Mutation of a conserved stromal C-terminus threonine residue of Photosystem II subunit S slows down NPQ induction and speeds up relaxation.

In order to prevent damage by excess light, photosystem antennae can switch to an energy dissipative mode (termed non-photochemical quenching, NPQ). In higher plants, this switch is facilitated by the presence of Photosystem II subunit S (PsbS) protein, which was discovered 25 years ago. While the role of PsbS in induction of NPQ was soon found to require protonation of key glutamate residues facing the thylakoid lumen, a complete understanding of how NPQ is subsequently initiated is still lacking. Recent work on Norway spruce suggests that reversible phosphorylation at a few key residues of PsbS may coincide with the induction of a sustained dissipative state. Here we used Arabidopsis thaliana mutant lines to assess the functional implications of phosphorylation at threonine-259, one of the implicated phosphorylatable residues. Using a set of residue changes expressed in the background of PsbS knock-out mutant, npq4, we show that neither phosphomimetic, phosphosubstitution, nor phosphonull complementations could rescue NPQ activity to the level of the unperturbed protein. Instead, all residue substitutions at threonine-259 gave rise to significantly impaired induction and accelerated NPQ recovery, while protein accumulation and thylakoid membrane localisation were not affected. We suggest that these results are consistent with a role for the C-terminus in the propensity or stability of hydrophobic interactions between PsbS monomers to form homodimers, or between PsbS and other LHCII proteins to initiate the quenched state. HighlightReversible phosphorylation of threonine-259 on PsbS has previously been suggested to play a role in sustained NPQ during cold-acclimation in evergreen conifers. Here we show that both phosphomimetic and phosphonull substitutions at T259 lead to significantly impaired NPQ induction and faster recovery, which may be explained by their impact on hydrophobic interactions between PsbS and other LHCII proteins.