Genetics of FAS in Mouse Component

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Heavy prenatal alcohol exposure can cause fetal alcohol syndrome (FAS) characterized by growth deficits, facial dysmorphology, brain damage, and neurobehavioral dysfunction. These abnormalities vary widely across a range now considered to reflect a fetal alcohol spectrum disorder (FASD). Variation in FASD is linked to individual differences in patterns of drinking during pregnancy together with genetic and biological factors influencing vulnerability. Little is understood about the molecular mechanisms of FASD pathogenesis, or about genetic differences that contribute to the diverse FASD presentations. Our long-term objectives are to identify alcohol effects on gene expression that can link mechanistically to abnormal brain development, and to identify genetic factors that influence susceptibility to those pathogenesis mechanisms. We have used embryonic cultures to expose C56BL/6 (B6) and DBA/2 (D2) inbred mice to alcohol during early neurulation. The B6 and D2 embryos showed different patterns of developmental delays and structural defects after 44 hours of exposure to high alcohol concentrations (300-400 mg/dl). In the B6 embryos, DMA microarray expression profiling, followed by hypothesis-driven bioinformatics analysis of differentially expressed genes, revealed over-representation of genes in gene sets regulating specific functions. Prominent down regulation of many genes critical for neural specification, neural typing, and neural patterning was evident, and confirmed for several. Our goal now is to generate more detailed temporal and spatial analyses of the disrupted expression of gene cohorts and their regulators, to identify correlations with dysmorphology and differential outcomes in the two inbred strains. Aim 1 will identify gene expression changes in B6 and D2 embryos after either 4 or 20 hrs of alcohol exposure, using DNA microarray and hypothesis-driven bioinformatics analysis to identify effects in gene sets in specific functional pathways. Patterns of dysmorphology will be compared with localization of changes seen with in situ hybridization of selected genes. Aim 2 will assess the relevance of observed down regulation of selected mouse genes to vertebrate neural development, by using novel zebrafish morpholino knockdown technology to carry out rapid screening of loss-of-function effects of orthologous genes. Aim 3 will use informatics to determine whether alcohol effects may be due to actions on functions of specific transcription factors (TF), by identifying TFs predicted from TF binding sites common to alcohol-affected gene cohorts in the mouse models. These studies will provide insights into prenatal alcohol effects on molecular regulators of embryonic brain development, along with genetic factors that influence the type or extent of damage associated with FAS. This knowledge will add important information for the identification, intervention, or treatment in pregnancies at-risk for FASD.