Plastids are essential, semi-autonomous organelles in plants that carry out a multitude of functions during development. Plastids existing in different subtypes are derived from proplastids progenitors and interconvert in response to environmental and growth cues. Most efforts focus on the differentiation from proplastid to other forms. However, the studies of proplastid development are insufficient and whether proplastid biogenesis affects plant growth is yet to be determined. Arabidopsis TIC236, a translocon component at the inner membrane of the chloroplast envelope, is critical for importing chloroplast-targeted preproteins and chloroplast division. In this study, we uncovered the fundamental influence of proplastid biogenesis on embryo development by exploring the function of TIC236 during embryogenesis. Widespread and strong expression of TIC236 was observed in leaves and embryos. The null mutant tic236 had an embryo-lethal phenotype, with cell division in the mutant embryos delayed starting at the octant stage and arrested at the globular stage. Transmission electron microscopy revealed enlarged proplastids with an aberrant inner structure at the dermatogen and globular stages that ultimately did not differentiate into chloroplasts. Additionally, the fluorescence signal distribution patterns of tic236 embryos carrying the pDR5rev::3xVENUS-N7, pPIN1::PIN1-GFP, pWOX5::GFP, and pSCR::H2B-YFP reporter systems were altered. Together, we provide genetic evidence supporting proplastid biogenesis plays a vital role in embryo development and TIC236 is identified as an indispensable player, ensuring normal proplastid development.

Alfalfa mosaic virus (AMV) is one of the most widely distributed viruses; it often exhibits combined infection with white clover mosaic virus (WCMV). Even so, little is known about the effects of co-infection with AMV and WCMV on plants. To determine whether there is a synergistic effect of AMV and WCMV co-infection, virus co-infection was studied by electron microscopy, the double-antibody sandwich enzyme-linked immunosorbent assay (DAS-ELISA), and real-time fluorescence quantitative PCR (RT-qPCR) of AMV and WCMV co-infection in Nicotiana benthamiana. Meanwhile, measurements were carried out on the photosynthetic pigments, photosynthetic gas exchange parameters, and chlorophyll fluorescence parameters. The results showed that the most severe disease development was induced by AMV and WCMV co-infection, and the disease grade was scale 7. N. benthamiana leaves induced mottled yellow-green alternating patterns, leaf wrinkling, and chlorosis, and chloroplasts were observed to be on the verge of disintegration. The relative accumulation of AMV CP and WCMV CP was significantly increased by 15.44-fold and 10.04-fold upon co-infection compared to that with AMV and WCMV single infection at 21 dpi. In addition, chlorophyll a, chlorophyll b, total chlorophyll, the net photosynthetic rate, the water use efficiency, the apparent electron transport rate, the PSII maximum photochemical efficiency, the actual photochemical quantum yield, and photochemical quenching were significantly reduced in leaves co-infected with AMV and WCMV compared to AMV- or WCMV-infected leaves and CK. On the contrary, the carotenoid content, transpiration rate, stomatal conductance, intercellular CO2 concentration, minimal fluorescence value, and non-photochemical quenching were significantly increased. These findings suggest that there was a synergistic effect between AMV and WCMV, and AMV and WCMV co-infection severely impacted the normal function of photosynthesis in N. benthamiana.

UNASSIGNED: Allopolyploidy-a hybridization-induced whole-genome duplication event-has been a major driver of plant diversification. The extent to which chromosomes pair with their proper homolog vs. with their homoeolog in allopolyploids varies across taxa, and methods to detect homoeologous gene flow (HGF) are needed to understand how HGF has shaped polyploid lineages.
UNASSIGNED: The ABBA-BABA test represents a classic method for detecting introgression between closely related species, but here we developed a modified use of the ABBA-BABA test to characterize the extent and direction of HGF in allotetraploid Coffea arabica.
UNASSIGNED: We found that HGF is abundant in the C. arabica genome, with both subgenomes serving as donors and recipients of variation. We also found that HGF is highly maternally biased in plastid-targeted-but not mitochondrial-targeted-genes, as would be expected if plastid-nuclear incompatibilities exist between the two parent species.
UNASSIGNED: Together, our analyses provide a simple framework for detecting HGF and new evidence consistent with selection favoring overwriting of paternally derived alleles by maternally derived alleles to ameliorate plastid-nuclear incompatibilities. Natural selection therefore appears to shape the direction and intensity of HGF in allopolyploid coffee, indicating that cytoplasmic inheritance has long-term consequences for polyploid lineages.

Narcissus pseudonarcissus L. is one of the most iconic plants of the European flora. It is a species of great horticultural interest, but also an endangered and protected plant in the wild as a consequence of loss of natural habitats. Complete plastid genome was assembled from next-generation sequencing data obtaining a circular genome of 160,008 bp long assembly. It comprises a pair of inverted repeat regions, a large single-copy region (108,400 bp), and a small single-copy region (16,434 bp). It encodes 131 genes, including 87 protein coding genes, 37 tRNA genes and seven rRNA genes. Phylogeny showed the strict relationship between N. pseudonarcissus and Narcissus poeticus L. The complete plastome will provide a useful genetic resource for future conservation programmes, phylogenetic studies and horticultural applications.

The chloroplast (cp.) genome, also known as plastome, plays crucial roles in plant survival, adaptation, and evolution. The stable genetic structure of cp. genomes provides an ideal system for investigating species evolution. We sequenced three complete cp. genome sequences of Capsicum species and analyzed them using sequences of various Capsicum species retrieved from the NCBI database. The cp. genome of Capsicum species maintains a well-preserved quadripartite structure consisting of two inverted repeats (IRs) flanked by a large single copy (LSC) region and a small single copy (SSC) region. The sizes of cp. genome sequences ranged from 156,583 bp (C. lycianthoides) to 157,390 bp (C.pubescens). A total of 127-132 unique genes, including 83-87 protein-coding, 36-37 tRNA, and eight rRNA genes, were predicted. Comparison of cp. genomes of 10 Capsicum species revealed high sequence similarity in genome-wide organization and gene arrangements. Fragments of trnT-UGU/trnL-UAA, ccsA, ndhD, rps12, and ycf1 were identified as variable regions, and nucleotide variability of LSC and SSC was higher than that of IR. Phylogenetic speciation analysis showed that the major domesticated C. annuum species were the most extensively divergent species and closely related to C. tovarii and C. frutescens. Analysis of divergent times suggested that a substantial range of speciation events started occurring ~ 25.79 million years ago (Mya). Overall, comparative analysis of cp. genomes of Capsicum species not only offers new insights into their genetic variation and phylogenetic relationships, but also lays a foundation for evolutionary history, genetic diversity, conservation, and biological breeding of Capsicum species.

Styrax japonicus is a medicinal and ornamental shrub belonging to the Styracaceae family. To explore the diversity and characteristics of the chloroplast genome of S. japonicus, we conducted sequencing and comparison of the chloroplast genomes of four naturally distributed S. japonicus. The results demonstrated that the four chloroplast genomes (157,914-157,962 bp) exhibited a typical quadripartite structure consisting of a large single copy (LSC) region, a small single copy (SSC) region, and a pair of reverse repeats (IRa and IRb), and the structure was highly conserved. DNA polymorphism analysis revealed that three coding genes (infA, psbK, and rpl33) and five intergene regions (petA-psbJ, trnC-petN, trnD-trnY, trnE-trnT, and trnY-trnE) were identified as mutation hotspots. These genetic fragments have the potential to be utilized as DNA barcodes for future identification purposes. When comparing the boundary genes, a small contraction was observed in the IR region of four S. japonicus. Selection pressure analysis indicated positive selection for ycf1 and ndhD. These findings collectively suggest the adaptive evolution of S. japonicus. The phylogenetic structure revealed conflicting relationships among several S. japonicus, indicating divergent evolutionary paths within this species. Our study concludes by uncovering the genetic traits of the chloroplast genome in the differentiation of S. japonicus variety, offering fresh perspectives on the evolutionary lineage of this species.

Cephaleuros species are well-known as plant pathogens that cause red rust or algae spot diseases in many economically cultivated plants that grow in shady and humid environments. Despite their prevalence, the adaptive evolution of these pathogens remains poorly understood. We sequenced and characterized three Cephaleuros (Cephaleuros lagerheimii, Cephaleuros diffusus, and Cephaleuros virescens) chloroplast genomes, and compared them with seven previously reported chloroplast genomes. The chloroplast sequences of C. lagerheimii, C. diffusus, and C. virescens were 480,613 bp, 383,846 bp, and 472,444 bp in length, respectively. These chloroplast genomes encoded 94 genes, including 27 tRNA genes, 3 rRNA genes, and 64 protein-coding genes. Comparative analysis uncovered that the variation in genome size was principally due to the length of intergenic spacer sequences, followed by introns. Furthermore, several highly variable regions (trnY-GTA, trnL-TAG, petA, psbT, trnD-GTC, trnL-TAA, ccsA, petG, psaA, psaB, rps11, rps2, and rps14) were identified. Codon bias analysis revealed that the codon usage pattern of Cephaleuros is predominantly shaped by natural selection. Additionally, six chloroplast protein-coding genes (atpF, chlN, psaA, psaB, psbA, and rbcL) were determined to be under positive selection, suggesting they may play a vital roles in the adaptation of Cephaleuros to low-light intensity habitats.

Climate change poses a significant threat to global agriculture, necessitating innovative solutions. Plant synthetic biology, particularly chloroplast engineering, holds promise as a viable approach to this challenge. Chloroplasts present a variety of advantageous traits for genetic engineering, but the development of genetic tools and genetic part characterization in these organelles is hindered by the lengthy time scales required to generate transplastomic organisms. To address these challenges, we have established a versatile protocol for generating highly active chloroplast-based cell-free gene expression (CFE) systems derived from a diverse range of plant species, including wheat (monocot), spinach, and poplar trees (dicots). We show that these systems work with conventionally used T7 RNA polymerase as well as the endogenous chloroplast polymerases, allowing for detailed characterization and prototyping of regulatory sequences at both transcription and translation levels. To demonstrate the platform for characterization of promoters and 5\' and 3\' untranslated regions (UTRs) in higher plant chloroplast gene expression, we analyze a collection of 23 5\'UTRs, 10 3\'UTRs, and 6 chloroplast promoters, assessed their expression in spinach and wheat extracts, and found consistency in expression patterns, suggesting cross-species compatibility. Looking forward, our chloroplast CFE systems open new avenues for plant synthetic biology, offering prototyping tools for both understanding gene expression and developing engineered plants, which could help meet the demands of a changing global climate.

Drought stress is one of the most critical threats to crop productivity and global food security. This review addresses the multiple effects of drought on the process of photosynthesis in major food crops. Affecting both light-dependent and light-independent reactions, drought leads to severe damage to photosystems and blocks the electron transport chain. Plants face a CO2 shortage provoked by stomatal closure, which triggers photorespiration; not only does it reduce carbon fixation efficiency, but it also causes lower overall photosynthetic output. Drought-induced oxidative stress generates reactive oxygen species (ROS) that damage cellular structures, including chloroplasts, further impairing photosynthetic productivity. Plants have evolved a variety of adaptive strategies to alleviate these effects. Non-photochemical quenching (NPQ) mechanisms help dissipate excess light energy as heat, protecting the photosynthetic apparatus under drought conditions. Alternative electron pathways, such as cyclical electron transmission and chloroplast respiration, maintain energy balance and prevent over-reduction of the electron transport chain. Hormones, especially abscisic acid (ABA), ethylene, and cytokinin, modulate stomatal conductance, chlorophyll content, and osmotic adjustment, further increasing the tolerance to drought. Structural adjustments, such as leaf reordering and altered root architecture, also strengthen tolerance. Understanding these complex interactions and adaptive strategies is essential for developing drought-resistant crop varieties and ensuring agricultural sustainability.

Pentatricopeptide repeat (PPR) proteins constitute one of the largest protein families in land plants, with over 300 members in various species. Nearly all PPR proteins are nuclear-encoded and targeted to the chloroplast and mitochondria, modulating organellar gene expression by participating in RNA metabolism, including mRNA stability, RNA editing, RNA splicing, and translation initiation. Organelle RNA metabolism significantly influences chloroplast and mitochondria functions, impacting plant photosynthesis, respiration, and environmental responses. Over the past decades, PPR proteins have emerged as a research focus in molecular biology due to their diverse roles throughout plant life. This review summarizes recent progress in understanding the roles and molecular mechanisms of PPR proteins, emphasizing their functions in fertility, abiotic and biotic stress, grain quality, and chloroplast development in rice. Furthermore, we discuss prospects for PPR family research in rice, aiming to provide a theoretical foundation for future investigations and applications.