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CRTC1 gene is differentially methylated in the human hippocampus in Alzheimer's disease.

Mendioroz M, Celarain N, Altuna M, Sánchez-Ruiz de Gordoa J, Zelaya MV, Roldán M, Rubio I, Larumbe R, Erro ME, Méndez I, Echávarri C - Alzheimers Res Ther (2016)

Bottom Line: Moreover, a 1.54-fold decrease in CRTC1 mRNA levels was observed in hippocampus of AD cases compared to controls (p < 0.05) supporting the notion that CRTC1 is downregulated in the AD hippocampus.DNA methylation levels within two distinct promoter regions of the CRTC1 gene were decreased in human hippocampus affected by AD compared with controls and methylation within Prom1 showed a strong inverse correlation with p-tau deposition.Further studies are guaranteed to elucidate the precise role that CRTC1 methylation plays in AD pathophysiology.

View Article: PubMed Central - PubMed

Affiliation: NeuroEpigenetics Laboratory, Navarrabiomed- IdiSNA (Navarra Institute for Health Research), c/ Irunlarrea, 3, Pamplona, Navarra, 31008, Spain. maitemendilab@gmail.com.

ABSTRACT

Background: CRTC1 (CREB regulated transcription coactivator 1) gene plays a role in synaptic plasticity, learning and long-term memory formation in the hippocampus. Recently, CRTC1 has been shown to be downregulated in Alzheimer's disease (AD). Nevertheless, the mechanisms underlying CRTC1 dysregulation in AD remain unclear.

Methods: To understand better the epigenetic mechanisms regulating CRTC1 expression that may be altered in AD, we profiled DNA methylation at CpG site resolution by bisulfite cloning sequencing in two promoter regions (referred to as Prom1 and Prom2) of the CRTC1 gene in human hippocampus from controls and AD cases. Next, we correlated DNA methylation levels with AD-related pathology, i.e., β-amyloid and phosphorylated-tau (p-tau) burden and also measured CRTC1 mRNA levels by RT-qPCR.

Results: Methylation levels were lower in AD cases as compared to controls within both promoter regions (Prom1: 0.95% vs. 5%, p-value < 0.01 and Prom2: 2.80% vs. 17.80%, p-value < 0.001). Interestingly, CRTC1 methylation levels inversely correlated with AD-related neuropathological changes, particularly with p-tau deposition (rSpearman = -0.903, p < 0.001). Moreover, a 1.54-fold decrease in CRTC1 mRNA levels was observed in hippocampus of AD cases compared to controls (p < 0.05) supporting the notion that CRTC1 is downregulated in the AD hippocampus.

Conclusions: DNA methylation levels within two distinct promoter regions of the CRTC1 gene were decreased in human hippocampus affected by AD compared with controls and methylation within Prom1 showed a strong inverse correlation with p-tau deposition. Further studies are guaranteed to elucidate the precise role that CRTC1 methylation plays in AD pathophysiology.

No MeSH data available.


Related in: MedlinePlus

DNA methylation levels within two promoter regions of CRTC1 in Alzheimer’s disease (AD) and control hippocampus. a The graph shows genomic position of the two amplicons (Prom1 and Prom2) within the two promoter regions of the CRTC1 gene that were examined by bisulfite cloning sequencing. At the bottom of the graph, predicted functional elements are shown for each of nine human cell lines explored by chromatin immunoprecipitation (ChIP) combined with massively parallel DNA sequencing. Boxes represent promoter regions (red), enhancers (yellow), transcriptional transition and elongation (dark green) and weak transcribed regions (light green). CpG islands are also represented by green boxes. The track was obtained from Chromatin State Segmentation by HMM from ENCODE/Broad track shown at the UCSC Genome Browser. Next, dot plot charts and representative examples of the methylation graphs for Prom1 (b) and Prom2 (c), respectively, are shown. A decrease in methylation was observed within both promoters in AD hippocampus compared to controls. Horizontal lines within dot plots represent median methylation values for each group. Boxes on the right represent individual patients. Black and white circles denote methylated and unmethylated cytosines, respectively. Each column symbolizes a unique CpG site in the examined amplicon and each line represents an individual DNA clone. *p-value < 0.05; ** p-value < 0.005. CpG cytosine-phosphate-guanine
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Fig1: DNA methylation levels within two promoter regions of CRTC1 in Alzheimer’s disease (AD) and control hippocampus. a The graph shows genomic position of the two amplicons (Prom1 and Prom2) within the two promoter regions of the CRTC1 gene that were examined by bisulfite cloning sequencing. At the bottom of the graph, predicted functional elements are shown for each of nine human cell lines explored by chromatin immunoprecipitation (ChIP) combined with massively parallel DNA sequencing. Boxes represent promoter regions (red), enhancers (yellow), transcriptional transition and elongation (dark green) and weak transcribed regions (light green). CpG islands are also represented by green boxes. The track was obtained from Chromatin State Segmentation by HMM from ENCODE/Broad track shown at the UCSC Genome Browser. Next, dot plot charts and representative examples of the methylation graphs for Prom1 (b) and Prom2 (c), respectively, are shown. A decrease in methylation was observed within both promoters in AD hippocampus compared to controls. Horizontal lines within dot plots represent median methylation values for each group. Boxes on the right represent individual patients. Black and white circles denote methylated and unmethylated cytosines, respectively. Each column symbolizes a unique CpG site in the examined amplicon and each line represents an individual DNA clone. *p-value < 0.05; ** p-value < 0.005. CpG cytosine-phosphate-guanine

Mentions: To begin to ask whether DNA methylation within CRTC1 is altered in AD hippocampus, two different promoters of the CRTC1 gene were examined by bisulfite cloning sequencing. Promoters were identified by using chromatin immunoprecipitation sequencing (ChIP-seq) data from the track Chromatin State Segmentation by HMM (Hidden Markov Model) from ENCODE project and Broad Institute, shown at the UCSC Genome Browser [33]. This track displays chromatin state segmentation data for each of nine human cell types (Fig. 1a, Additional file 1: Figure S3). We designed primers to amplify two independent amplicons. Promoter 1 (Prom1) amplicon was placed at the 5’ end of CRTC1, partially overlapping exon 1 and the beginning of intron 1, and contained 37 CpG sites within the southern shore of a 628 bp-CpG island (chr19:18794192–18794819). Whereas promoter 2 (Prom2) amplicon was located within intron 1 and encompassed 15 CpG sites at the southern shore of a 230 bp-CpG island (chr19:18811562–18811791) (Fig. 1a, Additional file 1: Figure S3). To sum up, a total of 52 individual CpG sites, representing 63.8 % and 88.2 % of each CpG island, respectively, were analyzed in a subset of five control and eight AD cases. To calculate the average methylation level, DNA methylation percentage was measured at CpG site resolution and further averaged across all the CpG sites for each promoter region and subject. Interestingly, we found that average methylation level at Prom1 was decreased in AD cases compared to controls (0.95 % vs. 5 %, p-value < 0.01) (Fig. 1b). Similarly, average methylation level was significantly decreased within Prom2 in AD cases compared to controls (2.80 % vs. 17.80 %, p-value < 0.001) (Fig. 1c).Fig. 1


CRTC1 gene is differentially methylated in the human hippocampus in Alzheimer's disease.

Mendioroz M, Celarain N, Altuna M, Sánchez-Ruiz de Gordoa J, Zelaya MV, Roldán M, Rubio I, Larumbe R, Erro ME, Méndez I, Echávarri C - Alzheimers Res Ther (2016)

DNA methylation levels within two promoter regions of CRTC1 in Alzheimer’s disease (AD) and control hippocampus. a The graph shows genomic position of the two amplicons (Prom1 and Prom2) within the two promoter regions of the CRTC1 gene that were examined by bisulfite cloning sequencing. At the bottom of the graph, predicted functional elements are shown for each of nine human cell lines explored by chromatin immunoprecipitation (ChIP) combined with massively parallel DNA sequencing. Boxes represent promoter regions (red), enhancers (yellow), transcriptional transition and elongation (dark green) and weak transcribed regions (light green). CpG islands are also represented by green boxes. The track was obtained from Chromatin State Segmentation by HMM from ENCODE/Broad track shown at the UCSC Genome Browser. Next, dot plot charts and representative examples of the methylation graphs for Prom1 (b) and Prom2 (c), respectively, are shown. A decrease in methylation was observed within both promoters in AD hippocampus compared to controls. Horizontal lines within dot plots represent median methylation values for each group. Boxes on the right represent individual patients. Black and white circles denote methylated and unmethylated cytosines, respectively. Each column symbolizes a unique CpG site in the examined amplicon and each line represents an individual DNA clone. *p-value < 0.05; ** p-value < 0.005. CpG cytosine-phosphate-guanine
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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Fig1: DNA methylation levels within two promoter regions of CRTC1 in Alzheimer’s disease (AD) and control hippocampus. a The graph shows genomic position of the two amplicons (Prom1 and Prom2) within the two promoter regions of the CRTC1 gene that were examined by bisulfite cloning sequencing. At the bottom of the graph, predicted functional elements are shown for each of nine human cell lines explored by chromatin immunoprecipitation (ChIP) combined with massively parallel DNA sequencing. Boxes represent promoter regions (red), enhancers (yellow), transcriptional transition and elongation (dark green) and weak transcribed regions (light green). CpG islands are also represented by green boxes. The track was obtained from Chromatin State Segmentation by HMM from ENCODE/Broad track shown at the UCSC Genome Browser. Next, dot plot charts and representative examples of the methylation graphs for Prom1 (b) and Prom2 (c), respectively, are shown. A decrease in methylation was observed within both promoters in AD hippocampus compared to controls. Horizontal lines within dot plots represent median methylation values for each group. Boxes on the right represent individual patients. Black and white circles denote methylated and unmethylated cytosines, respectively. Each column symbolizes a unique CpG site in the examined amplicon and each line represents an individual DNA clone. *p-value < 0.05; ** p-value < 0.005. CpG cytosine-phosphate-guanine
Mentions: To begin to ask whether DNA methylation within CRTC1 is altered in AD hippocampus, two different promoters of the CRTC1 gene were examined by bisulfite cloning sequencing. Promoters were identified by using chromatin immunoprecipitation sequencing (ChIP-seq) data from the track Chromatin State Segmentation by HMM (Hidden Markov Model) from ENCODE project and Broad Institute, shown at the UCSC Genome Browser [33]. This track displays chromatin state segmentation data for each of nine human cell types (Fig. 1a, Additional file 1: Figure S3). We designed primers to amplify two independent amplicons. Promoter 1 (Prom1) amplicon was placed at the 5’ end of CRTC1, partially overlapping exon 1 and the beginning of intron 1, and contained 37 CpG sites within the southern shore of a 628 bp-CpG island (chr19:18794192–18794819). Whereas promoter 2 (Prom2) amplicon was located within intron 1 and encompassed 15 CpG sites at the southern shore of a 230 bp-CpG island (chr19:18811562–18811791) (Fig. 1a, Additional file 1: Figure S3). To sum up, a total of 52 individual CpG sites, representing 63.8 % and 88.2 % of each CpG island, respectively, were analyzed in a subset of five control and eight AD cases. To calculate the average methylation level, DNA methylation percentage was measured at CpG site resolution and further averaged across all the CpG sites for each promoter region and subject. Interestingly, we found that average methylation level at Prom1 was decreased in AD cases compared to controls (0.95 % vs. 5 %, p-value < 0.01) (Fig. 1b). Similarly, average methylation level was significantly decreased within Prom2 in AD cases compared to controls (2.80 % vs. 17.80 %, p-value < 0.001) (Fig. 1c).Fig. 1

Bottom Line: Moreover, a 1.54-fold decrease in CRTC1 mRNA levels was observed in hippocampus of AD cases compared to controls (p < 0.05) supporting the notion that CRTC1 is downregulated in the AD hippocampus.DNA methylation levels within two distinct promoter regions of the CRTC1 gene were decreased in human hippocampus affected by AD compared with controls and methylation within Prom1 showed a strong inverse correlation with p-tau deposition.Further studies are guaranteed to elucidate the precise role that CRTC1 methylation plays in AD pathophysiology.

View Article: PubMed Central - PubMed

Affiliation: NeuroEpigenetics Laboratory, Navarrabiomed- IdiSNA (Navarra Institute for Health Research), c/ Irunlarrea, 3, Pamplona, Navarra, 31008, Spain. maitemendilab@gmail.com.

ABSTRACT

Background: CRTC1 (CREB regulated transcription coactivator 1) gene plays a role in synaptic plasticity, learning and long-term memory formation in the hippocampus. Recently, CRTC1 has been shown to be downregulated in Alzheimer's disease (AD). Nevertheless, the mechanisms underlying CRTC1 dysregulation in AD remain unclear.

Methods: To understand better the epigenetic mechanisms regulating CRTC1 expression that may be altered in AD, we profiled DNA methylation at CpG site resolution by bisulfite cloning sequencing in two promoter regions (referred to as Prom1 and Prom2) of the CRTC1 gene in human hippocampus from controls and AD cases. Next, we correlated DNA methylation levels with AD-related pathology, i.e., β-amyloid and phosphorylated-tau (p-tau) burden and also measured CRTC1 mRNA levels by RT-qPCR.

Results: Methylation levels were lower in AD cases as compared to controls within both promoter regions (Prom1: 0.95% vs. 5%, p-value < 0.01 and Prom2: 2.80% vs. 17.80%, p-value < 0.001). Interestingly, CRTC1 methylation levels inversely correlated with AD-related neuropathological changes, particularly with p-tau deposition (rSpearman = -0.903, p < 0.001). Moreover, a 1.54-fold decrease in CRTC1 mRNA levels was observed in hippocampus of AD cases compared to controls (p < 0.05) supporting the notion that CRTC1 is downregulated in the AD hippocampus.

Conclusions: DNA methylation levels within two distinct promoter regions of the CRTC1 gene were decreased in human hippocampus affected by AD compared with controls and methylation within Prom1 showed a strong inverse correlation with p-tau deposition. Further studies are guaranteed to elucidate the precise role that CRTC1 methylation plays in AD pathophysiology.

No MeSH data available.


Related in: MedlinePlus