Orthologous Pairs Research Articles

A variable gene family encoding nodule-specific cysteine-rich peptides in pea (Pisum sativum L.)

Various legume plants form root nodules in which symbiotic bacteria (rhizobia) fix atmospheric nitrogen after differentiation into a symbiotic form named bacteroids. In some legume species, bacteroid differentiation is promoted by defensin-like nodule-specific cysteine-rich (NCR) peptides. NCR peptides have best been studied in the model legume Medicago truncatula Gaertn., while in many other legumes relevant information is still fragmentary. Here, we characterize the NCR gene family in pea (Pisum sativum L.) using genomic and transcriptomic data. We found 360 genes encoding NCR peptides that are expressed in nodules. The sequences of pea NCR genes and putative peptides are highly variable and differ significantly from NCR sequences of M. truncatula. Indeed, only one pair of orthologs (PsNCR47–MtNCR312) has been identified. The NCR genes in the pea genome are located in clusters, and the expression patterns of NCR genes from one cluster tend to be similar. These data support the idea of independent evolution of NCR genes by duplication and diversification in related legume species. We also described spatiotemporal expression profiles of NCRs and identified specific transcription factor (TF) binding sites in promoters of “early” and “late” NCR genes. Further, we studied the expression of NCR genes in nodules of Fix– mutants and predicted potential regulators of NCR gene expression, one among them being the TF ERN1 involved in the early steps of nodule organogenesis. In general, this study contributes to understanding the functions of NCRs in legume nodules and contributes to understanding the diversity and potential antibiotic properties of pea nodule-specific antimicrobial molecules.

Genome-Wide Identification and Characterization of G2-Like Transcription Factor Genes in Moso Bamboo (Phyllostachys edulis).

G2-like (GLK) transcription factors contribute significantly and extensively in regulating chloroplast growth and development in plants. This study investigated the genome-wide identification, phylogenetic relationships, conserved motifs, promoter cis-elements, MCScanX, divergence times, and expression profile analysis of PeGLK genes in moso bamboo (Phyllostachys edulis). Overall, 78 putative PeGLKs (PeGLK1–PeGLK78) were identified and divided into 13 distinct subfamilies. Each subfamily contains members displaying similar gene structure and motif composition. By synteny analysis, 42 orthologous pairs and highly conserved microsynteny between regions of GLK genes across moso bamboo and maize were found. Furthermore, an analysis of the divergence times indicated that PeGLK genes had a duplication event around 15 million years ago (MYA) and a divergence happened around 38 MYA between PeGLK and ZmGLK. Tissue-specific expression analysis showed that PeGLK genes presented distinct expression profiles in various tissues, and many members were highly expressed in leaves. Additionally, several PeGLKs were significantly up-regulated under cold stress, osmotic stress, and MeJA and GA treatment, implying that they have a likelihood of affecting abiotic stress and phytohormone responses in plants. The results of this study provide a comprehensive understanding of the moso bamboo GLK gene family, as well as elucidating the potential functional characterization of PeGLK genes.

Adaptive radiation in Orinus, an endemic alpine grass of the Qinghai-Tibet Plateau, based on comparative transcriptomic analysis

The species of Orinus (Poaceae) are important alpine plants with a variety of phenotypic traits and potential usages in molecular breeding toward drought-tolerant forage crops. However, the genetic basis of evolutionary adaption and diversification in the genus is still unclear. In the present study, we obtained transcriptomes for the two most divergent species, O. thoroldii and O. kokonoricus, using the Illumina platform and de novo assembly. In total, we generated 23,029 and 24,086 unigenes with N50 values of 1188 and 1203 for O. thoroldii and O. kokonoricus respectively, and identified 19,005 pairs of putative orthologs between the two species of Orinus. For these orthologs, estimations of non-synonymous/synonymous substitution rate ratios indicated that 568 pairs may be under strongly positive selection (Ka/Ks > 1), and Gene Ontogeny (GO) enrichment analysis revealed that significantly enriched pathways were in DNA repair and resistance to abiotic stress. Meanwhile, the divergence times of species between O. thoroldii and O. kokonoricus occurred 3.2 million years ago (Mya), and the recent evolutionary branch is an allotetraploid species, Cleistogenes songorica. We also detected a Ks peak of ∼0.60 for Orinus. Additionally, we identified 188 pairs of differentially expressed genes (DEGs) between the two species of Orinus, which were significantly enrich in stress resistance and lateral root development. Thus, we considered that the species diversification and evolutionary adaption of this genus was initiated by environmental selection, followed by phenotypic differentiation, finally leading to niche separation in the Qinghai-Tibet Plateau.

Genome-wide identification of the pectin methylesterase inhibitor genes in Brassica napus and expression analysis of selected members.

Pectin methylesterase inhibitors (PMEIs) modulate the status of pectin methylesterification by inhibiting the activity of pectin methylesterase (PME). Recent advances indicate PMEIs play an important role in regulating plant cell wall properties and defense responses. In this study, a genome-wide analysis of PMEI gene family in Brassica napus (B. napus) was conducted and the expression patterns of PMEI genes in response to Sclerotinia sclerotiorum (S. sclerotiorum) was investigated. A total of 190 PMEI proteins were identified from the genome of B. napus. Chromosomal location, gene structure and properties of the PMEI family were analyzed, and these features were compared with Arabidopsis thaliana (A. thaliana). A total of 123 syntenic ortholog pairs were detected from BnPMEI family by synteny analysis. Results showed the expansion of BnPMEI genes was likely predominately from whole-genome duplication (WGD) or segmental duplications. Multiple cis-elements related to plant growth and development, environmental stress responses, hormone responses were detected in the promoters of BnPMEI genes, implying they were regulated by both internal and external factors. Furthermore, expression analysis of transcriptome data combined with quantitative RT-PCR (qRT-PCR) validation identified several candidates that were strongly responsive to S. sclerotiorum infection. These BnPMEI genes are candidates for manipulation to breed novel and improved genotypes that are more resistant to sclerotinia stem rot (SSR). Extensive interactions were detected among 30 BnPMEI proteins, forming complex protein-protein interaction networks. Besides, 48 BnPMEIs showed interactions with other proteins including a range of cell wall structure-related enzymes. This study provides new insights into the evolution and function of PMEIs in B. napus and lays a foundation for breeding novel genotypes for crop improvement.

Genome-Wide Identification and Expression Analysis of WRKY Genes during Anthocyanin Biosynthesis in the Mango (Mangifera indica L.)

The WRKY family is one of the largest transcription factor (TF) families in plants and is involved in the regulation of plant physiological processes, such as anthocyanin accumulation. However, little information is known regarding the WRKY genes in the mango. In this study, a total of 87 mango WRKY genes were identified and named MiWRKY1 to MiWRKY87. Phylogenetic results showed that the 87 MiWRKYs could be divided into three groups (I, II, III) and five subgroups of group II (II-a, II-b, II-c, II-d, II-e), with high similarity in exon–intron structures and WRKY domain and motif compositions within the same group and subgroup. One tandem duplication (MiWRKY76 and MiWRKY82) and 97 pairs of segmental duplicates were identified in the mango genome. Syntenic analysis showed that mango MiWRKY genes had 52 and 69 orthologous pairs with Arabidopsis and citrus, respectively. Promoter cis-acting element analysis revealed that MiWRKYs contain a large number of elements associated with light signaling, hormonal response, environmental stress, and plant development. Tissue specific expression profiles showed that the expression of MiWRKY genes displayed tissue preference. Quantitative-PCR analysis showed that high expression levels of MiWRKY1, MiWRKY3, MiWRKY5, MiWRKY81, and MiWRKY84 were detected in the skin of red mango cultivar, and the expressions of MiWRKY1 and MiWRKY81 were up-regulated during light-induced anthocyanin accumulation in the mango, indicating these genes might regulate anthocyanin biosynthesis in the mango. This study provides comprehensive genetic information on the MiWRKYs in mango fruit.

Comparative Transcriptomics and Metabolites Analysis of Two Closely Related Euphorbia Species Reveal Environmental Adaptation Mechanism and Active Ingredients Difference.

Roots of Euphorbia fischeriana and Euphorbia ebracteolata are recorded as the source plant of traditional Chinese medicine “Langdu,” containing active ingredients with anticancer and anti-AIDS activity. However, the two species have specific patterns in the graphic distribution. Compared with E. ehracteolata, E. fischeriana distributes in higher latitude and lower temperature areas and might have experienced cold stress adaptation. To reveal the molecular mechanism of environmental adaptation, RNA-seq was performed toward the roots, stems, and leaves of E. fischeriana and E. ehracteolata. A total of 6,830 pairs of putative orthologs between the two species were identified. Estimations of non-synonymous or synonymous substitution rate ratios for these orthologs indicated that 533 of the pairs may be under positive selection (Ka/Ks > 0.5). Functional enrichment analysis revealed that significant proportions of the orthologs were in the TCA cycle, fructose and mannose metabolism, starch and sucrose metabolism, fatty acid biosynthesis, and terpenoid biosynthesis providing insights into how the two closely related Euphorbia species adapted differentially to extreme environments. Consistent with the transcriptome, a higher content of soluble sugars and proline was obtained in E. fischeriana, reflecting the adaptation of plants to different environments. Additionally, 5 primary or secondary metabolites were screened as the biomarkers to distinguish the two species. Determination of 4 diterpenoids was established and performed, showing jolkinolide B as a representative component in E. fischeriana, whereas ingenol endemic to E. ebracteolate. To better study population genetics, EST-SSR markers were generated and tested in 9 species of Euphorbia. A total of 33 of the 68 pairs were screened out for producing clear fragments in at least four species, which will furthermore facilitate the studies on the genetic improvement and phylogenetics of this rapidly adapting taxon. In this study, transcriptome and metabolome analyses revealed the evolution of genes related to cold stress tolerance, biosynthesis of TCA cycle, soluble sugars, fatty acids, and amino acids, consistent with the molecular strategy that genotypes adapting to environment. The key active ingredients of the two species were quantitatively analyzed to reveal the difference in pharmacodynamic substance basis and molecular mechanism, providing insights into rational crude drug use.

Structure and Mechanism of a Cold-Adapted Bacterial Lipase.

The structural origin of enzyme cold-adaptation has been the subject of considerable research efforts in recent years. Comparative studies of orthologous mesophilic–psychrophilic enzyme pairs found in nature are an obvious strategy for solving this problem, but they often suffer from relatively low sequence identity of the enzyme pairs. Small bacterial lipases adapted to distinctly different temperatures appear to provide an excellent model system for these types of studies, as they may show a very high degree of sequence conservation. Here, we report the first crystal structures of lipase A from the psychrophilic bacterium Bacillus pumilus, which confirm the high structural similarity to the mesophilic Bacillus subtilis enzyme, as indicated by their 81% sequence identity. We further employ extensive QM/MM calculations to delineate the catalytic reaction path and its energetics. The computational prediction of a rate-limiting deacylation step of the enzymatic ester hydrolysis reaction is verified by stopped-flow experiments, and steady-state kinetics confirms the psychrophilic nature of the B. pumilus enzyme. These results provide a useful benchmark for examining the structural basis of cold-adaptation and should now make it possible to disentangle the effects of the 34 mutations between the two enzymes on catalytic properties and thermal stability.

Genome wide characterization of R2R3 MYB transcription factor from Apocynum venetum revealed potential stress tolerance and flavonoid biosynthesis genes.

MYB transcription factors are crucial in regulating stress tolerance and expression of major genes involved in flavonoid biosynthesis. The functions of MYBs is well explored in a number of plants, yet no study is reported in Apocynum venetum. We identified a total of 163 MYB candidates, that comprised of 101 (61.96%) R2R3, 6 3R, 1 4R and 55 1R. Syntenic analysis of A. venetum R2R3 (AvMYBs) showed highest orthologous pairs with Vitis vinifera MYBs followed by Arabidopsis thaliana among the four species evaluated. Thirty segmental duplications and 6 tandem duplications were obtained among AvMYB gene pairs signifying their role in the MYB gene family expansion. Nucleotide substitution analysis (Ka/Ks) showed the AvMYBs to be under the influence of strong purifying selection. Expression analysis of selected AvMYBs under low temperature and cadmium stresses resulted in the identification of AvMYB48, AvMYB97, AvMYB8, AvMYB4 as potential stress responsive genes and AvMYB10 and AvMYB11 in addition, proanthocyanidin biosynthesis regulatory genes which is consistent with their annotated homologues in Arabidopsis. Tissue specific expression profile analysis of the AvMYBs further supported the qPCR analysis result. MYBs with higher transcript levels in root, stem and leaf like AvMYB4 for example, was downregulated under the stresses and such with low transcript level such as AvMYB48 which had low transcript in the leaf was upregulated under both stresses. Transcriptome and phylogenetic analyses suggested AvMYB42 as a potential regulator of anthocyanin biosynthesis. Thus, this study provided valuable information on AvR2R3-MYB gene family with respect to stress tolerance and flavonoid biosynthesis.

Satellitome comparison of two oedipodine grasshoppers highlights the contingent nature of satellite DNA evolution

BackgroundThe full catalog of satellite DNA (satDNA) within a same genome constitutes the satellitome. The Library Hypothesis predicts that satDNA in relative species reflects that in their common ancestor, but the evolutionary mechanisms and pathways of satDNA evolution have never been analyzed for full satellitomes. We compare here the satellitomes of two Oedipodine grasshoppers (Locusta migratoria and Oedaleus decorus) which shared their most recent common ancestor about 22.8 Ma ago.ResultsWe found that about one third of their satDNA families (near 60 in every species) showed sequence homology and were grouped into 12 orthologous superfamilies. The turnover rate of consensus sequences was extremely variable among the 20 orthologous family pairs analyzed in both species. The satDNAs shared by both species showed poor association with sequence signatures and motives frequently argued as functional, except for short inverted repeats allowing short dyad symmetries and non-B DNA conformations. Orthologous satDNAs frequently showed different FISH patterns at both intra- and interspecific levels. We defined indices of homogenization and degeneration and quantified the level of incomplete library sorting between species.ConclusionsOur analyses revealed that satDNA degenerates through point mutation and homogenizes through partial turnovers caused by massive tandem duplications (the so-called satDNA amplification). Remarkably, satDNA amplification increases homogenization, at intragenomic level, and diversification between species, thus constituting the basis for concerted evolution. We suggest a model of satDNA evolution by means of recursive cycles of amplification and degeneration, leading to mostly contingent evolutionary pathways where concerted evolution emerges promptly after lineages split.

The Legionella genus core effectors display functional conservation among orthologs by themselves or combined with an accessory protein.

The intracellular pathogen Legionella pneumophila, as well as other Legionella species, utilize the Icm/Dot type-IV secretion system to translocate an exceptionally large and diverse repertoire of effectors into their host cells. However, only nine core effectors were found to be present in all analyzed Legionella species. In this study, we investigated the core effectors, and used intracellular growth complementation to determine whether orthologs of core effectors perform the same function in different Legionella species. We found that three out of the nine L. pneumophila core effectors are required for maximal intracellular growth. Examination of orthologous core effectors from four Legionella species spread over the Legionella phylogenetic tree revealed that most of them perform the same function. Nevertheless, some of the orthologs of the core effector LegA3 did not complement the L. pneumophila legA3 deletion mutant for intracellular growth. LegA3 is encoded as part of an operon together with another gene, which we named legA3C, encoding a non-translocated protein. We found that LegA3 and LegA3C physically interact with each other, are both required for maximal intracellular growth, and the LegA3-LegA3C orthologous pairs from all the Legionella species examined fully complement the L. pneumophila legA3 deletion mutant for intracellular growth. Our results indicate that the Legionella core effectors orthologs generally perform the same function and establish that LegA3 requires LegA3C to fulfill its conserved function.

Charge reversal mutations in mesophilic-thermophilic orthologous protein pairs and their role in enhancing coulombic interaction energy

Proteins from thermophilic organisms are a matter of immense interest for decades because of its application in fields like de-novo protein design, thermostable variants of biocatalysts etc. Previous studies have found several sequence and structural adaptations related to thermal stability, while charge reversal study remains ignored. Here we address whether charge reversal mutations naturally occur in mesophilic-thermophilic/hyperthermophilic orthologous proteins. Do they contribute to thermal stability? Our systematic study on 1550 mesophilic-thermophilic/hyperthermophilic orthologous protein pairs with remarkable structural and topological similarity, shows gain in coulombic interaction energy in thermophilic/hyperthermophilic proteins at short range associated with partially exposed and buried charge reversal mutations, which may enhance thermostability. Our findings call forth its application in future protein engineering studies. Communicated by Ramaswamy H. Sarma

Transcriptomic evidence for dense peptidergic networks within forebrains of four widely divergent tetrapods.

The primary function common to every neuron is communication with other neurons. Such cell-cell signaling can take numerous forms, including fast synaptic transmission and slower neuromodulation via secreted messengers, such as neuropeptides, dopamine, and many other diffusible small molecules. Individual neurons are quite diverse, however, in all particulars of both synaptic and neuromodulatory communication. Neuron classification schemes have therefore proven very useful in exploring the emergence of network function, behavior, and cognition from the communication functions of individual neurons. Recently published single-cell mRNA sequencing data and corresponding transcriptomic neuron classifications from turtle, songbird, mouse, and human provide evidence for a long evolutionary history and adaptive significance of localized peptidergic signaling. Across all four species, sets of at least twenty orthologous cognate pairs of neuropeptide precursor protein and receptor genes are expressed in individually sparse but heavily overlapping patterns suggesting that all forebrain neuron types are densely interconnected by local peptidergic signals.

Systematic genome-wide analysis of the ethylene-responsive ACS gene family: Contributions to sex form differentiation and development in melon and watermelon.

Ethylene is an important regulatory phytohormone for sex differentiation and flower development. As the rate-limiting enzyme encoding genes in ethylene biosynthesis, ACS gene family has been well studied in cucumber; however, little is known in other cucurbit crops, such as melon and watermelon, which show diverse sex types in the field. Here, we identified and characterized eight ACS genes each in the genomes of melon and watermelon. According to the conserved serine residues at C-terminal, all the ACS genes could be characterized into three groups, which were supported by the exon-intron organizations and conserved motif distributions. ACS genes displayed diverse tissue-specific expression patterns among four melon and three watermelon sex types. Furthermore, a comparative expression analysis in the shoot apex identified orthologous pairs with potential functions in sex determination, e.g., ACS1s and ACS6s. All ACS orthologs in melon and watermelon exhibited similar expression patterns in monoecious and gynoecious genotypes, except for ACS11s and ACS12s. As expected, the majority of ACS genes were responsive to exogenous ethephon; however, some orthologs exhibited opposite expression patterns, such as ACS1s, ACS9s, and ACS10s. Collectively, our findings provide valuable ACS candidates related to flower development in various sex types of melon and watermelon.

Identification and expression analysis of candidate genes related to seed dormancy and germination in the wheat GATA family

GATA transcription factors have been reported to function in plant growth and development and during various biotic/abiotic stresses in Arabidopsis and rice. However, the functions of wheat GATAs, particularly in the regulation of seed dormancy and germination, remain unclear. Here, we identified 78 TaGATAs in wheat and divided them into five subfamilies. Sixty-four paralogous pairs and 52 orthologous pairs were obtained, and Ka/Ks ratios showed that the TaGATAs had undergone strong purifying election during the evolutionary process. Triplet analysis indicated that a high homologue retention rate could explain the large number of TaGATAs in wheat. Gene structure analysis revealed that most members of the same subfamily had similar structures, and subcellular localization prediction indicated that most TaGATAs were located in the nucleus. Gene ontology annotation results showed that most TaGATAs had molecular functions in DNA and zinc binding, and promoter analysis suggested that they may play important roles in growth, development, and biotic/abiotic stress response. We combined three microarray datasets with qRT-PCR expression data from wheat varieties of contrasting dormancy and pre-harvest sprouting resistance levels during imbibition in order to identify ten candidate genes (TaGATA17/-25/-34/-37/-40/-46/-48/-51/-72/-73) that may be involved in the regulation of seed dormancy and germination in wheat. These findings provide valuable information for further dissection of TaGATA functions in the regulation of seed dormancy and germination, thereby enabling the improvement of wheat pre-harvest sprouting resistance by gene pyramiding.

Comprehensive Comparative Analysis of the GATA Transcription Factors in Four Rosaceae Species and Phytohormonal Response in Chinese Pear (Pyrus bretschneideri) Fruit.

The GATA gene family is one of the most important transcription factors (TFs). It extensively exists in plants, contributes to diverse biological processes such as the development process, and responds to environmental stress. Although the GATA gene family has been comprehensively and systematically studied in many species, less is known about GATA genes in Chinese pears (Pyrus bretschneideri). In the current study, the GATA gene family in the four Rosaceae genomes was identified, its structural characteristics identified, and a comparative analysis of its properties was carried out. Ninety-two encoded GATA proteins were authenticated in the four Rosaceae genomes (Pyrus bretschneideri, Prunus avium, Prunus mume, and Prunus persica) and categorized into four subfamilies (Ⅰ–Ⅳ) according to phylogeny. The majority of GATA genes contained one to two introns and conserved motif composition analysis revealed their functional divergence. Whole-genome duplications (WGDs) and dispersed duplication (DSD) played a key role in the expansion of the GATA gene family. The microarray indicated that, among P. bretschneideri, P. avium, P. mume and P. persica, GATA duplicated regions were more conserved between Pyrus bretschneideri and Prunus persica with 32 orthologous genes pairs. The physicochemical parameters, duplication patterns, non-synonymous (ka), and synonymous mutation rate (ks) and GO annotation ontology were performed using different bioinformatics tools. cis-elements respond to various phytohormones, abiotic/biotic stress, and light-responsive were found in the promoter regions of GATA genes which were induced via stimuli. Furthermore, subcellular localization of the PbGATA22 gene product was investigated, showing that it was present in the nucleus of tobacco (Nicotiana tabacum) epidermal cells. Finally, in silico analysis was performed on various organs (bud, leaf, stem, ovary, petal, and sepal) and different developmental stages of fruit. Subsequently, the expression profiles of PbGATA genes were extensively expressed under exogenous hormonal treatments of SA (salicylic acid), MeJA (methyl jasmonate), and ABA (abscisic acid) indicating that play important role in hormone signaling pathways. A comprehensive analysis of GATA transcription factors was performed through systematic biological approaches and comparative genomics to establish a theoretical base for further structural and functional investigations in Rosaceae species.

Transcriptomic analysis reveals innate immune mechanisms of an underlying parasite-resistant grouper hybrid (Epinephelus fuscogutatus × Epinephelus lanceolatus)

Hybridization is an artificial breeding strategy for generating potentially desirable offspring. Recently, a novel Hulong grouper hybrid (Epinephelus fuscogutatus × Epinephelus lanceolatus) yielded significant growth superiority over its parent. Improved innate immunity is considered as another desirable feature during hybridization. However, whether this Hulong grouper achieved disease resistance has not yet been revealed. In this study, we first examine the infection intensity of C. irritans in the Hulong grouper, and found that the Hulong grouper is less susceptible to C. irritans primary infection. A higher immobilization titer was found in the infected Hulong grouper at Day 2 when compared with the control grouper. Furthermore, severe hyperplasia was observed in the orange-spotted grouper, but not in the Hulong grouper's skin epidermis. To further understand the innate immune mechanism against C. irritans, we conducted a comparative transcriptome analysis of the Hulong grouper during the infection. There are 6464 differentially expressed genes (DEGs) identified in the skin between the control and infected Hulong grouper. This indicates that the innate immune components, such as the complement system, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, Interleukin 17 (IL-17) signaling pathway, and Toll-like receptor (TLR) signaling pathway were up-regulated during the infection. These results show that the C. irritans infection can induce a remarkable inflammatory response in the Hulong grouper. Moreover, a total of 75 pairs of orthologs with the ratio of nonsynonymous (Ka) to synonymous (Ks) substitutions ＞1, considered rapidly evolving genes (REGs), was identified between the Hulong and orange-spotted grouper. More critically, most REGs were enriched in the immune system, suggesting that rapid evolution of the immune system might occur in the Hulong grouper. These results provide a more comprehensive understanding of the innate immunity mechanism of the hybrid Hulong grouper.

Genome-Wide Comparative Analysis of R2R3 MYB Gene Family in Populus and Salix and Identification of Male Flower Bud Development-Related Genes.

The MYB transcription factor (TF) family is one of the largest plant transcription factor gene family playing vital roles in plant growth and development, including defense, cell differentiation, secondary metabolism, and responses to biotic and abiotic stresses. As a model tree species of woody plants, in recent years, the identification and functional prediction of certain MYB family members in the poplar genome have been reported. However, to date, the characterization of the gene family in the genome of the poplar’s sister species willow has not been done, nor are the differences and similarities between the poplar and willow genomes understood. In this study, we conducted the first genome-wide investigation of the R2R3 MYB subfamily in the willow, identifying 216 R2R3 MYB gene members, and combined with the poplar R2R3 MYB genes, performed the first comparative analysis of R2R3 MYB genes between the poplar and willow. We identified 81 and 86 pairs of R2R3 MYB paralogs in the poplar and willow, respectively. There were 17 pairs of tandem repeat genes in the willow, indicating active duplication of willow R2R3 MYB genes. A further 166 pairs of poplar and willow orthologs were identified by collinear and synonymous analysis. The findings support the duplication of R2R3 MYB genes in the ancestral species, with most of the R2R3 MYB genes being retained during the evolutionary process. The phylogenetic trees of the R2R3 MYB genes of 10 different species were drawn. The functions of the poplar and willow R2R3 MYB genes were predicted using reported functional groupings and clustering by OrthoFinder. Identified 5 subgroups in general expanded in woody species, three subgroups were predicted to be related to lignin synthesis, and we further speculate that the other two subgroups also play a role in wood formation. We analyzed the expression patterns of the GAMYB gene of subgroup 18 (S18) related to pollen development in the male flower buds of poplar and willow at different developmental stages by qRT-PCR. The results showed that the GAMYB gene was specifically expressed in the male flower bud from pollen formation to maturity, and that the expression first increased and then decreased. Both the specificity of tissue expression specificity and conservation indicated that GAMYB played an important role in pollen development in both poplar and willow and was an ideal candidate gene for the analysis of male flower development-related functions of the two species.

Multi-tissue transcriptome analysis of two Begonia species reveals dynamic patterns of evolution in the chalcone synthase gene family

Begonia is an important horticultural plant group, as well as one of the most speciose Angiosperm genera, with over 2000 described species. Genus wide studies of genome size have shown that Begonia has a highly variable genome size, and analysis of paralog pairs has previously suggested that Begonia underwent a whole genome duplication. We address the contribution of gene duplication to the generation of diversity in Begonia using a multi-tissue RNA-seq approach. We chose to focus on chalcone synthase (CHS), a gene family having been shown to be involved in biotic and abiotic stress responses in other plant species, in particular its importance in maximising the use of variable light levels in tropical plants. We used RNA-seq to sample six tissues across two closely related but ecologically and morphologically divergent species, Begonia conchifolia and B. plebeja, yielding 17,012 and 19,969 annotated unigenes respectively. We identified the chalcone synthase gene family members in our Begonia study species, as well as in Hillebrandia sandwicensis, the monotypic sister genus to Begonia, Cucumis sativus, Arabidopsis thaliana, and Zea mays. Phylogenetic analysis suggested the CHS gene family has high duplicate turnover, all members of CHS identified in Begonia arising recently, after the divergence of Begonia and Cucumis. Expression profiles were similar within orthologous pairs, but we saw high inter-ortholog expression variation. Sequence analysis showed relaxed selective constraints on some ortholog pairs, with substitutions at conserved sites. Evidence of pseudogenisation and species specific duplication indicate that lineage specific differences are already beginning to accumulate since the divergence of our study species. We conclude that there is evidence for a role of gene duplication in generating diversity through sequence and expression divergence in Begonia.

Genome-wide investigation of the bZIP transcription factor gene family in Prunus mume: Classification, evolution, expression profile and low-temperature stress responses

Prunus mume is an important woody plant that has high ornamental and economic value, widely distributed and used in landscape architecture in East Asia. In plants, basic (region) leucine zipper (bZIP) transcription factors play important regulatory roles in growth, development, dormancy and abiotic stress. To date, bZIP transcription factors have not been systematically studied in P. mume. In this study, 49 bZIP genes were first identified in P. mume, and the PmbZIP family was divided into 12 groups according to the grouping principles for the Arabidopsis thaliana bZIP family. For the first time, we constructed a detailed model of the PmbZIP domains (R-x3N-x7-R/K-x2-K-x6-L-x6-L-x6-L). Phylogenetic and synteny analyses showed that PmbZIPs duplication events might have occurred during the large-scale genome duplication events. A relatively short time of speciation and the finding that 91.84% of the bZIP genes formed orthologous pairs between P. mume and Prunus armeniaca provided evidence of a close relationship. Gene expression patterns were analysed in different tissues and periods, indicating that PmbZIP genes with the same motifs exhibited similar expression patterns. The gene expression results showed that PmbZIP31/36/41 genes played a more prominent role in the response to freezing stress than cold stress. The expression level of almost all subset III genes was upregulated under freezing treatment, especially after cold exposure. We analysed the gene expression patterns of PmbZIP12/31/36/41/48 and their responses to low-temperature stress, which provided useful resources for future studies on the cold/freezing-tolerant molecular breeding of P. mume.

Genome-Wide Investigation of Spliceosomal SM/LSM Genes in Wheat (Triticum aestivum L.) and Its Progenitors

The SSM/SLSM (spliceosomal Smith (SM)/SM-like (LSM)) genes are the central components of the spliceosome in eukaryotes, which play an important role in regulating RNA splicing, participating in diverse biological processes. Although it has been detected in Arabidopsis and rice etc. plants, the members and significance of the SSM/SLSM gene family in wheat are still not reported. In this study, we identified the SSM/SLSM genes in wheat and its progenitors at genome-scale, where 57 SSM/SLSM genes were identified in wheat, together with 41, 17and 19 found in Triticum dicoccoides, Triticum urartu, and Aegilops tauschii. Furthermore, their phylogenetic relationship, gene structures, conserved motifs, and cis-regulatory elements were systematically analyzed. By synteny analysis, good collinearity of SSM/SLSM genes was found among bread wheat and its progenitors’ genomes, and the distribution of SMD2 genes in wheat chromosome 5A, 4B and 4D located in the 4AL-5AL-7BS chromosome model, due to the translocation. Then, the positively selected genes were further investigated based on the non-synonymous to synonymous (dN/dS) analysis of the orthologous pairs. Finally, the expression profiles of the SSM/SLSM genes were detected using RNA-seq datasets, and eight stress-responsive candidate genes were selected to validate their expression through qPCR (real-time quantitative polymerase chain reaction). According to the co-expression network analysis, the correlation between the LSM7-7A gene and related genes was illustrated through Gene Ontology (GO) enrichment analysis. Furthermore, the LSM7-7A gene was related to the Arabidopsis homologous salt tolerance gene RCY1. This investigation systematically identified the complete candidates of SSM/SLSM genes and their characters in wheat and its progenitors, and provided clues to a better understanding of their contribution during the wheat polyploidy process.