Embryonic Exposure to the Monoterpenes Thymol and p‑Cymene Induces Social Deficits through KDM5 Inhibition

Through a proteome-wide inverse virtual screening, followed by <i>in silico</i>, <i>in vitro</i>, and <i>in vivo</i> validations, we discovered lysine demethylase 5 (KDM5) family proteins, in particular KDM5B, KDM5C, and KDM5D, as predicted binding targets of TH and PC. Both <i>KDM5B</i> and <i>KDM5C</i> genes have been previously associated with ASD. The KDM5 family proteins are histone demethylases that selectively remove methyl groups from histone H3 lysine 4 (H3K4me3/2/1), which are epigenetic marks typically associated with active gene transcription.

Both compounds directly inhibited KDM5 demethylase activity in nuclear extracts from zebrafish larvae and from human cells, and embryonic exposure increased the global level of H3K4me3 <i>in vivo</i>. Consistent with elevated H3K4me3, RNA-sequencing detected profound up-regulation of genes associated with genetic risk factors of ASD following TH and PC exposure. Furthermore, chemical inhibition of the opposing H3K4 methyltransferase complex, MLL/WDR5, effectively rescued social deficits induced by TH and PC.

In summary, this work discovered thymol and <i>p</i>-cymene as risk factors for social behavioral development and establishes KDM5 inhibition as a mechanism of their neurodevelopmental toxicity.