Chloroplast Protein Research Articles

A chloroplast localized heavy metal-associated domain containing protein regulates grain calcium accumulation in rice

Calcium (Ca) is an essential mineral nutrient and plays a crucial signaling role in all living organisms. Increasing Ca content in staple foods such as rice is vital for improving Ca nutrition of humans. Here we map a quantitative trait locus that controls Ca concentration in rice grains and identify the causal gene as GCSC1 (Grain Ca and Sr Concentrations 1), which encodes a chloroplast vesicle localized homo-oligomeric protein. GCSC1 exhibits Ca2+ transport activity in heterologous assays in yeast and Xenopus laevis oocytes and is involved in the efflux of Ca2+ from the chloroplast to the cytosol. Knockout of GCSC1 results in increased chloroplast Ca concentration, lower stomatal conductance in leaves and enhanced Ca allocation to grains. Natural variation in grain Ca concentration is attributed to the variable expression of GCSC1 resulting from its promoter sequence variation. Our study identifies a chloroplast localized heavy metal-associated domain containing protein that regulates chloroplast Ca2+ efflux and provides a way to biofortify Ca in rice to benefit human nutrition.

An effector protein of Fusarium graminearum targets chloroplasts and suppresses cyclic photosynthetic electron flow.

Chloroplasts are important photosynthetic organelles that regulate plant immunity, growth, and development. However, the role of fungal secretory proteins in linking the photosystem to the plant immune system remains largely unknown. Our systematic characterization of 17 chloroplast-targeting secreted proteins of Fusarium graminearum indicated that Fg03600 is an important virulence factor. Fg03600 translocation into plant cells and accumulation in chloroplasts depended on its chloroplast transit peptide. Fg03600 interacted with the wheat (Triticum aestivum L.) proton gradient regulation 5-like protein 1 (TaPGRL1), a part of the cyclic photosynthetic electron transport chain, and promoted TaPGRL1 homo-dimerization. Interestingly, TaPGRL1 also interacted with ferredoxin (TaFd), a chloroplast ferredoxin protein that transfers cyclic electrons to TaPGRL1. TaFd competed with Fg03600 for binding to the same region of TaPGRL1. Fg03600 expression in plants decreased cyclic electron flow (CEF) but increased the production of chloroplast-derived reactive oxygen species (ROS). Stably silenced TaPGRL1 impaired resistance to Fusarium head blight (FHB) and disrupted CEF. Overall, Fg03600 acts as a chloroplast-targeting effector to suppress plant CEF and increase photosynthesis-derived ROS for FHB development at the necrotrophic stage by promoting homo-dimeric TaPGRL1 or competing with TaFd for TaPGRL1 binding.

Uracil phosphoribosyltransferase is required to establish a functional cytochrome b6f complex.

Arabidopsis uracil phosphoribosyltransferase (UPP) is an essential enzyme and plants lacking this enzyme are strongly compromised in chloroplast function. Our analysis of UPP amiRNA mutants has confirmed that this vital function is crucial to establish a fully functional photosynthesis as the RIESKE iron sulfur protein (PetC) is almost absent, leading to a block in photosynthetic electron transport. Interestingly, this function appears to be unrelated to nucleotide homeostasis since nucleotide levels were not altered in the studied mutants. Transcriptomics and proteomic analysis showed that protein homeostasis but not gene expression is most likely responsible for this observation and high light provoked an upregulation of protease levels, including thylakoid filamentation temperature-sensitive 1, 5 (FtsH), caseinolytic protease proteolytic subunit 1 (ClpP1), and processing peptidases, as well as components of the chloroplast protein import machinery in UPP amiRNA lines. Strongly reduced PetC amounts were not only detected by immunoblot from mature plants but in addition in a de-etiolation experiment with young seedlings and are causing reduced high light-induced non-photochemical quenching Φ(NPQ) but increased unregulated energy dissipation Φ(NO). This impaired photosynthesis results in an inability to induce flavonoid biosynthesis. In addition, the levels of the osmoprotectants raffinose, proline, and fumarate were found to be reduced. In sum, our work suggests that UPP assists in stabilization PetC during import, processing or targeting to the thylakoid membrane, or protects it against proteolytic degradation.

Exploring Potential Aquaculture-Immunostimulant-Peptides Derived from Chlorella sorokiniana

Chlorella sorokiniana is a microalgae with an outstanding nutritional profile and numerous therapeutic substances that can be used as an immunostimulant, including in aquaculture. This research aimed to investigate and characterize peptides isolated from C. sorokiniana protein using TCA digestion and hydrolyzed enzymatically with trypsin. Peptides were then subsequently identified using Tandem LC-MS/MS and Mascot Distiller. Results showed that the percentage of pure protein yield following TCA digestion was 54.66%, and 12 peptides with lengths ranging from 7 to 23 sequences were discovered after trypsin digestion. These peptides originated from various enzymes and chloroplast proteins, including protein synthesis elongation factor TU, photosystem I iron-sulfur center, photosystem II 43 kDa, Ycf4, ATP-dependent zinc metalloprotease FtsH homolog, nitrate reductase, chloroplastic glucose-6-phospate dehydrogenase, and ATP synthase CF1 alpha chain. These findings demonstrated that C. sorokiniana might serve as a source of immunostimulant peptides and proteins, particularly for aquaculture biota.

Chloroplast elongation factors break the growth-immunity trade-off by simultaneously promoting yield and defence.

Chloroplasts regulate plant development and immunity. Here we report that potato chloroplast elongation factors StTuA and StTuB, targeted by Phytophthora infestans RXLR effector Pi22926, positively regulate immunity and growth. Plants expressing Pi22926, or silenced for TuA/B, show increased P. infestans susceptibility and decreased photosynthesis, plant growth and tuber yield. By contrast, StTuA/B overexpression reduces susceptibility, elevates chloroplast-derived reactive oxygen species production and increases photosynthesis and potato tuber yield by enhancing chloroplast protein translation. Another plant target of Pi22926, StMAP3Kβ2, interacts with StTuB, phosphorylating it to promote its translocation into chloroplasts. However, Pi22926 attenuates StTuB association with StMAP3Kβ2 and phosphorylation. This reduces StTuB translocation into chloroplasts, leading to its proteasome-mediated turnover in the cytoplasm. We uncover new mechanisms by which a pathogen effector inhibits immunity by disrupting key chloroplast functions. This work shows that StTuA/B break the growth-immunity trade-off, promoting both disease resistance and yield, revealing the enormous potential of chloroplast biology in crop breeding.

Overexpression of SIMK in menadione-treated alfalfa enhances antioxidant machinery and leads to oxidative stress resistance

Mitogen-activated protein kinases (MAPKs) transduce stress and developmental signals related to the production of reactive oxygen species (ROS). Alfalfa (Medicago sativa L.) is a valuable forage and human nutrition crop, however, the involvement of MAPKs in plant resistance to oxidative stress is poorly understood in this species. Therefore, we elucidated the role of STRESS-INDUCED MAPK (SIMK) in alfalfa response to menadione, a compound inducing ROS generation, exploiting transgenic alfalfa lines with contrasting SIMK abundance. SIMK was activated by short-term menadione treatment and relocated from the nucleus to the cytoplasm. Proteomic analysis revealed that menadione caused changes in the abundance of proteins involved in metabolism, oxidative stress, biotic stress response, detoxification of carbonyl species, glutathione homeostasis, chloroplast protein turnover, photosynthesis, and membrane trafficking. Genetic manipulations of SIMK altered the abundance of proteins involved in mitochondrial and chloroplast protein import and processing, as well as GLUTATHIONE S-TRANSFERASES (GSTs). Increased GST abundance and activity in roots, and modifications in mitochondrial and chloroplast protein turnover might be responsible for the elevated oxidative stress resistance of alfalfa line overexpressing SIMK. This was supported by the reduced ROS levels in this line. These results reveal a complex nature of plant stress response and suggest a new role of SIMK in the alfalfa resistance to menadione-induced oxidative stress.

Photosynthetic ROS and retrograde signaling pathways.

Sessile plants harness mitochondria and chloroplasts to sense and adapt to diverse environmental stimuli. These complex processes involve the generation of pivotal signaling molecules, including reactive oxygen species (ROS), phytohormones, volatiles, and diverse metabolites. Furthermore, the specific modulation of chloroplast proteins, through activation or deactivation, significantly enhances the plant's capacity to engage with its dynamic surroundings. While existing reviews have extensively covered the role of plastidial retrograde modules in developmental and light signaling, our focus lies in investigating how chloroplasts leverage photosynthetic ROS to navigate environmental fluctuations and counteract oxidative stress, thereby sustaining primary metabolism. Unraveling the nuanced interplay between photosynthetic ROS and plant stress responses holds promise for uncovering new insights that could reinforce stress resistance and optimize net photosynthesis rates. This exploration aspires to pave the way for innovative strategies to enhance plant resilience and agricultural productivity amidst changing environmental conditions.

Plastid HSP90C C-terminal extension region plays a regulatory role in chaperone activity and client binding.

HSP90Cs are essential molecular chaperones localized in the plastid stroma that maintain protein homeostasis and assist the import and thylakoid transport of chloroplast proteins. While HSP90C contains all conserved domains as an HSP90 family protein, it also possesses a unique feature in its variable C-terminal extension (CTE) region. This study elucidated the specific function of this HSP90C CTE region. Our phylogenetic analyses revealed that this intrinsically disordered region contains a highly conserved DPW motif in the green lineages. With biochemical assays, we showed that the CTE is required for the chaperone to effectively interact with client proteins PsbO1 and LHCB2 to regulate ATP-independent chaperone activity and to effectuate its ATP hydrolysis. The CTE truncation mutants could support plant growth and development reminiscing the wild type under normal conditions except for a minor phenotype in cotyledon when expressed at a level comparable to wild type. However, higher HSP90C expression was observed to correlate with a stronger response to specific photosystem II inhibitor DCMU, and CTE truncations dampened the response. Additionally, when treated with lincomycin to inhibit chloroplast protein translation, CTE truncation mutants showed a delayed response to PsbO1 expression repression, suggesting its role in chloroplast retrograde signaling. Our study therefore provides insights into the mechanism of HSP90C in client protein binding and the regulation of green chloroplast maturation and function, especially under stress conditions.

Architecture of the ATP-driven motor for protein import into chloroplasts

Thousands of nuclear-encoded proteins are transported into chloroplasts through the TOC-TIC translocon spanning the chloroplast envelope membranes. A motor complex pulls the translocated proteins out of the TOC-TIC complex into the chloroplast stroma by hydrolyzing ATP. The Orf2971-FtsHi complex was suggested to serve as the ATP-hydrolyzing motor in Chlamydomonas reinhardtii, but little is known about its architecture and assembly. Here, we report the 3.2-Å resolution structure of the Chlamydomonas Orf2971-FtsHi complex. The 20-subunit complex spans the chloroplast inner envelope with two bulky modules protruding into the intermembrane space and stromal matrix. Six subunits form a hetero-hexamer potentially providing the pulling force through ATP hydrolysis. The remaining subunits, including potential enzymes/chaperones, likely facilitate the complex assembly and regulate its proper function. Our results provide the structural foundation for mechanistic understanding of chloroplast protein translocation.

Participation of the stress-responsive CDSP32 thioredoxin in the modulation of chloroplast ATP-synthase activity in Solanum tuberosum.

Plant thioredoxins (TRXs) are involved in numerous metabolic and signalling pathways, such as light-dependent regulation of photosynthesis. The atypical TRX CDSP32, chloroplastic drought-induced stress protein of 32 kDa, includes two TRX-fold domains and participates in responses to oxidative stress as an electron donor to other thiol reductases. Here, we further characterised potato lines modified for CDSP32 expression to clarify the physiological roles of the TRX. Upon high salt treatments, modified lines displayed changes in the abundance and redox status of CDSP32 antioxidant partners, and exhibited sensitivity to combined saline-alkaline stress. In non-stressed plants overexpressing CDSP32, a lower abundance of photosystem II subunits and ATP-synthase γ subunit was noticed. The CDSP32 co-suppressed line showed altered chlorophyll a fluorescence induction and impaired regulation of the transthylakoid membrane potential during dark/light and light/dark transitions. These data, in agreement with the previously reported interaction between CDSP32 and ATP-synthase γ subunit, suggest that CDSP32 affects the redox regulation of ATP-synthase activity. Consistently, modelling of protein complex 3-D structure indicates that CDSP32 could constitute a suitable partner of ATP-synthase γ subunit. We discuss the roles of the TRX in the regulation of both photosynthetic activity and enzymatic antioxidant network in relation with environmental conditions.

STIC2 selectively binds ribosome-nascent chain complexes in the cotranslational sorting of Arabidopsis thylakoid proteins.

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 D1 nascent chains. Stalled RNCs translating the soluble ribosomal subunit uS2c were used for comparison. Quantitative tandem-mass spectrometry of the purified RNCs identified around 140 proteins specifically associated with D1 RNCs, mainly involved in protein and cofactor biogenesis, including chlorophyll biosynthesis, and other metabolic pathways. Functional analysis of STIC2, a newly identified D1 RNC interactor, revealed its cooperation with chloroplast protein SRP54 in the de novo biogenesis and repair of D1, and potentially other cotranslationally-targeted reaction center subunits of PSII and PSI. The primary binding interface between STIC2 and the thylakoid insertase Alb3 and its homolog Alb4 was mapped to STIC2's β-sheet region, and the conserved Motif III in the C-terminal regions of Alb3/4.

Recruitment of Cdc48 to chloroplasts by a UBX-domain protein in chloroplast-associated protein degradation

The translocon at the outer chloroplast membrane (TOC) is the gateway for chloroplast protein import and so is vital for photosynthetic establishment and plant growth. Chloroplast-associated protein degradation (CHLORAD) is a ubiquitin-dependent proteolytic system that regulates TOC. In CHLORAD, cytosolic Cdc48 provides motive force for the retrotranslocation of ubiquitinated TOC proteins to the cytosol but how Cdc48 is recruited is unknown. Here, we identify plant UBX-domain protein PUX10 as a component of the CHLORAD machinery. We show that PUX10 is an integral chloroplast outer membrane protein that projects UBX and ubiquitin-associated domains into the cytosol. It interacts with Cdc48 via its UBX domain, bringing it to the chloroplast surface, and with ubiquitinated TOC proteins via its ubiquitin-associated domain. Genetic analyses in Arabidopsis revealed a requirement for PUX10 during CHLORAD-mediated regulation of TOC function and plant development. Thus, PUX10 coordinates ubiquitination and retrotranslocation activities of CHLORAD to enable efficient TOC turnover.

MdBAM17, a novel member of the β-amylase gene family, positively regulates starch degradation in ALA-induced stomatal opening in apple (Malus × domestica)

5-Aminolevulinic acid (ALA) is a novel plant growth regulator that has shown outstanding capability to promote stomatal opening. Starch degradation, catalyzed by β-amylase (EC3.2.1.2, BAM), plays an important role in stomatal opening. However, whether the starch breakdown is involved in ALA-regulating stomatal movement is unclear. In the current study, we found that exogenous ALA effectively stimulated the starch breakdown in guard cells, increased β-amylase activity and promoted stomatal opening in leaves of apple (Malus × domestica). Based on genome-wide identification, we identified a total of 119 members of BAM gene family in ten commonly Rosaceae crops. Analyses of gene structure, motif identification, and gene pair collinearity revealed relative conservation among members within the same group or subgroup. Among these genes, MdBAM17 and other 12 genes were identified as the orthologous genes of AtBAM1, which is responsible for starch degradation to modulate the stomatal movement in Arabidopsis. RT-qPCR analysis revealed a positive correlation between the expressions of MdBAM17 and stomatal aperture, as well as β-amylase activity, whereas a negative correlation was observed with the starch content. Subcellular localization analysis confirmed that MdBAM17 is a chloroplast protein, consistent with the AtBAM1. MdBAM17 was mainly expressed in guard cells and responsive to exogenous ALA. Overexpressing MdBAM17 increased β-amylase activity and promoted starch breakdown, leading to stomatal opening, which was further strengthened by ALA. RNA-interfering MdBAM17 decreased β-amylase activity, resulting in starch accumulation, and impairing the stomatal opening by ALA. However, modulation of MdBAM17 expression did not affect the levels of flavonols and H2O2 in guard cells, suggesting that MdBAM17-promoted starch degradation may function at downstream of ROS signaling in the ALA-regulated stomatal opening. Our findings provide new insights into the mechanisms of ALA-regulated stomatal movement.

Structural insights into the chloroplast protein import in land plants

Chloroplast proteins are imported via the translocon at the outer chloroplast membrane (TOC)-translocon at the inner chloroplast membrane (TIC) supercomplex, driven by an ATPase motor. The Ycf2-FtsHi complex has been identified as the chloroplast import motor. However, its assembly and cooperation with the TIC complex during preprotein translocation remain unclear. Here, we present the structures of the Ycf2-FtsHi and TIC complexes from Arabidopsis and an ultracomplex formed between them from Pisum. The Ycf2-FtsHi structure reveals a heterohexameric AAA+ ATPase motor module with characteristic features. Four previously uncharacterized components of Ycf2-FtsHi were identified, which aid in complex assembly and anchoring of the motor module at a tilted angle relative to the membrane. When considering the structures of the TIC complex and the TIC-Ycf2-FtsHi ultracomplex together, it becomes evident that the tilted motor module of Ycf2-FtsHi enables its close contact with the TIC complex, thereby facilitating efficient preprotein translocation. Our study provides valuable structural insights into the chloroplast protein import process in land plants.

Conservation and specialization of the Ycf2-FtsHi chloroplast protein import motor in green algae

The protein import motor in chloroplasts plays a pivotal role in their biogenesis and homeostasis by driving the translocation of preproteins into chloroplasts. While the Ycf2-FtsHi complex serves as the import motor in land plants, its evolutionary conservation, specialization, and mechanisms across photosynthetic organisms are largely unexplored. Here, we isolated and determined the cryogenic electron microscopy (cryo-EM) structures of the native Ycf2-FtsHi complex from Chlamydomonas reinhardtii, uncovering a complex composed of up to 19 subunits, including multiple green-algae-specific components. The heterohexameric AAA+ ATPase motor module is tilted, potentially facilitating preprotein handover from the translocon at the inner chloroplast membrane (TIC) complex. Preprotein interacts with Ycf2-FtsHi and enhances its ATPase activity invitro. Integrating Ycf2-FtsHi and translocon at the outer chloroplast membrane (TOC)-TIC supercomplex structures reveals insights into their physical and functional interplay during preprotein translocation. By comparing these findings with those from land plants, our study establishes a structural foundation for understanding the assembly, function, evolutionary conservation, and diversity of chloroplast protein import motors.

A cargo sorting receptor mediates chloroplast protein trafficking through the secretory pathway.

Nucleus-encoded chloroplast proteins can be transported via the secretory pathway. The molecular mechanisms underlying the trafficking of chloroplast proteins between the intracellular compartments are largely unclear, and a cargo sorting receptor has not previously been identified in the secretory pathway. Here, we report a cargo sorting receptor that is specifically present in Viridiplantae and mediates the transport of cargo proteins to the chloroplast. Using a forward genetic analysis, we identified a gene encoding a transmembrane protein (MtTP930) in barrel medic (Medicago truncatula). Mutation of MtTP930 resulted in impaired chloroplast function and a dwarf phenotype. MtTP930 is highly expressed in the aerial parts of the plant and is localized to the endoplasmic reticulum (ER) exit sites and Golgi. MtTP930 contains typical cargo sorting receptor motifs, interacts with Sar1, Sec12, and Sec24, and participates in coat protein complex II vesicular transport. Importantly, MtTP930 can recognize the cargo proteins plastidial N-glycosylated nucleotide pyrophosphatase/phosphodiesterase (MtNPP) and α-carbonic anhydrase (MtCAH) in the ER and then transport them to the chloroplast via the secretory pathway. Mutation of a homolog of MtTP930 in Arabidopsis (Arabidopsis thaliana) resulted in a similar dwarf phenotype. Furthermore, MtNPP-GFP failed to localize to chloroplasts when transgenically expressed in Attp930 protoplasts, implying that these cargo sorting receptors are conserved in plants. These findings fill a gap in our understanding of the mechanism by which chloroplast proteins are sorted and transported via the secretory pathway.

Molecular and Biological Characteristics of a Peach Latent Mosaic Viroid PC Isolate in Peach from China: Base Mutations in Hairpin Stems and Implications for Symptomatology.

Peach latent mosaic viroid (PLMVd) infects peach trees in China and induces a conspicuous albino phenotype (peach calico, PC) that is closely associated with variants containing a 12-to-14 nucleotide hairpin insertion capped by a U-rich loop. Initially, PC disease distribution was limited to parts of Italy, and it was first detected in the field in China in 2019. To explore the molecular and biological characteristics of PLMVd PC isolates in peach in China, we conducted a comprehensive analysis of disease phenotype development and investigated the data-associated pathogenicity and in vivo dynamics of the Chinese isolate PC-A2 using slash-inoculation into GF-305 peach seedlings. Inoculated seedlings displayed PC symptoms much earlier following topping treatment, and PLMVd infectivity was further assessed using bioassay and semiquantitative RT-PCR experiments. Evolutionary analysis showed that the PC isolate and its progeny variants clustered into a single phylogroup distinct from reference PC-C40 isolates from Italy and PC-K1 and PC-K2 from South Korea. Some PC-A2 progeny variants from green leaves of PC-expressing seedlings showed unbalanced point mutations in hairpin stems compared with the PC-C40 reference sequence and constituted a new stem insertion type. The results reveal associations between the recessive phenotypes of peach albino symptoms and base variation in hairpin stem insertions relative to the PC-C40/chloroplastic heat shock protein 90 reference sequence.

Quantitative phosphoproteomics reveals molecular pathway network in wheat resistance to stripe rust

Protein phosphorylation plays an important role in immune signaling transduction in plant resistance to pathogens. Wheat stripe rust, caused by Puccinia striiformis f. sp. tritici (Pst), severely devastates wheat production. Nonetheless, the molecular mechanism of wheat resistance to stripe rust remains limited. In this study, quantitative phosphoproteomics was employed to investigate the protein phosphorylation changes in wheat challenged by Pst. A total of 1537 and 2470 differentially accumulated phosphoproteins (DAPs) were identified from four early infection stage (6, 12, 18 and 24 h post-inoculation) in incompatible and compatible wheat-Pst interactions respectively. KEGG analysis revealed that Oxidative Phosphorylation, Phosphatidylinositol Signaling, and MAPK signaling processes are distinctively enriched in incompatible interaction, while Biosynthesis of secondary metabolites and RNA degradation process were significantly enriched in compatible interactions. In particular, abundant changes in phosphorylation levels of chloroplast proteins were identified, suggesting the regulatory role of photosynthesis in wheat-Pst interaction, which is further emphasized by protein-protein interaction (PPI) network analysis. Motif-x analysis identified [xxxxSPxxxx] motif, likely phosphorylation sites for defensive response-related kinases, and a new [xxxxSSxxxx] motif significantly enriched in incompatible interaction. The results shed light on the early phosphorylation events contributing to wheat resistance against Pst. Moreover, our study demonstrated that the phosphorylation levels of Nucleoside diphosphate kinase TaNAPK1 are upregulated at 12 hpi with CYR23 and at 24 hpi with CYR31. Transient silencing of TaNAPK1 was able to attenuate wheat resistance to CYR23 and CYR31. Our study provides new insights into the mechanisms underlying Pst-wheat interactions and may provide database to find potential targets for the development of new resistant varieties.

SlPGR5/SlPGRL1 pathway-dependent cyclic electron transport regulates photoprotection and chloroplast quality in tomato plants

The essential photoprotective role of proton gradient regulation 5 (PGR5)-dependent cyclic electron flow (CEF) has been reported in Arabidopsis, rice, and algae. However, its functional assessment has not been performed in tomato yet. In this study, we focused on elucidate the function of SlPGR5 and SlPGR5-like photosynthetic phenotype 1 (PGRL1) in tomato. We performed RNA interference and found that SlPGR5/SlPGRL1-suppressed transformants exhibited extremely low CO2 assimilation capacity, their photosystem I (PSI) and PSII were severely photoinhibited and chloroplasts were obviously damaged. The SlPGR5/SlPGRL1-suppressed plants almost completely inhibited CEF and Y(ND), and PSII photoinhibition may be directly related to the inability to produce sufficient proton motive force to induce NPQ. The transgenic plants overexpressing SlPGR5 and SlPGRL1 driven by 35S promoter capable alleviate photoinhibition of plants under low night temperature. The transcriptomic and proteomic analyses suggested that the nuclear gene transcription and turnover of chloroplast proteins, including the plastoglobule-related proteins, were closely related to SlPGR5/SlPGRL1 pathway dependent CEF. The bridge relationship between CEF and chloroplast quality maintenance was a novel report to our knowledge. In conclusion, these results revealed the regulatory mechanism of the SlPGR5/SlPGRL1 pathway in photoprotection and maintenance of chloroplast function in tomato, which is crucial for reduce yield loss, especially under adverse environmental conditions.

Temporal dynamics of chloroplast biogenesis revealed initiation of photosynthesis-related gene expression and protein complexes during alfalfa seed germination

The chloroplast biogenesis occurs in cotyledon during alfalfa seed germination before true leaf formation, and is extremely important for the followed plant development and growth. In this study, we conducted a simulation of alfalfa seed germination in the soil by using tin foil and focused on 10 pivotal time points of chloroplast biogenesis in cotyledons before and after light exposure, which showed significant differences in multispectral images, and covered the whole process of chloroplast biogenesis from proplastid, etioplast to mature chloroplast. We revealed three phases that referred to the programmed involvements of photosynthesis promotion, ultrastructure maturity, transcriptomic expression, and protein complex construction, and observed distinct transcriptional expressions of genes from nuclear and chloroplast genomes. In phase I at dark germination before light exposure, chloroplast-encoded genes showed up-regulated expressions together with the importation of chloroplast proteins. In phase II for the first day after light exposure, nuclear-encoded genes’ expressions were initiated at 2 h after light exposure (E2h), followed by swift assembly of chloroplast thylakoid membrane protein complexes, and roaring Fv/Fm and contents of chlorophyll a, chlorophyll b and carotenoid. The initiation at E2h was pronounced by the observation of gradual accumulation of single lamella, and facilitated the formation of granum stacks (thylakoid) at E8h in phase II. In phase III from the second day after light exposure, chloroplast became gradually complete with the fully established photosynthetic capacity. Altogether, our results layed a theoretical foundation for enhancing potential photosynthetic efficiency in alfalfa and related species.