DBG's Eduard Strasburger Workshop - Co- and posttranslational control in chloroplasts

Photosynthesis is a fundamental process in biology as it converts solar energy into chemical energy and thus, directly or indirectly, fuels all life on earth. The chemical energy is used to fix atmospheric CO2 and produce reduced carbon compounds in the Calvin-Benson-Bassham cycle. The key enzyme for this process in all photosynthetic organisms is ribulose-1,5-bisphosphate...

Rubisco is the central CO2 fixing enzyme of the Cavin cycle and responsible for the vast majority of all CO2 fixation on our planet today. In plants, Rubisco undergoes an elaborate set of steps involving the sequential action of at least 6 different dedicated folding and assembly chaperones to assemble into its enzymatically active form. This complexity evolved from much simpler Rubisco...

Carmela GIGLIONE
Institute for Integrative Biology of the Cell (I2BC), University Paris-Saclay, CNRS, 1 avenue de la Terrasse, Gif-sur-Yvette, France

Protein modifications are emerging as key regulators of numerous essential cellular processes. Virtually all proteins undergo co- and/or post-translational modifications (CTMs, PTMs). However, a comprehensive understanding of the full range...

Rubisco is a unique catalyst involved in biological carbon dioxide fixation. It is also the most abundant protein on Earth which accumulation rises up to 50% the protein content of leaves. As a major storage protein, nitrogen recycling from Rubisco is essential during degradative processes such as plant senescence or starvation. In land plants, Rubisco functions as a large multi-subunit...

Acetylation on amino groups is a common modification seen in proteins across various organisms. This process involves N-terminal acetyltransferases (NATs) and lysine acetyltransferases (KATs) that transfer acetyl groups from acetyl-Coenzyme A (acetyl-CoA) to the N-terminal amino groups and to the side chains of lysine residues, respectively. In the case of plants, the majority of plastid...

A dynamin-like protein FZL is one of the few known thylakoid remodeling proteins and has been demonstrated to mediate thylakoid membrane fusion. However, its physiological functions have not been characterized well. First, to determine the sub-chloroplast localization of FZL, we sub-fractionated isolated chloroplasts/thylakoids and observed GFP-fused proteins, showing that FZL is enriched on...

In plants, photosynthesis is a critical process for survival, and copper is one of the essential micronutrients required for this process. Under abiotic stress conditions such as drought, plants typically reduce photosynthesis to balance their competitive ability with stress tolerance, ensuring survival. This study investigates Arabis nemorensis and Arabis sagittata, two species growing in...

Light is the primary energy source for photosynthesis and plays a crucial role in regulating numerous developmental processes in plants. However, high light (HL) conditions that exceed chloroplast energy requirements can trigger oxidative stress, necessitating protective responses within plant nuclei. The transcription factor ELONGATED HYPOCOTYL5 (HY5) operates downstream of multiple...

Yuwei Wang, Alexander Gehm, Patrick Schall and Bernhard Grimm
Humboldt-Universität zu Berlin, Institute of Biology/Plant Physiology, Philippstraße 13 (Building 12), 10115 Berlin, Germany, www2.hu-berlin.de/biologie/plantphys/

Co-translational N-terminal modifications of plastid genome-encoded proteins also belong to key mechanisms for the stability and function of these proteins. Previous...

Chloroplast Acetyltransferases in Arabidopsis thaliana are a part of the General control non-repressible 5-related N-Acetyltransferase superfamily (GNAT), which is characterized by a high structural conservancy. Among the acetylated chloroplast proteins, those involved in photosynthesis make up a large proportion, indicating that the GNATs may be important regulators of photosynthesis (Hartl...

In photosynthesis, light drives electron transfer reactions that conserve reducing power and energy. Electron transfer is coupled to proton translocation into the chloroplast thylakoid lumen, building up a proton motive force (PMF) that drives ATP synthesis. Regulating PMF is essential for rapidly adjusting photosynthesis to fluctuations in light intensity, enabling a dynamic balance between...

The structural plasticity of photosystem I – light harvesting complex I (PSI-LHCI) is reflected in the recent resolution of diverse PSI-LHCI macromolecular organization states in the green alga Chlamydomonas reinhardtii: PSI-LHCI1,2, PSI-LHCI – light-harvesting complex II (LHCII)3,4 and dimeric PSI-LHCI5. It has been proposed that dynamic changes in PSI subunit composition and LHCI...

With the energy from the sunlight, plants assimilate atmospheric CO2 via photosynthesis and produce sugars. However, photosynthesis-associated genes are downregulated when sugars accumulate beyond the storage or transport capacity in leaves to reduce carbon assimilation. While various sugar signalling pathways have been proposed to control this downregulation, the molecular mechanism of how...

Lysine acetylation is a crucial post-translational modification involved in plant development and responses to environmental stimuli. While much attention has been focused on the role of HDACs in histone acetylation, their involvement in deacetylating non-histone proteins remains less explored. Among the 18 HDACs in Arabidopsis, HDA14 stands out for its dual localization in plastids and...

Lysine acetylation is an important post-translational protein modification that plays a vital role in plant development and in responses to different environmental stimuli. Histone deacetylases (HDACs) are responsible for removing lysine acetylation on various proteins. While most work has focussed on the role of Arabidopsis HDACs on histone acetylation, their role in the deacetylation of...

Protein acetylation is a key co- and post-translational modification. How different types of acetylation respond to environmental stress is still unknown. A member of the newly discovered family of plastid acetyltransferases, which is featuring both lysine- and N-terminal acetyltransferase activities, was used to obtain a holistic multi-omics acetylation-dependent view of the acclimation of...

Proteolysis is an essential process to maintain cellular homeostasis. One pathway that mediates selective protein degradation is the N-degron pathway, which relates the in vivo half-life of a protein to its N-terminal amino acid residue. In the cytosol of eukaryotes and prokaryotes, N terminal residues are major determinants of protein stability. While the eukaryotic N-degron pathway depends...

In course of the day, the environmental conditions are changing, so that the plants have different needs of proteins and their functions. As a result, the proteome needs to change constantly and in a dynamic way. Post-translational modifications (PTM´s) change the properties of present proteins and have a massive impact on their functions, structures and activities. Such a modified protein has...

During the oxygenic photosynthesis in plants, algae and cyanobacteria, atmospheric carbon dioxide (CO2) is assimilated into carbohydrates making photosynthetic organisms autotrophic. Rubisco catalyzes this step of carbon dioxide and oxygen uptake. Interestingly, the catalytic subunit of Rubisco (RbcL) undergoes a unique maturation pathway leading to unique N-terminal modifications. This...

Histone deacetylases (HDACs) are a pivotal enzyme in the removing of lysine acetylation on various proteins. While most work has focused on the role of Arabidopsis HDACs on histone acetylation, their role in the deacetylation of non-histone proteins is much less known, although proteins of many different organelles have been found to be lysine-acetylated. From the 18 HDACs found in...

The N-degron pathway may relate stability of proteins to the biochemical features of its amino (N)terminal stretch or even only the very first residue at this end and its posttranslational modifications (PTMs). Often, these apparently crucial modifications have actually not been shown. To spotlight on these decisive biochemical events, we attempt to demonstrate their existence in vitro and in...

Chloroplast functionality requires the post-translational import of plastid-destined nuclear-encoded proteins. Chloroplast precursor protein availability in the cytosol and import into the plastid is tightly regulated to maintain chloroplast biogenesis and functionality, respectively. One of these regulatory mechanisms is the co-translational modification of the precursor proteins by...

The pH within the thylakoids is essential for photosynthesis. As part of the photosynthetic light reactions, proton translocation across the thylakoid membrane generates a proton motive force that consists of a pH gradient (ΔpH) and an electrical gradient (ΔΨ) to drive ATP synthase. While genetically encoded pH biosensors have been instrumental in dissecting pH dynamics of different plant cell...

Cyclic electron flow (CEF) in photosynthesis has been studied for decades, yet many aspects of its components, mechanisms, and regulation remain unclear. PROTON GRADIENT REGULATION5 (PGR5) is thought to play a crucial role in promoting CEF. The absence of PGR5 disrupts photosynthetic control and increases the sensitivity of photosystem I (PSI) to light damage, resulting in plant death under...

Post-translational modifications (PTMs) control protein stability, localization, turn-over and interactions, thus represent a fine-tuned regulatory mechanism to reversibly adjust cellular physiology.
In photosynthetic eukaryotes, light-dependent phosphorylation of thylakoid proteins dynamically regulates the light-harvesting apparatus in response to environmental fluctuations. In the...

In this study, we examined plastocyanin (PC) binding and electron transfer with both photosystem I (PSI) and cytochrome b6f (cyt b6f), and show the synergetic adaptations between these three enzymes. Furthermore, we explored the effects of PC phosphorylation on these interactions. To do so, we generated several recombinant variants of PC, in which we genetically engineered two of the...

Fixing CO2 via photosynthesis requires ATP and NADPH. Linear electron transfer (LET) supplies both metabolites, yet depending on environmental conditions, additional ATP is required which can be generated by cyclic electron transfer (CET). Recent studies suggest that CET, which draws electrons from the ferredoxin pool affects the redox regulation of the Calvin-Benson cycle enzymes involved in...

Acetylation is one of the most common chemical modifications affecting a variety of molecules ranging from metabolites to proteins. Recent development of enrichment techniques and mass spectrometry has revealed that acetylation is a prevalent modification also in plants, and that in addition to cytosolic and nuclear proteins also numerous chloroplast proteins are acetylated. We have...

Plants are exposed to a constantly changing environment, which requires fast acclimation strategies. Post-translational modifications (PTMs) of proteins allow cells to respond rapidly to varying environmental conditions and have the potential to alter localization, interactions, or enzymatic activities of proteins. Protein acetylation is one of the most abundant co- and post-translational...

The photosystem II (PSII) supercomplex (SC) of higher plants is a (multi)protein-pigment assembly performing the initial steps of photosynthetic electron transport consisting of water oxidation and photochemical plastoquinone reduction. In angiosperms, the PSII SC is equipped with three LHCB4-6 monomeric pigment-binding proteins which connect the dimeric core complex with the peripheral...

In plant chloroplasts, protein kinases regulate photosynthetic acclimation by phosphorylation of thylakoid membrane proteins allowing rapid short-term acclimation to changing light conditions. This type of phosphorylation control is mediated by the light-regulated kinases STN7/STN8 at the thylakoid membrane system. Recent data suggested furthermore that STN7 may be involved in long-term...

Regulation of Rubisco activity by light in tomato
Joana Amaral, Dawn Worrall, Elizabete Carmo-Silva
Lancaster Environment Centre, Lancaster University, Lancaster LA1 4YQ, UK

Rubisco is the most abundant protein in the chloroplast, fixing CO₂ into sugars during photosynthesis. Its activity is finely adjusted in response to changes in the environment. The regulation of Rubisco by...

Most plant cell compartments possess oxidoreductases that catalyze the interconversion between malate and oxaloacetate (OAA) in a reversible reaction. These malate dehydrogenases (MDHs) are either NAD- or NADP-specific.
Together with malate/OAA translocators, MDHs enable the indirect exchange of reducing equivalents across subcellular membranes. The exported malate can then be used as a...

Tetrapyrrole biosynthesis (TPB) in plants consists of more than twenty enzymatic steps and is tightly controlled due to the synthesis of photoreactive intermediates and the different spatial and temporal requirements of their end products chlorophyll and heme. At the TPB hotspots, many complementary post-translational control mechanisms act on specific enzymes: The two enzymes glutamyl-tRNA...

The pyrimidine nucleotide cofactors NAD and NADP operate at the critical intersection between the cellular metabolic network, Cys-based systems and redox regulation. In plants, the redox states of the NAD and NADP pools of the cytosol and the chloroplast stroma are physically separated, but connected by metabolic shuttles, such as the malate/oxaloacetate and the triose-phosphate shuttle....

Plants undergo daily dark-light transitions, leading to dynamic changes of the metabolic and physiological status of their cells. Those changes, which include cofactors, ions and other small molecules such as ATP, NAD(P) redox status, ROS, Ca2+ and pH, are a prerequisite for tuning protein functions through post-translational modifications (PTMs). However, it has been notoriously difficult to...

Arabidopsis plants can produce photosynthetic tissue with active chloroplasts at temperatures as low as 4°C, and this process depends on the presence of the nuclear-encoded, chloroplast-localized RNA-binding protein CP29A. We demonstrate that CP29A undergoes phase separation in vitro and in vivo in a temperature-dependent manner, which is mediated by a prion-like domain (PLD) located between...

ABSTRACT Different proteases and peptidases are present within chloroplasts and non-photosynthetic plastids to process precursor proteins and to degrade cleaved chloroplast transit peptides and damaged, misfolded, or otherwise unwanted proteins. Collectively, these proteases and peptidases form a proteolysis network, with complementary activities and hierarchies, and build-in redundancies [1]....

The development and operation of chloroplasts (or other members of the plastid family of plant organelles) requires the participation of thousands of different organellar proteins. Most chloroplast proteins are nucleus-encoded and synthesized in the cytosol in precursor form. These preproteins pass through multiprotein import machines in the organelle’s outer and inner envelope membranes –...

Chloroplast-encoded multi-span thylakoid membrane proteins are crucial for photosynthetic complexes, yet the coordination of their biogenesis remains poorly understood. To identify factors that specifically support the cotranslational biogenesis of the reaction center protein D1 of photosystem (PS) II, we generated and affinity-purified stalled ribosome-nascent chain complexes (RNCs) bearing...

Chloroplasts, crucial organelles in autotrophic organisms, possess distinctive regulatory pathways to control molecular processes that respond dynamically to environmental cues. Protein synthesis in these organelles relies on transfer RNAs (tRNAs) encoded by the chloroplast genome, which undergo extensive post-transcriptional modifications. These modifications play critical roles in accurate...

Accurate protein translation is a hallmark for cell function. It guarantees an efficient proteome while minimizing detrimental unfolded proteins and concomitant energy loss. Surprisingly, protein mistranslation happens quite frequently in vivo (error rates 10-2 to 10-4) mainly due to transfer RNA (tRNA) mis-decoding and tRNA mis-acylation. Bacteria, mitochondria and plastids synthesize...

The N-termini of chloroplast proteins are a common site of co- and post-translational protein modifications, including N-terminal acetylation, transit peptide cleavage and subsequent proteolytic processing, that result in distinct proteoforms that may differ in activity, interactions and location. However, protein N-terminal peptides are often missed in standard shotgun proteomics...

Cysteine redox modulation in proteins is a reversible adaptation to changing environments. For example, activity adjustment of the chloroplast ATP synthase (CF1FO) ensure full activity in the light and deactivation during night. The latter is believed to prevent ATP hydrolysis and build-up of excessive proton motive force (pmf). The adjustment is realised by a cysteine couple in the central...

Photosynthesis must be dynamically regulated in response to environmental conditions to prevent photoinhibition and photodamage to the photosystems. Various post-translational modifications of proteins are known to play important roles in modulating photosynthetic efficiency during plant acclimation responses. Alongside phosphorylation and dithiol-based modifications, acetylation of amino...

In plant chloroplasts, protein kinases regulate photosynthetic acclimation by phosphorylation of thylakoid membrane proteins allowing rapid short-term acclimation to changing light conditions. This type of phosphorylation control is mediated by the light-regulated kinases STN7/STN8 at the thylakoid membrane system. Recent data suggested furthermore that STN7 may be involved in long-term...