Gene-Environment interactions in Autism

摘要

ABSTRACT Autism spectrum disorder (ASD) is group of neurodevelopmental conditions characterized by impaired social interactions, repetitive or restrictive behaviors, and difficulties with communication. ASD is highly prevalent, affecting 1 in 54 children in the US. Whole genome and exome sequencing studies identified 192 high confidence ASD-associated genes, many of which are expressed early in various cell lineages during brain cortex development, including neural progenitors, immature and maturing neurons, and glial cells. In addition, GWAS studies suggest the existence of non-coding genome variants that contribute to ASD phenotypes. Exposure of mice to chemicals present in the environment, including bisphenol A (BPA), result in ASD-like phenotypes, alterations in the cellular composition of the brain cortex, and changes in the binding of transcription factors (TFs) in genes implicated in ASD. Based on these observations we hypothesize that sequence variants present in the non-coding genome of different individuals, when altering regulatory sequences, may influence the interaction of TFs with their target sites in response to environmental chemicals. Phenotypic effects may be weak or undetectable in individuals carrying specific sequence variants but exposure to environmental chemicals may amplify the effect of these variants on their interaction with TFs and the ensuing phenotypes. To test these hypotheses, we propose to use a collection of iPSCs obtained from normal and ASD individuals from different sex, age, and racial backgrounds. These iPSCs will be used to grow cerebral cortical organoids, which will be exposed to BPA at different times during the differentiation process to alter gene expression in different cell types of neural lineages. Single nucleus (sn) RNA-seq and snATAC- seq will be employed to analyze TF occupancy and gene expression in specific cell populations during the differentiation of brain organoids in the presence or absence of BPA. This will allow us to monitor the effect of BPA on differentiation pathways and relative ratios of different neural cell lineages. We will then identify differential BPA-responsive ATAC-seq peaks among brain organoids arising from different iPSCs that correlate with cellular differentiation and gene expression phenotypes related to ASD. We expect that these differential ATAC-seq peaks will correspond to sequence variants present in regulatory sequences of different iPSC lines that affect the expression of specific genes involved in ASD. This will be tested using massively parallel reporter assays (MPRAs) in cell lines corresponding to the affected cell type, and cerebral organoids. The role of specific SNPs in gene expression will be further tested using single-base scarless genome editing. Finally, the possible contribution of these BPA-responsive SNPs to autism phenotypes will be analyzed by performing snATAC-seq in post-mortem brain samples from ASD patients. These results will fill an important gap in our knowledge of the fundamental principles by which genome variants can respond to chemicals present in the environment to affect lineage commitment of neural cells and elicit ASD symptoms.