Functional divergence of <scp>LhcSR</scp> and <scp>PsbS</scp> in zeaxanthin‐mediated non‐photochemical quenching

To protect photosystem II from excess light, non-photochemical quenching (NPQ) dissipates excess energy as heat. NPQ relies on trigger proteins, LhcSR in algae and PsbS in vascular plants, and the light-regulated xanthophyll cycle, which interconverts violaxanthin and zeaxanthin through the opposite activities of violaxanthin de-epoxidase and zeaxanthin epoxidase (ZEP). Despite extensive research, the molecular mechanisms and the differences in NPQ triggers across lineages remain unclear. In this study, we used the moss Physcomitrium patens, an evolutionary intermediate possessing both LhcSR and PsbS, to dissect their contributions to zeaxanthin-mediated quenching by the application of in vivo fast spectroscopy analysis. In the zep knockout (KO) mutant, constitutive zeaxanthin accumulation poises the photosynthetic apparatus in a pre-quenched state even in the dark, resulting in sustained NPQ upon illumination. Multiple zep KO mutants reveal that this constitutive zeaxanthin-driven quenching is mediated by LhcSR, while PsbS-dependent quenching is strictly light-activated. Our findings show that the two NPQ triggers, PsbS and LhcSR, have distinct molecular mechanisms suggesting that the evolutionary shift toward PsbS dominance in vascular plants reflects the need for tighter and more energy-efficient photoprotection.