Though it is clear that autism spectrum disorder is highly heritable, the genetic basis of the disorder is quite complex, and it is likely caused by many genes and gene-environment interactions. Fragile X syndrome (FXS), a neurodevelopmental disorder caused by mutations in the FMR1 gene, is the most common single-gene disorder associated with ASD. Approximately 25-50% of children with FXS meet diagnostic criteria for ASD, and approximately 3% of children with ASD also have FXS. Recent studies have suggested that FMRP, the protein encoded by the FMR1 gene, is involved in the molecular pathways associated with ASD. Studying the features of ASD in FXS and the FMR1 premutation can help clarify what characteristics might be tied to the molecular mechanisms of the FMR1 gene. Pragmatic language impairment is a hallmark of ASD and FXS, and adults who carry the FMR1 gene in its premutation state exhibit subtle pragmatic differences as well. However, no studies have directly compared pragmatic language in children with ASD, FXS and the FMR1 premutation. Furthermore, it is not clear whether shared characteristics stem from shared underlying mechanisms, though there is growing evidence that autonomic hyperarousal may contribute to the social deficits of autism and FXS. The objective of this project is to investigate pragmatic language and autonomic arousal in autism, FXS, and the FMR1 premutation, and to relate these profiles to variation in FMRP expression. The rate and quality of pragmatic language impairments will be assessed with a semi-naturalistic measure of pragmatic language. Innovative eye tracking technology will be used to measure pupil dilation (an index of autonomic arousal) in response to social-emotional stimuli, and the relationship between autonomic arousal and pragmatic language difficulties will be examined. A recently-developed, highly sensitive assay will be used to measure FMRP in the blood, and the relationship between FMRP expression, pragmatic language, and autonomic arousal will be investigated. Through these complementary and innovative approaches, this project aims to inform future genetic studies and targeted treatment research by identifying and defining the characteristics associated with FMRP variation.