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Summary: Isoxaben is a pre‐emergent herbicide used to control broadleaf weeds. While the phytotoxic mechanism is not completely understood, isoxaben interferes with cellulose synthesis. Certain mutations in cellulose synthase complex proteins can confer isoxaben tolerance; however, these mutations can cause compromised cellulose synthesis and perturbed plant growth, rendering them unsuitable as herbicide tolerance traits. We conducted a genetic screen to identify new genes associated with isoxaben tolerance by screening a selection of Arabidopsis thaliana T‐DNA mutants. We found that mutations in a FERREDOXIN‐NADP(+) OXIDOREDUCTASE‐LIKE (FNRL) gene enhanced tolerance to isoxaben, exhibited as a reduction in primary root stunting, reactive oxygen species accumulation and ectopic lignification. The fnrl mutant did not exhibit a reduction in cellulose levels following exposure to isoxaben, indicating that FNRL operates upstream of isoxaben‐induced cellulose inhibition. In line with these results, transcriptomic analysis revealed a highly reduced response to isoxaben treatment in fnrl mutant roots. The fnrl mutants displayed constitutively induced mitochondrial retrograde signalling, and the observed isoxaben tolerance is partially dependent on the transcription factor ANAC017, a key regulator of mitochondrial retrograde signalling. Moreover, FNRL is highly conserved across all plant lineages, implying conservation of its function. Notably, fnrl mutants did not show a growth penalty in shoots, making FNRL a promising target for biotechnological applications in breeding isoxaben tolerance in crops. [ABSTRACT FROM AUTHOR]

Summary Fate determination is indispensable for the accurate shaping and specialization of plant organs, a process critical to the structural and functional diversity in plant kingdom. The TEOSINTE BRANCHED 1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP) family of transcription factors has been recognized for its significant contributions to plant organogenesis and morphogenesis. Recent research has shed light on the pivotal roles that TCPs play in fate determination. In this review, we delve into the current understanding of TCP functions, emphasizing their critical influence on fate determination from the organelle to the cell and organ levels. We also consolidate the molecular mechanisms through which TCPs exert their regulatory effects on fate determination. Additionally, we highlight intriguing points of TCPs that warrant further exploration in future research endeavors. [ABSTRACT FROM AUTHOR]

SUMMARY Citral, a naturally occurring acyclic monoterpene aldehyde, is present in the essential oils of various plants, but only a few produce it in abundance. Despite its importance as a key aroma molecule, knowledge regarding the in‐planta biosynthesis of citral and its metabolic origin remains limited. Here, we have elucidated the functions of an alcohol dehydrogenase (CfADH1) and an aldoketo‐reductase (CfAKR2b) in citral biosynthesis in lemongrass (Cymbopogon flexuosus), one of the most cultivated aromatic crops for its citral‐rich essential oil. Expression of both CfADH1 and CfAKR2b showed correlation with citral accumulation in different developmental stages. Recombinant CfADH1 and CfAKR2b, despite their sequence unrelatedness, catalyzed citral formation from geraniol with NADP cofactor. Virus‐induced gene silencing in lemongrass and transient expression in lemon balm (Melissa officinalis) demonstrated the in‐planta involvement of CfADH1 and CfAKR2b in citral biosynthesis. While CfADH1 exhibited a dual cytosolic/plastidial localization, CfAKR2b was localized to the cytosol. This was supported by higher citral‐forming activity in the cytosolic fraction than in the chloroplast fraction of lemongrass leaf extract. Moreover, feeding lemongrass seedlings with inhibitors specific to the cytosolic mevalonate pathway and the plastidial methylerythritol phosphate pathway, combined with volatile profiling, supported the involvement of both pathways in citral formation. Taken together, our results indicate that high citral production has evolved in lemongrass through the recruitment of phylogenetically distant enzymes localized in both the cytosol and plastids. [ABSTRACT FROM AUTHOR]

Mechanisms that allocate cellular space to organelles are of fundamental importance to biology but remain poorly understood. A detailed understanding of mechanisms that allocate cellular space to plastids, such as chloroplasts, will lead to high-yielding crops with enhanced nutritional value. The HIGH PIGMENT (HP) genes in tomato contribute to regulated proteolysis and abscisic acid metabolism. The HP1 gene was the first gene reported to influence the amount of cellular space occupied by chloroplasts and chromoplasts almost 20 years ago. Recently, our knowledge of mechanisms that allocate cellular space to plastids was enhanced by new information on the influence of cell type on the amount of cellular space occupied by plastids and the identification of new genes that help to allocate cellular space to plastids. These genes encode proteins with unknown and diverse biochemical functions. Several transcription factors were recently reported to regulate the numbers and sizes of chloroplasts in fleshy fruit. If these transcription factors do not induce compensating effects on cell size, they should affect the amount of cellular space occupied by plastids. Although we can now propose more detailed models for the network that allocates cellular space to plastids, many gaps remain in our knowledge of this network and the genes targeted by this network. Nonetheless, these recent breakthroughs provide optimism for future progress in this field. [ABSTRACT FROM AUTHOR]

Complex plastids, characterized by more than two bounding membranes, still present an evolutionary puzzle for the traditional endosymbiotic theory. Unlike primary plastids that directly evolved from cyanobacteria, complex plastids originated from green or red algae. The Chromalveolata hypothesis proposes a single red alga endosymbiosis that involved the ancestor of all the Chromalveolata lineages: cryptophytes, haptophytes, stramenopiles, and alveolates. As extensive phylogenetic analyses contradict the monophyly of Chromalveolata, serial plastid endosymbiosis models were proposed, suggesting a single secondary red alga endosymbiosis within Cryptophyta, followed by subsequent plastid transfers to other chromalveolates. Our findings based on 97 plastid-encoded markers, 112 species, and robust phylogenetic methods challenge all the existing models. They reveal two independent secondary endosymbioses, one within Cryptophyta and one within stramenopiles, precisely the phylum Ochrophyta, with two different groups of red algae. Consequently, we propose a new model for the emergence of red alga plastid–containing lineages and, through molecular clock analyses, estimate their ages. [ABSTRACT FROM AUTHOR]

BACKGROUND: Late‐ripening citrus plays an important role in the stability of the global citrus industry. However, the regreening phenomenon in Valencia oranges impacts the peel color and commercial value. Ethylene degreening is an effective technique to improve the color of citrus fruits, but this effect may be delayed in regreened oranges. To better clarify this phenomenon, plastid morphology, pigment and phytohormone content in ethephon‐degreened Midknight Valencia oranges harvested in different stages were evaluated. RESULTS: Results showed that in fruits harvested at the turning stage, ethephon degreening treatment induced a chloroplast‐to‐chromoplast transition, and chlorophyll degradation and carotenoid accumulation were accelerated. Conversely, in fruits harvested at the regreening stage, the changes in plastid morphology were minimal, with delayed changes in chlorophyll and carotenoids. Genes related to ethylene biosynthesis and signaling pathways supported these responses. Variations in endogenous auxin, jasmonic acid, abscisic acid and gibberellins could partially explain this phenomenon. CONCLUSION: The response of Midknight Valencia oranges to ethephon degreening was delayed in the regreening stage, possibly due to the dynamic variations in endogenous phytohormones. © 2024 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Repeats can mediate rearrangements and recombination in plant mitochondrial genomes and plastid genomes. While repeat accumulations are linked to heightened evolutionary rates and complex structures in specific lineages, debates persist regarding the extent of their influence on sequence and structural evolution. In this study, 75 Plantago plastomes were analyzed to investigate the relationships between repeats, nucleotide substitution rates, and structural variations. Extensive repeat accumulations were associated with significant rearrangements and inversions in the large inverted repeats (IRs), suggesting that repeats contribute to rearrangement hotspots. Repeats caused infrequent recombination that potentially led to substoichiometric shifting, supported by long-read sequencing. Repeats were implicated in elevating evolutionary rates by facilitating localized hypermutation, likely through DNA damage and repair processes. This study also observed a decrease in nucleotide substitution rates for loci translocating into IRs, supporting the role of biased gene conversion in maintaining lower substitution rates. Combined with known parallel changes in mitogenomes, it is proposed that potential dysfunction in nuclear-encoded genes associated with DNA replication, recombination, and repair may drive the evolution of Plantago organellar genomes. These findings contribute to understanding how repeats impact organellar evolution and stability, particularly in rapidly evolving plant lineages. [ABSTRACT FROM AUTHOR]

SUMMARY: Redox changes of pyridine nucleotides in cellular compartments are highly dynamic and their equilibria are under the influence of various reducing and oxidizing reactions. To obtain spatiotemporal data on pyridine nucleotides in living plant cells, typical biochemical approaches require cell destruction. To date, genetically encoded fluorescent biosensors are considered to be the best option to bridge the existing technology gap, as they provide a fast, accurate, and real‐time readout. However, the existing pyridine nucleotides genetically encoded fluorescent biosensors are either sensitive to pH change or slow in dissociation rate. Herein, we employed the biosensors which generate readouts that are pH stable for in planta measurement of NADH/NAD+ ratio and NADPH level. We generated transgenic Arabidopsis lines that express these biosensors in plastid stroma and cytosol of whole plants and pollen tubes under the control of CaMV 35S and LAT52 promoters, respectively. These transgenic biosensor lines allow us to monitor real‐time dynamic changes in NADH/NAD+ ratio and NADPH level in the plastids and cytosol of various plant tissues, including pollen tubes, root hairs, and mesophyll cells, using a variety of fluorescent instruments. We anticipate that these valuable transgenic lines may allow improvements in plant redox biology studies. Significant Statement: Monitoring dynamic changes in pyridine nucleotides (e.g., NADPH and NADH/NAD+ ratio) in plants at the subcellular level is a major obstacle in plant bioenergetics research. Genetically encoded fluorescent biosensors are considered to be the best option so far because they provide fast, accurate, and real‐time readouts. Here, we present a detailed study on the application of pyridine nucleotide biosensors, and we anticipate that these transgenic lines will be useful in plant redox biology studies. [ABSTRACT FROM AUTHOR]

DNA cytosine methylation is an important epigenetic mechanism in genomic DNA. In most land plants, it is absent in the chloroplast DNA. We detected methylation in the chloroplast DNA of the kelp Saccharina latissima, a non‐model macroalgal species of high ecological and economic importance. Since the functional role of the chloroplast methylome is yet largely unknown, this fundamental research assessed the chloroplast DNA cytosine methylation in wild and laboratory raised kelp from different climatic origins (High‐Arctic at 79° N, and temperate at 54° N), and in laboratory samples from these origins raised at different temperatures (5, 10 and 15°C). Results suggest genome‐wide differences in methylated sites and methylation level between the origins, while rearing temperature had only weak effects on the chloroplast methylome. Our findings point at the importance of matching conditions to origin in restoration and cultivation processes to be valid even on plastid level. [ABSTRACT FROM AUTHOR]

SUMMARY: Maize plastid terminal oxidase1 (ZmPTOX1) plays a pivotal role in seed development by upholding redox balance within seed plastids. This study focuses on characterizing the white kernel mutant 3735 (wk3735) mutant, which yields pale‐yellow seeds characterized by heightened protein but reduced carotenoid levels, along with delayed germination compared to wild‐type (WT) seeds. We successfully cloned and identified the target gene ZmPTOX1, responsible for encoding maize PTOX—a versatile plastoquinol oxidase and redox sensor located in plastid membranes. While PTOX's established role involves regulating redox states and participating in carotenoid metabolism in Arabidopsis leaves and tomato fruits, our investigation marks the first exploration of its function in storage organs lacking a photosynthetic system. Through our research, we validated the existence of plastid‐localized ZmPTOX1, existing as a homomultimer, and established its interaction with ferredoxin‐NADP+ oxidoreductase 1 (ZmFNR1), a crucial component of the electron transport chain (ETC). This interaction contributes to the maintenance of redox equilibrium within plastids. Our findings indicate a propensity for excessive accumulation of reactive oxygen species (ROS) in wk3735 seeds. Beyond its known role in carotenoids' antioxidant properties, ZmPTOX1 also impacts ROS homeostasis owing to its oxidizing function. Altogether, our results underscore the critical involvement of ZmPTOX1 in governing seed development and germination by preserving redox balance within the seed plastids. Significance Statement: Aside from its established role as a vital regulator of redox states and its involvement in carotenoid metabolism, as observed in Arabidopsis leaves and tomato fruits, this study marks the inaugural exploration into PTOX's function within storage organs lacking a photosynthetic system. Our findings elucidate how ZmPTOX1 significantly impacts protein and starch accumulation in seeds, along with the pace of seed germination. Moreover, it actively maintains redox equilibrium across various stages of seed development and germination. [ABSTRACT FROM AUTHOR]

Summary: Endosperm is the main storage organ in cereal grain and determines grain yield and quality. The molecular mechanisms of heat shock proteins in regulating starch biosynthesis and endosperm development remain obscure.Here, we report a rice floury endosperm mutant flo24 that develops abnormal starch grains in the central starchy endosperm cells. Map‐based cloning and complementation test showed that FLO24 encodes a heat shock protein HSP101, which is localized in plastids. The mutated protein FLO24T296I dramatically lost its ability to hydrolyze ATP and to rescue the thermotolerance defects of the yeast hsp104 mutant.The flo24 mutant develops more severe floury endosperm when grown under high‐temperature conditions than normal conditions. And the FLO24 protein was dramatically induced at high temperature. FLO24 physically interacts with several key enzymes required for starch biosynthesis, including AGPL1, AGPL3 and PHO1. Combined biochemical and genetic evidence suggests that FLO24 acts cooperatively with HSP70cp‐2 to regulate starch biosynthesis and endosperm development in rice.Our results reveal that FLO24 acts as an important regulator of endosperm development, which might function in maintaining the activities of enzymes involved in starch biosynthesis in rice. [ABSTRACT FROM AUTHOR]

SUMMARY: Knotted1‐like homeodomain (KNOX) proteins are essential in regulating plant organ differentiation. Land plants, including tomato (Solanum lycopersicum), have two classes of the KNOX protein family, namely, class I (KNOX I) and class II KNOX (KNOX II). While tomato KNOX I proteins are known to stimulate chloroplast development in fruit, affecting fruit coloration, the role of KNOX II proteins in this context remains unclear. In this study, we employ CRISPR/Cas9 to generate knockout mutants of the KNOX II member, SlKN5. These mutants display increased leaf complexity, a phenotype commonly associated with reduced KNOX II activity, as well as enhanced accumulation of chloroplasts and chlorophylls in smaller cells within young, unripe fruit. RNA‐seq data analyses indicate that SlKN5 suppresses the transcriptions of genes involved in chloroplast biogenesis, chlorophyll biosynthesis, and gibberellin catabolism. Furthermore, protein–protein interaction assays reveal that SlKN5 physically interacts with three transcriptional repressors from the BLH1‐clade of BEL1‐like homeodomain (BLH) protein family, SlBLH4, SlBLH5, and SlBLH7, with SlBLH7 showing the strongest interaction. CRISPR/Cas9‐mediated knockout of these SlBLH genes confirmed their overlapping roles in suppressing chloroplast biogenesis, chlorophyll biosynthesis, and lycopene cyclization. Transient assays further demonstrate that the SlKN5–SlBLH7 interaction enhances binding capacity to regulatory regions of key chloroplast‐ and chlorophyll‐related genes, including SlAPRR2‐like1, SlCAB‐1C, and SlGUN4. Collectively, our findings elucidate that the KNOX II SlKN5‐SlBLH regulatory modules serve to inhibit fruit greening and subsequently promote lycopene accumulation, thereby fine‐tuning the color transition from immature green fruit to mature red fruit. Significance Statement: This study shed light on regulatory mechanisms of plastid development in fruits by demonstrating that the class II knotted‐like homeodomain protein SlKN5 negatively regulates chloroplast number and chlorophyll accumulation in tomato fruit development. SlKN5 interacts with BEL1‐like homeodomain protein SlBLH7 and its paralogs to suppress SlAPRR2‐like1 and chlorophyll biosynthetic genes, promoting the developmental shift from growth phase to onset of ripening. This highlights the crucial role of the SlKN5‐BLH module in the regulation of tomato fruit coloration. [ABSTRACT FROM AUTHOR]

The chloroplast thylakoid membrane is composed of membrane lipids and photosynthetic protein complexes, and the orchestration of thylakoid lipid biosynthesis and photosynthesis-associated protein accumulation is considered important for thylakoid development. Galactolipids consist of ∼80% of the thylakoid lipids, and their biosynthesis is fundamental for chloroplast development. We previously reported that the suppression of galactolipid biosynthesis decreased the expression of photosynthesis-associated nuclear-encoded genes (PhAPGs) and photosynthesis-associated plastid-encoded genes (PhAPGs). However, the mechanism for coordinative regulation between galactolipid biosynthesis in plastids and the expression of PhANGs and PhAPGs remains largely unknown. To elucidate this mechanism, we investigated the gene expression patterns in galactolipid-deficient Arabidopsis seedlings during the de-etiolation process. We found that galactolipids are crucial for inducing both the transcript accumulation of PhANGs and PhAPGs and the accumulation of plastid-encoded photosynthesis-associated proteins in developing chloroplasts. Genetic analysis indicates the contribution of the GENOMES UNCOUPLED1 (GUN1)–mediated plastid-to-nucleus signaling pathway to PhANG regulation in response to galactolipid levels. Previous studies suggested that the accumulation of GUN1 reflects the state of protein homeostasis in plastids and alters the PhANG expression level. Thus, we propose a model that galactolipid biosynthesis determines the protein homeostasis in plastids in the initial phase of de-etiolation and optimizes GUN1-dependent signaling to regulate the PhANG expression. This mechanism might contribute to orchestrating the biosynthesis of lipids and proteins for the biogenesis of functional chloroplasts in plants. [ABSTRACT FROM AUTHOR]

Although plastid genome (plastome) structure is highly conserved across most seed plants, investigations during the past two decades have revealed several disparately related lineages that experienced substantial rearrangements. Most plastomes contain a large inverted repeat and two single-copy regions, and a few dispersed repeats; however, the plastomes of some taxa harbour long repeat sequences (>300 bp). These long repeats make it challenging to assemble complete plastomes using short-read data, leading to misassemblies and consensus sequences with spurious rearrangements. Single-molecule, long-read sequencing has the potential to overcome these challenges, yet there is no consensus on the most effective method for accurately assembling plastomes using long-read data. We generated a pipeline, plastid Genome Assembly Using Long-read data (ptGAUL), to address the problem of plastome assembly using long-read data from Oxford Nanopore Technologies (ONT) or Pacific Biosciences platforms. We demonstrated the efficacy of the ptGAUL pipeline using 16 published long-read data sets. We showed that ptGAUL quickly produces accurate and unbiased assemblies using only ~50× coverage of plastome data. Additionally, we deployed ptGAUL to assemble four new Juncus (Juncaceae) plastomes using ONT long reads. Our results revealed many long repeats and rearrangements in Juncus plastomes compared with basal lineages of Poales. The ptGAUL pipeline is available on GitHub: https://github.com/Bean061/ptgaul.

This article discusses the development of new monomeric TALE-linked base editors for editing the plastid genome in rice. The researchers explored different cytidine deaminases to generate these base editors, including a human APOBEC3A variant and an improved cytidine deaminase based on TadA. They evaluated the editing efficiency of these base editors in rice calli and transgenic rice plants, finding that the monomeric TALE-linked base editors induced high-efficiency C-to-T conversions. The researchers also examined the off-target activity of these base editors and discussed the advantages they offer over other base editors. Overall, this study demonstrates the potential of these monomeric TALE-linked base editors for efficient plastid genome editing in rice. [Extracted from the article]

In plants, C-to-U RNA editing mainly occurs in plastid and mitochondrial transcripts, which contributes to a complex transcriptional regulatory network. More evidence reveals that RNA editing plays critical roles in plant growth and development. However, accurate detection of RNA editing sites using transcriptome sequencing data alone is still challenging. In the present study, we develop PlantC2U, which is a convolutional neural network, to predict plastid C-to-U RNA editing based on the genomic sequence. PlantC2U achieves >95% sensitivity and 99% specificity, which outperforms the PREPACT tool, random forests, and support vector machines. PlantC2U not only further checks RNA editing sites from transcriptome data to reduce possible false positives, but also assesses the effect of different mutations on C-to-U RNA editing based on the flanking sequences. Moreover, we found the patterns of tissue-specific RNA editing in the mangrove plant Kandelia obovata , and observed reduced C-to-U RNA editing rates in the cold stress response of K. obovata , suggesting their potential regulatory roles in plant stress adaptation. In addition, we present RNAeditDB, available online at https://jasonxu.shinyapps.io/RNAeditDB/. Together, PlantC2U and RNAeditDB will help researchers explore the RNA editing events in plants and thus will be of broad utility for the plant research community. [ABSTRACT FROM AUTHOR]

Pyrenoids are microcompartments that are universally found in the photosynthetic plastids of various eukaryotic algae. They contain ribulose-1,5-bisphosphate carbox-ylase/oxygenase (Rubisco) and play a pivotal role in facilitating CO2 assimilation via CO2-concentrating mechanisms (CCMs). Recent investigations involving model algae have revealed that pyrenoid-associated proteins participate in pyrenoid biogenesis and CCMs. However, these organisms represent only a small part of algal lineages, which limits our comprehensive understanding of the diversity and evolution of pyrenoid-based CCMs. Here we report a pyrenoid proteome of the chlorarachniophyte alga Amorphochlora amoebiformis, which possesses complex plastids acquired through secondary endosymbiosis with green algae. Proteomic analysis using mass spectrom-etry resulted in the identification of 154 potential pyrenoid components. Subsequent localization experiments demonstrated the specific targeting of eight proteins to pyrenoids. These included a putative Rubisco-binding linker, carbonic anhydrase, membrane transporter, and uncharacterized GTPase proteins. Notably, most of these proteins were unique to this algal lineage. We suggest a plausible scenario in which pyrenoids in chlorarachniophytes have evolved independently, as their components are not inherited from green algal pyrenoids. [ABSTRACT FROM AUTHOR]

Corylopsis Siebold & Zucc. (Hamamelidaceae) is widely used as a horticultural plant and comprises approximately 25 species in East Asia. Molecular research is essential to distinguish Corylopsis species, which are morphologically similar. Molecular research has been conducted using a small number of genes but not in Corylopsis. Plastid genomes of Corylopsis species (Corylopsis gotoana, Corylopsis pauciflora, and Corylopsis sinensis) were sequenced using next-generation sequencing techniques. Repeats and nucleotide diversity that could be used as DNA markers were also investigated. A phylogenetic investigation was carried out using 79 protein-coding genes to infer the evolutionary relationships within the genus Corylopsis. By including new plastomes, the overall plastid genome structure of Corylopsis was similar. Simple sequence repeats of 73–106 SSRs were identified in the protein-coding genes of the plastid genomes, and 33–40 long repeat sequences were identified in the plastomes. The Pi value of the rpl33_rps18 region, an intergenic spacer, was the highest. Phylogenetic analysis demonstrated that Corylopsis is a monophyletic group and Loropetalum is closely related to Corylopsis. C. pauciflora, C. gotoana, and C. spicata formed a clade distributed in Japan, whereas C. sinensis, C. glandulifera, and C. velutina formed a clade that was distributed in China. [ABSTRACT FROM AUTHOR]

The interaction and coevolution between nuclear and cytoplasmic genomes are one of the fundamental hallmarks of eukaryotic genome evolution and, 2 billion yr later, are still major contributors to the formation of new species. Although many studies have investigated the role of cytonuclear interactions following allopolyploidization, the relative magnitude of the effect of subgenome dominance versus cytonuclear interaction on genome evolution remains unclear. The Brassica triangle of U features 3 diploid species that together have formed 3 separate allotetraploid species on similar evolutionary timescales, providing an ideal system for understanding the contribution of the cytoplasmic donor to hybrid polyploid. Here, we investigated the evolutionary pattern of organelle-targeted genes in Brassica carinata (BBCC) and 2 varieties of Brassica juncea (AABB) at the whole-genome level, with particular focus on cytonuclear enzyme complexes. We found partial evidence that plastid-targeted genes experience selection to match plastid genomes, but no obvious corresponding signal in mitochondria-targeted genes from these 2 separately formed allopolyploids. Interestingly, selection acting on plastid genomes always reduced the retention rate of plastid-targeted genes encoded by the B subgenome, regardless of whether the Brassica nigra (BB) subgenome was contributed by the paternal or maternal progenitor. More broadly, this study illustrates the distinct selective pressures experienced by plastid- and mitochondria-targeted genes, despite a shared pattern of inheritance and natural history. Our study also highlights an important role for subgenome dominance in allopolyploid genome evolution, even in genes whose function depends on separately inherited molecules. [ABSTRACT FROM AUTHOR]

AbstractPetrocosmea qinlingensis is a protected wild plant endemic in China, inhabiting low-light limestone cliffs but the complete chloroplast genome has not been reported. In this study, we first sequenced and assembled the complete chloroplast genome of P. qinlingensis. The total size of this genome was 153,865 bp, including a large single-copy (LSC) region (84,737 bp), a small single-copy (SSC) region (18,244 bp), and two inverted repeats (IRs) regions (25,442 bp). This genome encoded 111 uniquegenes, consisted of 77 protein-coding genes, four ribosomal RNA genes, and 30 transfer RNA genes. Phylogenomic analysis based on the chloroplast protein-coding genes and showed that the genus Petrocosmea was the closest relative to Raphiocarpus. Our results will support further phylogeographic, population genetic studies of this species.

Typical (peridinin-containing) dinoflagellates possess plastid genomes composed of small plasmids named "minicircles". Despite the ecological importance of dinoflagellate photosynthesis in corals and marine ecosystems, the structural characteristics, replication dynamics, and evolutionary forcing of dinoflagellate plastid genomes remain poorly understood. Here, we sequenced the plastid genome of the symbiodiniacean species Fugacium kawagutii and conducted comparative analyses. We identified psbT -coding minicircles, features previously not found in Symbiodiniaceae. The copy number of F. kawagutii minicircles showed a strong diel dynamics, changing between 3.89 and 34.3 copies/cell and peaking in mid-light period. We found that F. kawagutii minicircles are the shortest among all dinoflagellates examined to date. Besides, the core regions of the minicircles are highly conserved within genus in Symbiodiniaceae. Furthermore, the codon usage bias of the plastid genomes in Heterocapsaceae, Amphidiniaceae, and Prorocentraceae species are greatly influenced by selection pressure, and in Pyrocystaceae, Symbiodiniaceae, Peridiniaceae, and Ceratiaceae species are influenced by both natural selection pressure and mutation pressure, indicating a family-level distinction in codon usage evolution in dinoflagellates. Phylogenetic analysis using 12 plastid-encoded proteins and five nucleus-encoded plastid proteins revealed accelerated evolution trend of both plastid- and nucleus-encoded plastid proteins in peridinin- and fucoxanthin-dinoflagellate plastids compared to plastid proteins of nondinoflagellate algae. These findings shed new light on the structure and evolution of plastid genomes in dinoflagellates, which will facilitate further studies on the evolutionary forcing and function of the diverse dinoflagellate plastids. The accelerated evolution documented here suggests plastid-encoded sequences are potentially useful for resolving closely related dinoflagellates. [ABSTRACT FROM AUTHOR]

Petrocosmea qinlingensis is a protected wild plant endemic in China, inhabiting low-light limestone cliffs but the complete chloroplast genome has not been reported. In this study, we first sequenced and assembled the complete chloroplast genome of P. qinlingensis. The total size of this genome was 153,865 bp, including a large single-copy (LSC) region (84,737 bp), a small single-copy (SSC) region (18,244 bp), and two inverted repeats (IRs) regions (25,442 bp). This genome encoded 111 uniquegenes, consisted of 77 protein-coding genes, four ribosomal RNA genes, and 30 transfer RNA genes. Phylogenomic analysis based on the chloroplast protein-coding genes and showed that the genus Petrocosmea was the closest relative to Raphiocarpus. Our results will support further phylogeographic, population genetic studies of this species. [ABSTRACT FROM AUTHOR]

Diatoms are important primary producers in the world's oceans, yet their growth is constrained in large regions by low bioavailable iron (Fe). Low‐Fe stress‐induced limitation of primary production is due to requirements for Fe in components of essential metabolic pathways including photosynthesis and other chloroplast plastid functions. Studies have shown that under low‐Fe stress, diatoms alter plastid‐specific processes, including components of electron transport. These physiological changes suggest changes of protein content and in protein abundances within the diatom plastid. While in silico predictions provide putative information on plastid‐localized proteins, knowledge of diatom plastid proteins remains limited in comparison to well‐studied model photosynthetic organisms. To address this, we employed shotgun proteomics to investigate the proteome of subcellular plastid‐enriched fractions from Thalassiosira pseudonana to gain a better understanding of how the plastid proteome is remodeled in response to Fe limitation. Using mass spectrometry‐based peptide identification and quantification, we analyzed T. pseudonana grown under Fe‐replete and ‐limiting conditions. Through these analyses, we inferred the relative quantities of each protein, revealing that Fe limitation regulates major metabolic pathways in the plastid, including the Calvin cycle. Additionally, we observed changes in the expression of light‐harvesting proteins. In silico localization predictions of proteins identified in this plastid‐enriched proteome allowed for an in‐depth comparison of theoretical versus observed plastid‐localization, providing evidence for the potential of additional protein import pathways into the diatom plastid. [ABSTRACT FROM AUTHOR]

Photosynthesis allows for the formation of biomass from inorganic carbon and therefore greatly enhances the amount of organic material on planet Earth. Especially, oxygenic photosynthesis removed a major bottleneck in the formation of biomass by utilising ubiquitous water (H2O) and CO2 molecules as raw materials for organic molecules. This, over billions of years, shaped the world into the form we know today, with an oxygen-containing atmosphere, largely oxygenated water bodies and landmasses consisting of sediment rocks. Oxygenic photosynthesis furthermore enabled the evolution of aerobic energy metabolism, and it would be very difficult to imagine animal (including human) life in the absence of molecular oxygen as an electron acceptor. Oxygenic photosynthesis first, and exclusively, evolved in cyanobacteria. However, eukaryotes also learned to photosynthesise, albeit with a trick, which is the integration of formerly free-living cyanobacteria into the eukaryotic cell. There, the former bacteria became endosymbionts, and from these endosymbionts, the photosynthetic organelles (termed plastids) evolved. In almost all major groups of eukaryotes, plastid-containing members are found. At the same time, plastid-related features also indicate that these plastids form a monophyletic group. This can be explained by the transfer of plastids between the eukaryotic super-groups, leading to plastids being found in groups that are otherwise non-photosynthetic. In this chapter, we discuss the evolutionary origin of plastids, with a special emphasis on the evolution of plankton algae, such as diatoms or dinoflagellates, who acquired their plastids from other photosynthetic eukaryotes. [ABSTRACT FROM AUTHOR]

BACKGROUND: By expressing double‐stranded RNA (dsRNA) in potato plastids targeting the β‐Actin (ACT) gene of the Colorado potato beetle (CPB), transplastomic plants can trigger the beetle's RNA interference response to kill the CPB larvae. High expression of dsACT driven by rrn16 promoter (Prrn) in the leaf chloroplasts of transplastomic plants confers strong resistance to CPB. However, there are still residual amounts of dsRNA in the tubers, which are unnecessary for CPB control and may raise a potential food exposure issue. RESULTS: In order to reduce dsRNA accumulation in the tubers while maintaining stable resistance to CPB, we selected two promoters (PrbcL and PpsbD) from potato plastid‐encoded rbcL and psbD genes and compared their activities with Prrn promoter for dsRNA synthesis in the leaf chloroplasts and tuber amyloplasts. We found that the dsACT accumulation levels in leaves of transplastomic plants St‐PrbcL‐ACT and St‐PpsbD‐ACT were significantly reduced when compared to St‐Prrn‐ACT, but they still maintained high resistance to CPB. By contrast, a few amounts of dsACT were still accumulated in the tubers of St‐PrbcL‐ACT, whereas no dsACT accumulation in tubers was detectable in St‐PpsbD‐ACT. CONCLUSION: We identified PpsbD as a useful promoter to reduce dsRNA accumulation in potato tubers while maintaining the high resistance of the potato leaves to CPB. © 2023 Society of Chemical Industry. [ABSTRACT FROM AUTHOR]

Chloroplasts and photosynthesis are the physiologically fateful arenas of salinity stress. Morphological and anatomical alterations in the leaf tissue, ultrastructural changes in the chloroplast, compromise in the integrity of the three-layered chloroplast membrane system, and defects in the light and dark reactions during the osmotic, ionic, and oxidative phases of salt stress are conversed in detail to bring the salinity-mediated physiological alterations in the chloroplast on to a single platform. Chloroplasts of salt-tolerant plants have evolved highly regulated salt-responsive pathways. Thylakoid membrane remodeling, ion homeostasis, osmoprotection, upregulation of chloroplast membrane and stromal proteins, chloroplast ROS scavenging, efficient retrograde signalling, and differential gene and metabolite abundance are the key attributes of optimal photosynthesis in tolerant species. This review throws light into the comparative mechanism of chloroplast and photosynthetic response to salinity in sensitive and tolerant plant species.

Dinoflagellates possess many cellular characteristics with unresolved evolutionary histories. These include nuclei with greatly expanded genomes and chromatin packaged using histone-like proteins and dinoflagellate-viral nucleoproteins instead of histones, highly reduced mitochondrial genomes with extensive RNA editing, a mix of photosynthetic and cryptic secondary plastids, and tertiary plastids. Resolving the evolutionary origin of these traits requires understanding their ancestral states and early intermediates. Several early-branching dinoflagellate lineages are good candidates for such reconstruction, however these cells tend to be delicate and environmentally sparse, complicating such analyses. Here, we employ transcriptome sequencing from manually isolated and microscopically documented cells to resolve the placement of two cells of one such genus, Abedinium, collected by remotely operated vehicle in deep waters off the coast of Monterey Bay, CA. One cell corresponds to the only described species, Abedinium dasypus, whereas the second cell is distinct and formally described as Abedinium folium, sp. nov. Abedinium has classically been assigned to the early-branching dinoflagellate subgroup Noctilucales, which is weakly supported by phylogenetic analyses of small subunit ribosomal RNA, the single characterized gene from any member of the order. However, an analysis based on 221 proteins from the transcriptome places Abedinium as a distinct lineage, separate from and basal to Noctilucales and the rest of the core dinoflagellates. The transcriptome also contains evidence of a cryptic plastid functioning in the biosynthesis of isoprenoids, iron-sulfur clusters, and heme, a mitochondrial genome with all three expected protein-coding genes (cob, cox1, and cox3), and the presence of some but not all dinoflagellate-specific chromatin packaging proteins.

Tubular extensions emerging from plastids, termed stromules, have received renewed attention due to advancements in imaging techniques. Stromules are widespread in plant and algal species; however, their role in organelle communication and physiology is yet to be elucidated. Initially, stromules were thought to facilitate interplastid communication; however, this proposition is still debated. Stromules with diameters of 0.3-0.8 µm enable protein movement via diffusion and Adenosine Triphosphate (ATP)-dependent transport. Stromule formation is more evident in non-photosynthetic plastids and is induced by various biotic and abiotic stresses, suggesting the involvement of stress-triggered signal transduction via phytohormones and redox changes. Recent studies have emphasized the significance of stromules in plant immunity, especially in response to viral and bacterial effectors, where they serve as conduits for the transport of retrograde signaling molecules from the plastids to the nucleus. Peroxules and matrixules, extending from peroxisomes and mitochondria, respectively, are parallel tubular extensions that were originally found in plant cells, while similar structures also exist in mammalian cells. The response of these extensions to stress may contribute to the management of Reactive Oxygen Species (ROS) and organelle proliferation. This short review discusses the potential roles of the organelle extensions in retrograde signaling pathways.

Iron–sulfur (Fe–S) clusters are ancient protein cofactors ubiquitously exist in organisms. They are involved in many important life processes. Plastids are semi-autonomous organelles with a double membrane and it is believed to originate from a cyanobacterial endosymbiont. By learning form the research in cyanobacteria, a Fe–S cluster biosynthesis and delivery pathway has been proposed and partly demonstrated in plastids, including iron uptake, sulfur mobilization, Fe–S cluster assembly and delivery. Fe–S clusters are essential for the downstream Fe–S proteins to perform their normal biological functions. Because of the importance of Fe–S proteins in plastid, researchers have made a lot of research progress on this pathway in recent years. This review summarizes the detail research progress made in recent years. In addition, the scientific problems remained in this pathway are also discussed. [ABSTRACT FROM AUTHOR]

The interaction between the nuclear and chloroplast genomes in plants is crucial for preserving essential cellular functions in the face of varying rates of mutation, levels of selection, and modes of transmission. Despite this, identifying nuclear genes that coevolve with chloroplast genomes at a genome-wide level has remained a challenge. In this study, we conducted an evolutionary rate covariation analysis to identify candidate nuclear genes coevolving with chloroplast genomes in Juglandaceae. Our analysis was based on 4,894 orthologous nuclear genes and 76 genes across seven chloroplast partitions in nine Juglandaceae species. Our results indicated that 1,369 (27.97%) of the nuclear genes demonstrated signatures of coevolution, with the Ycf1/2 partition yielding the largest number of hits (765) and the ClpP1 partition yielding the fewest (13). These hits were found to be significantly enriched in biological processes related to leaf development, photoperiodism, and response to abiotic stress. Among the seven partitions, AccD, ClpP1, MatK, and RNA polymerase partitions and their respective hits exhibited a narrow range, characterized by d N /d S values below 1. In contrast, the Ribosomal, Photosynthesis, Ycf1/2 partitions and their corresponding hits, displayed a broader range of d N /d S values, with certain values exceeding 1. Our findings highlight the differences in the number of candidate nuclear genes coevolving with the seven chloroplast partitions in Juglandaceae species and the correlation between the evolution rates of these genes and their corresponding chloroplast partitions. [ABSTRACT FROM AUTHOR]

Tubular extensions emerging from plastids, termed stromules, have received renewed attention due to advancements in imaging techniques. Stromules are widespread in plant and algal species; however, their role in organelle communication and physiology is yet to be elucidated. Initially, stromules were thought to facilitate interplastid communication; however, this proposition is still debated. Stromules with diameters of 0.3-0.8 µm enable protein movement via diffusion and Adenosine Triphosphate (ATP)-dependent transport. Stromule formation is more evident in nonphotosynthetic plastids and is induced by various biotic and abiotic stresses, suggesting the involvement of stress-triggered signal transduction via phytohormones and redox changes. Recent studies have emphasized the significance of stromules in plant immunity, especially in response to viral and bacterial effectors, where they serve as conduits for the transport of retrograde signaling molecules from the plastids to the nucleus. Peroxules and matrixules, extending from peroxisomes and mitochondria, respectively, are parallel tubular extensions that were originally found in plant cells, while similar structures also exist in mammalian cells. The response of these extensions to stress may contribute to the management of Reactive Oxygen Species (ROS) and organelle proliferation. This short review discusses the potential roles of the organelle extensions in retrograde signaling pathways. [ABSTRACT FROM AUTHOR]

Cannabis glandular trichomes produce and store an abundance of lipidic specialised metabolites (e.g. cannabinoids and terpenes) that are consumed by humans for medicinal and recreational purposes. Due to a lack of genetic resources and inherent autofluorescence of cannabis glandular trichomes, our knowledge of cannabinoid trafficking and secretion is limited to transmission electron microscopy (TEM). Advances in cryofixation methods has resulted in ultrastructural observations closer to the 'natural state' of the living cell, and recent reports of cryofixed cannabis trichome ultrastructure challenge the long‐standing model of cannabinoid trafficking proposed by ultrastructural reports using chemically fixed samples. Here, we compare the ultrastructural morphology of cannabis glandular trichomes preserved using conventional chemical fixation and ultrarapid cryofixation. We show that chemical fixation results in amorphous metabolite inclusions surrounding the organelles of glandular trichomes that were not present in cryofixed samples. Vacuolar morphology in cryofixed samples exhibited homogenous electron density, while chemically fixed samples contained a flocculent electron dense periphery and electron lucent lumen. In contrast to the apparent advantages of cryopreservation, fine details of cell wall fibre orientation could be observed in chemically fixed glandular trichomes that were not seen in cryofixed samples. Our data suggest that chemical fixation results in intracellular artefacts that impact the interpretation of lipid production and trafficking, while enabling greater detail of extracellular polysaccharide organisation. LAY DESCRIPTION: For millennia humans have grown and consumed cannabis flowers for use in traditional medicine and intoxication, due to the plant's active ingredients called cannabinoids. The most well‐known cannabinoid is tetrahydrocannabidiol (THC). Despite this long history, and recent relaxation of cannabis uses for medicinal and recreational consumption in many parts of the world, little is known about how the plant makes the cannabinoids in its cells. Small mushroom‐shaped structures on the flower surface, called glandular trichomes, produce and store the cannabinoids. To image inside the cells of THC‐rich cannabis flowers, it is necessary to use electron microscopy, and this has revealed complex internal membrane networks and specialised cell structures in these tiny biofactories. In this study, we explored different ways to preserve the cannabis glandular trichomes for electron microscopy, using either cryogenic preservation or chemical crosslinking, in order to determine the benefits and drawbacks of each method. The chemical crosslinking method produced dramatic re‐arrangement of cell contents that were not seen in the cryofixation, where the near‐instantaneous immobilisation of the cell structure led to exceptional preservation of the THC‐producing cells. Despite the benefits of using cryofixation for preserving the complex cell structures, it was not as useful for seeing details of the cell walls surrounding the glandular trichome. Together, our data show stark differences in cannabis glandular trichome cell structure that result from cryofixation and chemical fixation. This is important because the chemical fixation protocol was historically used and informed previous models of cannabinoid production and trafficking. Cryofixation produces exciting new models of THC production in cannabis, due to its high‐fidelity preservation of cell structures. In contrast, studies of cell walls may still benefit from the traditional chemical cross‐linking technique of sample preservation for electron microscopy. https://www.cell.com/current‐biology/fulltext/S0960‐9822(22)01115‐0 [ABSTRACT FROM AUTHOR]

Plastid genomes (plastomes) of angiosperms are well known for their relative stability in size, structure, and gene content. However, little is known about their heredity and variations in wide crossing. To such an end, the plastomes of five representative rice backcross inbred lines (BILs) developed from crosses of O. glaberrima/O. sativa were analyzed. We found that the size of all plastomes was about 134,580 bp, with a quadripartite structure that included a pair of inverted repeat (IR) regions, a small single-copy (SSC) region and a large single-copy (LSC) region. They contained 76 protein genes, 4 rRNA genes, and 30 tRNA genes. Although their size, structure, and gene content were stable, repeat-mediated recombination, gene expression, and RNA editing were extensively changed between the maternal line and the BILs. These novel discoveries demonstrate that wide crossing causes not only nuclear genomic recombination, but also plastome variation in plants, and that the plastome plays a critical role in coordinating the nuclear–cytoplasmic interaction. [ABSTRACT FROM AUTHOR]

The methylerythritol phosphate (MEP) pathway is responsible for producing isoprenoids, metabolites with essential functions in the bacterial kingdom and plastid-bearing organisms including plants and Apicomplexa. Additionally, the MEP-pathway intermediate methylerythritol cyclodiphosphate (MEcPP) serves as a plastid-to-nucleus retrograde signal. A suppressor screen of the high MEcPP accumulating mutant plant (ceh1) led to the isolation of 3 revertants (designated Rceh1-3) resulting from independent intragenic substitutions of conserved amino acids in the penultimate MEP-pathway enzyme, hydroxymethylbutenyl diphosphate synthase (HDS). The revertants accumulate varying MEcPP levels, lower than that of ceh1, and exhibit partial or full recovery of MEcPP-mediated phenotypes, including stunted growth and induced expression of stress response genes and the corresponding metabolites. Structural modeling of HDS and ligand docking spatially position the substituted residues at the MEcPP binding pocket and cofactor binding domain of the enzyme. Complementation assays confirm the role of these residues in suppressing the ceh1 mutant phenotypes, albeit to different degrees. In vitro enzyme assays of wild type and HDS variants exhibit differential activities and reveal an unanticipated mismatch between enzyme kinetics and the in vivo MEcPP levels in the corresponding Rceh lines. Additional analyses attribute the mismatch, in part, to the abundance of the first and rate-limiting MEP-pathway enzyme, DXS, and further suggest MEcPP as a rheostat for abundance of the upstream enzyme instrumental in fine-tuning of the pathway flux. Collectively, this study identifies critical residues of a key MEP-pathway enzyme, HDS, valuable for synthetic engineering of isoprenoids, and as potential targets for rational design of antiinfective drugs.

Abstract Background Chlorophyll and carotenoids, the most widely distributed lipophilic pigments in plants, contribute to fruit coloration during development and ripening. These pigments are assembled with pigment-protein complexes localized at plastid membrane. Pigment-protein complexes are essential for multiple cellular processes, however, their identity and composition in fruit have yet to be characterized. Results By using BN-PAGE technique in combination with microscopy, we studied pigment-protein complexes and plastid transformation in the purified plastids from the exocarp of citrus fruit. The discontinuous sucrose gradient centrifugation was used to isolate total plastids from kumquat fruit, and the purity of isolated plastids was assessed by microscopy observation and western blot analysis. The isolated plastids at different coloring stages were subjected to pigment autofluorescence observation, western blot, two-dimensional electrophoresis analysis and BN-PAGE assessment. Our results demonstrated that (i) chloroplasts differentiate into chromoplasts during fruit coloring, and this differentiation is accompanied with a decrease in the chlorophyll/carotenoid ratio; (ii) BN-PAGE analysis reveals the profiles of macromolecular protein complexes among different types of plastids in citrus fruit; and (iii) the degradation rate of chlorophyll-protein complexes varies during the transition from chloroplasts to chromoplasts, with the stability generally following the order of LHCII > PS II core > LHC I > PS I core. Conclusions Our optimized methods for both plastid separation and BN-PAGE assessment provide an opportunity for developing a better understanding of pigment-protein complexes and plastid transitions in plant fruit. These attempts also have the potential for expanding our knowledge on the sub-cellular level synchronism of protein changes and pigment metabolism during the transition from chloroplasts to chromoplasts.

Terpenes are the largest and most diverse class of plant specialized metabolites. Sesterterpenes (C25), which are derived from the plastid methylerythritol phosphate pathway, were recently characterized in plants. In Arabidopsis thaliana, four genes encoding geranylfarnesyl diphosphate synthase (GFPPS) (AtGFPPS1 to 4) are responsible for the production of GFPP, which is the common precursor for sesterterpene biosynthesis. However, the interplay between sesterterpenes and other known terpenes remain elusive. Here, we first provide genetic evidence to demonstrate that GFPPSs are responsible for sesterterpene production in Arabidopsis. Blockage of the sesterterpene pathway at the GFPPS step increased the production of geranylgeranyl diphosphate (GGPP)‐derived terpenes. Interestingly, co‐expression of sesterTPSs in GFPPS‐OE (overexpression) plants rescued the phenotypic changes of GFPPS‐OE plants by restoring the endogenous GGPP. We further demonstrated that, in addition to precursor (DMAPP/IPP) competition by GFPPS and GGPP synthase (GGPPS) in plastids, GFPPS directly decreased the activity of GGPPS through protein‐protein interaction, ultimately leading to GGPP deficiency in planta. Our study provides a new regulatory mechanism of the plastidial terpenoid network in plant cells. [ABSTRACT FROM AUTHOR]

Main conclusion: Most species in Cuscuta subgenus Grammica retain many photosynthesis-related plastid genes, generally under purifying selection. A group of holoparasitic species in section Subulatae may have lost their plastid genomes entirely. The c. 153 species of plants belonging to Cuscuta subgenus Grammica are all obligate stem parasites. However, some have completely lost the ability to conduct photosynthesis while others retain photosynthetic machinery and genes. The plastid genome that primarily encodes key photosynthesis genes functions as a bellwether for how reliant plants are on primary production. This research assembles and analyses 17 plastomes across Cuscuta subgenus Grammica with the aim of characterizing the state of the plastome in each of its sections. By comparing the structure and content of plastid genomes across the subgenus, as well as by quantifying the selection acting upon each gene, we reconstructed the patterns of plastome change within the phylogenetic context for this group. We found that species in 13 of the 15 sections that comprise Grammica retain the bulk of plastid photosynthesis genes and are thus hemiparasitic. The complete loss of photosynthesis can be traced to two clades: the entire section Subulatae and a complex of three species within section Ceratophorae. We were unable to recover any significant plastome sequences from section Subulatae, suggesting that plastomes in these species are either drastically reduced or lost entirely. [ABSTRACT FROM AUTHOR]

Retrograde signaling modulates the expression of nuclear genome-encoded organelle proteins to adjust organelle function in response to environmental cues. MULTIPLE ORGANELLAR RNA EDITING FACTOR 2 (MORF2) was initially recognized as a plastidial RNA-editing factor but recently shown to interact with GUN1. Given the central role of GUN1 in chloroplast retrograde signaling and the unviable phenotype of morf2 mutants that is inconsistent with many viable mutants involved in RNA editing, we hypothesized that MORF2 has functions either dosage dependent or beyond RNA editing. Using an inducible Clustered Interspaced Short Palindromic Repeat interference (iCRISPRi) approach, we were able to reduce the MORF2 transcripts in a controlled manner. In addition to MORF2-dosage dependent RNA-editing errors, we discovered that reducing MORF2 by iCRISPRi stimulated the expression of stress responsive genes, triggered plastidial retrograde signaling, repressed ethylene signaling and skotomorphogenesis, and increased accumulation of hydrogen peroxide. These findings along with previous discoveries suggest that MORF2 is an effective regulator involved in plastidial metabolic pathways whose reduction can readily activate multiple retrograde signaling molecules possibly involving reactive oxygen species to adjust plant growth. In addition, our newly developed iCRISPRi approach provided a novel genetic tool for quantitative reverse genetics studies on hub genes in plants. [ABSTRACT FROM AUTHOR]

Initial morphometric investigations into the differences between Leucobryum glaucum (Hedw.) Ångstr. and L. juniperoideum (Brid.) Müll.Hal. in Britain showed that some populations could not be clearly assigned to either species. An intensive morphological and molecular study was therefore undertaken to understand the diagnostic characters of the European species of the genus, including L. albidum (Brid. ex P.Beauv.) Lindb., whose reported presence in Europe was surrounded by uncertainty. There has been little agreement among previous authors on the relative taxonomic importance of the different morphological characters, or on their metrics. Molecular sequencing using nuclear and chloroplast markers, in conjunction with previously accessioned data, allowed judgement of the taxonomic importance of various methods for morphological discrimination. These methods included both existing published techniques and newly discovered diagnostic characters. This integrated process was cyclical, the morphological analysis being directed and confirmed by the sequencing of selected specimens. Data are presented demonstrating the correlation between molecular identification and certain morphological characters in the three European species of Leucobryum. Leucobryum albidum is confirmed as a locally frequent member of the British flora and it is also shown to be present in Ireland and western part of continental Europe. Leucocyst pore morphology is described and its value as a diagnostic character is discussed. An identification key is provided for the three species. Plants from two sites are shown to have a discordance between nuclear and plastid molecular markers and we consider the possibility that horizontal gene transfer could explain the observed patterns. [ABSTRACT FROM AUTHOR]

Retrograde signaling modulates the expression of nuclear genome-encoded organelle proteins to adjust organelle function in response to environmental cues. MULTIPLE ORGANELLAR RNA EDITING FACTOR 2 (MORF2) was initially recognized as a plastidial RNA-editing factor but recently shown to interact with GUN1. Given the central role of GUN1 in chloroplast retrograde signaling and the unviable phenotype of morf2 mutants that is inconsistent with many viable mutants involved in RNA editing, we hypothesized that MORF2 has functions either dosage dependent or beyond RNA editing. Using an inducible Clustered Interspaced Short Palindromic Repeat interference (iCRISPRi) approach, we were able to reduce the MORF2 transcripts in a controlled manner. In addition to MORF2-dosage dependent RNA-editing errors, we discovered that reducing MORF2 by iCRISPRi stimulated the expression of stress responsive genes, triggered plastidial retrograde signaling, repressed ethylene signaling and skotomorphogenesis, and increased accumulation of hydrogen peroxide. These findings along with previous discoveries suggest that MORF2 is an effective regulator involved in plastidial metabolic pathways whose reduction can readily activate multiple retrograde signaling molecules possibly involving reactive oxygen species to adjust plant growth. In addition, our newly developed iCRISPRi approach provided a novel genetic tool for quantitative reverse genetics studies on hub genes in plants.

The spectacular diversity of plastids in non-green organs such as flowers, fruits, roots, tubers, and senescing leaves represents a Universe of metabolic processes in higher plants that remain to be completely characterized. The endosymbiosis of the plastid and the subsequent export of the ancestral cyanobacterial genome to the nuclear genome, and adaptation of the plants to all types of environments has resulted in the emergence of diverse and a highly orchestrated metabolism across the plant kingdom that is entirely reliant on a complex protein import and translocation system. The TOC and TIC translocons, critical for importing nuclear-encoded proteins into the plastid stroma, remain poorly resolved, especially in the case of TIC. From the stroma, three core pathways (cpTat, cpSec, and cpSRP) may localize imported proteins to the thylakoid. Non-canonical routes only utilizing TOC also exist for the insertion of many inner and outer membrane proteins, or in the case of some modified proteins, a vesicular import route. Understanding this complex protein import system is further compounded by the highly heterogeneous nature of transit peptides, and the varying transit peptide specificity of plastids depending on species and the developmental and trophic stage of the plant organs. Computational tools provide an increasingly sophisticated means of predicting protein import into highly diverse non-green plastids across higher plants, which need to be validated using proteomics and metabolic approaches. The myriad plastid functions enable higher plants to interact and respond to all kinds of environments. Unraveling the diversity of non-green plastid functions across the higher plants has the potential to provide knowledge that will help in developing climate resilient crops.

Autophagy is a highly conserved system that delivers cytoplasmic components to lysosomes/vacuoles. Plastids are also degraded through autophagy for nutrient recycling and quality control; however, the involvement of autophagic degradation of plastids in plant cellular differentiation remains unclear. Here, we investigated whether spermiogenesis, the differentiation of spermatids into spermatozoids, in the liverwort Marchantia polymorpha involves autophagic degradation of plastids. Spermatozoids of M. polymorpha possess one cylindrical plastid at the posterior end of the cell body. By fluorescently labeling and visualizing plastids, we detected dynamic morphological changes during spermiogenesis. We found that a portion of the plastid was degraded in the vacuole in an autophagy-dependent manner during spermiogenesis, and impaired autophagy resulted in defective morphological transformation and starch accumulation in the plastid. Furthermore, we found that autophagy was dispensable for the reduction in plastid number and plastid DNA elimination. These results demonstrate a critical but selective role of autophagy in plastid reorganization during spermiogenesis in M. polymorpha. [ABSTRACT FROM AUTHOR]

Chloroplasts and photosynthesis are the physiologically fateful arenas of salinity stress. Morphological and anatomical alterations in the leaf tissue, ultrastructural changes in the chloroplast, compromise in the integrity of the three-layered chloroplast membrane system, and defects in the light and dark reactions during the osmotic, ionic, and oxidative phases of salt stress are conversed in detail to bring the salinity-mediated physiological alterations in the chloroplast on to a single platform. Chloroplasts of salt-tolerant plants have evolved highly regulated salt-responsive pathways. Thylakoid membrane remodeling, ion homeostasis, osmoprotection, upregulation of chloroplast membrane and stromal proteins, chloroplast ROS scavenging, efficient retrograde signalling, and differential gene and metabolite abundance are the key attributes of optimal photosynthesis in tolerant species. This review throws light into the comparative mechanism of chloroplast and photosynthetic response to salinity in sensitive and tolerant plant species. [ABSTRACT FROM AUTHOR]

Chloroplasts are the defining plant organelles with responsibility for photosynthesis and other vital functions. To deliver these functions, they possess a complex proteome comprising thousands of largely nucleus-encoded proteins. Composition of the proteome is controlled by diverse processes affecting protein translocation and degradation—our focus here. Most chloroplast proteins are imported from the cytosol via multiprotein translocons in the outer and inner envelope membranes (the TOC and TIC complexes, respectively), or via one of several noncanonical pathways, and then sorted by different systems to organellar subcompartments. Chloroplast proteolysis is equally complex, involving the concerted action of internal proteases of prokaryotic origin and the nucleocytosolic ubiquitin–proteasome system (UPS). The UPS degrades unimported proteins in the cytosol and chloroplast-resident proteins via chloroplast-associated protein degradation (CHLORAD). The latter targets the TOC apparatus to regulate protein import, as well as numerous internal proteins directly, to reconfigure chloroplast functions in response to developmental and environmental signals. [ABSTRACT FROM AUTHOR]

Enhanced carotenoid accumulation in plants is crucial for the nutritional and health demands of the human body since these beneficial substances are acquired through dietary intake. Plastids are the major organelles to accumulate carotenoids in plants and it is reported that manipulation of a single plastid phosphate transporter gene enhances carotenoid accumulation. Amongst all phosphate transport proteins including phosphate transporters (PHTs), plastidial phosphate translocators (pPTs), PHOSPHATE1 (PHO1), vacuolar phosphate efflux transporter (VPE), and Sulfate transporter [SULTR]- like phosphorus distribution transporter (SPDT) in plants, plastidic PHTs (PHT2 & PHT4) are found as the only clade that is plastid located, and manipulation of which affects carotenoid accumulation. Manipulation of a single chromoplast PHT (PHT4;2) enhances carotenoid accumulation, whereas manipulation of a single chloroplast PHT has no impact on carotenoid accumulation. The underlying mechanism is mainly attributed to their different effects on plastid orthophosphate (Pi) concentration. PHT4;2 is the only chromoplast Pi efflux transporter, and manipulating this single chromoplast PHT significantly regulates chromoplast Pi concentration. This variation subsequently modulates the carotenoid accumulation by affecting the supply of glyceraldehyde 3-phosphate, a substrate for carotenoid biosynthesis, by modulating the transcript abundances of carotenoid biosynthesis limited enzyme genes, and by regulating chromoplast biogenesis (facilitating carotenoid storage). However, at least five orthophosphate influx PHTs are identified in the chloroplast, and manipulating one of the five does not substantially modulate the chloroplast Pi concentration in a long term due to their functional redundancy. This stable chloroplast Pi concentration upon one chloroplast PHT absence, therefore, is unable to modulate Pi-involved carotenoid accumulation processes and finally does affect carotenoid accumulation in photosynthetic tissues. Despite these advances, several cases including the precise location of plastid PHTs, the phosphate transport direction mediated by these plastid PHTs, the plastid PHTs participating in carotenoid accumulation signal pathway, the potential roles of these plastid PHTs in leaf carotenoid accumulation, and the roles of these plastid PHTs in other secondary metabolites are waiting for further research. The clarification of the above-mentioned cases is beneficial for breeding high-carotenoid accumulation plants (either in photosynthetic or non-photosynthetic edible parts of plants) through the gene engineering of these transporters. [ABSTRACT FROM AUTHOR]

Pollen tubes consume a tremendous amount of energy and are the fastest-growing cells known in plants. Mitochondria are key organelles that supply energy and play important roles in modulating cellular redox homeostasis. Here, we found that endogenous NAD(P)H in Arabidopsis pollen tubes was spatially highly correlated with the distribution of mitochondria, both peaking in the subapex region. A weak association was also observed between the NAD(P)H levels and pollen plastids. Further studies using Class XI myosin mutants confirmed that altered mitochondrial distribution and trafficking concomitantly affected intracellular NAD(P)H zoning in pollen tubes. By targeting the NADPH- and NADH/NAD+-specific biosensors to the pollen tube cytosol of the myo11c1/myo11c2 double mutants, we showed that the growing pollen tubes in the double mutants possessed a lower level of cytosolic NADPH but a higher cytosolic NADH/NAD+ ratio than the WT. We also found that the knockout of Myo11C1 and Myo11C2 led to fragmented mitochondria with reduced motility. Therefore, altered cytosolic NAD(P)H levels may be secondary to changes in mitochondrial mobility, positioning, or morphology. Our results suggest that the spatial distribution and movement of mitochondria and plastids affect NAD(P)H zoning in Arabidopsis growing pollen tubes and that their movements depend on Class XI myosins. • Endogenous NAD(P)H of Arabidopsis pollen tube was spatially correlated with mitochondria and plastids. • Loss of Myo11C1 and Myo11C2 caused altered mitochondrial distribution and abnormal intracellular NAD(P)H zoning. • Mobility of pollen mitochondria and plastids was compromised in myo11c1/c2. • Myo11c1/c2 pollen tubes possessed reduced levels of cytosolic NADPH but higher cytosolic NADH/NAD+ ratios. [ABSTRACT FROM AUTHOR]