title: Maternal smoking impacts key biological pathways in newborns through epigenetic modification in Utero.
abstract: Children exposed to maternal smoking during pregnancy exhibit increased risk for many adverse health effects. Maternal smoking influences methylation in newborns at specific CpG sites (CpGs). Here, we extend evaluation of individual CpGs to gene-level and pathway-level analyses among 1062 participants in the Norwegian Mother and Child Cohort Study (MoBa) using the Illumina 450 K platform to measure methylation in newborn DNA and maternal smoking in pregnancy, assessed using the biomarker, plasma cotinine. We used novel implementations of bioinformatics tools to collapse epigenome-wide methylation data into gene- and pathway-level effects to test whether exposure to maternal smoking in utero differentially methylated CpGs in genes enriched in biologic pathways. Unlike most pathway analysis applications, our approach allows replication in an independent cohort.
  
  
  
  Data on 485,577 CpGs, mapping to a total of 20,199 genes, were used to create gene scores that were tested for association with maternal plasma cotinine levels using Sequence Kernel Association Test (SKAT), and 15 genes were found to be associated (q < 0.25). Six of these 15 genes (GFI1, MYO1G, CYP1A1, RUNX1, LCTL, and AHRR) contained individual CpGs that were differentially methylated with regards to cotinine levels (p < 1.06 × 10-7). Nine of the 15 genes (FCRLA, MIR641, SLC25A24, TRAK1, C1orf180, ITLN2, GLIS1, LRFN1, and MIR451) were associated with cotinine at the gene-level (q < 0.25) but had no genome-wide significant individual CpGs (p > 1.06 × 10-7). Pathway analyses using gene scores resulted in 51 significantly associated pathways, which we tested for replication in an independent cohort (q < 0.05). Of those 32 replicated in an independent cohort, which clustered into six groups. The largest cluster consisted of pathways related to cancer, cell cycle, ERα receptor signaling, and angiogenesis. The second cluster, organized into five smaller pathway groups, related to immune system function, such as T-cell regulation and other white blood cell related pathways.
  
  
  
  Here we use novel implementations of bioinformatics tools to determine biological pathways impacted through epigenetic changes in utero by maternal smoking in 1062 participants in the MoBa, and successfully replicate these findings in an independent cohort. The results provide new insight into biological mechanisms that may contribute to adverse health effects from exposure to tobacco smoke in utero.
authors.affiliation: Bioinformatics Research Center, North Carolina State University, Raleigh, NC, USA.|Department of Statistics, North Carolina State University, Raleigh, NC, USA.
authors.forename: Daniel M
authors.initials: DM
authors.lastname: Rotroff
authors.affiliation: Division of Intramural Research, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, PO Box 12233, MD A3-05, Research Triangle Park, NC, 27709, USA.
authors.forename: Bonnie R
authors.initials: BR
authors.lastname: Joubert
authors.affiliation: Bioinformatics Research Center, North Carolina State University, Raleigh, NC, USA.
authors.forename: Skylar W
authors.initials: SW
authors.lastname: Marvel
authors.affiliation: Norwegian Institute of Public Health, Oslo, Norway.
authors.forename: Siri E
authors.initials: SE
authors.lastname: Håberg
authors.affiliation: Public Health Sciences Division, Fred Hutchinson Cancer Research Center, Seattle, WA, USA.
authors.forename: Michael C
authors.initials: MC
authors.lastname: Wu
authors.affiliation: Centre for Clinical Research, Haukeland University Hospital, Bergen, Norway.
authors.forename: Roy M
authors.initials: RM
authors.lastname: Nilsen
authors.affiliation: Department of Clinical Science, University of Bergen, Bergen, Norway.|Laboratory of Clinical Biochemistry, Haukeland University Hospital, Bergen, Norway.
authors.forename: Per M
authors.initials: PM
authors.lastname: Ueland
authors.affiliation: Norwegian Institute of Public Health, Oslo, Norway.
authors.forename: Wenche
authors.initials: W
authors.lastname: Nystad
authors.affiliation: Division of Intramural Research, National Institute of Environmental Health Sciences, National Institutes of Health, Department of Health and Human Services, PO Box 12233, MD A3-05, Research Triangle Park, NC, 27709, USA. London2@niehs.nih.gov.
authors.forename: Stephanie J
authors.initials: SJ
authors.lastname: London
authors.affiliation: Bioinformatics Research Center, North Carolina State University, Raleigh, NC, USA.|Department of Statistics, North Carolina State University, Raleigh, NC, USA.|Center for Comparative Medicine and Translational Research, North Carolina State University, Raleigh, NC, USA.
authors.forename: Alison
authors.initials: A
authors.lastname: Motsinger-Reif
mesh_terms: D016000:Cluster Analysis; D018899:CpG Islands; D019175:DNA Methylation; D044127:Epigenesis, Genetic; D005260:Female; D005786:Gene Expression Regulation; D006801:Humans; D018811:Maternal Exposure; D011247:Pregnancy; D011297:Prenatal Exposure Delayed Effects; D015398:Signal Transduction; D012907:Smoking / Q000009:adverse effects*
keywords: Cancer; Epigenetics; In utero; Pathway analysis; Smoking