A new species of Populus and the extensive hybrid speciation arising from it on the Qinghai-Tibet Plateau.

While the diversity of species formation is broadly acknowledged, significant debate exists regarding the universal nature of hybrid species formation. Through an 18-year comprehensive study of all Populus species on the Qinghai-Tibet Plateau, 23 previously recorded species and 8 new species were identified. Based on morphological characteristics, these can be classified into three groups: species in section Leucoides, species with large leaves, and species with small leaves in section Tacamahaca. By conducting whole-genome re-sequencing of 150 genotypes from these 31 species, 2.28 million single nucleotide polymorphisms (SNPs) were identified. Phylogenetic analysis utilizing these SNPs not only revealed a highly intricate evolutionary network within the large-leaf species of section Tacamahaca but also confirmed that a new species, P. curviserrata, naturally hybridized with P. cathayana, P. szechuanica, and P. ciliata, resulting in 11 hybrid species. These findings indicate the widespread occurrence of hybrid species formation within this genus, with hybridization serving as a key evolutionary mechanism for Populus on the plateau. A novel hypothesis, "Hybrid Species Exterminating Their Ancestral Species (HSEAS)," is introduced to explain the mechanisms of hybrid species formation at three different scales: the entire plateau, the southeastern mountain region, and individual river valleys.

Drivers of genomic landscapes of differentiation across a Populus divergence gradient.

Speciation, the continuous process by which new species form, is often investigated by looking at the variation of nucleotide diversity and differentiation across the genome (hereafter genomic landscapes). A key challenge lies in how to determine the main evolutionary forces at play shaping these patterns. One promising strategy, albeit little used to date, is to comparatively investigate these genomic landscapes as progression through time by using a series of species pairs along a divergence gradient. Here, we resequenced 201 whole-genomes from eight closely related Populus species, with pairs of species at different stages along the divergence gradient to learn more about speciation processes. Using population structure and ancestry analyses, we document extensive introgression between some species pairs, especially those with parapatric distributions. We further investigate genomic landscapes, focusing on within-species (i.e. nucleotide diversity and recombination rate) and among-species (i.e. relative and absolute divergence) summary statistics of diversity and divergence. We observe relatively conserved patterns of genomic divergence across species pairs. Independent of the stage across the divergence gradient, we find support for signatures of linked selection (i.e. the interaction between natural selection and genetic linkage) in shaping these genomic landscapes, along with gene flow and standing genetic variation. We highlight the importance of investigating genomic patterns on multiple species across a divergence gradient and discuss prospects to better understand the evolutionary forces shaping the genomic landscapes of diversity and differentiation.

Molecular Karyotyping on Populus simonii × P. nigra and the Derived Doubled Haploid.

The molecular karyotype could represent the basic genetic make-up in a cell nucleus of an organism or species. A doubled haploid (DH) is a genotype formed from the chromosome doubling of haploid cells. In the present study, molecular karyotype analysis of the poplar hybrid Populus simonii × P. nigra (P. xiaohei) and the derived doubled haploids was carried out with labeled telomeres, rDNA, and two newly repetitive sequences as probes by fluorescence in situ hybridization (FISH). The tandem repeats, pPC349_XHY and pPD284_XHY, with high-sequence homology were used, and the results showed that they presented the colocalized distribution signal in chromosomes. For P. xiaohei, pPD284_XHY produced hybridizations in chromosomes 1, 5, 8, and 9 in the hybrid. The combination of pPD284_XHY, 45S rDNA, and 5S rDNA distinctly distinguished six pairs of chromosomes, and the three pairs of chromosomes showed a significant difference in the hybridization between homologous chromosomes. The repeat probes used produced similar FISH hybridizations in the DH; nevertheless, pPD284_XHY generated an additional hybridization site in the telomere region of chromosome 14. Moreover, two pairs of chromosomes showed differential hybridization distributions between homologous chromosomes. Comparisons of the distinguished chromosomes between hybrid and DH poplar showed that three pairs of chromosomes in the DH presented hybridization patterns that varied from those of the hybrid. The No. 8 chromosome in DH and one of the homologous chromosomes in P. xiaohei shared highly similar FISH patterns, which suggested the possibility of intact or mostly partial transfer of the chromosome between the hybrid and DH. Our study will contribute to understanding the genetic mechanism of chromosomal variation in P. xiaohei and derived DH plants.

Genome-Wide Comprehensive Analysis of the GASA Gene Family in Populus.

Gibberellic acid-stimulated Arabidopsis (GASA) proteins, as cysteine-rich peptides (CRPs), play roles in development and reproduction and biotic and abiotic stresses. Although the GASA gene family has been identified in plants, the knowledge about GASAs in Populus euphratica, the woody model plant for studying abiotic stress, remains limited. Here, we referenced the well-sequenced Populus trichocarpa genome, and identified the GASAs in the whole genome of P. euphratica and P. trichocarpa. 21 candidate genes in P. trichocarpa and 19 candidate genes in P. euphratica were identified and categorized into three subfamilies by phylogenetic analysis. Most GASAs with signal peptides were located extracellularly. The GASA genes in Populus have experienced multiple gene duplication events, especially in the subfamily A. The evolution of the subfamily A, with the largest number of members, can be attributed to whole-genome duplication (WGD) and tandem duplication (TD). Collinearity analysis showed that WGD genes played a leading role in the evolution of GASA genes subfamily B. The expression patterns of P. trichocarpa and P. euphratica were investigated using the PlantGenIE database and the real-time quantitative PCR (qRT-PCR), respectively. GASA genes in P. trichocarpa and P. euphratica were mainly expressed in young tissues and organs, and almost rarely expressed in mature leaves. GASA genes in P. euphratica leaves were also widely involved in hormone responses and drought stress responses. GUS activity assay showed that PeuGASA15 was widely present in various organs of the plant, especially in vascular bundles, and was induced by auxin and inhibited by mannitol dramatically. In summary, this present study provides a theoretical foundation for further research on the function of GASA genes in P. euphratica.

Combined transcriptome and proteome analysis provides insights into anthocyanin accumulation in the leaves of red-leaved poplars.

KEY MESSAGE: Anthocyanin was highly accumulated in the leaves of red-leaved poplars; Many structural genes involved in anthocyanin synthesis were significantly up-regulated in 'Quanhong' and 'Xuanhong'; TTG2, HYH, and HY5 may be directly involved in the regulation of anthocyanin synthesis in both red-leaved poplars. The red-leaved poplar cultivars 'Quanhong' and 'Xuanhong' are bud mutations of Populus deltoides cv. 'Zhonglin 2025'. These cultivars are valued for their beautiful shape, lack of flying catkins, and ornamental leaf colors. However, the understanding of the molecular mechanism of anthocyanin accumulation in the leaves of red-leaved poplars is still unclear. Here, we profiled the changes of pigment content, transcriptome and proteome expression in the leaves of three poplar cultivars and the results showed that the ratios of anthocyanin to total chlorophyll in both red-leaved poplars were higher than that in 'Zhonglin 2025', indicating that the anthocyanin was highly accumulated in the leaves of red-leaved poplars. Based on the results of combined transcriptome and proteome analysis, 15 and 11 differentially expressed genes/proteins involved in anthocyanin synthesis were screened in 'Quanhong' and 'Xuanhong', respectively, indicating that the two red-leaved poplar cultivars have slightly different patterns of regulating anthocyanin biosynthesis. Among the 120 transcription factors, 3 (HY5, HYH, and TTG2), may be directly involved in the regulation of anthocyanin synthesis in both red-leaved poplars. This study screens the candidate genes involved in anthocyanin accumulation in the leaves of red-leaved poplars and lays a foundation for further exploring the molecular mechanism of leaf red coloration in red-leaved poplars.

Genome-wide identification of polar auxin transporter gene families reveals a possible new polar auxin flow in inverted cuttings of Populus yunnanensis.

Inverted cuttings of Populus yunnanensis are characterized by enlarged stems and dwarfed new shoots, and phytohormones play a crucial role in the response to inversion. The polar auxin transport (PAT) system is distinct from the transport systems of other hormones and is controlled by three major transporter gene families: pin-formed (PIN), auxin-resistant/like aux (AUX/LAX) and ATP-binding cassette transporters of the B class (ABCB). Here, we identified these three families in P. trichocarpa, P. euphratica and P. yunnanensis through a genome-wide analysis. The Populus PIN, AUX/LAX and ABCB gene families comprised 15, 8 and 31 members, respectively. Most PAT genes in Populus and Arabidopsis were identified as clear sister pairs, and some had unique motifs. Transcriptome profiling revealed that the expression of most PAT genes was unrelated to cutting inversion and that only several genes showed altered expression when cuttings were inverted. The auxin content difference at positions was opposite in upright and inverted cutting bodies during rooting, which obeyed the original plant polarity. However, during plant growth, the two direction types exhibited similar auxin movements in the cutting bodies, and the opposite auxin changes were observed in new shoots. Four PAT genes with a positive response to cutting inversion, PyuPIN10, PyuPIN11, PyuLAX6 and PyuABCB27, showed diverse expression patterns between upright and inverted cuttings during rooting and plant growth. Furthermore, PAT gene expression retained its polarity, which differs from the results found for auxin flow during plant growth. The inconformity indicated that a new downward auxin flow in addition to the old upward flow might be established during the growth of inverted cuttings. Some highly polar PAT genes were involved in the maintenance of original auxin polarity, which might cause the enlarged stems of inverted cuttings. This work lays a foundation for understanding the roles of auxin transport in plant responses to inversion.

Roots play a key role in drought-tolerance of poplars as suggested by reciprocal grafting between male and female clones.

Drought stress influences the growth of plants and thus grafting has been widely used to improve tolerance to abiotic stresses. Poplars possess sex-specific responses to drought stress, but how male or female rootstock affect the grafted plant is little known. To explore the mechanisms underlying changes in drought tolerance caused by grafting, we investigated the changes in growth, leaf traits, gas exchange and antioxidant enzyme activities of reciprocally grafted seedlings between Populus euramericana cv. "Nanlin895" (NL-895) (female) and Populus deltiodes cv."3412" (NL-3412) (male) under water deficit stress with 30% field capacity for 30 d. Results showed that drought stress affected adversely growth, morphological, and physiological characteristics in all seedlings studied. Grafted seedlings with male roots can effectively alleviated the inhibition of growth induced by drought stress, as shown by higher WUE, activities of SOD, POD and CAT, and lower levels of lipid peroxidation. Male seedlings with female roots were found to be less tolerance to drought than non-grafted male clones and female scions with male roots, but more tolerance than non-grafted female clones. This results suggested that drought tolerance of grafted seedlings is primarily caused by the rootstock, although the scion also affects the grafted plant. Thus, paying attention on the root genotype can provide an important means of improving the drought tolerance of poplars.

Chloroplast (Cp) Transcriptome of P. davidiana Dode×P. bolleana Lauch provides insight into the Cp drought response and Populus Cp phylogeny.

BACKGROUND: Raw second-generation (2G) lignocellulosic biomass materials have the potential for development into a sustainable and renewable source of energy. Poplar is regarded as a promising 2G material (P. davidiana Dode×P. bolleana Lauch, P. bolleana, P. davidiana, P. euphratica, et al). However, their large-scale commercialization still faces many obstacles. For example, drought prevents sufficient irrigation or rainfall, which can reduce soil moisture and eventually destroy the chloroplast, the plant photosynthetic organelle. Heterosis is widely used in the production of drought-tolerant materials, such as the superior clone "Shanxinyang" selected from the offspring of Populus davidiana Dode×Populus bolleana Lauch. Because it produces good wood and is easily genetically transformed, "Shanxinyang" has become a promising material for use in tree genetics. It is also one of the most abundant biofuel plants in northern China. Understanding the genetic features of chloroplasts, the cp transcriptome and physiology is crucial to elucidating the chloroplast drought-response model. RESULTS: In this study, the whole genome of "Shanxinyang" was sequenced. The chloroplast genome was assembled, and chloroplast structure was analysed and compared with that of other popular plants. Chloroplast transcriptome analysis was performed under drought conditions. The total length of the "Shanxinyang" chloroplast genome was 156,190 bp, the GC content was 36.75%, and the genome was composed of four typical areas (LSC, IRa, IRb, and SSC). A total of 114 simple repeats were detected in the chloroplast genome of "Shanxinyang". In cp transcriptome analysis, we found 161 up-regulated and 157 down-regulated genes under drought, and 9 cpDEGs was randomly selected to conduct reverse transcription (RT)-qPCR., in which the Log2 (fold change) was significantly consistent with the qPCR results. The analysis of chloroplast transcription under drought provided clues for understanding chloroplast function under drought. The phylogenetic position of "Shanxinyang" within Populus was analysed by using the chloroplast genome sequences of 23 Populus plants, showing that "Shanxinyang" belongs to Sect. Populus and is sister to Populus davidiana. Further, mVISTA analysis showed that the variation in non-coding (regulatory) regions was greater than that in coding regions, which suggests that further attention should be paid to the chloroplast in order to obtain new evolutionary or functional insights related to aspects of plant biology. CONCLUSIONS: Our findings indicate that complex prokaryotic genome regulation occurs when processing transcripts under drought stress. The results not only offer clues for understanding the chloroplast genome and transcription features in woody plants but also serve as a basis for future molecular studies on poplar species.

Evidence for widespread selection in shaping the genomic landscape during speciation of Populus.

Increasing our understanding of how evolutionary processes drive the genomic landscape of variation is fundamental to a better understanding of the genomic consequences of speciation. However, genome-wide patterns of within- and between- species variation have not been fully investigated in most forest tree species despite their global ecological and economic importance. Here, we use whole-genome resequencing data from four Populus species spanning the speciation continuum to reconstruct their demographic histories and investigate patterns of diversity and divergence within and between species. Using Populus trichocarpa as an outgroup species, we further infer the genealogical relationships and estimate the extent of ancient introgression among the three aspen species (Populus tremula, Populus davidiana and Populus tremuloides) throughout the genome. Our results show substantial variation in these patterns along the genomes with this variation being strongly predicted by local recombination rates and the density of functional elements. This implies that the interaction between recurrent selection and intrinsic genomic features has dramatically sculpted the genomic landscape over long periods of time. In addition, our findings provide evidence that, apart from background selection, recent positive selection and long-term balancing selection have also been crucial components in shaping patterns of genome-wide variation during the speciation process.

Genetic diversity and population structure of black cottonwood (Populus deltoides) revealed using simple sequence repeat markers.

BACKGROUND: Black cottonwood (Populus deltoides) is one of the keystone forest tree species, and has become the main breeding parents in poplar hybrid breeding. However, the genetic diversity and population structure of the introduced resources are not fully understood. RESULTS: In the present study, five loci containing null alleles were excluded and 15 pairs of SSR (simple sequence repeat) primers were used to analyze the genetic diversity and population structure of 384 individuals from six provenances (Missouri, Iowa, Washington, Louisiana, and Tennessee (USA), and Quebec in Canada) of P. deltoides. Ultimately, 108 alleles (Na) were detected; the expected heterozygosity (He) per locus ranged from 0.070 to 0.905, and the average polymorphic information content (PIC) was 0.535. The provenance 'Was' had a relatively low genetic diversity, while 'Que', 'Lou', and 'Ten' provenances had high genetic diversity, with Shannon's information index (I) above 1.0. The mean coefficient of genetic differentiation (Fst) and gene flow (Nm) were 0.129 and 1.931, respectively. Analysis of molecular variance (AMOVA) showed that 84.88% of the genetic variation originated from individuals. Based on principal coordinate analysis (PCoA) and STRUCTURE cluster analysis, individuals distributed in the Mississippi River Basin were roughly classified as one group, while those distributed in the St. Lawrence River Basin and Columbia River Basin were classified as another group. The cluster analysis based on the population level showed that provenance 'Iow' had a small gene flow and high degree of genetic differentiation compared with the other provenances, and was classified into one group. There was a significant relationship between genetic distance and geographical distance. CONCLUSIONS: P. deltoides resources have high genetic diversity and there is a moderate level of genetic differentiation among provenances. Geographical isolation and natural conditions may be the main factors causing genetic differences among individuals. Individuals reflecting population genetic information can be selected to build a core germplasm bank. Meanwhile, the results could provide theoretical support for the scientific management and efficient utilization of P. deltoides genetic resources, and promote the development of molecular marker-assisted breeding of poplar.

Element content and expression of genes of interest in guard cells are connected to spatiotemporal variations in stomatal conductance.

Element content and expression of genes of interest on single cell types, such as stomata, provide valuable insights into their specific physiology, improving our understanding of leaf gas exchange regulation. We investigated how far differences in stomatal conductance (gs ) can be ascribed to changes in guard cells functioning in amphistomateous leaves. gs was measured during the day on both leaf sides, on well-watered and drought-stressed trees (two Populus euramericana Moench and two Populus nigra L. genotypes). In parallel, guard cells were dissected for element content and gene expressions analyses. Both were strongly arranged according to genotype, and drought had the lowest impact overall. Normalizing the data by genotype highlighted a structure on the basis of leaf sides and time of day both for element content and gene expression. Guard cells magnesium, phosphorus, and chlorine were the most abundant on the abaxial side in the morning, where gs was at the highest. In contrast, genes encoding H+ -ATPase and aquaporins were usually more abundant in the afternoon, whereas genes encoding Ca2+ -vacuolar antiporters, K+ channels, and ABA-related genes were in general more abundant on the adaxial side. Our work highlights the unique physiology of each leaf side and their analogous rhythmicity through the day.

PtrARF2.1 Is Involved in Regulation of Leaf Development and Lignin Biosynthesis in Poplar Trees.

Auxin response factors (ARFs) are important regulators modulating the expression of auxin-responsive genes in various biological processes in plants. In the Populus genome, a total of 39 ARF members have been identified, but their detailed functions are still unclear. In this study, six poplar auxin response factor 2 (PtrARF2) members were isolated from P. trichocarpa. Expression pattern analysis showed that PtrARF2.1 is highly expressed in leaf tissues compared with other PtrARF2 genes and significantly repressed by exogenous auxin treatment. PtrARF2.1 is a nuclear-localized protein without transcriptional activation activity. Knockdown of PtrARF2.1 by RNA interference (RNAi) in poplars led to the dwarf plant, altered leaf shape, and reduced size of the leaf blade, while overexpression of PtrARF2.1 resulted in a slight reduction in plant height and the similar leaf phenotype in contrast to the wildtype. Furthermore, histological staining analysis revealed an ectopic deposition of lignin in leaf veins and petioles of PtrARF2.1-RNAi lines. RNA-Seq analysis showed that 74 differential expression genes (DEGs) belonging to 12 transcription factor families, such as NAM, ATAF and CUC (NAC), v-myb avian myeloblastosis viral oncogene homolog (MYB), ethylene response factors (ERF) and basic helix-loop-helix (bHLH), were identified in PtrARF2.1-RNAi leaves and other 24 DEGs were associated with the lignin biosynthetic pathway. Altogether, the data indicate that PtrARF2.1 plays an important role in regulating leaf development and influences the lignin biosynthesis in poplars.

Comparative Proteomic Analysis of Plant-Pathogen Interactions in Resistant and Susceptible Poplar Ecotypes Infected with Botryosphaeria dothidea.

Poplar are important forestry species in China, but the Botryosphaeria dothidea pathogen causes serious economic losses worldwide. To identify candidate B. dothidea resistance proteins and explore the molecular mechanisms involved in poplar-pathogen interactions, proteomic responses of stem samples from resistant and susceptible poplar ecotypes to B. dothidea were investigated using nanoflow liquid chromatography-tandem mass spectrometry with label-free quantitative analysis. We identified 588 proteins, divided into 21 biological process categories including 48 oxidoreductases, 72 hydrolytic enzymes, 80 metabolic enzymes, and 29 proteins of unknown function. Differential proteome analysis revealed large differences between resistant Populus tomentosa Carr and susceptible Populus beijingensis Hsu ecotypes before and after inoculation. Among 102 identified proteins, 22 were highly upregulated in the resistant genotype but downregulated in the susceptible genotype. Proteins induced in P. tomentosa Carr in response to B. dothidea are associated with plant defenses including oxidoreductase activity (catalase, isocitrate dehydrogenase, and superoxide dismutase), phenylpropanoid biosynthesis and phenylalanine metabolism (alcohol dehydrogenase), photosynthesis (ATP synthase subunit alpha, ATP synthase gamma chain, photosystem I P700 chlorophyll a apoprotein A2, photosystem II CP47 chlorophyll apoprotein), carbon fixation (pyruvate kinase, triosephosphate isomerase, malic enzyme, phosphoglycerate kinase, ribulose-1,5-bisphosphate carboxylase, and ribulose bisphosphate carboxylase small chain), and glycolysis/gluconeogenesis (fructose-bisphosphate aldolase). Kyoto Encyclopedia of Genes and Genomes pathway analysis identified 168 proteins related to metabolic pathways, 41 proteins related to the biosynthesis of phenylpropanoids, and 36 proteins related to the biosynthesis of plant hormones, the biosynthesis of alkaloids derived from ornithine, lysine, and nicotinic acid, and photosynthesis in response to B. dothidea. Our findings provide insight into plant-pathogen interactions in resistant and susceptible poplar ecotypes infected with B. dothidea and could assist the development of novel strategies for fighting poplar canker disease.

The poplar pangenome provides insights into the evolutionary history of the genus.

The genus Populus comprises a complex amalgam of ancient and modern species that has become a prime model for evolutionary and taxonomic studies. Here we sequenced the genomes of 10 species from five sections of the genus Populus, identified 71 million genomic variations, and observed new correlations between the single-nucleotide polymorphism-structural variation (SNP-SV) density and indel-SV density to complement the SNP-indel density correlation reported in mammals. Disease resistance genes (R genes) with heterozygous loss-of-function (LOF) were significantly enriched in the 10 species, which increased the diversity of poplar R genes during evolution. Heterozygous LOF mutations in the self-incompatibility genes were closely related to the self-fertilization of poplar, suggestive of genomic control of self-fertilization in dioecious plants. The phylogenetic genome-wide SNPs tree also showed possible ancient hybridization among species in sections Tacamahaca, Aigeiros, and Leucoides. The pangenome resource also provided information for poplar genetics and breeding.

Comparative analysis of the complete chloroplast genomes of seven Populus species: Insights into alternative female parents of Populus tomentosa.

Populus tomentosa, of section Populus, is distributed mainly in northern China. This species has high resistance to many diseases and insects, and it plays key roles in shelterbelts and urban afforestation in northern China. It has long been suspected to be a hybrid, but its parents remain unknown. In the present study, we report four newly sequenced complete cp genomes from section Populus and comparative genomic analyses of these new sequences and three published cp genome sequences. The seven cp genomes ranged from 155,853 bp (P. tremula var. davidiana) to 156,746 bp (P. adenopoda) in length, and their gene orders, gene numbers and GC contents were similar. We analyzed SNPs, indels, SSRs and repeats among the seven cp genomes, and eight small inversions were detected in the ndhC-trnV, rbcL-accD, petA-psbJ, trnW-trnP, rpl16-rps3, trnL-ycf15, ycf15-trnL, and ndhF-trnL intergenic regions. Furthermore, seven divergent regions (trnH-psbA, matK, psbM-psbD, ndhC-trnV, ycf1, ndhF-ccsA and ccsA-ndhD) were found in more highly variable regions. The phylogenetic tree reveals that P. tomentosa is closely related to P. alba and P. alba var. pyramidalis. Hence, P. alba was involved in the formation of P. tomentosa.

A New Propolis Type from Changbai Mountains in North-east China: Chemical Composition, Botanical Origin and Biological Activity.

Propolis is a bee product with a wide range of biological activities and its chemical compounds depend highly on the type of plant accessible to the bees. The Changbai Mountains are a major mountain range in Northeast China and are one of the major bee product-producing areas in China. In this study, we evaluated the total phenolic acids and flavonoid contents as well as the antioxidant activity of propolis sampled from the Changbai Mountains area (CBM). We identified the major compounds and qualified their contents by HPLC-ESI/MS and HPLC-UV, and found that the content of p-coumaric acid and an unknown peak (CBE) in CBM propolis was higher than in propolis from other parts of China. The unknown compound CBE was isolated, purified, and identified as benzyl p-coumarate by MS and NMR. Possible plant sources of CBM propolis are Populus davidiana dode and Populus simonii Carr, which widely distributed in the Changbai Mountains area. CBM propolis is a new propolis type, that could be an excellent raw material for health foods and pharmaceuticals.

De novo assembly of white poplar genome and genetic diversity of white poplar population in Irtysh River basin in China.

The white poplar (Populus alba) is widely distributed in Central Asia and Europe. There are natural populations of white poplar in Irtysh River basin in China. It also can be cultivated and grown well in northern China. In this study, we sequenced the genome of P. alba by single-molecule real-time technology. De novo assembly of P. alba had a genome size of 415.99 Mb with a contig N50 of 1.18 Mb. A total of 32,963 protein-coding genes were identified. 45.16% of the genome was annotated as repetitive elements. Genome evolution analysis revealed that divergence between P. alba and Populus trichocarpa (black cottonwood) occurred ~5.0 Mya (3.0, 7.1). Fourfold synonymous third-codon transversion (4DTV) and synonymous substitution rate (ks) distributions supported the occurrence of the salicoid WGD event (~ 65 Mya). Twelve natural populations of P. alba in the Irtysh River basin in China were sequenced to explore the genetic diversity. Average pooled heterozygosity value of P. alba populations was 0.170±0.014, which was lower than that in Italy (0.271±0.051) and Hungary (0.264±0.054). Tajima's D values showed a negative distribution, which might signify an excess of low frequency polymorphisms and a bottleneck with later expansion of P. alba populations examined.

Identification and Characterization of an OSH1 Thiol Reductase from Populus Trichocarpa.

Interferon gamma-induced lysosomal thiol reductase (GILT) is abundantly expressed in antigen-presenting cells and participates in the treatment and presentation of antigens by major histocompatibility complex II. Also, GILT catalyzes the reduction of disulfide bonds, which plays an important role in cellular immunity. (1) Background: At present, the studies of GILT have mainly focused on animals. In plants, GILT homologous gene (Arabidopsis thalianaOSH1: AtOSH1) was discovered in the forward screen of mutants with compromised responses to sulphur nutrition. However, the complete properties and functions of poplar OSH1 are unclear. In addition, CdCl2 stress is swiftly engulfing the limited land resources on which humans depend, restricting agricultural production. (2) Methods: A prokaryotic expression system was used to produce recombinant PtOSH1 protein, and Western blotting was performed to identify its activity. In addition, a simplified version of the floral-dip method was used to transform A. thaliana. (3) Results: Here, we describe the identification and characterization of OSH1 from Populus trichocarpa. The deduced PtOSH1 sequence contained CQHGX2ECX2NX4C and CXXC motifs. The transcript level of PtOSH1 was increased by cadmium (Cd) treatment. In addition, recombinant PtOSH1 reduced disulfide bonds. A stress assay showed that PtOSH1-overexpressing (OE) A. thaliana lines had greater resistance to Cd than wild-type (WT) plants. Also, the activities of superoxide dismutase (SOD), peroxidase (POD), and catalase (CAT) in PtOSH1-OE plants were significantly higher than those in WT A. thaliana. These results indicate that PtOSH1 likely plays an important role in the response to Cd by regulating the reactive oxygen species (ROS)-scavenging system. (4) Conclusions: PtOSH1 catalyzes the reduction of disulfide bonds and behaves as a sulfhydryl reductase under acidic conditions. The overexpression of PtOSH1 in A. thaliana promoted root development, fresh weight, and dry weight; upregulated the expression levels of ROS scavenging-related genes; and improved the activity of antioxidant enzymes, enhancing plant tolerance to cadmium (Cd) stress. This study aimed to provide guidance that will facilitate future studies of the function of PtOSH1 in the response of plants to Cd stress.

Analyzing the phylogeny of poplars based on molecular data.

Methods for constructing trees using DNA sequences, known as molecular phylogenetics, have been applied to analyses of phylogenetic origin, evolutionary relatedness and taxonomic classification. Combining data sequenced in this study and downloaded from GenBank, we sampled 112 (chloroplast data) / 122 (ITS data) specimens belonging to 49 (chloroplast data) / 46 (ITS data) poplar species or hybrids from six (chloroplast data) / five sections (ITS data). Maximum parsimony and Bayesian inference were used to analyze phylogenetic relationships within the genus Populus based on eight chloroplast combinations and ITS regions. The results suggested that Bayesian inference might be more suitable for the phylogenetic reconstruction of Populus. All Populus species could be divided into two clades: clade 1, including subclades 1 and 2, and clade 2, including subclades 3 and 4. Species within clade 1, involving five sections except for Leuce, clustered coinciding with their two specific main geographical distribution areas: China (subclade 1) and North America (subclade 2). Clustering in subclade 3, section Leuce was confirmed to be of monophyletic origin and independent evolution. Its two subsections, namely Albidae and Trepidae, could be separated by chloroplast data but had frequent gene flow based on ITS data. Phylogeny analysis based on chloroplast data demonstrated once more that section Aigeiros was paraphyletic and further showed that the P. deltoides lineage is restricted in subclade 2 and that P. nigra lineage, located in subclade 3, originated from a hybrid of which an Albidae ancestor species was the material parent. Similarly, section Tacamahaca was found to be paraphyletic and had two lineages: a clade 1 lineage, such as P. cathayana, and a clade 2 lineage, such as P. simonii. Section Leucoides was paraphyletic and closely linked to section Tacamahaca. Their section boundaries were not conclusively delimitated by sequencing information.

Adaptive introgression and maintenance of a trispecies hybrid complex in range-edge populations of Populus.

In hybrid zones occurring in marginal environments, adaptive introgression from one species into the genomic background of another may constitute a mechanism facilitating adaptation at range limits. Although recent studies have improved our understanding of adaptive introgression in widely distributed tree species, little is known about the dynamics of this process in populations at the margins of species ranges. We investigated the extent of introgression between three species of the genus Populus sect. Tacamahaca (P. balsamifera, P. angustifolia and P. trichocarpa) at the margins of their distributions in the Rocky Mountain region of the United States and Canada. Using genotyping by sequencing (GBS), we analysed ~ 83,000 single nucleotide polymorphisms genotyped in 296 individuals from 29 allopatric and sympatric populations of the three species. We found a trispecies hybrid complex present throughout the zone of range overlap, including early as well as advanced generation backcross hybrids, indicating recurrent gene flow in this hybrid complex. Using genomic cline analysis, we found evidence of non-neutral patterns of introgression at 23% of loci in hybrids, of which 47% and 8% represented excess ancestry from P. angustifolia and P. balsamifera, respectively. Gene ontology analysis suggested these genomic regions were enriched for genes associated with photoperiodic regulation, metal ion transport, maintenance of redox homeostasis and cell wall metabolites involved in regulation of seasonal dormancy. Our study demonstrates the role of adaptive introgression in a multispecies hybrid complex in range-edge populations and has implications for understanding the evolutionary dynamics of adaptation in hybrid zones, especially at the margins of species distributions.