Why can Mikania micrantha cover trees quickly during invasion?

The invasion of Mikania micrantha by climbing and covering trees has rapidly caused the death of many shrubs and trees, seriously endangering forest biodiversity. In this study, M. micrantha seedlings were planted together with local tree species (Cryptocarya concinna) to simulate the process of M. micrantha climbing under the forest. We found that the upper part of the M. micrantha stem lost its support after climbing to the top of the tree, grew in a turning and creeping manner, and then grew branches rapidly to cover the tree canopy. Then, we simulated the branching process through turning treatment. We found that a large number of branches had been formed near the turning part of the M. micrantha stem (TP). Compared with the upper part of the main stem (UP), the contents of plant hormones (auxin, cytokinin, gibberellin), soluble sugars (sucrose, glucose, fructose) and trehalose-6-phosphate (T6P) were significantly accumulated at TP. Further combining the transcriptome data of different parts of the main stem under erect or turning treatment, a hypothetical regulation model to illustrate how M. micrantha can quickly cover trees was proposed based on the regulation of sugars and hormones on plant branching; that is, the lack of support after ascending the top of the tree led to turning growth of the main stem, and the enhancement of sugars and T6P levels in the TP may first drive the release of nearby dormant buds. Plant hormone accumulation may regulate the entrance of buds into sustained growth and maintain the elongation of branches together with sugars to successfully covering trees.

Genome-Wide Characterization of PIN Auxin Efflux Carrier Gene Family in Mikania micrantha.

Mikania micrantha, recognized as one of the world's top 10 pernicious weeds, is a rapidly spreading tropical vine that has invaded the coastal areas of South China, causing serious economic losses and environmental damage. Rapid stem growth is an important feature of M. micrantha which may be related to its greater number of genes involved in auxin signaling and transport pathways and its ability to synthesize more auxin under adverse conditions to promote or maintain stem growth. Plant growth and development is closely connected to the regulation of endogenous hormones, especially the polar transport and asymmetric distribution of auxin. The PIN-FORMED (PIN) auxin efflux carrier gene family plays a key role in the polar transport of auxin and then regulates the growth of different plant tissues, which could indicate that the rapid growth of M. micrantha is closely related to this PIN-dependent auxin regulation. In this study, 11 PIN genes were identified and the phylogenetic relationship and structural compositions of the gene family in M. micrantha were analyzed by employing multiple bioinformatic methods. The phylogenetic analysis indicated that the PIN proteins could be divided into five distinct clades. The structural analysis revealed that three putative types of PIN (canonical, noncanonical and semi-canonical) exist among the proteins according to the length and the composition of the hydrophilic domain. The majority of the PINs were involved in the process of axillary bud differentiation and stem response under abiotic stress, indicating that M. micrantha may regulate its growth, development and stress response by regulating PIN expression in the axillary bud and stem, which may help explain its strong growth ability and environmental adaptability. Our study emphasized the structural features and stress response patterns of the PIN gene family and provided useful insights for further study into the molecular mechanism of auxin-regulated growth and control in M. micrantha.

Adaptation of the Invasive Plant (Sphagneticola trilobata L. Pruski) to a High Cadmium Environment by Hybridizing With Native Relatives.

Invasive species can evolve rapidly in the invasion areas to adapt to new habitats. Sphagneticola trilobata L. Pruski, an invasive species, was studied for its tolerance to cadmium (Cd) in the soil and compared with its natural hybrid. From the perspective of photosynthetic physiology, antioxidant characteristics, and leaf hormone levels, the differences between the leaves of the two species before and after Cd treatment were compared. The results showed that the hybrid had stronger tolerance to Cd stress than invasive species. After Cd stress, the indexes of gas-exchange [net photosynthetic rate (Pn), intercellular CO2 concentration (Ci), stomatal conductance (Gs), and transpiration rate (Tr)] of the hybrid was higher than invasive species, while the content of non-enzymatic antioxidants (flavonoids and total phenols) and antioxidant enzyme activities [peroxidase (POD) and superoxide dismutase (SOD)] was lower in hybrid than in invasive species. The changes in the content of plant hormones [auxin (IAA) and abscisic acid (ABA)] under Cd stress showed that hybrid can still maintain growth and prevent leaf senescence. Furthermore, the differences in gene expression between hybrid and invasive species in photosynthetic physiology, the antioxidant capacity of leaves, and endogenous hormone (IAA and ABA) synthesis pathway also showed that hybrid has stronger Cd tolerance than invasive species. This suggests that invasive species will realize the invasion through hybridization with the native relatives to overcome the stress from environmental factors. The study implied that hybridization between invasive species and native relatives is an important way for invasive species to spread in a wider and new environment that invasive species have not experienced in the area of origin.

Responses of Sphagneticola trilobata, Sphagneticola calendulacea and Their Hybrid to Drought Stress.

Sphagneticola trilobata is an invasive plant in South China. A hybrid between S. trilobata and Sphagneticola calendulacea (a native related species) has also been found in South China. The drought resistance of S. calendulacea, S. trilobata and their hybrid was studied in this paper. Under drought stress, the leaves of S. trilobata synthesized more abscisic acid (ABA) than those of the other species to reduce stomatal opening and water loss. The activities of antioxidant enzymes were the highest in S. trilobata and the lowest in S. calendulacea. The leaves of S. calendulacea suffered the most serious damage, and their maximum photochemical efficiency was the lowest. RNA-sequencing ware used to analyze the expression levels of genes in ABA, antioxidant enzyme, sugar and proline synthesis and photosynthesis pathways. Further real-time PCR detection verified the RNA-sequence results, and the results were in accordance with the physiological data. The results showed that S. trilobata was the most drought tolerant, and the drought tolerance of the hybrid did not show heterosis but was higher than S. calendulacea. Therefore, compared with S. trilobata and the hybrid, the population number and distribution of S. calendulacea may be less in arid areas.

Strong Response of Stem Photosynthesis to Defoliation in Mikania micrantha Highlights the Contribution of Phenotypic Plasticity to Plant Invasiveness.

Phenotypic plasticity affords invasive plant species the ability to colonize a wide range of habitats, but physiological plasticity of their stems is seldom recognized. Investigation of the stem plasticity of invasive plant species could lead to a better understanding of their invasiveness. We performed pot experiments involving defoliation treatments and isolated culture experiments to determine whether the invasive species Mikania micrantha exhibits greater plasticity in the stems than do three non-invasive species that co-occur in southern China and then explored the mechanism underlying the modification of its stem photosynthesis. Our results showed that the stems of M. micrantha exhibited higher plasticity in terms of either net or gross photosynthetic rate in response to the defoliation treatment. These effects were positively related to an increased stem elongation rate. The enhancement of stem photosynthesis in M. micrantha resulted from the comprehensive action involving increases in the Chl a/b ratio, D1 protein and stomatal aperture, changes in chloroplast morphology and a decrease in anthocyanins. Increased plasticity of stem photosynthesis may improve the survival of M. micrantha under harsh conditions and allow it to rapidly recover from defoliation injuries. Our results highlight that phenotypic plasticity promotes the invasion success of alien plant invaders.

Interpretation of the difference in shade tolerance of two subtropical forest tree species of different successional stages at the transcriptome and physiological levels.

Differences in plant shade tolerance constitute a major mechanism driving the succession of forest communities in subtropical forests. However, the indirect effects of differences in light requirements on the growth of mid- and late-successional tree species are unclear, and this potential growth effect has not been explained at the transcriptome level. Here, a typical mid-successional dominant tree species, Schima superba Gardn. et Champ, and a typical late-successional dominant tree species, Cryptocarya concinna Hance were used as materials and planted under 100% full light (FL) and 30% FL (low light, LL) to explore the responses of tree species in different successional stages of subtropical forests to different light environments. Transcriptome sequencing was used to analyze the expression changes in genes related to growth and photoprotection under different light environments. The young leaves of S. superba accumulated more malondialdehyde (MDA) and superoxide radicals (${\mathrm{O}}_2^{{{}^{\bullet}}^{-}}$) under LL. A lower hormone content (auxin, cytokinin, gibberellin) in the young leaves, a weaker photosynthetic capacity in the mature leaves and significant downregulation of related gene expression were also found under LL, which resulted in the total biomass of S. superba under LL being lower than that under FL. The young leaves of C. concinna had less MDA and ${\mathrm{O}}_2^{{{}^{\bullet}}^{-}}$, and a higher hormone contents under LL than those under FL. There was no significant difference in photosynthetic capacity between mature leaves in contrasting light environments. Although the biomass of C. concinna under LL was less than that under FL, the height of C. concinna under LL was higher than that under FL, indicating that C. concinna could grow well under the two light environments. Our results describing the acclimatization of light at the physiological, molecular and transcriptome levels are important for a complete understanding of successional mechanisms.

Why Is the Invasive Plant Sphagneticola trilobata More Resistant to High Temperature than Its Native Congener?

Climate change and invasive alien species threaten biodiversity. High temperature is a worrying ecological factor. Most responses of invasive plants aimed at coping with adversity are focused on the physiological level. To explore the molecular mechanisms underlying the response of an invasive plant (Sphagneticola trilobata L.) to high temperature, using a native species (Sphagneticola calendulacea L.) as the control, relevant indicators, including photosynthetic pigments, gas exchange, chlorophyll fluorescence, the antioxidant system, and related enzyme-coding genes were measured. The results showed that the leaves of S. calendulacea turned yellow, photosynthetic pigment content (Chl a, Chl b, Car, Chl) decreased, gas exchange (Pn) and chlorophyll fluorescence parameters (Fv/Fm, ΦPSII) decreased under high temperature. It was also found that high temperature caused photoinhibition and a large amount of ROS accumulated, resulting in an increase in MDA and relative conductivity. Antioxidant enzymes (including SOD, POD, CAT, and APX) and antioxidants (including flavonoids, total phenols, and carotenoids) were decreased. The qPCR results further showed that the expression of the PsbP, PsbA, and RubiscoL, SOD, POD, CAT, and APX genes was downregulated, which was consistent with the results of physiological data. Otherwise, the resistance of S. trilobata to high temperature was better than that of S. calendulacea, which made it a superior plant in the invasion area. These results further indicated that the gradual warming of global temperature will greatly accelerate the invasion area of S. trilobata.

The major photoprotective role of anthocyanins in leaves of Arabidopsis thaliana under long-term high light treatment: antioxidant or light attenuator?

Anthocyanins are water-soluble pigments in plants known for their photoprotective role against photoinhibitory and photooxidative damage under high light (HL). However, it remains unclear whether light-shielding or antioxidant activity plays a major role in the photoprotection exerted by anthocyanins under HL stress. To shed light on this question, we analyzed the physiological and biochemical responses to HL of three Arabidopsis thaliana lines (Col, chi, ans) with different light absorption and antioxidant characteristics. Under HL, ans had the highest antioxidant capacity, followed by Col, and finally chi; Col had the strongest light attenuation capacity, followed by chi, and finally ans. The line ans had weaker physiological activity of chloroplasts and more severe oxidative damage than chi after HL treatment. Col with highest photoprotection of light absorption capacity had highest resistance to HL among the three lines. The line ans with high antioxidant capacity could not compensate for its disadvantages in HL caused by the absence of the light-shielding function of anthocyanins. In addition, the expression level of the Anthocyanin Synthase (ANS) gene was most upregulated after HL treatment, suggesting that the conversion of colorless into colored anthocyanin precursors was necessary under HL. The contribution of anthocyanins to flavonoids, phenols, and antioxidant capacity increased in the late period of HL, suggesting that plants prefer to synthesize red anthocyanins (a group of colored antioxidants) over other colorless antioxidants to cope with HL. These experimental observations indicate that the light attenuation role of anthocyanins is more important than their antioxidant role in photoprotection.

The relationship between anthocyanin accumulation and photoprotection in young leaves of two dominant tree species in subtropical forests in different seasons.

Increasing amounts of experimental evidence show that anthocyanins provide physiological protection to plants under stress. However, the difference in photoprotection mediated by anthocyanins and other photoprotective substances in different seasons is still uncertain. To determine the relationship between anthocyanin accumulation and the photoprotective effects in different seasons, Castanopsis chinensis and Acmena acuminatissima, whose anthocyanin accumulation patterns differ in different seasons, were used as materials to explain how plants adapt to different seasons; as such, their physiological and biochemical responses were analyzed. Young leaves of C. chinensis and A. acuminatissima presented different colors in the different seasons. In summer, the young leaves of C. chinensis were purplish red, while those of A. acuminatissima were light green. In winter, the young leaves of C. chinensis were light green, while those of A. acuminatissima were red. Compared with the young red leaves, the young light green leaves that did not accumulate anthocyanins had higher flavonoid and phenolics contents, total antioxidant capacity, non-photochemical quenching (NPQ), and relative membrane leakage, and a slower recovery rate in the maximum photochemical efficiency (Fv/Fm) after high-light treatment. In addition, the net photosynthesis rate (Pn), transpiration rate (Tr), stomatal conductance (gs), and the effective quantum yield of PSII (ΦPSII) of the young leaves in winter were significantly lower than those in summer, while the activities of catalase (CAT, EC 1.11.1.6), peroxidase (POD, EC 1.11.1.7), and superoxide dismutase (SOD, EC 1.15.1.1) were significantly higher than those in summer. These data indicate that to adapt to seasonal changes anthocyanins, other antioxidative substances and antioxidative enzymes, as well as components involved in the safe dissipation of excitation energy as heat need to cooperate with one another.

Photosynthetic compensation of non-leaf organ stems of the invasive species Sphagneticola trilobata (L.) Pruski at low temperature.

Biological invasion is a hot topic in ecological research. Most studies on the physiological mechanisms of plants focus on leaves, but few studies focus on stems. To study the tolerance of invasive plant (Sphagneticola trilobata L.) to low temperature, relevant physiological indicators (including anthocyanin and chlorophyll) in different organs (leaves and stems) were analyzed, using a native species (Sphagneticola calendulacea L.) as the control. The results showed that, upon exposure to low temperature for 15 days, the stems of two Sphagneticola species were markedly reddened, their anthocyanin content increased, chlorophyll and chlorophyll fluorescence parameters decreased, and the accumulation of reactive oxygen species in the stem increased. The percentage increases of antioxidants and total antioxidant capacities in stems were significantly higher in S. trilobata than in S. calendulacea. This showed that S. trilobata had higher cold tolerance in stems while leaves were opposite. To further verify the higher cold tolerance of the stem of S. trilobata, a defoliation experiment was designed. We found that the defoliated stem of S. trilobata reduced anthocyanin accumulation and increased chlorophyll content, while alleviating membrane lipid damage and electrical conductivity, and the defoliated stem still showed an increase in stem diameter and biomass under low temperature. The discovery of the physiological and adaptive mechanisms of the stem of S. trilobata to low temperature will provide a theoretical basis for explaining how S. trilobata maintains its annual growth in South China. This is of great significance for predicting the future spread of cloned and propagated invasive plants.

Comparative physiological and transcriptomic analyses of photosynthesis in Sphagneticola calendulacea (L.) Pruski and Sphagneticola trilobata (L.) Pruski.

Sphagneticola trilobata (L.) Pruski is one of the fast-growing malignant weeds in South China. It has severely influenced local biodiversity and native plant habitat. Photosynthesis is the material basis of plant growth and development. However, there are few reports on the photosynthetic transcriptome of S. trilobata. In this study, S. trilobata had a relatively large leaf area and biomass. The gas exchange parameters per unit area of leaves, including net photosynthetic capacity (Pn), intercellular CO2 (Ci), stomatal conductance (Gs), transpiration rate (Tr), water use efficiency (WUE), photosynthetic pigment and Rubisco protein content were higher than those of the native plant Sphagneticola calendulacea (L.) Pruski. On this basis, the differences in photosynthesis pathways between the two Sphagneticola species were analyzed by using the Illumina HiSeq platform. The sequencing results for S. trilobata and S. calendulacea revealed 159,366 and 177,069 unigenes, respectively. Functional annotation revealed 119,350 and 150,846 non-redundant protein database annotations (Nr), 96,637 and 115,711 Swiss-Prot annotations, 49,159 and 60,116 Kyoto Encyclopedia of Genes and Genomes annotations (KEGG), and 83,712 and 97,957 Gene Ontology annotations (GO) in S. trilobata and S. calendulacea, respectively. Additionally, our analysis showed that the expression of key protease genes involved in the photosynthesis pathway, particularly CP43, CP47, PsbA and PetC, had high expression levels in leaves of S. trilobata in comparison to native species. Physiological and transcriptomic analyses suggest the high expression of photosynthetic genes ensures the high photosynthetic capacity of leaves, which is one of the inherent advantages underlying the successful invasion by S. trilobata.

Mikania micrantha genome provides insights into the molecular mechanism of rapid growth.

Mikania micrantha is one of the top 100 worst invasive species that can cause serious damage to natural ecosystems and substantial economic losses. Here, we present its 1.79 Gb chromosome-scale reference genome. Half of the genome is composed of long terminal repeat retrotransposons, 80% of which have been derived from a significant expansion in the past one million years. We identify a whole genome duplication event and recent segmental duplications, which may be responsible for its rapid environmental adaptation. Additionally, we show that M. micrantha achieves higher photosynthetic capacity by CO2 absorption at night to supplement the carbon fixation during the day, as well as enhanced stem photosynthesis efficiency. Furthermore, the metabolites of M. micrantha can increase the availability of nitrogen by enriching the microbes that participate in nitrogen cycling pathways. These findings collectively provide insights into the rapid growth and invasive adaptation.

Sequencing of anthocyanin synthesis-related enzyme genes and screening of reference genes in leaves of four dominant subtropical forest tree species.

The young leaves generally accumulate a certain concentration anthocyanins in the dominant species of the subtropical forest, and the changes of anthocyanin synthesis-related enzyme genes expression levels had an important effect on the study photoprotection of anthocyanins in the young leaves of subtropical forests. The determination of anthocyanin synthesis-related enzyme gene sequences and the selection of appropriate reference genes provide a basis for analyzing the functional properties of anthocyanins. In this study, four dominant subtropical forest species (i.e., Schima superba, Castanopsis fissa, Acmena acuminatissima, Cryptocarya concinna) were taken as materials. To obtain the correct nucleotide sequences of anthocyanin-related enzymes, the nucleotide sequences of CHS, DFR and ANS in each dominant species were obtained by sequencing and comparison. Then, to select the most stable reference genes for leaves at different developmental stages and different light conditions, the expression levels of six reference genes, including 18S, Actin, GAPDH, TUB, EF1 and UBQ, were studied by real-time fluorescent quantitative PCR (qRT-PCR), and reference gene stability was analyzed by GeNorm and NormFinder software. The results showed that the expression level of Actin was the most stable in S. superba, A. acuminatissima and C. concinna, and the expression level of GAPDH was the most stable in C. fissa. Finally, the expression levels of the anthocyanin synthesis genes CHS, DFR and ANS were analyzed and found to be consistent with the accumulation trend of anthocyanins in leaves. This study has important theoretical and practical significance for future research into the expression of anthocyanin synthesis-related enzyme genes in the dominant tree species in subtropical forests and reveals that anthocyanin has a photoprotective effect for young leaves in high-light environments.

ANS-deficient Arabidopsis is sensitive to high light due to impaired anthocyanin photoprotection.

Light attenuation and antioxidation are the main mechanisms of photoprotection by anthocyanin under high light (HL) stress. Anthocyanin synthase (ANS) is the key enzyme in the downstream portion of anthocyanin synthetic pathways. To explore the role of ANS in photoprotection by anthocyanin under HL stress, homozygous ANS-deficient Arabidopsis mutants were screened from SALK_073183 and SALK_028793. Here, we obtained two deficient mutants, ans-1 and ans-2, which had ANS gene expression levels equal to 5.9 and 32.9% of that of Col respectively. By analysing their physiological and biochemical responses to HL stress, we found that there were positive correlations among ANS expression level, anthocyanin content and resistance to HL. The line with the lowest ANS expression level, ans-1, was also the most sensitive to HL, showing the lowest anthocyanin content, chlorophyll content, Fv/Fm ratio, and Rubisco content and the highest O2•- accumulation and membrane leakage rate, although it also had the highest antioxidant capacity. Experimental evidence suggests that ANS mainly regulated the light-attenuating function of anthocyanin in photoprotection under HL. Blocking excess light is an important function of anthocyanin that protects plants from HL stress, and a high antioxidant capacity cannot compensate for the absence of the light-shielding function of anthocyanin.