Epigenome-wide analysis identifies DNA methylation signatures associated with the infant pupillary light reflex, a candidate intermediate phenotype for autism.

The pupillary light reflex (PLR), the automatic constriction of the pupil in response to increased luminance, is a candidate early intermediate phenotype associated with autism, with potential to help understand early neurodevelopmental differences because it is controlled by relatively simple neural circuitry. We conducted epigenome-wide association analyses of PLR onset latency and constriction amplitude at 9, 14, and 24 months, with 51 male infants enriched for familial autism likelihood (~ 80% with a first-degree autistic relative), using buccal DNA collected at 9 months. We identified four epigenome-wide differentially methylated probes (p < 2.4 × 10⁻⁷) significantly associated with PLR latency at 14 and 24 months, and 14- to 24-month developmental change in latency. Probes linked to PLR amplitude were identified at a discovery threshold (p < 5 × 10⁻⁵). Regional analyses revealed multiple differentially methylated regions associated with both latency and amplitude. Associated probes were enriched for neurodevelopmental processes and autism-associated genes, including NR4A2, HNRNPU, and NAV2. While the findings are most directly relevant to male infants in whom PLR variability may be associated with familial autism likelihood, they provide novel evidence that DNAm contributes to early variation in PLR. These insights into the biological underpinnings of this reflex support PLR as an early intermediate phenotype associated with autism.