UGT1A9) and intestinal-type (UGT1A10 and UGT1A8) genes were examined in adult liver tissues (NLA and NL2), fetal liver tissue (NLF), and adult small intestinal tissue (NSI). In each panel, the open or closed circles located on the horizontal axis indicate the positions of CpG sites arranged in the 5′ to 3′ direction according to the relative distance of each CpG site. The closed circles indicate CpG sites in which tissue-specific expression could be explained by the DNA methylation status

Mentions: Next, the transcript levels of UGT1A isoforms were examined in different tissues using quantitative real-time PCR. The tissue-specific expression profiles shown in Additional file 7: Figure S7 were similar to the results of a previous study [10–12]. The UGT1A genes were then classified into two major groups according to the tissues in which they were dominantly expressed: hepatic type (i.e., UGT1A1, UGT1A3, UGT1A4, UGT1A6, and UGT1A9) and intestinal type (i.e., UGT1A5, UGT1A7, UGT1A8, and UGT1A10). Although the level of UGT1A8 expression in adult livers was somewhat higher than that in small intestines, UGT1A8, unlike the other hepatic-type UGT1As, showed relatively high expression in the small intestine as well. Therefore, we classified UGT1A8 as an intestinal-type gene. DNA methylation mapping on the UGT1A locus revealed that DNA methylation status was largely variable among different tissues (Additional file 8: Figure S8). We focused on the first exon of each isoform and found that the hepatic-type genes UGT1A1, UGT1A4, UGT1A6, and UGT1A9 tended to be methylated at higher levels in the small intestine than in the liver (Fig. 6). In order to perform statistical analysis, moreover, DNA methylation status in individual CpG sites were also compared between 18 adult livers and one normal small intestine (NSI) or one normal fetal liver (NLF). We further focused on the CpG sites most proximal to each transcription start site (excluding more distal CpG sites of TSS1500). In hepatic-type UGT1A genes, as a result, we found that β values of the NSI and NLF were statistically higher than medians of 18 individual β values of normal adult livers (Additional file 9: Table S1). In contrast, the methylation levels of the intestinal-type genes UGT1A10 and UGT1A8 were relatively higher in the liver than in the small intestine. This suggested that DNA methylation status around the first exon determined the splicing isoforms of the UGT1A gene and led to tissue-specific expression. In addition, higher levels of DNA methylation were observed in fetal livers than in adult livers. This tendency was found in hepatic-type genes rather than intestinal-type genes, implying that the low levels of expression observed in the fetal liver may result from downregulation by DNA methylation.Fig. 6


Analysis of DNA methylation landscape reveals the roles of DNA methylation in the regulation of drug metabolizing enzymes.

Habano W, Kawamura K, Iizuka N, Terashima J, Sugai T, Ozawa S - Clin Epigenetics (2015)

Representative results of DNA methylation mapping of UGT1A isoforms. The levels of the DNA methylation of hepatic-type (UGT1A1, UGT1A4, UGT1A6, and UGT1A9) and intestinal-type (UGT1A10 and UGT1A8) genes were examined in adult liver tissues (NLA and NL2), fetal liver tissue (NLF), and adult small intestinal tissue (NSI). In each panel, the open or closed circles located on the horizontal axis indicate the positions of CpG sites arranged in the 5′ to 3′ direction according to the relative distance of each CpG site. The closed circles indicate CpG sites in which tissue-specific expression could be explained by the DNA methylation status
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4587720&req=5

Fig6: Representative results of DNA methylation mapping of UGT1A isoforms. The levels of the DNA methylation of hepatic-type (UGT1A1, UGT1A4, UGT1A6, and UGT1A9) and intestinal-type (UGT1A10 and UGT1A8) genes were examined in adult liver tissues (NLA and NL2), fetal liver tissue (NLF), and adult small intestinal tissue (NSI). In each panel, the open or closed circles located on the horizontal axis indicate the positions of CpG sites arranged in the 5′ to 3′ direction according to the relative distance of each CpG site. The closed circles indicate CpG sites in which tissue-specific expression could be explained by the DNA methylation status
Mentions: Next, the transcript levels of UGT1A isoforms were examined in different tissues using quantitative real-time PCR. The tissue-specific expression profiles shown in Additional file 7: Figure S7 were similar to the results of a previous study [10–12]. The UGT1A genes were then classified into two major groups according to the tissues in which they were dominantly expressed: hepatic type (i.e., UGT1A1, UGT1A3, UGT1A4, UGT1A6, and UGT1A9) and intestinal type (i.e., UGT1A5, UGT1A7, UGT1A8, and UGT1A10). Although the level of UGT1A8 expression in adult livers was somewhat higher than that in small intestines, UGT1A8, unlike the other hepatic-type UGT1As, showed relatively high expression in the small intestine as well. Therefore, we classified UGT1A8 as an intestinal-type gene. DNA methylation mapping on the UGT1A locus revealed that DNA methylation status was largely variable among different tissues (Additional file 8: Figure S8). We focused on the first exon of each isoform and found that the hepatic-type genes UGT1A1, UGT1A4, UGT1A6, and UGT1A9 tended to be methylated at higher levels in the small intestine than in the liver (Fig. 6). In order to perform statistical analysis, moreover, DNA methylation status in individual CpG sites were also compared between 18 adult livers and one normal small intestine (NSI) or one normal fetal liver (NLF). We further focused on the CpG sites most proximal to each transcription start site (excluding more distal CpG sites of TSS1500). In hepatic-type UGT1A genes, as a result, we found that β values of the NSI and NLF were statistically higher than medians of 18 individual β values of normal adult livers (Additional file 9: Table S1). In contrast, the methylation levels of the intestinal-type genes UGT1A10 and UGT1A8 were relatively higher in the liver than in the small intestine. This suggested that DNA methylation status around the first exon determined the splicing isoforms of the UGT1A gene and led to tissue-specific expression. In addition, higher levels of DNA methylation were observed in fetal livers than in adult livers. This tendency was found in hepatic-type genes rather than intestinal-type genes, implying that the low levels of expression observed in the fetal liver may result from downregulation by DNA methylation.Fig. 6

Bottom Line: Moreover, tissue-specific and age-dependent expression of UDP-glucuronosyltransferase 1A splicing variants was associated with DNA methylation status of individual first exons.Some DME genes were regulated by DNA methylation, potentially resulting in inter- and intra-individual differences in drug metabolism.Analysis of DNA methylation landscape facilitated elucidation of the role of DNA methylation in the regulation of DME genes, such as mediator of inter-individual variability, guide for correct alternative splicing, and potential tumor-suppressor or housekeeper.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacodynamics and Molecular Genetics, School of Pharmacy, Iwate Medical University, 2-1-1 Nishitokuta, Yahaba-Cho, Shiwa-Gun 028-3694 Japan.

ABSTRACT

Background: Drug metabolizing enzymes (DMEs) exhibit dramatic inter- and intra-individual variability in expression and activity. However, the mechanisms determining this variability have not been fully elucidated. The aim of this study was to evaluate the biological significance of DNA methylation in the regulation of DME genes by genome-wide integrative analysis.

Results: DNA methylation and mRNA expression profiles of human tissues and hepatoma cells were examined by microarrays. The data were combined with GEO datasets of liver tissues, and integrative analysis was performed on selected DME genes. Detailed DNA methylation statuses at individual CpG sites were evaluated by DNA methylation mapping. From analysis of 20 liver tissues, highly variable DNA methylation was observed in 37 DME genes, 7 of which showed significant inverse correlations between DNA methylation and mRNA expression. In hepatoma cells, treatment with a demethylating agent resulted in upregulation of 5 DME genes, which could be explained by DNA methylation status. Interestingly, some DMEs were suggested to act as tumor-suppressor or housekeeper based on their unique DNA methylation features. Moreover, tissue-specific and age-dependent expression of UDP-glucuronosyltransferase 1A splicing variants was associated with DNA methylation status of individual first exons.

Conclusions: Some DME genes were regulated by DNA methylation, potentially resulting in inter- and intra-individual differences in drug metabolism. Analysis of DNA methylation landscape facilitated elucidation of the role of DNA methylation in the regulation of DME genes, such as mediator of inter-individual variability, guide for correct alternative splicing, and potential tumor-suppressor or housekeeper.

No MeSH data available.


Related in: MedlinePlus