The Role of Phosphorylation Dynamics of CURVATURE THYLAKOID 1B in Plant Thylakoid Membranes

Abstract

Thylakoid membranes in land plant chloroplasts are organized into appressed and nonappressed membranes, which contribute to the control of energy distribution between the two photosystems (PSI and PSII) from the associated light-harvesting complexes (LHCs). Under fluctuating light conditions, fast reversible phosphorylation of the N-terminal thylakoid protein domains and changes in electrostatic forces induce modifications in thylakoid organization. To gain insight into the role and dynamics of thylakoid protein phosphorylation, we used targeted proteomics to quantify amounts of the structural proteins CURVATURE THYLAKOID1 (CURT1), including the levels of CURT1B N terminus phosphorylation and acetylation, after short-term fluctuating light treatments of Arabidopsis (Arabidopsis thaliana). The CURT1B protein was localized to a specific curvature domain separated from the margin domain, and specifically depleted of chlorophyll-binding protein complexes. The acetylation and phosphorylation of the CURT1B N terminus were mutually exclusive. The level of CURT1B phosphorylation, but not of acetylation, increased upon light shifts that also led to an increase in PSII core protein phosphorylation. These dynamics were largely absent in the knockout mutant of PSII core protein kinase SER/THR PROTEIN KINASE8 (STN8). Moreover, in mutants impaired in interaction between phosphorylated LHCII and PSI, the phosphorylation dynamics of CURT1B and the amount of the other CURT1 proteins were misregulated, indicating a functional interaction between CURT1B and PSI-LHCII complexes in grana margins. The complex relationships between phosphorylation of PSII, LHCII, and CURT1B support the dynamics of thylakoid protein complexes that are crucial in the optimization of photosynthesis under fluctuating light intensities. The photosynthetic conversion of solar energy into reducing equivalents and chemical bonds is achieved in plant chloroplasts by the orchestrated action of two thylakoid membrane-integral pigment-binding protein complexes, PSI and PSII, together with cytochrome b 6 f (Cyt b 6 f) complex, ATP synthase, light harvesting complexes (LHCs) I and II, and several auxiliary proteins. The regulation of the photosynthetic apparatus is tightly connected to the structural arrangement of the thylakoid membrane with defined appressed and nonappressed regions. The two photosystems are spatially segregated due to defined lateral heterogeneity of the thylakoid membrane system, with PSII predominantly present in the appressed membranes (grana), while PSI is confined to the stroma-exposed nonappressed thylakoid membranes (lamellae; Andersson and Anderson, 1980). How exactly the thylakoid membrane is organized into grana and how the continuity with the nonappressed regions is guaranteed is still a matter of debate (

Comparative analysis of thylakoid protein complexes in state transition mutants nsi and stn7: focus on PSI and LHCII

“…These values are in accordance with the previously published results (e.g. Kyle et al 1983;Rantala et al 2017;Koochak et al 2019;Trotta et al 2019) and also with the results obtained by mechanical fractionation (e.g. Danielsson et al 2006;Suorsa et al 2014).…”

“…Since DM-solubilized thylakoids contain also the appressed grana thylakoids, which host the majority of the PSII complexes, the PSII proteins were more abundant in the PSI/PSII band of DM-solubilized samples as compared to digitonin-solubilized samples (Table 2), reaching up to half of the PSAA amount. This result is in line with earlier studies showing enrichment of PSI subunits in digitonin-solubilized thylakoids (Rantala et al 2017;Koochak et al 2019;Trotta et al 2019). Interestingly, the CP43/PSAA ratio in the PSI/PSII band of the DM-solubilized thylakoids was 30% lower in nsi as compared to stn7 (p = 0.023 and 0.085 for nsi-2 and nsi-1, respectively).…”

Mutation of the Atypical Kinase ABC1K3 Partially Rescues the PROTON GRADIENT REGULATION 6 Phenotype in Arabidopsis thaliana

“…We cannot exclude that protein phosphorylation has a positive feedback loop effect; in fact, the phosphorylation of the thylakoid proteins may favor their mobility in the membrane and by doing so increase also the mobility of the PQ between different portions of the thylakoid membrane (Kirchhoff et al, 2000;Fristedt et al, 2009;Kirchhoff, 2014). We also cannot exclude the involvement of the phosphorylation of other, less evident, targets such as CURT1b, the phosphorylation of which is also dependent on STN8 kinase (Trotta et al, 2019). By their phosphorylation level, the proteins imbedded in the thylakoid membrane may change the overall conformation of the membrane system and by this also the mobility and exchange of the PQ, and other lipids, between the photoactive pool and the reservoir (Armbruster et al, 2013).…”

Composition, phosphorylation and dynamic organization of photosynthetic protein complexes in plant thylakoid membrane

“…5,11 A recently characterized thylakoid fraction, obtained by digitonin solubilisation and subsequent centrifugation steps, was reported to be a combination of a highly curved grana margins, straight membranes at the interphase between grana and stroma as well as of the fret-like protrusions (the slits) between grana and stroma membranes. 12,13 Subsequent further optimization of thylakoid fractionation yielded a separation of the curved membrane domain from the rest of the margin domain, 14 and accordingly, the nomenclature adopted in Fig. 1 includes a distinction between the grana curvature and grana margin domains.…”

“…31 Cyt b 6 f is likely located in all thylakoid domains 32,33 and its distribution might be regulated according to light conditions. 34 The grana margins, which should not be confused with the curvature domain in the edge of the grana membranes, 14 are biochemically a fusion of grana and stroma thylakoids, and accommodate both photosystems. 5,[35][36][37][38][39] Composition and LHCII-mediated supramolecular arrangement of the photosystems…”