DNA methylation shows genome-wide association of NFIX, RAPGEF2 and MSRB3 with gestational age at birth

Center for Epigenetics, Johns Hopkins School of Medicine, Baltimore, MD 21205, USA.
International Journal of Epidemiology (Impact Factor: 9.18). 02/2012; 41(1):188-99. DOI: 10.1093/ije/dyr237
Source: PubMed


Gestational age at birth strongly predicts neonatal, adolescent and adult morbidity and mortality through mostly unknown mechanisms. Identification of specific genes that are undergoing regulatory change prior to birth, such as through changes in DNA methylation, would increase our understanding of developmental changes occurring during the third trimester and consequences of pre-term birth (PTB).
We performed a genome-wide analysis of DNA methylation (using microarrays, specifically CHARM 2.0) in 141 newborns collected in Baltimore, MD, using novel statistical methodology to identify genomic regions associated with gestational age at birth. Bisulphite pyrosequencing was used to validate significant differentially methylated regions (DMRs), and real-time PCR was performed to assess functional significance of differential methylation in a subset of newborns.
We identified three DMRs at genome-wide significance levels adjacent to the NFIX, RAPGEF2 and MSRB3 genes. All three regions were validated by pyrosequencing, and RAGPEF2 also showed an inverse correlation between DNA methylation levels and gene expression levels. Although the three DMRs appear very dynamic with gestational age in our newborn sample, adult DNA methylation levels at these regions are stable and of equal or greater magnitude than the oldest neonate, directionally consistent with the gestational age results.
We have identified three differentially methylated regions associated with gestational age at birth. All three nearby genes play important roles in the development of several organs, including skeletal muscle, brain and haematopoietic system. Therefore, they may provide initial insight into the basis of PTB's negative health outcomes. The genome-wide custom DNA methylation array technology and novel statistical methods employed in this study could constitute a model for epidemiologic studies of epigenetic variation.

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    • "To determine the adverse effects at the single locus level of the observed confounding between age, cell composition, and DNAm, we reexamined the CpGs reported in the literature to be associated with age [6-13] across several different measurement platforms (Additional file 4). For each of the CpGs reported to associate with age on the Illumina 450k array (n = 134,489), we tested between-to-within cell type variability on the sorted DNAm data and found that 86.7% of these had P < 0.05 across cell type (Figure S5 in Additional file 2). "
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    • "Genomic regions subject to DNA methylation change have been identified during gestation [20-22], neonatal development [23] and the entire lifespan [24-28]. The aim of this exploratory study was to assess genome-wide DNA methylation profiles of extremely preterm survivors compared with term controls at both birth and at 18 years of age, using a longitudinal case-control study design. "
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    • "Application of the CHARM method may be possible with future higher density designs of the Infinium array enabling clustering of MVPs. CHARM arrays have recently been technologically extended to encompass significantly larger numbers of CpGs up to 5.2 million CpG sites [56] and although this would not cover all 28 million CpGs in the human genome, this method may offer a suitable alternative that has some of the benefits of MeDIP-seq with the flexibility and cost efficiency of an array. "
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