Figure 1.
Location of analyzed CpG sites and global DNA methylation in human adipose tissue.
All CpG sites analyzed on the Infinium HumanMethylation450 BeadChip are mapped to gene regions based on functional genome distribution (A) and to CpG island regions based on CpG content and neighbourhood context (B). In the lower panels, global DNA methylation in human adipose tissue is shown for each gene region (C) and for CpG island regions (D). Global DNA methylation is calculated as average DNA methylation based on all CpG sites in each region on the chip, and presented separately for Infinium I and Infinium II assays, respectively. Data is presented as mean ± SD. TSS, proximal promoter, defined as 200 bp (basepairs) or 1500 bp upstream of the transcription start site; UTR, untranslated region; CpG island, 200 bp (or more) stretch of DNA with a C+G content of 50% and an observed CpG/expected CpG in excess of 0.6; Shelf, regions flanking island shores, i.e., covering 2000–4000 bp distant from the CpG island; Shore: the flanking region of CpG islands, 0–2000 bp. *Significant difference between average DNA methylation before versus after exercise, q<0.05.
Table 1.
Clinical characteristics of study participants (n = 23) with DNA methylation data both before (baseline) and after the exercise intervention.
Figure 2.
Analysis flowchart.
Figure 3.
DNA methylation of individual CpG sites.
The absolute change in DNA methylation of individual CpG sites with a significant difference after exercise compared with baseline (q<0.05) ranges from 0.2–10.9% (A and B). A) Number of sites with increased methylation in adipose tissue in response to exercise (n = 16,470). B) Number of sites with decreased DNA methylation in adipose tissue in response to exercise (n = 1,505). Panels C and D show the distribution of CpG sites with a significant change (q<0.05) and an absolute difference ≥5% in DNA methylation in adipose tissue before versus after exercise, in comparison to all analyzed sites on the Infinium HumanMethylation450 BeadChip. C) Distribution of significant CpG sites vs. all analyzed sites in relation to nearest gene regions. D) Distribution of significant CpG sites vs. all analyzed sites in relation to CpG island regions. *The overall distribution of significant CpG sites compared with all analyzed sites on the Infinium HumanMethylation450 BeadChip was analyzed using a chi2 test.
Table 2.
Changes in adipose tissue DNA methylation in response to a 6 months exercise intervention. Most significant CpG sites (q<0.005) with a difference in DNA methylation ≥5%.
Table 3.
Changes in adipose tissue DNA methylation in response to a 6 months exercise intervention. Significant CpG sites (q<0.05) with the biggest change in DNA methylation (>8%).
Figure 4.
DNA methylation of RALBP1 is associated with a decrease in gene expression.
A CpG site in the promoter region of RALBP1 showed A) increased DNA methylation in response to exercise as well as B) a decrease in mRNA expression. C) In vitro DNA methylation of the RALBP1 promoter decreased gene expression, as measured by luciferase activity. The result represents the mean of three independent experiments, and the values in each experiment are the mean of five replicates (background control subtracted). Data is presented as mean ± SEM.
Table 4.
Individual CpG sites located within/near candidate genes for obesity [3], with a significant change in DNA methylation in adipose tissue in response to exercise.
Table 5.
Individual CpG sites located within/near candidate genes for T2D [3], with a significant change in DNA methylation in adipose tissue in response to exercise.
Figure 5.
Silencing of Hdac4 and Ncor2 in 3T3-L1 adipocytes results in increased lipogenesis.
CpG sites in the promoter region of A) HDAC4 and B) NCOR2 showed increased DNA methylation in response to exercise as well as decreased mRNA expression (C–D). Knock-downs were verified either by E) Western blot analysis (for Hdac4) or F) by qRT-PCR (for Ncor2). Lipogenesis increased in 3T3-L1 adipocytes where G) Hdac4 (n = 5) or H) Ncor2 (n = 5) had been silenced. Data is presented as mean ± SEM.