Developmental and Functional Analyses of Neural Circuits Affected in Autism Spectrum Disorders

Background:  ASD are a spectrum of disorders likely resulting from aberrant development and function of neural circuits. Copy number variation analyses and whole-exome sequencing have identified multiple novel ASD genes and loci, while at the same highlighting extensive genetic heterogeneity. This complexity and the biological pleiotropy of known risk genes has complicated the identification of specific cell types and circuits underlying ASD pathogenesis. Combined analysis of complementary expression and regulatory networks in the developing brain promises to provide new insights and testable hypothesis about what cell types are involved, as well as where and when developmental perturbations occur in the brain. However, methods to rigorously test these predictions and translate them to an actionable understanding of neurobiology in ASD are needed. 

Objectives: To determine how genes associated with ASD shape the development and evolution of neuronal circuits of the human cerebral cortex.

Methods: Informed by spatiotemporal maps of gene expression, gene-regulatory interactions, and chromatin states, we have used traditional molecular biological and genetics tools to analyze the function and expression of ASD risk genes in the developing brain.

Results: Expression patterns of the diverse ASD risk genes in the developing brain provide novel insights into the underlying biology. One such example is the expression of the FMR1 gene,  which encodes an RNA-binding protein (FMRP) altered in Fragile X syndrome. We show that FMRP regulates translation of neuronal nitric oxide synthase 1 (NOS1) in the developing human neocortex. Whereas NOS1 mRNA is widely expressed, NOS1 protein is transiently co-expressed with FMRP during early synaptogenesis in layer- and region-specific pyramidal neurons. These include mid-fetal layer 5 subcortically projecting neurons arranged into alternating columns in the prospective Broca’s area and orofacial motor cortex. Human NOS1 translation is activated by FMRP via interactions with coding region-binding motifs absent from mouse Nos1 mRNA, which is expressed in mouse pyramidal neurons, but not efficiently translated. Correspondingly, neocortical NOS1 protein levels are severely reduced in developing human FXS cases, but not FMRP-deficient mice.

Conclusions: Our findings provide insights into cells and neural circuits affected in ASD and identify novel species-specific molecular mechanism altered in the leading monogenic cause of intellectual disability and autism, FXS. Importantly, this study illustrates a process of moving from genetic findings, to expression analysis, to functional characterization in order to expand the understanding of human-specific molecular mechanisms that may be compromised in ASD.