Limits...
Genome-Wide DNA Methylation Analysis Identifies Novel Hypomethylated Non-Pericentromeric Genes with Potential Clinical Implications in ICF Syndrome.

Simo-Riudalbas L, Diaz-Lagares A, Gatto S, Gagliardi M, Crujeiras AB, Matarazzo MR, Esteller M, Sandoval J - PLoS ONE (2015)

Bottom Line: Our results demonstrate that ICF-specific DNMT3B variants A603T/STP807ins and V699G/R54X cause global DNA hypomethylation compared to wild-type protein.Among the identified hypomethylated CpG-island associated genes, we confirmed the overexpression of three selected genes, BOLL, SYCP2 and NCRNA00221, in ICF compared to healthy controls, which are supposed to be expressed in germ line and silenced in somatic tissues.In conclusion, this study contributes in clarifying the direct relationship between DNA methylation defect and gene expression impairment in ICF syndrome, identifying novel direct target genes of DNMT3B.

View Article: PubMed Central - PubMed

Affiliation: Cancer Epigenetics and Biology Program (PEBC), Bellvitge Biomedical Research Institute (IDIBELL), Barcelona, Catalonia, Spain.

ABSTRACT

Introduction and results: Immunodeficiency, centromeric instability and facial anomalies syndrome (ICF) is a rare autosomal recessive disease, characterized by severe hypomethylation in pericentromeric regions of chromosomes (1, 16 and 9), marked immunodeficiency and facial anomalies. The majority of ICF patients present mutations in the DNMT3B gene, affecting the DNA methyltransferase activity of the protein. In the present study, we have used the Infinium 450K DNA methylation array to evaluate the methylation level of 450,000 CpGs in lymphoblastoid cell lines and untrasformed fibroblasts derived from ICF patients and healthy donors. Our results demonstrate that ICF-specific DNMT3B variants A603T/STP807ins and V699G/R54X cause global DNA hypomethylation compared to wild-type protein. We identified 181 novel differentially methylated positions (DMPs) including subtelomeric and intrachromosomic regions, outside the classical ICF-related pericentromeric hypomethylated positions. Interestingly, these sites were mainly located in intergenic regions and inside the CpG islands. Among the identified hypomethylated CpG-island associated genes, we confirmed the overexpression of three selected genes, BOLL, SYCP2 and NCRNA00221, in ICF compared to healthy controls, which are supposed to be expressed in germ line and silenced in somatic tissues.

Conclusions: In conclusion, this study contributes in clarifying the direct relationship between DNA methylation defect and gene expression impairment in ICF syndrome, identifying novel direct target genes of DNMT3B. A high percentage of the DMPs are located in the subtelomeric regions, indicating a specific role of DNMT3B in methylating these chromosomal sites. Therefore, we provide further evidence that hypomethylation in specific non-pericentromeric regions of chromosomes might be involved in the molecular pathogenesis of ICF syndrome. The detection of DNA hypomethylation at BOLL, SYCP2 and NCRNA00221 may pave the way for the development of specific clinical biomarkers with the aim to facilitate the identification of ICF patients.

No MeSH data available.


Related in: MedlinePlus

Identification of Differentially methylated CpGs.(A) Unsupervised hierarchical clustering and heatmap of four cord blood donors (purple), three unrelated healthy donors (blue) and two ICF patients (orange) using 5000 random selected CpGs. DNA Methylation levels scale is shown. Each column represents patients and each row represents the different CpGs. (B) Supervised cluster and heatmap representing the distinctive 181 CpGs corresponding to the comparison between ICF patients (orange) and all control samples (dark blue).
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4498748&req=5

pone.0132517.g002: Identification of Differentially methylated CpGs.(A) Unsupervised hierarchical clustering and heatmap of four cord blood donors (purple), three unrelated healthy donors (blue) and two ICF patients (orange) using 5000 random selected CpGs. DNA Methylation levels scale is shown. Each column represents patients and each row represents the different CpGs. (B) Supervised cluster and heatmap representing the distinctive 181 CpGs corresponding to the comparison between ICF patients (orange) and all control samples (dark blue).

Mentions: Since we observed that ICF patients with DNMT3B mutations show a global reduction of DNA methylation, we focused on studying the CpG positions with loss of methylation. To gain robustness and reliability, we added to the previous set of ICF1 samples four new controls of peripheral mononuclear cells obtained from cord blood samples (CB10, CB13, CB20 and CB76), being aware of the limitation of the cell type heterogeneity in these samples. The unsupervised hierarchical clustering, using 5000 random CpGs mimicking the global methylome, shows that samples are grouped based on their tissue type. The methylome of the four peripheral mononuclear derived cells samples is homogeneous and clustered together. The immortalized cell lines derived from controls and ICF patients clusterized in the same group, but in a separate subgroup. Finally, fibroblast cells clustered in an independent group (Fig 2A). These results indicate that the distance between samples is mainly due to tissue type.


Genome-Wide DNA Methylation Analysis Identifies Novel Hypomethylated Non-Pericentromeric Genes with Potential Clinical Implications in ICF Syndrome.

Simo-Riudalbas L, Diaz-Lagares A, Gatto S, Gagliardi M, Crujeiras AB, Matarazzo MR, Esteller M, Sandoval J - PLoS ONE (2015)

Identification of Differentially methylated CpGs.(A) Unsupervised hierarchical clustering and heatmap of four cord blood donors (purple), three unrelated healthy donors (blue) and two ICF patients (orange) using 5000 random selected CpGs. DNA Methylation levels scale is shown. Each column represents patients and each row represents the different CpGs. (B) Supervised cluster and heatmap representing the distinctive 181 CpGs corresponding to the comparison between ICF patients (orange) and all control samples (dark blue).
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4498748&req=5

pone.0132517.g002: Identification of Differentially methylated CpGs.(A) Unsupervised hierarchical clustering and heatmap of four cord blood donors (purple), three unrelated healthy donors (blue) and two ICF patients (orange) using 5000 random selected CpGs. DNA Methylation levels scale is shown. Each column represents patients and each row represents the different CpGs. (B) Supervised cluster and heatmap representing the distinctive 181 CpGs corresponding to the comparison between ICF patients (orange) and all control samples (dark blue).
Mentions: Since we observed that ICF patients with DNMT3B mutations show a global reduction of DNA methylation, we focused on studying the CpG positions with loss of methylation. To gain robustness and reliability, we added to the previous set of ICF1 samples four new controls of peripheral mononuclear cells obtained from cord blood samples (CB10, CB13, CB20 and CB76), being aware of the limitation of the cell type heterogeneity in these samples. The unsupervised hierarchical clustering, using 5000 random CpGs mimicking the global methylome, shows that samples are grouped based on their tissue type. The methylome of the four peripheral mononuclear derived cells samples is homogeneous and clustered together. The immortalized cell lines derived from controls and ICF patients clusterized in the same group, but in a separate subgroup. Finally, fibroblast cells clustered in an independent group (Fig 2A). These results indicate that the distance between samples is mainly due to tissue type.

Bottom Line: Our results demonstrate that ICF-specific DNMT3B variants A603T/STP807ins and V699G/R54X cause global DNA hypomethylation compared to wild-type protein.Among the identified hypomethylated CpG-island associated genes, we confirmed the overexpression of three selected genes, BOLL, SYCP2 and NCRNA00221, in ICF compared to healthy controls, which are supposed to be expressed in germ line and silenced in somatic tissues.In conclusion, this study contributes in clarifying the direct relationship between DNA methylation defect and gene expression impairment in ICF syndrome, identifying novel direct target genes of DNMT3B.

View Article: PubMed Central - PubMed

Affiliation: Cancer Epigenetics and Biology Program (PEBC), Bellvitge Biomedical Research Institute (IDIBELL), Barcelona, Catalonia, Spain.

ABSTRACT

Introduction and results: Immunodeficiency, centromeric instability and facial anomalies syndrome (ICF) is a rare autosomal recessive disease, characterized by severe hypomethylation in pericentromeric regions of chromosomes (1, 16 and 9), marked immunodeficiency and facial anomalies. The majority of ICF patients present mutations in the DNMT3B gene, affecting the DNA methyltransferase activity of the protein. In the present study, we have used the Infinium 450K DNA methylation array to evaluate the methylation level of 450,000 CpGs in lymphoblastoid cell lines and untrasformed fibroblasts derived from ICF patients and healthy donors. Our results demonstrate that ICF-specific DNMT3B variants A603T/STP807ins and V699G/R54X cause global DNA hypomethylation compared to wild-type protein. We identified 181 novel differentially methylated positions (DMPs) including subtelomeric and intrachromosomic regions, outside the classical ICF-related pericentromeric hypomethylated positions. Interestingly, these sites were mainly located in intergenic regions and inside the CpG islands. Among the identified hypomethylated CpG-island associated genes, we confirmed the overexpression of three selected genes, BOLL, SYCP2 and NCRNA00221, in ICF compared to healthy controls, which are supposed to be expressed in germ line and silenced in somatic tissues.

Conclusions: In conclusion, this study contributes in clarifying the direct relationship between DNA methylation defect and gene expression impairment in ICF syndrome, identifying novel direct target genes of DNMT3B. A high percentage of the DMPs are located in the subtelomeric regions, indicating a specific role of DNMT3B in methylating these chromosomal sites. Therefore, we provide further evidence that hypomethylation in specific non-pericentromeric regions of chromosomes might be involved in the molecular pathogenesis of ICF syndrome. The detection of DNA hypomethylation at BOLL, SYCP2 and NCRNA00221 may pave the way for the development of specific clinical biomarkers with the aim to facilitate the identification of ICF patients.

No MeSH data available.


Related in: MedlinePlus