| 1. |
- Flood, Padraic, et al.
(författare)
-
Natural variation in phosphorylation of photosystem II proteins in Arabidopsis thaliana: is it caused by genetic variation in the STN kinases?
- 2014
-
Ingår i: Philosophical Transactions of the Royal Society of London. Biological Sciences. - : The Royal Society. - 0962-8436 .- 1471-2970. ; 369:1640
-
Tidskriftsartikel (refereegranskat)abstract
- Reversible phosphorylation of photosystem II (PSII) proteins is an important regulatory mechanism that can protect plants from changes in ambient light intensity and quality. We hypothesized that there is natural variation in this process in Arabidopsis (Arabidopsis thaliana), and that this results from genetic variation in the STN7 and STN8 kinase genes. To test this, Arabidopsis accessions of diverse geographical origins were exposed to two light regimes, and the levels of phospho-D1 and phospho-light harvesting complex II (LHCII) proteins were quantified by western blotting with anti-phosphothreonine antibodies. Accessions were classified as having high, moderate or low phosphorylation relative to Col-0. This variation could not be explained by the abundance of the substrates in thylakoid membranes. In genotypes with atrazine-resistant forms of the D1 protein, low D1 and LHCII protein phosphorylation was observed, which may be due to low PSII efficiency, resulting in reduced activation of the STN kinases. In the remaining genotypes, phospho-D1 levels correlated with STN8 protein abundance in high-light conditions. In growth light, D1 and LHCII phosphorylation correlated with longitude and in the case of LHCII phosphorylation also with temperature variability. This suggests a possible role of natural variation in PSII protein phosphorylation in the adaptation of Arabidopsis to diverse environments.
|
|
| 2. |
- Yin, Lan, 1979
(författare)
-
Molecular Mechanisms Optimizing Photosynthesis during High Light Stress in Plants
- 2014
-
Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Oxygenic photosynthesis is the process by which plants, algae and cyanobacteria use solar energy to convert water and carbon dioxide into molecular oxygen and carbohydrates. Photosynthesis sustains life on Earth since it provides not only energy for individual growth, but also represents the starting point of the food chain for most living organisms. Sunlight is essential for driving photosynthesis, but it is also known that in excess it can be stressful with severe consequences for plant growth. In this thesis I have used the model plant Arabidopsis thaliana to study molecular mechanisms optimizing photosynthesis during high light stress. One of these mechanisms is the reversible phosphorylation of proteins in the water-oxidizing photosystem II (PSII) complex. The serine/threonine-protein kinases STN7 and STN8 are involved in the phosphorylation of the PSII light-harvesting complex (LHCII) and core proteins, respectively. In Paper II, I found variation in the phosphorylation levels of these proteins in Arabidopsis natural accessions. In high light conditions, I found a correlation between the STN8 protein abundance and the D1 protein phosphorylation level. In growth light conditions, D1 and LHCII phosphorylation correlated with longitude, and in the case of LHCII phosphorylation with temperature variability as well. Another molecular mechanism for plants to overcome high light stress is via PSII repair. STN8- mediated PSII core phosphorylation is an early and crucial step for efficient PSII repair, since it alters the folding of the thylakoid membrane in a manner facilitating lateral migration of complexes to the sites of repair. Among three laboratory Arabidopsis accessions studied, Ws-4 displayed a reduced STN8 level resulting in decreased PSII core protein phosphorylation (Paper I). Nevertheless, the downstream steps in PSII repair proceeded normal or slightly faster. This phenomenon is probably due to compensatory mechanisms involving additional lipids and carotenoids to increase membrane fluidity and thus lateral migration of complexes. The thylakoid ATP/ADP carrier (TAAC) transports ATP into the thylakoid lumen for nucleotide-dependent reactions. In Paper III, I have found that TAAC- deficient plants displayed wild-type levels of PSII protein phosphorylation but slower disassembly of complexes and slower D1 protein degradation. I propose that ATP supplied by TAAC into the lumen is used for nucleotide-dependent reactions with roles in various steps of PSII repair. I have also found that, via its transport activity, TAAC may consume part of the proton gradient across the thylakoid membrane, which is critical for the initiation of photoprotective mechanisms. In a proteomics study of the stroma thylakoid membrane from Arabidopsis (Paper IV), I identified 58 proteins, including previously known ones as well as new putative thylakoid proteins with roles in photosynthesis transport, translation, protein fate, metabolism, stress response and signaling. This thesis deepens our understanding of photosynthetic regulation at the molecular level and improves the biochemical overview of the chloroplast thylakoid membrane.
|
|
| 3. |
- Yin, Lan, 1979, et al.
(författare)
-
Photosystem II Function and Dynamics in Three Widely Used Arabidopsis thaliana Accessions
- 2012
-
Ingår i: PLoS One. - : Public Library of Science (PLoS). - 1932-6203. ; 7:9
-
Tidskriftsartikel (refereegranskat)abstract
- Columbia-0 (Col-0), Wassilewskija-4 (Ws-4), and Landsberg erecta-0 (Ler-0) are used as background lines for many public Arabidopsis mutant collections, and for investigation in laboratory conditions of plant processes, including photosynthesis and response to high-intensity light (HL). The photosystem II (PSII) complex is sensitive to HL and requires repair to sustain its function. PSII repair is a multistep process controlled by numerous factors, including protein phosphorylation and thylakoid membrane stacking. Here we have characterized the function and dynamics of PSII complex under growth-light and HL conditions. Ws-4 displayed 30% more thylakoid lipids per chlorophyll and 40% less chlorophyll per carotenoid than Col-0 and Ler-0. There were no large differences in thylakoid stacking, photoprotection and relative levels of photosynthetic complexes among the three accessions. An increased efficiency of PSII closure was found in Ws-4 following illumination with saturation flashes or continuous light. Phosphorylation of the PSII D1/D2 proteins was reduced by 50% in Ws-4 as compared to Col-0 and Ler-0. An increase in abundance of the responsible STN8 kinase in response to HL treatment was found in all three accessions, but Ws-4 displayed 50% lower levels than Col-0 and Ler-0. Despite this, the HL treatment caused in Ws-4 the lagest extent of PSII inactivation, disassembly, D1 protein degradation, and the largest decrease in the size of stacked thylakoids. The dilution of chlorophyll-protein complexes with additional lipids and carotenoids in Ws-4 may represent a mechanism to facilitate lateral protein traffic in the membrane, thus compensating for the lack of a full complement of STN8 kinase. Nevertheless, additional PSII damage occurs in Ws-4, which exceeds the D1 protein synthesis capacity, thus leading to enhanced photoinhibition. Our findings are valuable for selection of appropriate background line for PSII characterization in Arabidopsis mutants, and also provide the first insights into natural variation of PSII protein phosphorylation.
|
|
| 4. |
- Yin, Lan, 1979, et al.
(författare)
-
The membrane proteome of stroma thylakoids from Arabidopsis thaliana studied by successive in-solution and in-gel digestion.
- 2015
-
Ingår i: Physiologia Plantarum : An International Journal for Plant Biology. - : Wiley. - 0031-9317 .- 1399-3054. ; 154:3, s. 433-446
-
Tidskriftsartikel (refereegranskat)abstract
- From individual localization and large-scale proteomic studies, we know that stroma-exposed thylakoid membranes harbor part of the machinery performing the light-dependent photosynthetic reactions. The minor components of the stroma thylakoid proteome, regulating and maintaining the photosynthetic machinery, are in the process of being unraveled. In this study, we developed in-solution and in-gel proteolytic digestion methods, and used them to identify minor membrane proteins, e.g. transporters, in stroma thylakoids prepared from Arabidopsis thaliana (L.) Heynh Columbia-0 leaves. In-solution digestion with chymotrypsin yielded the largest number of peptides, but in combination with methanol extraction resulted in identification of the largest number of membrane proteins. Although less efficient in extracting peptides, in-gel digestion with trypsin and chymotrypsin led to identification of additional proteins. We identified a total of 58 proteins including 44 membrane proteins. Almost half are known thylakoid proteins with roles in photosynthetic light reactions, proteolysis and import. The other half, including many transporters, are not known as chloroplast proteins, because they have been either curated (manually assigned) to other cellular compartments or not curated at all at the plastid protein databases. Transporters include ATP-binding cassette (ABC) proteins, transporters for K+ and other cations. Other proteins either have a role in processes probably linked to photosynthesis, namely translation, metabolism, stress and signaling or are contaminants. Our results indicate that all these proteins are present in stroma thylakoids; however, individual studies are required to validate their location and putative roles. This study also provides strategies complementary to traditional methods for identification of membrane proteins from other cellular compartments.
|
|
| 5. |
- Örtegren Kugelberg, Unn, 1975-, et al.
(författare)
-
Separation and characterization of caveolae subclasses in the plasma membrane of primary adipocytes : segregation of specific proteins and functions
- 2006
-
Ingår i: The FEBS Journal. - : Wiley. - 1742-464X .- 1742-4658. ; 273:14, s. 3381-3392
-
Tidskriftsartikel (refereegranskat)abstract
- Caveolae are nearly ubiquitous plasma membrane domains that in adipocytes vary in size between 25 and 150 nm. They constitute sites of entry into the cell as well as platforms for cell signalling. We have previously reported that plasma membrane-associated caveolae that lack cell surface access can be identified by electron microscopy. We now report the identification, after density gradient ultracentrifugation, of a subclass of very high-density apparently closed caveolae that were not labelled by cell surface protein labelling of intact cells. These caveolae contained caveolin-1 and caveolin-2. Another class of high-density caveolae contained caveolin-1, caveolin-2 and specifically fatty acid transport protein-1, fatty acid transport protein-4, fatty acyl-CoA synthetase, hormone-sensitive lipase, perilipin, and insulin-regulated glucose transporter-4. This class of caveolae was specialized in fatty acid uptake and conversion to triacylglycerol. A third class of low-density caveolae contained the insulin receptor, class B scavenger receptor-1, and insulin-regulated glucose transporter-4. Small amounts of these proteins were also detected in the high-density caveolae. In response to insulin, the insulin receptor autophosphorylation and the amount of insulin-regulated glucose transporter-4 increased in these caveolae. The molar ratio of cholesterol to phospholipid in the three caveolae classes varied considerably, from 0.4 in very high-density caveolae to 0.9 in low-density caveolae. There was no correlation between the caveolar contents of caveolin and cholesterol. The low-density caveolae, with the highest cholesterol concentration, were particularly enriched with the cholesterol-rich lipoprotein receptor class B scavenger receptor-1, which mediated cholesteryl ester uptake from high-density lipoprotein and generation of free cholesterol in these caveolae, suggesting a specific role in cholesterol uptake/metabolism. These findings demonstrate a segregation of functions in caveolae subclasses.
|
|
