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Alcohol Clin Exp Res. 2013 Oct;37(10):1657-67. doi: 10.1111/acer.12139. Epub 2013 Jun 25.

Suppression and epigenetic regulation of MiR-9 contributes to ethanol teratology: evidence from zebrafish and murine fetal neural stem cell models.

Author information

1
Department of Neuroscience and Experimental Therapeutics, Texas A&M Health Science Center, College of Medicine, Bryan, Texas.

Abstract

BACKGROUND:

Fetal alcohol exposure produces multiorgan defects, making it difficult to identify underlying etiological mechanisms. However, recent evidence for ethanol (EtOH) sensitivity of the miRNA miR-9 suggests one mechanism, whereby EtOH broadly influences development. We hypothesized that loss of miR-9 function recapitulates aspects of EtOH teratology.

METHODS:

Zebrafish embryos were exposed to EtOH during gastrulation, or injected with anti-miR-9 or nonsense control morpholinos during the 2-cell stage of development and collected between 24 and 72 hours postfertilization (hpf). We also assessed the expression of developmentally important, and known miR-9 targets, FGFR-1, FOXP2, and the nontargeted transcript, MECP2. Methylation at CpG islands of mammalian miR-9 genes was assessed in fetal murine neural stem cells (mNSCs) by methylation-specific PCR, and miRNA processing assessed by qRT-PCR for pre-miR-9 transcripts.

RESULTS:

EtOH treatment and miR-9 knockdown resulted in similar cranial defects including microcephaly. Additionally, EtOH transiently suppressed miR-9, as well as FGFR-1 and FOXP2, and alterations in miR-9 expression were correlated with severity of EtOH-induced teratology. In mNSCs, EtOH increased CpG dinucleotide methylation at the miR-9-2 locus and accumulation of pre-miR-9-3.

CONCLUSIONS:

EtOH exerts regulatory control at multiple levels of miR-9 biogenesis. Moreover, early embryonic loss of miR-9 function recapitulated the severe range of teratology associated with developmental EtOH exposure. EtOH also disrupts the relationship between miR-9 and target gene expression, suggesting a nuanced relationship between EtOH and miRNA regulatory networks in the developing embryo. The implications of these data for the expression and function of mature miR-9 warrant further investigation.

KEYWORDS:

Embryonic Development; Ethanol; Methylation; MiR-9; Zebrafish

PMID:
23800254
PMCID:
PMC3785568
DOI:
10.1111/acer.12139
[Indexed for MEDLINE]
Free PMC Article

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