Photorespiration is linked to DNA methylation by formate as one-carbon source.

DNA methylation is a key epigenetic modification regulating genome organization, stability, and gene expression. Stable DNA methylation critically relies on methyl groups provided through folate-mediated one-carbon (C1) metabolism, yet the origin and regulation of C1 supply remain elusive. Here we demonstrate that photorespiration serves as a major C1 source for DNA methylation in Arabidopsis. We show that C1 from formate, a photorespiratory byproduct, is incorporated into 5-methyl-cytosine via the reductive cytosolic folate pathway. This occurs predominantly during the day, negatively regulating serine utilization as alternative C1 source. Consequently, suppression of photorespiration under elevated CO₂ levels alters the DNA methylation landscape, an effect exacerbated when regulation of C1 metabolism by the formate-dependent pathway is impaired. Thus, our findings link the fundamental metabolic process of photorespiration to epigenetic stability, highlighting how rising atmospheric CO₂ levels can induce DNA methylation changes.