Carotenoid composition affects thylakoid morphology and membrane fluidity

Thylakoids are the core of photosynthesis in the chloroplasts of higher plants. They are lipidic membrane compartments organized in appressed piles called grana intercalated with monolayered lamellae. Grana and lamellae differ in their composition, with photosystem I (PSI) preferentially localized in the lamellaeand photosystem II (PSII) preferentially localized in the grana (Andersson and Anderson, 1980). However, thylakoid organization is fluid. For example, phosphorylation of the light-harvesting complex of PSII triggers its detachment and subsequent connection to PSI, to maximize energy funneling to the two photosystems according to the light quality available for the plant (Minagawa, 2011).

Carotenoids are important pigments that sustain the photosynthetic process. Most of the carotenoid pool is associated with the antennae complexes, where they are necessary for light harvesting, for dissipation of reactive oxygen species, and for nonphotochemical quenching (Hashimoto et al., 2016), while the remaining 15% of carotenoids are freely dispersed in the thylakoid membrane (Dall’Osto et al., 2010).

Although the impact of lipids and protein complexes on thylakoid structure and morphology has recently received much attention (Mazur et al., 2019; Zsiros et al., 2020), the role of carotenoids has so far been neglected. To fill this void and reveal the structural role of carotenoids on the thylakoid network, in this issue of Plant Physiology, Michał Bykowski et al. (2021) describe a thorough investigation of 2D and 3D morphology of four Arabidopsis thaliana mutants with altered carotenoid contents, by combining confocal laser scanning microscopy and spectroscopic measurements.

All four mutants showed some degree of morphological aberrations at the thylakoid level, but the strongest phenotypes were observed in those mutants presenting higher lutein contents: szl1-1 and szl1-1 npq1-2 (suppressor of zeaxanthin-less 1-1 and suppressor of zeaxanthin-less1-1 nonphotochemical quenching1-2; the latter also lacks zeaxantine). These mutants also presented a decrease in neoxantine and violaxantine, while α and β carotene, which are key components of antennae, failed to accumulate; concomitantly, their precursors γ and δ carotene increased and probably accumulated in plastoglobules (Figure 1). The alterations in thylakoid morphology in both lutein-accumulating mutants included fewer stacks per granum, smaller area of grana cross-sections, higher distance between stacking repeats and more regular grana margins.

Figure 1.

Scheme summarizing the thylakoid morphology of lutein and carotenoid accumulating-mutants (szl1-1 and szl1-1 npq1-2): main phenotypes include: reduced layers of thylakoid grana, increased plastoglobule size (black circles) with accumulation of δ- and γ-carotene, and accumulation of nonphosphorylated low-mass PSII supercomplexes. There is evidence for association of RUBISCO with the grana as well. Modified from Bykowski et al. (2021).

The authors also observed that the thylakoid membrane fluidity was severely compromised in the lutein-accumulating mutants. The excessive stiffness of the membranes might be explained by a reduction in α and β carotene (Gruszecki and Strzałka, 2005), as well as an increased level of free lutein in the lipidic space. Unlike α and β carotene, which are entirely hydrophobic, lutein has polar hydroxyl groups at each end, limiting its orientation to aligning parallel or perpendicular to the lipid bilayer, which could contribute to the loss of membrane fluidity. Conversely, lut5-1, which displays a higher level of α and β carotene, shows increased membrane fluidity, suggesting that α and β carotene directly affect membrane fluidity.

Blue-native electrophoresis revealed that both lutein-accumulating mutants were associated with an increased amount of low-molecular-weight supercomplexes, and a reduction in the levels of light-harvesting complex II trimers (LHCII trimers), cytochrome b6f, and PSI-LHCI. Indeed, low amount of carotene affects the accumulation of PSII complexes (Sheng et al., 2018), but the lutein-dependent increase in membrane rigidity might also be responsible for the formation of low-mass PSII supercomplexes. Unsurprisingly, these changes are accompanied by altered photosynthetic parameters, in particular, diminished capabilities to dissipate excess energy by nonphotochemical quenching.

The results of Bykowski et al. are an accurate and detailed characterization of the morphological and physiological phenotypes associated with an altered carotenoid profile (Figure 1), which will provide a valuable foothold for further research on the importance of pigments and membrane fluidity for plant acclimation to light changes, and to distinguish between direct and indirect effects of carotenoids on thylakoid morphology.