Peck, L. D.; Sork, V.

Trees experience decades-to-centuries of environmental change within a single lifetime, requiring molecular mechanisms that enable rapid physiological and transcriptional adjustment without genetic adaptation across generations. Increasing drought frequency provides one important example of the environmental challenges faced by long-lived species. DNA methylation is a candidate regulator of such responses, but whether environmentally induced methylation is associated with transcription remains an outstanding question. To address this, we integrated methylome and transcriptome data from valley oak (Quercus lobata) seedlings exposed to drought and well-watered treatments. Drought conditions induced widespread CHH methylation that targeted the same gene-proximal transposable elements (TEs) across successive drought exposures despite turnover of individual methylated cytosines. This response was concentrated within specific TE families, and genes associated with CHH-methylated upstream TEs showed increased transcription under drought and were enriched for drought-response pathways, including abscisic acid signaling. Nonetheless, the magnitude of transcriptional activation declined with increasing CHH methylation. Despite little overall change in the TE transcriptome, greater CHH methylation was associated with reduced expression of intragenic TEs, consistent with maintenance of local TE repression. These findings support a model in which repeated drought recruits CHH methylation to reproducible gene-proximal TEs, where it is associated with maintenance of local TE repression despite continued activation of neighboring stress-responsive genes. Increasing methylation load is associated with progressively weaker transcriptional responses, suggesting that high methylation levels may simultaneously suppress TE activity and constrain nearby gene expression. Such a mechanism may influence how long-lived trees repeatedly adjust transcriptional responses to fluctuating climates throughout their lifespan.