On the importance of sequence alignment inspections in plastid phylogenomics - an example from revisiting the relationships of the water-lilies.

The water-lily clade represents the second earliest-diverging branch of angiosperms. Most of its species belong to Nymphaeaceae, of which the "core Nymphaeaceae"-comprising the genera Euryale, Nymphaea and Victoria-is the most diverse clade. Despite previous molecular phylogenetic studies on the core Nymphaeaceae, various aspects of their evolutionary relationships have remained unresolved. The length-variable introns and intergenic spacers are known to contain most of the sequence variability within the water-lily plastomes. Despite the challenges with multiple sequence alignment, any new molecular phylogenetic investigation on the core Nymphaeaceae should focus on these noncoding plastome regions. For example, a new plastid phylogenomic study on the core Nymphaeaceae should generate DNA sequence alignments of all plastid introns and intergenic spacers based on the principle of conserved sequence motifs. In this investigation, we revisit the phylogenetic history of the core Nymphaeaceae by employing such an approach. Specifically, we use a plastid phylogenomic analysis strategy in which all coding and noncoding partitions are separated and then undergo software-driven DNA sequence alignment, followed by a motif-based alignment inspection and adjustment. This approach allows us to increase the reliability of the character base compared to the default practice of aligning complete plastomes through software algorithms alone. Our approach produces significantly different phylogenetic tree reconstructions for several of the plastome regions under study. The results of these reconstructions underscore that Nymphaea is paraphyletic in its current circumscription, that each of the five subgenera of Nymphaea is monophyletic, and that the subgenus Nymphaea is sister to all other subgenera of Nymphaea. Our results also clarify many evolutionary relationships within the Nymphaea subgenera Brachyceras, Hydrocallis and Nymphaea. In closing, we discuss whether the phylogenetic reconstructions obtained through our motif-based alignment adjustments are in line with morphological evidence on water-lily evolution.

Genome copy number predicts extreme evolutionary rate variation in plant mitochondrial DNA.

Nuclear and organellar genomes can evolve at vastly different rates despite occupying the same cell. In most bilaterian animals, mitochondrial DNA (mtDNA) evolves faster than nuclear DNA, whereas this trend is generally reversed in plants. However, in some exceptional angiosperm clades, mtDNA substitution rates have increased up to 5,000-fold compared with closely related lineages. The mechanisms responsible for this acceleration are generally unknown. Because plants rely on homologous recombination to repair mtDNA damage, we hypothesized that mtDNA copy numbers may predict evolutionary rates, as lower copy numbers may provide fewer templates for such repair mechanisms. In support of this hypothesis, we found that copy number explains 47% of the variation in synonymous substitution rates of mtDNA across 60 diverse seed plant species representing ~300 million years of evolution. Copy number was also negatively correlated with mitogenome size, which may be a cause or consequence of mutation rate variation. Both relationships were unique to mtDNA and not observed in plastid DNA. These results suggest that homologous recombinational repair plays a role in driving mtDNA substitution rates in plants and may explain variation in mtDNA evolution more broadly across eukaryotes. Our findings also contribute to broader questions about the relationships between mutation rates, genome size, selection efficiency, and the drift-barrier hypothesis.

Out of Place: Phylogenomics resolves the placement of Eurasian taxa and sheds light on origin of Thermopsideae in North America.

The North American Thermopsideae (Fabaceae: Papilionoideae), a monophyletic group comprising the North American endemic genus Baptisia, and the paraphyletic Eurasian-North American disjunct Thermopsis, is nested within the tribe Sophoreae. Previous phylogenetic studies have identified two East Asian taxa within the North American Thermopsideae, suggesting two independent dispersal events between North America-East Asia. More recent studies have also placed a third taxon, Vuralia turcica, an endemic species from Turkey, among the North American Thermopsideae. The presence of three geographically distant Eurasian taxa within a relatively young clade of North American origin is unprecedented among papilionoid legumes, and the biogeographic implications of this observation are not clear. To investigate this matter, 1540 low-copy nuclear genes and complete plastomes were obtained from 36 taxa across the core genistoids, including 26 newly sequenced taxa. Nuclear and plastome based maximum likelihood (ML) and ASTRAL analyses were conducted based on varying degrees of taxon coverage and read mapping consensus threshold values. Additional analyses were performed to estimate divergence times and to reconstruct biogeographic history. The results strongly support a previously undetected Old World clade, presently composed of V. turcica and T. chinensis, which diverged from the ancestor of the North American lineage during the mid to late Miocene. A single and recent North America-East Asia dispersal involving T. lupinoides is reported. Furthermore, the traditional inclusion of the genus Ammopiptanthus among Thermopsideae is not supported, and the monotypic generic status of Vuralia is called into question. A relatively high degree of cytonuclear discordance is reported within each sub-clade of the North American Thermopsideae. This finding is likely attributable to the high degree of interspecific hybridization reported within these groups and raises the need for more rigorous genome-scale testing to better delimit species within each of the reticulating subclades. Subjects: Biodiversity, Biogeography, Evolutionary Studies, Genetics, Plant Science.

Plastid Genome Assembly Using Long-read data.

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.

Early Fruit Development Regulation-Related Genes Concordantly Expressed with TCP Transcription Factors in Tomato (Solanum lycopersicum).

The tomato (Solanum lycopersicum L.) is considered one of the most important vegetable crops globally, both agronomically and economically; however, its fruit development regulation network is still unclear. The transcription factors serve as master regulators, activating many genes and/or metabolic pathways throughout the entire plant life cycle. In this study, we identified the transcription factors that are coordinated with TCP gene family regulation in early fruit development by making use of the high-throughput sequencing of RNA (RNAseq) technique. A total of 23 TCP-encoding genes were found to be regulated at various stages during the growth of the fruit. The expression patterns of five TCPs were consistent with those of other transcription factors and genes. There are two unique subgroups of this larger family: class I and class II TCPs. Others were directly associated with the growth and/or ripening of fruit, while others were involved in the production of the hormone auxin. Moreover, it was discovered that TCP18 had an expression pattern that was similar to that of the ethylene-responsive transcription factor 4 (ERF4). Tomato fruit set and overall development are under the direction of a gene called auxin response factor 5 (ARF5). TCP15 revealed an expression that was in sync with this gene. This study provides insight into the potential processes that help in acquiring superior fruit qualities by accelerating fruit growth and ripening.

Rate accelerations in plastid and mitochondrial genomes of Cyperaceae occur in the same clades.

Cyperaceae, the second largest family in the monocot order Poales, comprises >5500 species and includes the genus Eleocharis with âˆ¼ 250 species. A previous study of complete plastomes of two Eleocharis species documented extensive structural heteroplasmy, gene order changes, high frequency of dispersed repeats along with gene losses and duplications. To better understand the phylogenetic distribution of gene and intron content as well as rates and patterns of sequence evolution within and between mitochondrial and plastid genomes of Eleocharis and Cyperaceae, an additional 29 Eleocharis organelle genomes were sequenced and analyzed. Eleocharis experienced extensive gene loss in both genomes while loss of introns was mitochondria-specific. Eleocharis has higher rates of synonymous (dS) and nonsynonymous (dN) substitutions in the plastid and mitochondrion than most sampled angiosperms, and the pattern was distinct from other eudicot lineages with accelerated rates. Several clades showed higher dS and dN in mitochondrial genes than in plastid genes. Furthermore, nucleotide substitution rates of mitochondrial genes were significantly accelerated on the branch leading to Cyperaceae compared to most angiosperms. Mitochondrial genes of Cyperaceae exhibited dramatic loss of RNA editing sites and a negative correlation between RNA editing and dS values was detected among angiosperms. Mutagenic retroprocessing and dysfunction of DNA replication, repair and recombination genes are the most likely cause of striking rate accelerations and loss of edit sites and introns in Eleocharis and Cyperaceae organelle genomes.

Plastid phylogenomics uncovers multiple species in Medicago truncatula (Fabaceae) germplasm accessions.

Medicago truncatula is a model legume that has been extensively investigated in diverse subdisciplines of plant science. Medicago littoralis can interbreed with M. truncatula and M. italica; these three closely related species form a clade, i.e. TLI clade. Genetic studies have indicated that M. truncatula accessions are heterogeneous but their taxonomic identities have not been verified. To elucidate the phylogenetic position of diverse M. truncatula accessions within the genus, we assembled 54 plastid genomes (plastomes) using publicly available next-generation sequencing data and conducted phylogenetic analyses using maximum likelihood. Five accessions showed high levels of plastid DNA polymorphism. Three of these highly polymorphic accessions contained sequences from both M. truncatula and M. littoralis. Phylogenetic analyses of sequences placed some accessions closer to distantly related species suggesting misidentification of source material. Most accessions were placed within the TLI clade and maximally supported the interrelationships of three subclades. Two Medicago accessions were placed within a M. italica subclade of the TLI clade. Plastomes with a 45-kb (rpl20-ycf1) inversion were placed within the M. littoralis subclade. Our results suggest that the M. truncatula accession genome pool represents more than one species due to possible mistaken identities and gene flow among closely related species.

Effects of Salt Stress on Transcriptional and Physiological Responses in Barley Leaves with Contrasting Salt Tolerance.

Salt stress negatively impacts crop production worldwide. Genetic diversity among barley (Hordeum vulgare) landraces adapted to adverse conditions should provide a valuable reservoir of tolerance genes for breeding programs. To identify molecular and biochemical differences between barley genotypes, transcriptomic and antioxidant enzyme profiles along with several morpho-physiological features were compared between salt-tolerant (Boulifa) and salt-sensitive (Testour) genotypes subjected to salt stress. Decreases in biomass, photosynthetic parameters, and relative water content were low in Boulifa compared to Testour. Boulifa had better antioxidant protection against salt stress than Testour, with greater antioxidant enzymes activities including catalase, superoxide dismutase, and guaiacol peroxidase. Transcriptome assembly for both genotypes revealed greater accumulation of differentially expressed transcripts in Testour compared to Boulifa, emphasizing the elevated transcriptional response in Testour following salt exposure. Various salt-responsive genes, including the antioxidant catalase 3, the osmoprotectant betaine aldehyde dehydrogenase 2, and the transcription factors MYB20 and MYB41, were induced only in Boulifa. By contrast, several genes associated with photosystems I and II, and light receptor chlorophylls A and B, were more repressed in Testour. Co-expression network analysis identified specific gene modules correlating with differences in genotypes and morpho-physiological traits. Overall, salinity-induced differential transcript accumulation underlies the differential morpho-physiological response in both genotypes and could be important for breeding salt tolerance in barley.

Highly Resolved Papilionoid Legume Phylogeny Based on Plastid Phylogenomics.

Comprising 501 genera and around 14,000 species, Papilionoideae is not only the largest subfamily of Fabaceae (Leguminosae; legumes), but also one of the most extraordinarily diverse clades among angiosperms. Papilionoids are a major source of food and forage, are ecologically successful in all major biomes, and display dramatic variation in both floral architecture and plastid genome (plastome) structure. Plastid DNA-based phylogenetic analyses have greatly improved our understanding of relationships among the major groups of Papilionoideae, yet the backbone of the subfamily phylogeny remains unresolved. In this study, we sequenced and assembled 39 new plastomes that are covering key genera representing the morphological diversity in the subfamily. From 244 total taxa, we produced eight datasets for maximum likelihood (ML) analyses based on entire plastomes and/or concatenated sequences of 77 protein-coding sequences (CDS) and two datasets for multispecies coalescent (MSC) analyses based on individual gene trees. We additionally produced a combined nucleotide dataset comprising CDS plus matK gene sequences only, in which most papilionoid genera were sampled. A ML tree based on the entire plastome maximally supported all of the deep and most recent divergences of papilionoids (223 out of 236 nodes). The Swartzieae, ADA (Angylocalyceae, Dipterygeae, and Amburaneae), Cladrastis, Andira, and Exostyleae clades formed a grade to the remainder of the Papilionoideae, concordant with nine ML and two MSC trees. Phylogenetic relationships among the remaining five papilionoid lineages (Vataireoid, Dermatophyllum, Genistoid s.l., Dalbergioid s.l., and Baphieae + Non-Protein Amino Acid Accumulating or NPAAA clade) remained uncertain, because of insufficient support and/or conflicting relationships among trees. Our study fully resolved most of the deep nodes of Papilionoideae, however, some relationships require further exploration. More genome-scale data and rigorous analyses are needed to disentangle phylogenetic relationships among the five remaining lineages.

Born in the mitochondrion and raised in the nucleus: evolution of a novel tandem repeat family in Medicago polymorpha (Fabaceae).

Plant nuclear genomes harbor sequence elements derived from the organelles (mitochondrion and plastid) through intracellular gene transfer (IGT). Nuclear genomes also show a dramatic range of repeat content, suggesting that any sequence can be readily amplified. These two aspects of plant nuclear genomes are well recognized but have rarely been linked. Through investigation of 31 Medicago taxa we detected exceptionally high post-IGT amplification of mitochondrial (mt) DNA sequences containing rps10 in the nuclear genome of Medicago polymorpha and closely related species. The amplified sequences were characterized as tandem arrays of five distinct repeat motifs (2157, 1064, 987, 971, and 587 bp) that have diverged from the mt genome (mitogenome) in the M. polymorpha nuclear genome. The mt rps10-like arrays were identified in seven loci (six intergenic and one telomeric) of the nuclear chromosome assemblies and were the most abundant tandem repeat family, representing 1.6-3.0% of total genomic DNA, a value approximately three-fold greater than the entire mitogenome in M. polymorpha. Compared to a typical mt gene, the mt rps10-like sequence coverage level was 691.5-7198-fold higher in M. polymorpha and closely related species. In addition to the post-IGT amplification, our analysis identified the canonical telomeric repeat and the species-specific satellite arrays that are likely attributable to an ancestral chromosomal fusion in M. polymorpha. A possible relationship between chromosomal instability and the mt rps10-like tandem repeat family in the M. polymorpha clade is discussed.

Transcriptomic Analysis of Salt-Stress-Responsive Genes in Barley Roots and Leaves.

Barley is characterized by a rich genetic diversity, making it an important model for studies of salinity response with great potential for crop improvement. Moreover, salt stress severely affects barley growth and development, leading to substantial yield loss. Leaf and root transcriptomes of a salt-tolerant Tunisian landrace (Boulifa) exposed to 2, 8, and 24 h salt stress were compared with pre-exposure plants to identify candidate genes and pathways underlying barley's response. Expression of 3585 genes was upregulated and 5586 downregulated in leaves, while expression of 13,200 genes was upregulated and 10,575 downregulated in roots. Regulation of gene expression was severely impacted in roots, highlighting the complexity of salt stress response mechanisms in this tissue. Functional analyses in both tissues indicated that response to salt stress is mainly achieved through sensing and signaling pathways, strong transcriptional reprograming, hormone osmolyte and ion homeostasis stabilization, increased reactive oxygen scavenging, and activation of transport and photosynthesis systems. A number of candidate genes involved in hormone and kinase signaling pathways, as well as several transcription factor families and transporters, were identified. This study provides valuable information on early salt-stress-responsive genes in roots and leaves of barley and identifies several important players in salt tolerance.

Evaluating the contribution of osmotic and oxidative stress components on barley growth under salt stress.

Salt stress is considered one of the most devastating environmental stresses, affecting barley growth and leading to significant yield loss. Hence, there is considerable interest in investigating the most effective traits that determine barley growth under salt stress. The objective of this study was to elucidate the contribution of osmotic and oxidative stress components in leaves and roots growth under salt stress. Two distinct barley (Hordeum vulgare) salt-stress tolerant genotypes, Barrage Malleg (BM, tolerant) and Saouef (Sf, sensitive), were subjected to 200 mM NaCl at early vegetative stages. Stressed and control leaves and roots tissue were assessed for several growth traits, including fresh and dry weight and plant length, as well as the content of osmoprotectants proline and soluble sugars. In addition, malondialdehyde content and activities of superoxide dismutase (SOD), catalase (CAT) and ascorbate peroxidase (APX), as well as their corresponding gene expression patterns, were investigated. The results showed better performance of BM over Sf for leaf dry weight (LDW), root dry weight (RDW) and root length (RL). The salt-tolerant genotype (BM) had better osmoprotection against salt stress compared with the salt-sensitive genotype (Sf), with a higher accumulation of proline and soluble sugars in leaves and roots and a stronger antioxidant system as evidenced by higher activities of SOD, CAT and APX and more abundant Cu/Zn-SOD transcripts, especially in roots. Stepwise regression analysis indicated that under salt stress the most predominant trait of barley growth was Cu/Zn-SOD gene expression level, suggesting that alleviating oxidative stress and providing cell homeostasis is the first priority.

In and out: Evolution of viral sequences in the mitochondrial genomes of legumes (Fabaceae).

Plant specific mitoviruses in the 'genus' Mitovirus (Narnaviridae) and their integrated sequences (non-retroviral endogenous RNA viral elements or NERVEs) have been recently identified in various plant lineages. However, the sparse phylogenetic coverage of complete plant mitochondrial genome (mitogenome) sequences and the non-conserved nature of mitochondrial intergenic regions have hindered comparative studies on mitovirus NERVEs in plants. In this study, 10 new mitogenomes were sequenced from legumes (Fabaceae). Based on comparative genomic analysis of 27 total mitogenomes, we identified mitovirus NERVEs and transposable elements across the family. All legume mitogenomes included NERVEs and total NERVE length varied from ca. 2 kb in the papilionoid Trifolium to 35 kb in the mimosoid Acacia. Most of the NERVE integration sites were in highly variable intergenic regions, however, some were positioned in six cis-spliced mitochondrial introns. In the Acacia mitogenome, there were L1-like transposon sequences including an almost full-length copy with target site duplications (TSDs). The integration sites of NERVEs in four introns showed evidence of L1-like retrotransposition events. Phylogenetic analysis revealed that there were multiple instances of precise deletion of NERVEs between TSDs. This study provides clear evidence that a L1-like retrotransposition mechanism has a long history of contributing to the integration of viral RNA into plant mitogenomes while microhomology-mediated deletion can restore the integration site.

The chicken or the egg? Plastome evolution and an independent loss of the inverted repeat in papilionoid legumes.

The plastid genome (plastome), while surprisingly constant in gene order and content across most photosynthetic angiosperms, exhibits variability in several unrelated lineages. During the diversification history of the legume family Fabaceae, plastomes have undergone many rearrangements, including inversions, expansion, contraction and loss of the typical inverted repeat (IR), gene loss and repeat accumulation in both shared and independent events. While legume plastomes have been the subject of study for some time, most work has focused on agricultural species in the IR-lacking clade (IRLC) and the plant model Medicago truncatula. The subfamily Papilionoideae, which contains virtually all of the agricultural legume species, also comprises most of the plastome variation detected thus far in the family. In this study three non-papilioniods were included among 34 newly sequenced legume plastomes, along with 33 publicly available sequences, to assess plastome structural evolution in the subfamily. In an effort to examine plastome variation across the subfamily, approximately 20% of the sampling represents the IRLC with the remainder selected to represent the early-branching papilionoid clades. A number of IR-related and repeat-mediated changes were identified and examined in a phylogenetic context. Recombination between direct repeats associated with ycf2 resulted in intraindividual plastome heteroplasmy. Although loss of the IR has not been reported in legumes outside of the IRLC, one genistoid taxon was found to completely lack the typical plastome IR. The role of the IR and non-IR repeats in the progression of plastome change is discussed.

Plastid Genomes of Flowering Plants: Essential Principles.

The plastid genome (plastome ) has proved a valuable source of data for evaluating evolutionary relationships among angiosperms. Through basic and applied approaches, plastid transformation technology offers the potential to understand and improve plant productivity, providing food, fiber, energy, and medicines to meet the needs of a burgeoning global population. The growing genomic resources available to both phylogenetic and biotechnological investigations is allowing novel insights and expanding the scope of plastome research to encompass new species. In this chapter, we present an overview of some of the seminal and contemporary research that has contributed to our current understanding of plastome evolution and attempt to highlight the relationship between evolutionary mechanisms and the tools of plastid genetic engineering.

The evolutionary fate of rpl32 and rps16 losses in the Euphorbia schimperi (Euphorbiaceae) plastome.

Gene transfers from mitochondria and plastids to the nucleus are an important process in the evolution of the eukaryotic cell. Plastid (pt) gene losses have been documented in multiple angiosperm lineages and are often associated with functional transfers to the nucleus or substitutions by duplicated nuclear genes targeted to both the plastid and mitochondrion. The plastid genome sequence of Euphorbia schimperi was assembled and three major genomic changes were detected, the complete loss of rpl32 and pseudogenization of rps16 and infA. The nuclear transcriptome of E. schimperi was sequenced to investigate the transfer/substitution of the rpl32 and rps16 genes to the nucleus. Transfer of plastid-encoded rpl32 to the nucleus was identified previously in three families of Malpighiales, Rhizophoraceae, Salicaceae and Passifloraceae. An E. schimperi transcript of pt SOD-1-RPL32 confirmed that the transfer in Euphorbiaceae is similar to other Malpighiales indicating that it occurred early in the divergence of the order. Ribosomal protein S16 (rps16) is encoded in the plastome in most angiosperms but not in Salicaceae and Passifloraceae. Substitution of the E. schimperi pt rps16 was likely due to a duplication of nuclear-encoded mitochondrial-targeted rps16 resulting in copies dually targeted to the mitochondrion and plastid. Sequences of RPS16-1 and RPS16-2 in the three families of Malpighiales (Salicaceae, Passifloraceae and Euphorbiaceae) have high sequence identity suggesting that the substitution event dates to the early divergence within Malpighiales.

Clade-Specific Plastid Inheritance Patterns Including Frequent Biparental Inheritance in Passiflora Interspecific Crosses.

Plastid inheritance in angiosperms is presumed to be largely maternal, with the potential to inherit plastids biparentally estimated for about 20% of species. In Passiflora, maternal, paternal and biparental inheritance has been reported; however, these studies were limited in the number of crosses and progeny examined. To improve the understanding of plastid transmission in Passiflora, the progeny of 45 interspecific crosses were analyzed in the three subgenera: Passiflora, Decaloba and Astrophea. Plastid types were assessed following restriction digestion of PCR amplified plastid DNA in hybrid embryos, cotyledons and leaves at different developmental stages. Clade-specific patterns of inheritance were detected such that hybrid progeny from subgenera Passiflora and Astrophea predominantly inherited paternal plastids with occasional incidences of maternal inheritance, whereas subgenus Decaloba showed predominantly maternal and biparental inheritance. Biparental plastid inheritance was also detected in some hybrids from subgenus Passiflora. Heteroplasmy due to biparental inheritance was restricted to hybrid cotyledons and first leaves with a single parental plastid type detectable in mature plants. This indicates that in Passiflora, plastid retention at later stages of plant development may not reflect the plastid inheritance patterns in embryos. Passiflora exhibits diverse patterns of plastid inheritance, providing an excellent system to investigate underlying mechanisms in angiosperms.

Transcriptional analysis of Rhazya stricta in response to jasmonic acid

BACKGROUND: Jasmonic acid (JA) is a signal transducer molecule that plays an important role in plant development and stress response; it can also efficiently stimulate secondary metabolism in plant cells. RESULTS: RNA-Seq technology was applied to identify differentially expressed genes and study the time course of gene expression in Rhazya stricta in response to JA. Of more than 288 million total reads, approximately 27% were mapped to genes in the reference genome. Genes involved during the secondary metabolite pathways were up- or downregulated when treated with JA in R. stricta. Functional annotation and pathway analysis of all up- and downregulated genes identified many biological processes and molecular functions. Jasmonic acid biosynthetic, cell wall organization, and chlorophyll metabolic processes were upregulated at days 2, 6, and 12, respectively. Similarly, the molecular functions of calcium-transporting ATPase activity, ADP binding, and protein kinase activity were also upregulated at days 2, 6, and 12, respectively. Time-dependent transcriptional gene expression analysis showed that JA can induce signaling in the phenylpropanoid and aromatic acid pathways. These pathways are responsible for the production of secondary metabolites, which are essential for the development and environmental defense mechanism of R. stricta during stress conditions. CONCLUSIONS: Our results suggested that genes involved in flavonoid biosynthesis and aromatic acid synthesis pathways were upregulated during JA stress. However, monoterpenoid indole alkaloid (MIA) was unaffected by JA treatment. Hence, we can postulate that JA plays an important role in R. stricta during plant development and environmental stress conditions.

Extensive variation in nucleotide substitution rate and gene/intron loss in mitochondrial genomes of Pelargonium.

Geraniaceae organelle genomes have been shown to exhibit several highly unusual features compared to most other photosynthetic angiosperms. This includes massively rearranged plastomes with considerable size variation, extensive gene and intron loss, accelerated rates of nucleotide substitutions in both mitogenomes and plastomes, and biparental inheritance and cytonuclear incompatibility of the plastome. Most previous studies have focused on plastome evolution with mitogenome comparisons limited to only a few taxa or genes. In this study, mitogenomes and transcriptomes were examined for 27 species of Geraniales, including 13 species of Pelargonium. Extensive gene and intron losses were detected across the Geraniales with Pelargonium representing the most gene depauperate lineage in the family. Plotting these events on the Geraniaceae phylogenetic tree showed that gene losses occurred multiple times, whereas intron losses more closely reflected the relationships among taxa. In addition, P. australe acquired an intron by horizontal transfer. Comparisons of nucleotide substitution rates in Pelargonium showed that synonymous changes in nuclear genes were much lower than in mitochondrial genes. This is in contrast to the previously published studies that indicated that nuclear genes have 16 fold higher rates than mitochondrial genes across angiosperms. Elevated synonymous substitutions occurred for each mitochondrial gene in Pelargonium with the highest values 783 and 324 times higher than outgroups and other Geraniaceae, respectively. Pelargonium is one of four unrelated genera of angiosperms (Ajuga, Plantago and Silene) that have experienced highly accelerated nucleotide substitutions in mitogenomes. It is distinct from most angiosperms in also having elevated substitution rates in plastid genes but the cause of rate accelerations in Pelargonium plastomes and mitogenomes may be different.

Nucleotide substitution rates of diatom plastid encoded protein genes are positively correlated with genome architecture.

Diatoms are the largest group of heterokont algae with more than 100,000 species. As one of the single-celled photosynthetic organisms that inhabit marine, aquatic and terrestrial ecosystems, diatoms contribute ~ 45% of global primary production. Despite their ubiquity and environmental significance, very few diatom plastid genomes (plastomes) have been sequenced and studied. This study explored patterns of nucleotide substitution rates of diatom plastids across the entire suite of plastome protein-coding genes for 40 taxa representing the major clades. The highest substitution rate was lineage-specific within the araphid 2 taxon Astrosyne radiata and radial 2 taxon Proboscia sp. Rate heterogeneity was also evident in different functional classes and individual genes. Similar to land plants, proteins genes involved in photosynthetic metabolism have lower synonymous and nonsynonymous substitutions rates than those involved in transcription and translation. Significant positive correlations were identified between substitution rates and measures of genomic rearrangements, including indels and inversions, which is a similar result to what was found in legume plants. This work advances the understanding of the molecular evolution of diatom plastomes and provides a foundation for future studies.