Loss of qE Does Not Necessarily Lead to Photoinhibition: Sustained Non‐Photochemical Quenching in the Absence of PsbS and Zeaxanthin

Photosynthetic light-harvesting complexes mediate light absorption and energy dissipation. By modulating the photosystems' absorption cross-section, they affect both photosynthetic activity and non-photochemical quenching (NPQ). These processes are often studied by spectrally integrated chlorophyll fluorescence, masking their associated spectral information. We explore in Aspen and Arabidopsis npq mutants how qE affects the development of NPQ spectra under two contrasting conditions: in the absence and the presence of photoinhibition. We introduce a new parameter, the development of new emitting species (NESD), during time- and spectrally resolved NPQ inductions, and develop a pipeline to resolve PSII energy-partitioning heterogeneity. LHCII, PsbS, and zeaxanthin are required for NESD. Combining gas exchange, P700 oxidation, and spectrally resolved kinetics, we show that under photoinhibitory conditions, NES can develop even without PsbS or zeaxanthin, producing sustained quenching independent of photoinhibition of PSII or PSI. Furthermore, the absence of LHCII and CURVATURE THYLAKOID 1 leads to increased photoinhibition, indicating that long-term photoprotection relies on LHCII and thylakoid plasticity, whereas PsbS and zeaxanthin mainly facilitate LHCII-dependent quenching. Finally, we show the limitations of traditional parameters in discriminating between photoinhibition and photoprotective sustained quenching and propose time-resolved monitoring of CO₂ assimilation and Y(II) for their accurate assessment.