Natural variation in the prolyl 4-hydroxylase gene PtoP4H9 contributes to perennial stem growth in Populus.

Perennial trees must maintain stem growth throughout their entire lifespan to progressively increase in size as they age. The overarching question of the molecular mechanisms that govern stem perennial growth in trees remains largely unanswered. Here we deciphered the genetic architecture that underlies perennial growth trajectories using genome-wide association studies (GWAS) for measures of growth traits across years in a natural population of Populus tomentosa. By analyzing the stem growth trajectory, we identified PtoP4H9, encoding prolyl 4-hydroxylase 9, which is responsible for the natural variation in the growth rate of diameter at breast height (DBH) across years. Quantifying the dynamic genetic contribution of PtoP4H9 loci to stem growth showed that PtoP4H9 played a pivotal role in stem growth regulation. Spatiotemporal expression analysis showed that PtoP4H9 was highly expressed in cambium tissues of poplars of various ages. Overexpression and knockdown of PtoP4H9 revealed that it altered cell expansion to regulate cell wall modification and mechanical characteristics, thereby promoting stem growth in Populus. We showed that natural variation in PtoP4H9 occurred in a BASIC PENTACYSTEINE transcription factor PtoBPC1-binding promoter element controlling PtoP4H9 expression. The geographic distribution of PtoP4H9 allelic variation was consistent with the modes of selection among populations. Altogether, our study provides important genetic insights into dynamic stem growth in Populus, and we confirmed PtoP4H9 as a potential useful marker for breeding or genetic engineering of poplars.

PPGR: a comprehensive perennial plant genomes and regulation database.

Perennial woody plants hold vital ecological significance, distinguished by their unique traits. While significant progress has been made in their genomic and functional studies, a major challenge persists: the absence of a comprehensive reference platform for collection, integration and in-depth analysis of the vast amount of data. Here, we present PPGR (Resource for Perennial Plant Genomes and Regulation; https://ngdc.cncb.ac.cn/ppgr/) to address this critical gap, by collecting, integrating, analyzing and visualizing genomic, gene regulation and functional data of perennial plants. PPGR currently includes 60 species, 847 million protein-protein/TF (transcription factor)-target interactions, 9016 transcriptome samples under various environmental conditions and genetic backgrounds. Noteworthy is the focus on genes that regulate wood production, seasonal dormancy, terpene biosynthesis and leaf senescence representing a wealth of information derived from experimental data, literature mining, public databases and genomic predictions. Furthermore, PPGR incorporates a range of multi-omics search and analysis tools to facilitate browsing and application of these extensive datasets. PPGR represents a comprehensive and high-quality resource for perennial plants, substantiated by an illustrative case study that demonstrates its capacity in unraveling gene functions and shedding light on potential regulatory processes.

Computational dissection of genetic variation modulating the response of multiple photosynthetic phenotypes to the light environment.

BACKGROUND: The expression of biological traits is modulated by genetics as well as the environment, and the level of influence exerted by the latter may vary across characteristics. Photosynthetic traits in plants are complex quantitative traits that are regulated by both endogenous genetic factors and external environmental factors such as light intensity and CO2 concentration. The specific processes impacted occur dynamically and continuously as the growth of plants changes. Although studies have been conducted to explore the genetic regulatory mechanisms of individual photosynthetic traits or to evaluate the effects of certain environmental variables on photosynthetic traits, the systematic impact of environmental variables on the dynamic process of integrated plant growth and development has not been fully elucidated. RESULTS: In this paper, we proposed a research framework to investigate the genetic mechanism of high-dimensional complex photosynthetic traits in response to the light environment at the genome level. We established a set of high-dimensional equations incorporating environmental regulators to integrate functional mapping and dynamic screening of gene‒environment complex systems to elucidate the process and pattern of intrinsic genetic regulatory mechanisms of three types of photosynthetic phenotypes of Populus simonii that varied with light intensity. Furthermore, a network structure was established to elucidate the crosstalk among significant QTLs that regulate photosynthetic phenotypic systems. Additionally, the detection of key QTLs governing the response of multiple phenotypes to the light environment, coupled with the intrinsic differences in genotype expression, provides valuable insights into the regulatory mechanisms that drive the transition of photosynthetic activity and photoprotection in the face of varying light intensity gradients. CONCLUSIONS: This paper offers a comprehensive approach to unraveling the genetic architecture of multidimensional variations in photosynthetic phenotypes, considering the combined impact of integrated environmental factors from multiple perspectives.

Allelic variations of WAK106-E2Fa-DPb1-UGT74E2 module regulate fibre properties in Populus tomentosa.

Wood formation, intricately linked to the carbohydrate metabolism pathway, underpins the capacity of trees to produce renewable resources and offer vital ecosystem services. Despite their importance, the genetic regulatory mechanisms governing wood fibre properties in woody plants remain enigmatic. In this study, we identified a pivotal module comprising 158 high-priority core genes implicated in wood formation, drawing upon tissue-specific gene expression profiles from 22 Populus samples. Initially, we conducted a module-based association study in a natural population of 435 Populus tomentosa, pinpointing PtoDPb1 as the key gene contributing to wood formation through the carbohydrate metabolic pathway. Overexpressing PtoDPb1 led to a 52.91% surge in cellulose content, a reduction of 14.34% in fibre length, and an increment of 38.21% in fibre width in transgenic poplar. Moreover, by integrating co-expression patterns, RNA-sequencing analysis, and expression quantitative trait nucleotide (eQTN) mapping, we identified a PtoDPb1-mediated genetic module of PtoWAK106-PtoDPb1-PtoE2Fa-PtoUGT74E2 responsible for fibre properties in Populus. Additionally, we discovered the two PtoDPb1 haplotypes that influenced protein interaction efficiency between PtoE2Fa-PtoDPb1 and PtoDPb1-PtoWAK106, respectively. The transcriptional activation activity of the PtoE2Fa-PtoDPb1 haplotype-1 complex on the promoter of PtoUGT74E2 surpassed that of the PtoE2Fa-PtoDPb1 haplotype-2 complex. Taken together, our findings provide novel insights into the regulatory mechanisms of fibre properties in Populus, orchestrated by PtoDPb1, and offer a practical module for expediting genetic breeding in woody plants via molecular design.

Temporal dynamics of genetic architecture governing leaf development in Populus.

Leaf development is a multifaceted and dynamic process orchestrated by a myriad of genes to shape the proper size and morphology. The dynamic genetic network underlying leaf development remains largely unknown. Utilizing a synergistic genetic approach encompassing dynamic genome-wide association study (GWAS), time-ordered gene co-expression network (TO-GCN) analyses and gene manipulation, we explored the temporal genetic architecture and regulatory network governing leaf development in Populus. We identified 42 time-specific and 18 consecutive genes that displayed different patterns of expression at various time points. We then constructed eight TO-GCNs that covered the cell proliferation, transition, and cell expansion stages of leaf development. Integrating GWAS and TO-GCN, we postulated the functions of 27 causative genes for GWAS and identified PtoGRF9 as a key player in leaf development. Genetic manipulation via overexpression and suppression of PtoGRF9 revealed its primary influence on leaf development by modulating cell proliferation. Furthermore, we elucidated that PtoGRF9 governs leaf development by activating PtoHB21 during the cell proliferation stage and attenuating PtoLD during the transition stage. Our study provides insights into the dynamic genetic underpinnings of leaf development and understanding the regulatory mechanism of PtoGRF9 in this dynamic process.

Rare variations within the serine/arginine-rich splicing factor PtoRSZ21 modulate stomatal size to determine drought tolerance in Populus.

Rare variants contribute significantly to the 'missing heritability' of quantitative traits. The genome-wide characteristics of rare variants and their roles in environmental adaptation of woody plants remain unexplored. Utilizing genome-wide rare variant association study (RVAS), expression quantitative trait loci (eQTL) mapping, genetic transformation, and molecular experiments, we explored the impact of rare variants on stomatal morphology and drought adaptation in Populus. Through comparative analysis of five world-wide Populus species, we observed the influence of mutational bias and adaptive selection on the distribution of rare variants. RVAS identified 75 candidate genes correlated with stomatal size (SS)/stomatal density (SD), and a rare haplotype in the promoter of serine/arginine-rich splicing factor PtoRSZ21 emerged as the foremost association signal governing SS. As a positive regulator of drought tolerance, PtoRSZ21 can recruit the core splicing factor PtoU1-70K to regulate alternative splicing (AS) of PtoATG2b (autophagy-related 2). The rare haplotype PtoRSZ21hap2 weakens binding affinity to PtoMYB61, consequently affecting PtoRSZ21 expression and SS, ultimately resulting in differential distribution of Populus accessions in arid and humid climates. This study enhances the understanding of regulatory mechanisms that underlie AS induced by rare variants and might provide targets for drought-tolerant varieties breeding in Populus.

Enhanced genome-wide association reveals the role of YABBY11-NGATHA-LIKE1 in leaf serration development of Populus.

Leaf margins are complex plant morphological features that contribute to leaf shape diversity, which affects plant structure, yield, and adaptation. Although several leaf margin regulators have been identified to date, the genetic basis of their natural variation has not been fully elucidated. In this study, we profiled two distinct leaf morphology types (serrated and smooth) using the persistent homology mathematical framework (PHMF) in two poplar species (Populus tomentosa and Populus simonii, respectively). A combined genome-wide association study (GWAS) and expression quantitative trait nucleotide (eQTN) mapping were applied to create a leaf morphology control module using data from P. tomentosa and P. simonii populations. Natural variation in leaf margins was associated with YABBY11 (YAB11) transcript abundance in poplar. In P. tomentosa, PtoYAB11 carries a premature stop codon (PtoYAB11PSC), resulting in the loss of its positive regulation of NGATHA-LIKE1 (PtoNGAL-1) and RIBULOSE BISPHOSPHATE CARBOXYLASE LARGE SUBUNIT (PtoRBCL). Overexpression of PtoYAB11PSC promoted serrated leaf margins, enlarged leaves, enhanced photosynthesis, and increased biomass. Overexpression of PsiYAB11 in P. tomentosa promoted smooth leaf margins, higher stomatal density, and greater light damage repair ability. In poplar, YAB11-NGAL1 is sensitive to environmental conditions, acts as a positive regulator of leaf margin serration, and may also link environmental signaling to leaf morphological plasticity.

Growth-regulating factor 15-mediated gene regulatory network enhances salt tolerance in poplar.

Soil salinity is an important determinant of crop productivity and triggers salt stress response pathways in plants. The salt stress response is controlled by transcriptional regulatory networks that maintain regulatory homeostasis through combinations of transcription factor (TF)-DNA and TF-TF interactions. We investigated the transcriptome of poplar 84 K (Populus alba × Populus glandulosa) under salt stress using samples collected at 4- or 6-h intervals within 2 days of salt stress treatment. We detected 24,973 differentially expressed genes, including 2,231 TFs that might be responsive to salt stress. To explore these interactions and targets of TFs in perennial woody plants, we combined gene regulatory networks, DNA affinity purification sequencing, yeast two-hybrid-sequencing, and multi-gene association approaches. Growth-regulating factor 15 (PagGRF15) and its target, high-affinity K+ transporter 6 (PagHAK6), were identified as an important regulatory module in the salt stress response. Overexpression of PagGRF15 and PagHAK6 in transgenic lines improved salt tolerance by enhancing Na+ transport and modulating H2O2 accumulation in poplar. Yeast two-hybrid assays identified more than 420 PagGRF15-interacting proteins, including ETHYLENE RESPONSE FACTOR TFs and a zinc finger protein (C2H2) that are produced in response to a variety of phytohormones and environmental signals and are likely involved in abiotic stress. Therefore, our findings demonstrate that PagGRF15 is a multifunctional TF involved in growth, development, and salt stress tolerance, highlighting the capability of a multifaceted approach in identifying regulatory nodes in plants.

Allelic variation in transcription factor PtoWRKY68 contributes to drought tolerance in Populus.

Drought stress limits woody species productivity and influences tree distribution. However, dissecting the molecular mechanisms that underpin drought responses in forest trees can be challenging due to trait complexity. Here, using a panel of 300 Chinese white poplar (Populus tomentosa) accessions collected from different geographical climatic regions in China, we performed a genome-wide association study (GWAS) on seven drought-related traits and identified PtoWRKY68 as a candidate gene involved in the response to drought stress. A 12-bp insertion and/or deletion and three nonsynonymous variants in the PtoWRKY68 coding sequence categorized natural populations of P. tomentosa into two haplotype groups, PtoWRKY68hap1 and PtoWRKY68hap2. The allelic variation in these two PtoWRKY68 haplotypes conferred differential transcriptional regulatory activities and binding to the promoters of downstream abscisic acid (ABA) efflux and signaling genes. Overexpression of PtoWRKY68hap1 and PtoWRKY68hap2 in Arabidopsis (Arabidopsis thaliana) ameliorated the drought tolerance of two transgenic lines and increased ABA content by 42.7% and 14.3% compared to wild-type plants, respectively. Notably, PtoWRKY68hap1 (associated with drought tolerance) is ubiquitous in accessions in water-deficient environments, whereas the drought-sensitive allele PtoWRKY68hap2 is widely distributed in well-watered regions, consistent with the trends in local precipitation, suggesting that these alleles correspond to geographical adaptation in Populus. Moreover, quantitative trait loci analysis and an electrophoretic mobility shift assay showed that SHORT VEGETATIVE PHASE (PtoSVP.3) positively regulates the expression of PtoWRKY68 under drought stress. We propose a drought tolerance regulatory module in which PtoWRKY68 modulates ABA signaling and accumulation, providing insight into the genetic basis of drought tolerance in trees. Our findings will facilitate molecular breeding to improve the drought tolerance of forest trees.

Long-term warming increased carbon sequestration capacity in a humid subtropical forest.

Tropical and subtropical forests play a crucial role in global carbon (C) pools, and their responses to warming can significantly impact C-climate feedback and predictions of future global warming. Despite earth system models projecting reductions in land C storage with warming, the magnitude of this response varies greatly between models, particularly in tropical and subtropical regions. Here, we conducted a field ecosystem-level warming experiment in a subtropical forest in southern China, by translocating mesocosms (ecosystem composed of soils and plants) across 600 m elevation gradients with temperature gradients of 2.1°C (moderate warming), to explore the response of ecosystem C dynamics of the subtropical forest to continuous 6-year warming. Compared with the control, the ecosystem C stock decreased by 3.8% under the first year of 2.1°C warming; but increased by 13.4% by the sixth year of 2.1°C warming. The increased ecosystem C stock by the sixth year of warming was mainly attributed to a combination of sustained increased plant C stock due to the maintenance of a high plant growth rate and unchanged soil C stock. The unchanged soil C stock was driven by compensating and offsetting thermal adaptation of soil microorganisms (unresponsive soil respiration and enzyme activity, and more stable microbial community), increased plant C input, and inhibitory C loss (decreased C leaching and inhibited temperature sensitivity of soil respiration) from soil drying. These results suggest that the humid subtropical forest C pool would not necessarily diminish consistently under future long-term warming. We highlight that differential and asynchronous responses of plant and soil C processes over relatively long-term periods should be considered when predicting the effects of climate warming on ecosystem C dynamics of subtropical forests.

Genomic insights into selection for heterozygous alleles and woody traits in Populus tomentosa.

Heterozygous alleles are widespread in outcrossing and clonally propagated woody plants. The variation in heterozygosity that underlies population adaptive evolution and phenotypic variation, however, remains largely unknown. Here, we describe a de novo chromosome-level genome assembly of Populus tomentosa, an economic and ecologically important native tree in northern China. By resequencing 302 natural accessions, we determined that the South subpopulation (Pop_S) encompasses the ancestral strains of P. tomentosa, while the Northwest subpopulation (Pop_NW) and Northeast subpopulation (Pop_NE) experienced different selection pressures during population evolution, resulting in significant population differentiation and a decrease in the extent of heterozygosity. Analysis of heterozygous selective sweep regions (HSSR) suggested that selection for lower heterozygosity contributed to the local adaptation of P. tomentosa by dwindling gene expression and genetic load in the Pop_NW and Pop_NE subpopulations. Genome-wide association studies (GWAS) revealed that 88 single nucleotide polymorphisms (SNPs) within 63 genes are associated with nine wood composition traits. Among them, the selection for the homozygous AA allele in PtoARF8 is associated with reductions in cellulose and hemicellulose contents by attenuating PtoARF8 expression, and the increase in lignin content is attributable to the selection for decreases in exon heterozygosity in PtoLOX3 during adaptive evolution of natural populations. This study provides novel insights into allelic variations in heterozygosity associated with adaptive evolution of P. tomentosa in response to the local environment and identifies a series of key genes for wood component traits, thereby facilitating genomic-based breeding of important traits in perennial woody plants.

Multifeature analysis of age-related microbiome structures reveals defense mechanisms of Populus tomentosa trees.

Root microbiota composition shifts during the development of most annual plants. Although some perennial plants can live for centuries, the host-microbiome partnerships and interaction mechanisms underlying their longevity remain unclear. To address this gap, we investigated age-related changes in the root metabolites, transcriptomes, and microbiome compositions of 1- to 35-yr-old Populus tomentosa trees. Ten co-response clusters were obtained according to their accumulation patterns, and members of each cluster displayed a uniform and clear pattern of abundance. Multi-omics network analysis demonstrated that the increased abundance of Actinobacteria with tree age was strongly associated with the flavonoid biosynthesis. Using genetic approaches, we demonstrate that the flavonoid biosynthesis regulator gene Transparent Testa 8 is associated with the recruitment of flavonoid-associated Actinobacteria. Further inoculation experiments of Actinobacteria isolates indicated that their colonization could significantly improve the host's phenotype. Site-directed mutagenesis revealed that the hyBl gene cluster, involved in biosynthesis of an aminocyclitol hygromycin B analog in Streptomyces isolate bj1, is associated with disease suppression. We hypothesize that interactions between perennial plants and soil microorganisms lead to gradual enrichment of a subset of microorganisms that may harbor a wealth of currently unknown functional traits.

Transcriptome and miRNAome analysis reveals components regulating tissue differentiation of bamboo shoots.

Primary thickening determines bamboo yield and wood property. However, little is known about the regulatory networks involved in this process. This study identified a total of 58,652 genes and 150 miRNAs via transcriptome and small RNA sequencing using the underground thickening shoot samples of wild-type (WT) Moso bamboo (Phyllostachys edulis) and a thick wall (TW) variant (P. edulis "Pachyloen") at five developmental stages (WTS1/TWS1-WTS5/TWS5). A total of 14,029 (65.17%) differentially expressed genes and 68 (45.33%) differentially expressed miRNAs were identified from the WT, TW, and WTTW groups. The first two groups were composed of four pairwise combinations, each between two successive stages (WTS2/TWS2_versus_WTS1/TWS1, WTS3/TWS3_versus_WTS2/TWS2, WTS4/TWS4_versus_WTS3/TWS3, and WTS5/TWS5_versus_WTS4/TWS4), and the WTTW group was composed of five combinations, each between two relative stages (TWS1-5_versus_WTS1-5). Additionally, among the phytohormones, zeatin showed more remarkable changes in concentrations than indole-3-acetic acid, gibberellic acid, and abscisic acid throughout the five stages in the WT and the TW groups. Moreover, 125 cleavage sites were identified for 387 miRNA-mRNA pairs via degradome sequencing (P < 0.05). The dual-luciferase reporter assay confirmed that 13 miRNAs bound to 12 targets. Fluorescence in situ hybridization localized miR166 and miR160 in the shoot apical meristem and the procambium of Moso bamboo shoots at the S1 stage. Thus, primary thickening is a complex process regulated by miRNA-gene-phytohormone networks, and the miRNAome and transcriptome dynamics regulate phenotypic plasticity. These findings provide insights into the molecular mechanisms underlying wood formation and properties and propose targets for bamboo breeding.

Local diversity of drought resistance and resilience in Populus tomentosa correlates with the variation of DNA methylation.

Little information is known about DNA methylation variation in shaping environment-specific drought resistance and resilience for tree adaptation. In this study, we leveraged RNA sequencing and whole-genome bisulfite sequencing data to dissect the distinction of epigenetic regulation under drought stress and rewater condition of Populus tomentosa accessions from three geographical regions. We demonstrated low resistance and high resilience for accessions from South. Non-CG methylation levels in promoter regions of Southern accessions were lower than accessions from higher latitudes and negatively regulated gene expression. CHH context methylation was more sensitive to drought stress, and the geographical-specific differentially methylated regions were scarcely changed by environmental fluctuation. We identified 60 conserved hub genes within the co-expression networks that correlate with photosynthetic and stomatal morphological traits. Epigenome-wide association studies and genome-wide association studies of these 60 hub genes revealed the interdependency between genetic and epigenetic variation in GATA9 and LECRK-VIII.2, which was associated with stomatal morphology and chlorophyll content. The natural epigenetic variation in GATA9 was also faithfully transmitted to progenies in two family-based F1 populations. This study indicates a functional relationship of DNA methylation diversity with drought resistance and resilience which offers new insights into plants' local adaptation to a stressful environment.

Leaf physiology variations are modulated by natural variations that underlie stomatal morphology in Populus.

Stomata are essential for photosynthesis and abiotic stress tolerance. Here, we used multiomics approaches to dissect the genetic architecture and adaptive mechanisms that underlie stomatal morphology in Populus tomentosa juvenile natural population (303 accessions). We detected 46 candidate genes and 15 epistatic gene-pairs, associated with 5 stomatal morphologies and 18 leaf development and photosynthesis traits, through genome-wide association studies. Expression quantitative trait locus mapping revealed that stomata-associated gene loci were significantly associated with the expression of leaf-related genes; selective sweep analysis uncovered significant differentiation in the allele frequencies of genes that underlie stomatal variations. An allelic regulatory network operating under drought stress and adequate precipitation conditions, with three key regulators (DUF538, TRA2 and AbFH2) and eight interacting genes, was identified that might regulate leaf physiology via modulation of stomatal shape and density. Validation of candidate gene variations in drought-tolerant and F1 hybrid populations of P. tomentosa showed that the DUF538, TRA2 and AbFH2 loci cause functional stabilisation of spatiotemporal regulatory, whose favourable alleles can be faithfully transmitted to offspring. This study provides insights concerning leaf physiology and stress tolerance via the regulation of stomatal determination in perennial plants.

Acidity of Soil and Water Decreases in Acid-Sensitive Forests of Tropical China.

Acid deposition in China has been declining since the 2000s. While this may help mitigate acidification in forest soils and water, little is known about the recovery of soils and water from previous severe acidification in tropical China. Here, we assessed the chemistry of mineral soils, water, and acid gases (SO2 and NOx) from three successional forest types in tropical China from 2000 to 2022. Our results showed that soil pH increased synchronously from 3.9 (2000-2015) to 4.2 (2016-2022) across all three forest types, with exchangeable acid initially decreasing and thereafter stabilizing. Surface and ground water pH also gradually increased throughout the monitoring period. Soil pH recovery was stronger in the primary than in the planted forest. However, soil pH recovery lagged behind the increase in rainfall pH by approximately a decade. The recovery of soil pH was likely related to the positive effects of the dissolution of Al/Fe-hydroxysulfate mineral and subsequent sulfur desorption on soil acid-neutralizing capacity, increased soil organic matter, and climate warming, but was likely moderated by increased exchangeable aluminum and potentially proton-producing hydroxysulfate mineral dissolution that caused the lagged soil pH recovery. Surface and ground water pH recovery was attributed to increased water acid-neutralizing capacity. Our study reports the potential for the recovery of acidified soil and water following decreased acid deposition and provides new insights into the functional recovery of acid-sensitive forests.

Design, synthesis, and STING-agonistic activity of benzo[b]thiophene-2-carboxamide derivatives.

STING is an important immune-associated protein that localizes in the endoplasmic reticulum membrane. Upon being activated by its agonists, STING triggers the IRF and NF-κB pathways, which generates type I interferons and proinflammatory cytokines, and ultimately primes the innate immune responses to achieve valid antitumor efficacy. We designed and synthesized a series of benzo[b]thiophene-2-carboxamide derivatives. Through STING-agonistic activity evaluation, compounds 12d and 12e exhibited marginal human STING-activating activities. Western blot analysis demonstrated that both 12d and 12e treatment increased the phosphorylation of the downstream signaling molecules (TBK1 and IRF3) of STING. The proposed binding mode of 12d/12e and STING protein displayed that two canonical hydrogen bonds, a π-π stacking interaction, as well as a π-cation interaction formed between the agonist and the CDN-binding domain of STING protein.

Dynamic physiological and transcriptome changes reveal a potential relationship between the circadian clock and salt stress response in Ulmus pumila.

Despite the important role the circadian clock plays in numerous critical physiological responses in plants, such as hypocotyl elongation, leaf movement, stomatal opening, flowering, and stress responses, there have been no investigations into the effect of the circadian clock on physiological and transcriptional networks under salt stress. Ulmus pumila L. has been reported to tolerate 100-150 mM NaCl treatment. We measured the diurnal variation in photosynthesis and chlorophyll fluorescence parameters and performed a time-course transcriptome analysis of 2-years-old U. pumila seedlings under salt treatment to dissect the physiological regulation and potential relationship between the circadian network and the salt stress response. Seedlings in 150 mM NaCl treatment exhibited salt-induced physiological enhancement compared to the control group. A total of 7009 differentially expressed unigenes (DEGs) were identified under salt stress, of which 16 DEGs were identified as circadian rhythm-related DEGs (crDEGs). Further analysis of dynamic expression changes revealed that DEGs involved in four crucial pathways-photosynthesis, thiamine metabolism, abscisic acid synthesis and metabolism, and the hormone-MAPK signal crosstalk pathway-are closely related to the circadian clock. Finally, we constructed a co-expression network between the circadian clock and these four crucial pathways. Our results help shed light on the molecular link between the circadian network and salt stress tolerance in U. pumila.

Evolutionary Origins of Pseudogenes and Their Association with Regulatory Sequences in Plants.

Pseudogenes (Ψs), nonfunctional relatives of functional genes, form by duplication or retrotransposition, and loss of gene function by disabling mutations. Evolutionary analysis provides clues to Ψ origins and effects on gene regulation. However, few systematic studies of plant Ψs have been conducted, hampering comparative analyses. Here, we examined the origin, evolution, and expression patterns of Ψs and their relationships with noncoding sequences in seven angiosperm plants. We identified ∼250,000 Ψs, most of which are more lineage specific than protein-coding genes. The distribution of Ψs on the chromosome indicates that genome recombination may contribute to Ψ elimination. Most Ψs evolve rapidly in terms of sequence and expression levels, showing tissue- or stage-specific expression patterns. We found that a surprisingly large fraction of nontransposable element regulatory noncoding RNAs (microRNAs and long noncoding RNAs) originate from transcription of Ψ proximal upstream regions. We also found that transcription factor binding sites preferentially occur in putative Ψ proximal upstream regions compared with random intergenic regions, suggesting that Ψs have conditioned genome evolution by providing transcription factor binding sites that serve as promoters and enhancers. We therefore propose that rapid rewiring of Ψ transcriptional regulatory regions is a major mechanism driving the origin of novel regulatory modules.

Mycorrhizal fungi alleviate acidification-induced phosphorus limitation: Evidence from a decade-long field experiment of simulated acid deposition in a tropical forest in south China.

South China has been experiencing very high rate of acid deposition and severe soil acidification in recent decades, which has been proposed to exacerbate the regional ecosystem phosphorus (P) limitation. We conducted a 10-year field experiment of simulated acid deposition to examine how acidification impacts seasonal changes of different soil P fractions in a tropical forest with highly acidic soils in south China. As expected, acid addition significantly increased occluded P pool but reduced the other more labile P pools in the dry season. In the wet season, however, acid addition did not change microbial P, soluble P and labile organic P pools. Acid addition significantly increased exchangeable Al3+ and Fe3+ and the activation of Fe oxides in both seasons. Different from the decline of microbial abundance in the dry season, acid addition increased ectomycorrhizal fungi and its ratio to arbuscular mycorrhiza fungi in the wet season, which significantly stimulated phosphomonoesterase activities and likely promoted the dissolution of occluded P. Our results suggest that, even in already highly acidic soils, the acidification-induced P limitation could be alleviated by stimulating ectomycorrhizal fungi and phosphomonoesterase activities. The differential responses and microbial controls of seasonal soil P transformation revealed here should be implemented into ecosystem biogeochemical model for predicting plant productivity under future acid deposition scenarios.