The pentatricopeptide repeat protein DG1 promotes the transition to bilateral symmetry during Arabidopsis embryogenesis through GUN1-mediated plastid signals.

During Arabidopsis embryogenesis, the transition of the embryo's symmetry from radial to bilateral between the globular and heart stage is a crucial event, involving the formation of cotyledon primordia and concurrently the establishment of a shoot apical meristem (SAM). However, a coherent framework of how this transition is achieved remains to be elucidated. In this study, we investigated the function of DELAYED GREENING 1 (DG1) in Arabidopsis embryogenesis using a newly identified dg1-3 mutant. The absence of chloroplast-localized DG1 in the mutants led to embryos being arrested at the globular or heart stage, accompanied by an expansion of WUSCHEL (WUS) and SHOOT MERISTEMLESS (STM) expression. This finding pinpoints the essential role of DG1 in regulating the transition to bilateral symmetry. Furthermore, we showed that this regulation of DG1 may not depend on its role in plastid RNA editing. Nevertheless, we demonstrated that the DG1 function in establishing bilateral symmetry is genetically mediated by GENOMES UNCOUPLED 1 (GUN1), which represses the transition process in dg1-3 embryos. Collectively, our results reveal that DG1 functionally antagonizes GUN1 to promote the transition of the Arabidopsis embryo's symmetry from radial to bilateral and highlight the role of plastid signals in regulating pattern formation during plant embryogenesis.

Confusing boundaries of the Labrador Tea species: dispersal history explains the lack of clear species structure.

BACKGROUND AND AIMS: The Labrador Teas (genus Rhododendron, subsection Ledum) are a complex of species widely distributed in the Northern Hemisphere. They occupy cold-resistant plant communities from highlands to forest understory and wetland habitats almost circumboreally and they are especially abundant in Northeast Asia (NE Asia) and northern North America (NN Am), still there are no clear species boundaries in this group. The genetic structure of species of the subsect. Ledum from Eurasia and North America as well as the dispersal history of the group require clarification. METHODS: Phylogeny and biogeography of the subsect. Ledum of the genus Rhododendron were assessed using phylogenetic trees constructed based on the analysis of variation in chloroplast petB-petD, trnV-ndhC, trnH-psbA, K2R-K707, atpB oligo2 - rbcL oligo5 and nuclear (ITS1) markers of four Eurasian and one American species (65 populations, 408 individuals). The data were evaluated with Maximum Parsimony and Bayesian analysis. Molecular dating and ancestral areas reconstruction were obtained. KEY RESULTS: Dense sampling revealed widespread presence of shared haplotypes and ribotypes among Ledum populations and species. Two American, three Eurasian and one mixed lineage diversified during the Neogene climate cooling and then rapidly dispersed during the Pleistocene. The ability to accumulate high genetic diversity and to preserve it across distribution ranges and generations prevented Ledum from lineage sorting. As a result, a species complex with a reserve of genetic variability appeared. CONCLUSIONS: Although no clear phylogenetic inference can be obtained at present, the plastid genealogy is consisted with the nuclear genealogy and demonstrates the processes involved in speciation in the Ledum species complex.

Species of the Sections Hedysarum and Multicaulia of the Genus Hedysarum (Fabaceae): Taxonomy, Distribution, Chromosomes, Genomes, and Phylogeny.

The genus Hedysarum L. (Fabaceae) includes about 200 species of annual and perennial herbs distributed in Asia, Europe, North Africa, and North America. Many species of this genus are valuable medicinal, melliferous, and forage resources. In this review, we consider the taxonomic history of the genus Hedysarum, the chromosomal organization of the species from the sections Hedysarum and Multicaulia, as well as phylogenetic relationships between these sections. According to morphological, genetic, and phylogenetic data, the genus Hedysarum is divided into three main sections: Hedysarum (= syn. Gamotion), Multicaulia, and Stracheya. In species of this genus, two basic chromosome numbers, x = 7 (section Hedysarum) and x = 8 (sections Multicaulia and Stracheya), were determined. The systematic positions of some species within the sections are still uncertain due to their morphological similarities. The patterns of distribution of molecular chromosomal markers (45S rDNA, 5S rDNA, and different satellite DNAs) in karyotypes of various Hedysarum species made it possible to determine their ploidy status and also specify genomic relationships within the sections Hedysarum and Multicaulia. Recent molecular phylogenetic studies clarified significantly the taxonomy and evolutionary development of the genus Hedysarum.

Morphology, behavior, and phylogenomics of Oxytoxum lohmannii, Dinoflagellata.

Dinoflagellates are an abundant and diverse group of protists representing a wealth of unique biology and ecology. While many dinoflagellates are photosynthetic or mixotrophic, many taxa are heterotrophs, often with complex feeding strategies. Compared to their photosynthetic counterparts, heterotrophic dinoflagellates remain understudied, as they are difficult to culture. One exception, a long-cultured isolate originally classified as Amphidinium but recently reclassified as Oxytoxum, has been the subject of a number of feeding, growth, and chemosensory studies. This lineage was recently determined to be closely related to Prorocentrum using phylogenetics of ribosomal RNA gene sequences, but the exact nature of this relationship remains unresolved. Using transcriptomes sequenced from culture and three single cells from the environment, we produce a robust phylogeny of 242 genes, revealing Oxytoxum is likely sister to the Prorocentrum clade, rather than nested within it. Molecular investigations uncover evidence of a reduced, nonphotosynthetic plastid and proteorhodopsin, a photoactive proton pump acquired horizontally from bacteria. We describe the ultrastructure of O. lohmannii, including densely packed trichocysts, and a new type of mucocyst. We observe that O. lohmannii feeds preferentially on cryptophytes using myzocytosis, but can also feed on various phytoflagellates using conventional phagocytosis. O. lohmannii is amenable to culture, providing an opportunity to better study heterotrophic dinoflagellate biology and feeding ecology.

Molecular and morphological evidence supports the resurrection of Chrysospleniumguangxiense H.G.Ye & Gui C.Zhang (Saxifragaceae).

Chrysospleniumguangxiense H.G.Ye & Gui C.Zhang was first described as a new species in 1994 but later synonymized in the Flora of China treatment with C.glossophyllum H.Hara. Plastid genomes and nrDNA sequences were used to infer the phylogenetic relationships of selected taxa in Chrysosplenium. Our phylogenetic analyses revealed that C.guangxiense belongs to sect. Alternifolia, is closely related to Chrysospleniumhydrocotylifolium H.Lév. & Vaniot but distant from C.glossophyllum. Morphologically, C.guangxiense could be easily distinguished from C.glossophyllum by having robust rhizomes, basal leaves with a long cuneate base and fewer teeth in the margin, curled sepal margins, and red, larger seeds. It could also be easily distinguished from C.hydrocotylifolium by possessing long elliptic leaves and a long cuneate leaf base. Along with the phylogenetic studies, the complete plastid genome of C.guangxiense was also reported. The plastid genome was 154,004 bp in length and comprised two inverted repeats (IRs) of 28,120 bp, separated by a large single-copy of 80,646 bp and a small single-copy of 17,118 bp. A total of 111 functional genes were discovered, comprising 78 protein-coding genes, 29 tRNA genes, and four rRNA genes. Based on assessment of morphological and molecular data Chrysospleniumguangxiense H.G.Ye & Gui C.Zhang is resurrected from C.glossophyllum H.Hara at species level. A global conservation assessment classifies C.guangxiense as Vulnerable (VU).

Chloroplast DNA methylation in the kelp Saccharina latissima is determined by origin and possibly influenced by cultivation.

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.

Evolutionary unraveling: new insights into the Persicaria amphibia complex.

The Persicaria amphibia complex exhibits significant morphological variation depending on its habitat, existing in either aquatic or terrestrial forms. Traditionally, four distinct elements have been recognized based on morphological features along with their distinct geographical distributions. Recent studies suggest that the Asian element may be genetically distinct from the European and American elements. However, a comprehensive study on the genetic differentiation among all four elements remains lacking. This study aimed to leverage whole plastid genome sequences and ITS2 haplotypes to comprehensively assess the genomic diversity within the P. amphibia complex. Notably, we included multiple individuals from New York State to resolve the ongoing debate regarding the taxonomic status of two American elements - whether they represent a single species or distinct entities. Our analysis revealed a well-supported monophyletic clade encompassing all four elements, endorsing their own section, Amphibia. Notably, the terrestrial form of the American element is sister to all other elements, suggesting it deserves its own species status. This reinstates its historical name, P. coccinea, separating it from the broader P. amphibia. Furthermore, distinct compositions of the ITS2 haplotypes differentiated the four elements, although the European element should be further investigated with more sampling. The most intriguing discovery is the identification of putative hybrids between the two American elements. In one population out of four putative hybrid populations, all three entities - the two parent species and their hybrid offspring - thrive together, showcasing a fascinating microcosm of ongoing evolutionary processes. Unraveling the intricate genetic tapestry within each American species and their hybrid populations remains a compelling next step. By delving deeper into their genetic makeup, we can gain a richer understanding of their evolutionary trajectories and the intricacies of their interactions. Finally, it is estimated that the two species of sect. Amphibia diverged approximately 4.02 million years ago during the Pliocene epoch, when there was a significant global cooling and drying trend.

Mitochondrial ATP Synthase beta-Subunit Affects Plastid Retrograde Signaling in Arabidopsis.

Plastid retrograde signaling plays a key role in coordinating the expression of plastid genes and photosynthesis-associated nuclear genes (PhANGs). Although plastid retrograde signaling can be substantially compromised by mitochondrial dysfunction, it is not yet clear whether specific mitochondrial factors are required to regulate plastid retrograde signaling. Here, we show that mitochondrial ATP synthase beta-subunit mutants with decreased ATP synthase activity are impaired in plastid retrograde signaling in Arabidopsis thaliana. Transcriptome analysis revealed that the expression levels of PhANGs were significantly higher in the mutants affected in the AT5G08670 gene encoding the mitochondrial ATP synthase beta-subunit, compared to wild-type (WT) seedlings when treated with lincomycin (LIN) or norflurazon (NF). Further studies indicated that the expression of nuclear genes involved in chloroplast and mitochondrial retrograde signaling was affected in the AT5G08670 mutant seedlings treated with LIN. These changes might be linked to the modulation of some transcription factors (TFs), such as LHY (Late Elongated Hypocotyl), PIF (Phytochrome-Interacting Factors), MYB, WRKY, and AP2/ERF (Ethylene Responsive Factors). These findings suggest that the activity of mitochondrial ATP synthase significantly influences plastid retrograde signaling.

Comparative Analysis of Plastome Sequences of Seven Tulipa L. (Liliaceae Juss.) Species from Section Kolpakowskianae Raamsd. Ex Zonn and Veldk.

Tulipa L. is a genus of significant economic, environmental, and cultural importance in several parts of the world. The exact number of species in the genus remains uncertain due to inherent taxonomic challenges. We utilized next-generation sequencing technology to sequence and assemble the plastid genomes of seven Tulipa species collected in Kazakhstan and conducted a comparative analysis. The total number of annotated genes was 136 in all seven studied Tulipa species, 114 of which were unique, including 80 protein-coding, 30 tRNA, and 4 rRNA genes. Nine regions (petD, ndhH, ycf2-ycf3, ndhA, rpl16, clpP, ndhD-ndhF, rpoC2, and ycf1) demonstrated significant nucleotide variability, suggesting their potential as molecular markers. A total of 1388 SSRs were identified in the seven Tulipa plastomes, with mononucleotide repeats being the most abundant (60.09%), followed by dinucleotide (34.44%), tetranucleotide (3.90%), trinucleotide (1.08%), pentanucleotide (0.22%), and hexanucleotide (0.29%). The Ka/Ks values of the protein-coding genes ranged from 0 to 3.9286, with the majority showing values <1. Phylogenetic analysis based on a complete plastid genome and protein-coding gene sequences divided the species into three major clades corresponding to their subgenera. The results obtained in this study may contribute to understanding the phylogenetic relationships and molecular taxonomy of Tulipa species.

Proximity driven plastid-nucleus relationships are facilitated by tandem plastid-ER dynamics.

Peri-nuclear clustering (PNC) of chloroplasts has largely been described in senescent and pathogen- or ROS- stressed cells. Stromules, tubular plastid extensions are also observed under similar conditions. Coincident observations of PNC and stromules associate the two phenomena in facilitating retrograde signaling between chloroplasts and the nucleus. However, PNC incidence in non-stressed cells under normal growth and developmental conditions, when stromules are usually not observed, remains unclear. Using transgenic Arabidopsis expressing different organelle-targeted fluorescent proteins we show that PNC is a dynamic subcellular phenomenon that continues in the absence of light and is not dependent on stromule formation. PNC is facilitated by tandem plastid-ER dynamics created through membrane contact sites between the two organelles. While PNC increases upon ER-membrane expansion, some plastids may remain in the peri-nuclear region due to their localization in ER-lined nuclear indentions. Moreover, some PNC plastids may sporadically extend stromules into ER-lined nuclear grooves. Our findings strongly suggest that PNC is not an exclusive response to stress caused by pathogens, high light or exogenous-H2O2 treatment and does not require stromule formation. However, morphological and behavioural alterations in ER and concomitant changes in tandem, plastid-ER dynamics play a major role in facilitating the phenomenon.

Integrated overview of stramenopile ecology, taxonomy, and heterotrophic origin.

Stramenopiles represent a significant proportion of aquatic and terrestrial biota. Most biologists can name a few, but these are limited to the phototrophic (e.g., diatoms and kelp) or parasitic species (e.g., oomycetes, Blastocystis), with free-living heterotrophs largely overlooked. Though our attention is slowly turning towards heterotrophs, we have only a limited understanding of their biology due to a lack of cultured models. Recent metagenomic and single-cell investigations have revealed the species richness and ecological importance of stramenopiles-especially heterotrophs. However, our lack of knowledge of the cell biology and behaviour of these organisms leads to our inability to match species to their particular ecological functions. Because photosynthetic stramenopiles are studied independently of their heterotrophic relatives, they are often treated separately in the literature. Here, we present stramenopiles as a unified group with shared synapomorphies and evolutionary history. We introduce the main lineages, describe their important biological and ecological traits, and provide a concise update on the origin of the ochrophyte plastid. We highlight the crucial role of heterotrophs and mixotrophs in our understanding of stramenopiles with the goal of inspiring future investigations in taxonomy and life history. To understand each of the many diversifications within stramenopiles-towards autotrophy, osmotrophy, or parasitism-we must understand the ancestral heterotrophic flagellate from which they each evolved. We hope the following will serve as a primer for new stramenopile researchers or as an integrative refresher to those already in the field.

The response of Midknight Valencia oranges to ethephon degreening varies in the turning and regreening stages.

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.

Complementary environmental analysis and functional characterization of lower glycolysis-gluconeogenesis in the diatom plastid.

Organic carbon fixed in chloroplasts through the Calvin-Benson-Bassham Cycle can be diverted towards different metabolic fates, including cyoplasmic and mitochondrial respiration, gluconeogenesis, and synthesis of diverse plastid metabolites via the pyruvate hub. In plants, pyruvate is principally produced via cytoplasmic glycolysis, although a plastid-targeted lower glycolytic pathway is known to exist in non-photosynthetic tissue. Here, we characterized a lower plastid glycolysis-gluconeogenesis pathway enabling the direct interconversion of glyceraldehyde-3-phosphate and phospho-enol-pyruvate in diatoms, ecologically important marine algae distantly related to plants. We show that two reversible enzymes required to complete diatom plastid glycolysis-gluconeogenesis, Enolase and bis-phospho-glycerate mutase (PGAM), originated through duplications of mitochondria-targeted respiratory isoforms. Through CRISPR-Cas9 mutagenesis, integrative 'omic analyses, and measured kinetics of expressed enzymes in the diatom Phaeodactylum tricornutum, we present evidence that this pathway diverts plastid glyceraldehyde-3-phosphate into the pyruvate hub, and may also function in the gluconeogenic direction. Considering experimental data, we show that this pathway has different roles dependent in particular on day length and environmental temperature, and show that the cpEnolase and cpPGAM genes are expressed at elevated levels in high latitude oceans where diatoms are abundant. Our data provide evolutionary, meta-genomic and functional insights into a poorly understood yet evolutionarily recurrent plastid metabolic pathway.

The chloroplast genome sequences of Ipomoea alba and I. obscura (Convolvulaceae): genome comparison and phylogenetic analysis.

Ipomoea species have diverse uses as ornamentals, food, and medicine. However, their genomic information is limited; I. alba and I. obscura were sequenced and assembled. Their chloroplast genomes were 161,353 bp and 159,691 bp, respectively. Both genomes exhibited a quadripartite structure, consisting of a pair of inverted repeat (IR) regions, which are separated by the large single-copy (LSC) and small single-copy (SSC) regions. The overall GC content was 37.5% for both genomes. A total of 104 and 93 simple sequence repeats, 50 large repeats, and 30 and 22 short tandem repeats were identified in the two chloroplast genomes, respectively. G and T were more preferred than C and A at the third base position based on the Parity Rule 2 plot analysis, and the neutrality plot revealed correlation coefficients of 0.126 and 0.105, indicating the influence of natural selection in shaping the codon usage bias in most protein-coding genes (CDS). Genome comparative analyses using 31 selected Ipomoea taxa from Thailand showed that their chloroplast genomes are rather conserved, but the presence of expansion or contraction of the IR region was identified in some of these Ipomoea taxa. A total of five highly divergent regions were identified, including the CDS genes accD, ndhA, and ndhF, as well as the intergenic spacer regions psbI-atpA and rpl32-ccsA. Phylogenetic analysis based on both the complete chloroplast genome sequence and CDS datasets of 31 Ipomoea taxa showed that I. alba is resolved as a group member for series (ser.) Quamoclit, which contains seven other taxa, including I. hederacea, I. imperati, I. indica, I. nil, I. purpurea, I. quamoclit, and I. × sloteri, while I. obscura is grouped with I. tiliifolia, both of which are under ser. Obscura, and is closely related to I. biflora of ser. Pes-tigridis. Divergence time estimation using the complete chloroplast genome sequence dataset indicated that the mean age of the divergence for Ipomoeeae, Argyreiinae, and Astripomoeinae, was approximately 29.99 Mya, 19.81 Mya, and 13.40 Mya, respectively. The node indicating the divergence of I. alba from the other members of Ipomoea was around 10.06 Mya, and the split between I. obscura and I. tiliifolia is thought to have happened around 17.13 Mya. The split between the I. obscura accessions from Thailand and Taiwan is thought to have taken place around 0.86 Mya.

The fnr-like mutants confer isoxaben tolerance by initiating mitochondrial retrograde signalling.

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.

Piece and parcel of gymnosperm organellar genomes.

MAIN CONCLUSION: Significant past, present, and potential future research into the organellar (plastid and mitochondrial) genomes of gymnosperms that can provide insight into the unknown origin and evolution of plants is highlighted. Gymnosperms are vascular seed plants that predominated the ancient world before their sister clade, angiosperms, took over during the Late Cretaceous. The divergence of gymnosperms and angiosperms took place around 300 Mya, with the latter evolving into the diverse group of flowering plants that dominate the plant kingdom today. Although gymnosperms have reportedly made some evolutionary innovations, the literature on their genome advances, particularly their organellar (plastid and mitochondrial) genomes, is relatively scattered and fragmented. While organellar genomes can shed light on plant origin and evolution, they are frequently overlooked, due in part to their limited contribution to gene expression and lack of evolutionary dynamics when compared to nuclear genomes. A better understanding of gymnosperm organellar genomes is critical because they reveal genetic changes that have contributed to their unique adaptations and ecological success, potentially aiding in plant survival, enhancement, and biodiversity conservation in the face of climate change. This review reveals significant information and gaps in the existing knowledge base of organellar genomes in gymnosperms, as well as the challenges and research needed to unravel their complexity.

Production of biologically active human basic fibroblast growth factor (hFGFb) using Nicotiana tabacum transplastomic plants.

MAIN CONCLUSION: We generated transplastomic tobacco lines that stably express a human Basic Fibroblast Growth Factor (hFGFb) in their chloroplasts stroma and purified a biologically active recombinant hFGFb. MAIN: The use of plants as biofactories presents as an attractive technology with the potential to efficiently produce high-value human recombinant proteins in a cost-effective manner. Plastid genome transformation stands out for its possibility to accumulate recombinant proteins at elevated levels. Of particular interest are recombinant growth factors, given their applications in animal cell culture and regenerative medicine. In this study, we produced recombinant human Fibroblast Growth Factor (rhFGFb), a crucial protein required for animal cell culture, in tobacco chloroplasts. We successfully generated two independent transplastomic lines that are homoplasmic and accumulate rhFGFb in their leaves. Furthermore, the produced rhFGFb demonstrated its biological activity by inducing proliferation in HEK293T cell lines. These results collectively underscore plastid genome transformation as a promising plant-based bioreactor for rhFGFb production.

Marchantia polymorpha GOLDEN2-LIKE transcriptional factor; a central regulator of chloroplast and plant vegetative development.

The GOLDEN2-LIKE (GLK) transcription factors act as a central regulatory node involved in both developmental processes and environmental responses. Marchantia polymorpha, a basal terrestrial plant with strategic evolutionary position, contains a single GLK representative that possesses an additional domain compared to spermatophytes. We analyzed the role of MpGLK in chloroplast biogenesis and development by altering its levels, preforming transcriptomic profiling and conducting chromatin immunoprecipitation. Decreased MpGLK levels impair chloroplast differentiation and disrupt the expression of photosynthesis-associated nuclear genes, while overexpressing MpGLK leads to ectopic chloroplast biogenesis. This demonstrates the MpGLK functions as a bona fide GLK protein, likely representing an ancestral GLK architecture. Altering MpGLK levels directly regulates the expression of genes involved in Chl synthesis and degradation, similar to processes observed in eudicots, and causes various developmental defects in Marchantia, including the formation of dorsal structures such as air pores and gemma cups. MpGLK, also directly activates MpMAX2 gene expression, regulating the timing of gemma cup development. Our study shows that MpGLK functions as a master regulator, potentially coupling chloroplast development with vegetative reproduction. This illustrates the complex regulatory networks governing chloroplast function and plant development communication and highlight the evolutionary conservation of GLK-mediated regulatory processes across plant species.

Long-read RNA sequencing can probe organelle genome pervasive transcription.

40 years ago, organelle genomes were assumed to be streamlined and, perhaps, unexciting remnants of their prokaryotic past. However, the field of organelle genomics has exposed an unparallel diversity in genome architecture (i.e. genome size, structure, and content). The transcription of these eccentric genomes can be just as elaborate - organelle genomes are pervasively transcribed into a plethora of RNA types. However, while organelle protein-coding genes are known to produce polycistronic transcripts that undergo heavy posttranscriptional processing, the nature of organelle noncoding transcriptomes is still poorly resolved. Here, we review how wet-lab experiments and second-generation sequencing data (i.e. short reads) have been useful to determine certain types of organelle RNAs, particularly noncoding RNAs. We then explain how third-generation (long-read) RNA-Seq data represent the new frontier in organelle transcriptomics. We show that public repositories (e.g. NCBI SRA) already contain enough data for inter-phyla comparative studies and argue that organelle biologists can benefit from such data. We discuss the prospects of using publicly available sequencing data for organelle-focused studies and examine the challenges of such an approach. We highlight that the lack of a comprehensive database dedicated to organelle genomics/transcriptomics is a major impediment to the development of a field with implications in basic and applied science.

Genetic Diversity and Phylogeography of the Relict Tree Fern Culcita macrocarpa: Influence of Clonality and Breeding System on Genetic Variation.

The tree fern Culcita macrocarpa, a threatened Iberian-Macaronesian endemism, represents the sole European species of the order Cyatheales. Considered a Tertiary relict of European Palaeotropical flora, its evolutionary history and genetic diversity, potentially influenced by presumed high clonal propagation, remain largely unknown. This study elucidates the phylogeographic history of C. macrocarpa, assessing the impact of vegetative reproduction on population dynamics and genetic variability. We provide genetic data from eight newly identified nuclear microsatellite loci and one plastid DNA region for 17 populations spanning the species' range, together with species distribution modeling data. Microsatellites reveal pervasive clonality in C. macrocarpa, which has varied among populations. We assess the impact of clonality on genetic diversity and evaluate how estimates of intra-population genetic diversity indices and genetic structuring are affected by the chosen definition of "individual" (focusing exclusively on genetically distinct individuals, genets, as opposed to considering all independent clonal replicates, ramets). We identify two main population groups, one in the northern Iberian Peninsula and the other in the Macaronesian archipelagos and southern Iberian Peninsula. Within each group, we found relict populations (in the Azores and the Cantabrian Cornice) as well as recent originated populations. This population structure suggests colonization dynamics in which recent populations originated from one or a few genets of relict populations and became established through intra-gametophytic self-fertilization and vegetative expansion. DAPC analysis facilitated the identification of alleles that most significantly contributed to the observed population structure. The current Andalusian populations appear to have originated from colonization events from the Azores and the Cantabrian Cornice. Our findings suggest that C. macrocarpa persisted through the Last Glacial Maximum in two refugia: the Azores and the Cantabrian Cornice. Colonization into new areas occurred presumably from these refuges, generating two large population groups with structured genetic diversity. This study underscores the significance of clonality in establishing new populations and shaping genetic structure.