Production of isoflavone genistein in transgenic IFS tobacco roots and its role in stimulating the development of arbuscular mycorrhiza

Flavonoids and isoflavonoids are secondary metabolites in plants. With the goal of obtaining isoflavonoids from a wide range of plants, a few key studies have proven that isoflavonoids can be produced in non-leguminous plants by transgenic engineering. Many earlier studies investigate genistein biosynthesis in leaves and petals of isoflavone synthase (IFS) transgenic tobacco. However, most reports do not attempt to analyze quantification of genistein or do not check the presence of genistein in transgenic plant roots. In addition, little is known about the influence of genistein on arbuscular mycorrhiza (AM). In this paper, we reported that genistein was obtained from transgenic IFS tobacco roots. In addition, we revealed that endogenous genistein and 10???g?g^?1 exogenous genistein enhanced the development of AM symbiosis. We also revealed the relative expression levels of pertinent genes during the development of AM symbiosis. Our results suggest that genistein plays a positive role in the development of AM symbiosis in tobacco roots.     

Sex-Related Responses of Populus cathayana Shoots and Roots to AM Fungi and Drought Stress

We investigated the impact of drought and arbuscular mycorrhizal (AM) fungi on the morphological structure and physiological function of shoots and roots of male and female seedlings of the dioecious plant Populus cathayana Rehder. Pot-grown seedlings were subjected to well watered or water-limiting conditions (drought) and were grown in soil that was either inoculated or not inoculated with the AM fungus Rhizophagus intraradices. No significant differences were found in the infection rates between the two sexes. Drought decreased root and shoot growth, biomass and root morphological characteristics, whereas superoxide radical (O₂–) and hydrogen peroxide content, peroxidase (POD) activity, malondialdehyde (MDA) concentration and proline content were significantly enhanced in both sexes. Male plants that formed an AM fungal symbiosis showed a significant increase in shoot and root morphological growth, increased proline content of leaves and roots, and increased POD activity in roots under both watering regimes; however, MDA concentration in the roots decreased. By contrast, AM fungi either had no effect or a slight negative effect on the shoot and root growth of female plants, with lower root biomass, total biomass and root/shoot ration under drought. In females, MDA concentration increased in leaves and roots under both watering regimes, and the proline content and POD activity of roots increased under drought conditions; however, POD activity significantly decreased under well-watered conditions. These findings suggest that AM fungi enhanced the tolerance of male plants to drought by improving shoot and root growth, biomass and the antioxidant system. Further investigation is needed to unravel the complex effects of AM fungi on the growth and antioxidant system of female plants.     

Functional adaptation of a plant receptor-kinase paved the way for the evolution of intracellular root symbioses with bacteria

Nitrogen-fixing root nodule symbioses (RNS) occur in two major forms—Actinorhiza and legume-rhizobium symbiosis—which differ in bacterial partner, intracellular infection pattern, and morphogenesis. The phylogenetic restriction of nodulation to eurosid angiosperms indicates a common and recent evolutionary invention, but the molecular steps involved are still obscure. In legumes, at least seven genes—including the symbiosis receptor-kinase gene SYMRK—are essential for the interaction with rhizobia bacteria and for the Arbuscular Mycorrhiza (AM) symbiosis with phosphate-acquiring fungi, which is widespread in occurrence and believed to date back to the earliest land plants. We show that SYMRK is also required for Actinorhiza symbiosis of the cucurbit Datisca glomerata with actinobacteria of the genus Frankia, revealing a common genetic basis for both forms of RNS. We found that SYMRK exists in at least three different structural versions, of which the shorter forms from rice and tomato are sufficient for AM, but not for functional endosymbiosis with bacteria in the legume Lotus japonicus. Our data support the idea that SYMRK sequence evolution was involved in the recruitment of a pre-existing signalling network from AM, paving the way for the evolution of intracellular root symbioses with nitrogen-fixing bacteria.     

H2O2参与AM真菌与烟草共生过程

以烟草((Nicotiana tabacum,品种CF90NF)为寄主,苗期接种丛枝菌根(AM)真菌摩西球囊霉(Glomus mosseae,G.m),测定G.m与烟草共生过程中烟草根部H2O2含量以及多胺氧化酶(PAO)和过氧化物酶(POD)活性;研究外源H2O2对G.m侵染烟草的影响以及H2O2清除剂和合成抑制剂对烟草侧根H2O2含量及烟草侧根和菌丝中H2O2荧光强度的影响,以探究H2O2在AM真菌侵染烟草过程中的作用。结果表明,接种G.m 20d后烟草侧根中出现H2O2含量的猝发,一定浓度的外源H2O2促进G.m对烟草的侵染,而H2O2清除剂抗坏血酸(AsA)显著削弱烟草侧根和菌丝中的H2O2荧光强度,降低G.m对烟草的侵染率,表明H2O2参与G.m与烟草共生过程;在G.m与烟草共生过程中,PAO和POD活性显著升高,PAO抑制剂二氨基十二烷(DADD)和POD抑制剂水杨羟肟酸(SHAM)显著降低烟草侧根中H2O2荧光强度,对菌丝中H2O2荧光强度无显著影响,表明烟草根部和G.m均可产生H2O2,PAO和POD参与烟草侧根中H2O2的合成,菌丝中可能存在其他来源的H2O2。     

Contribution of different arbuscular mycorrhizal fungal inoculum to <Emphasis Type="Italic">Elymus nutans</Emphasis> under nitrogen addition

Arbuscular mycorrhizal (AM) fungi are known to promote plant growth and nutrient uptake, but their role in nitrogen (N) uptake still remains unclear. Therefore, a pot experiment was set up to evaluate the impacts of N addition and AM inoculation (Diversispora eburnea, Claroideoglomus etunicatum, Paraglomus occultum, and their mixture) on AM root colonization, plant biomass, N and P nutrition in Elymus nutans. Our results showed that AM root colonization was unaffected by N addition but was significantly affected by different AM fungal species. D. eburnea and C. etunicatum showed significant higher root colonization than P. occultum. The E. nutans exhibited the highest biomass when inoculated with D. eburnea and significantly higher than non-mycorrhizal (the control) regardless of N addition. Under N addition treatment, D. eburnea significantly enhanced P content of roots, N content of shoots and roots, while AM mixture significantly enhanced shoot P content compared with non-mycorrhizal. However, N and P content in shoots and roots did not significantly vary among treatments when no N was added. In addition, inoculation with C. etunicatum and P. occultum showed no significant effect on plant biomass, N and P content regardless of N addition. In conclusion, this study revealed that the plant response to N addition depends on AM fungal species and also confirmed that significant functional diversity exists among AM fungal species.     

MycoRed: Betalain pigments enable in vivo real-time visualisation of arbuscular mycorrhizal colonisation

Arbuscular mycorrhiza (AM) are mutualistic interactions formed between soil fungi and plant roots. AM symbiosis is a fundamental and widespread trait in plants with the potential to sustainably enhance future crop yields. However, improving AM fungal association in crop species requires a fundamental understanding of host colonisation dynamics across varying agronomic and ecological contexts. To this end, we demonstrate the use of betalain pigments as in vivo visual markers for the occurrence and distribution of AM fungal colonisation by Rhizophagus irregularis in Medicago truncatula and Nicotiana benthamiana roots. Using established and novel AM-responsive promoters, we assembled multigene reporter constructs that enable the AM-controlled expression of the core betalain synthesis genes. We show that betalain colouration is specifically induced in root tissues and cells where fungal colonisation has occurred. In a rhizotron setup, we also demonstrate that betalain staining allows for the noninvasive tracing of fungal colonisation along the root system over time. We present MycoRed, a useful innovative method that will expand and complement currently used fungal visualisation techniques to advance knowledge in the field of AM symbiosis.

Arbuscular mycorrhiza are mutualistic interactions formed between soil fungi and plant roots. This study presents the MycoRed system, which uses red plant pigments derived from beetroot to reveal how fungi establish symbiosis with living legume and wild tobacco roots.     

Contrasting impacts of defoliation on root colonization by arbuscular mycorrhizal and dark septate endophytic fungi of Medicago sativa

Individual plants typically interact with multiple mutualists and enemies simultaneously. Plant roots encounter both arbuscular mycorrhizal (AM) and dark septate endophytic (DSE) fungi, while the leaves are exposed to herbivores. AMF are usually beneficial symbionts, while the functional role of DSE is largely unknown. Leaf herbivory may have a negative effect on root symbiotic fungi due to decreased carbon availability. However, evidence for this is ambiguous and no inoculation-based experiment on joint effects of herbivory on AM and DSE has been done to date. We investigated how artificial defoliation impacts root colonization by AM (Glomus intraradices) and DSE (Phialocephala fortinii) fungi and growth of Medicago sativa host in a factorial laboratory experiment. Defoliation affected fungi differentially, causing a decrease in arbuscular colonization and a slight increase in DSE-type colonization. However, the presence of one fungal species had no effect on colonization by the other or on plant growth. Defoliation reduced plant biomass, with this effect independent of the fungal treatments. Inoculation by either fungal species reduced root/shoot ratios, with this effect independent of the defoliation treatments. These results suggest AM colonization is limited by host carbon availability, while DSE may benefit from root dieback or exudation associated with defoliation. Reductions in root allocation associated with fungal inoculation combined with a lack of effect of fungi on plant biomass suggest DSE and AMF may be functional equivalent to the plant within this study. Combined, our results indicate different controls of colonization, but no apparent functional consequences between AM and DSE association in plant roots in this experimental setup.     

Mycorrhiza alters the profile of root hairs in trifoliate orange

Root hairs and arbuscular mycorrhiza (AM) coexist in root systems for nutrient and water absorption, but the relation between AM and root hairs is poorly known. A pot study was performed to evaluate the effects of four different AM fungi (AMF), namely, Claroideoglomus etunicatum, Diversispora versiformis, Funneliformis mosseae, and Rhizophagus intraradices on root hair development in trifoliate orange (Poncirus trifoliata) seedlings grown in sand. Mycorrhizal seedlings showed significantly higher root hair density than non-mycorrhizal seedlings, irrespective of AMF species. AMF inoculation generally significantly decreased root hair length in the first- and second-order lateral roots but increased it in the third- and fourth-order lateral roots. AMF colonization induced diverse responses in root hair diameter of different order lateral roots. Considerably greater concentrations of phosphorus (P), nitric oxide (NO), glucose, sucrose, indole-3-acetic acid (IAA), and methyl jasmonate (MeJA) were found in roots of AM seedlings than in non-AM seedlings. Levels of P, NO, carbohydrates, IAA, and MeJA in roots were correlated with AM formation and root hair development. These results suggest that AMF could alter the profile of root hairs in trifoliate orange through modulation of physiological activities. F. mosseae, which had the greatest positive effects, could represent an efficient AM fungus for increasing fruit yields or decreasing fertilizer inputs in citrus production.     

指数施肥对楸树无性系生物量分配和根系形态的影响

为探求楸树不同无性系生物量分配和根系形态的差异,2011年3-8月在甘肃省天水市小陇山林科所,以2年生楸树无性系1-4、7080和015-1组培苗为试验材料,设置了CK、6、10、14 g尿素/株4个处理,研究指数施肥对楸树无性系生物量分配和根系形态的影响.结果表明:(1)同一无性系中,10 g尿素/株的根、茎、叶生物量及总生物量、根长、根表面积、根体积和根平均直径均高于其它处理.无性系015-1的生物量和根系形态参数整体上高于无性系1-4和7080.无性系015-1在10 g尿素/株的根、茎、叶生物量及总生物量分别为89.44 g、61.30 g、79.97 g、230.71 g,是CK的1.48、1.52、2.09、1.66倍;根长、根表面积和根体积为22667 cm、6260 cm2、578.14 cm3,是CK的1.94、1.54、2.43倍.(2)指数施肥和无性系的遗传差异明显影响楸树不同无性系生物量的分配格局.适量施氮明显促进3个楸树无性系生物量的积累,而氮素缺乏或过量均不利于生物量的积累.8月同一无性系的根冠比均随施氮量的增加而降低;同一处理下无性系7080的根冠比高于无性系1-4和015-l.无性系1-4和7080的生物量主要向叶和茎分配,而无性系015-1主要向叶分配.(3)指数施肥在6月和7月主要促进细根根长和根表面积的增加;指数施肥在8月主要促进细根、中等根和粗根体积的急剧增加,分别比7月高达36.88％、124.96％、154.79％.这利于根系在中后期吸收更多养分,从而引起生物量分配格局的变化.(4)生物量参数和根系形态参数关系密切.根生物量、地上生物量、总生物量分别和根长、根表面积、根体积、根平均直径极显著正相关;根冠比和根长、根表面积、根体积、根平均直径极显著负相关;比根长和地上生物量、总生物量显著正相关,和根冠比极显著负相关.     

Early Root Herbivory Impairs Arbuscular Mycorrhizal Fungal Colonization and Shifts Defence Allocation in Establishing Plantago lanceolata

Research into plant-mediated indirect interactions between arbuscular mycorrhizal (AM) fungi and insect herbivores has focussed on those between plant shoots and above-ground herbivores, despite the fact that only below-ground herbivores share the same part of the host plant as AM fungi. Using Plantago lanceolata L., we aimed to characterise how early root herbivory by the vine weevil (Otiorhynchus sulcatus F.) affected subsequent colonization by AM fungi (Glomus spp.) and determine how the two affected plant growth and defensive chemistry. We exposed four week old P. lanceolata to root herbivory and AM fungi using a 2×2 factorial design (and quantified subsequent effects on plant biomass and iridoid glycosides (IGs) concentrations. Otiorhynchus sulcatus reduced root growth by c. 64%, whereas plant growth was unaffected by AM fungi. Root herbivory reduced extent of AM fungal colonization (by c. 61%). O. sulcatus did not influence overall IG concentrations, but caused qualitative shifts in root and shoot IGs, specifically increasing the proportion of the more toxic catalpol. These changes may reflect defensive allocation in the plant against further attack. This study demonstrates that very early root herbivory during plant development can shape future patterns of AM fungal colonization and influence defensive allocation in the plant.     

Arbuscular mycorrhizal symbiosis regulates physiology and performance of <Emphasis Type="Italic">Digitaria eriantha</Emphasis> plants subjected to abiotic stresses by modulating antioxidant and jasmonate levels

This study evaluates antioxidant responses and jasmonate regulation in Digitaria eriantha cv. Sudafricana plants inoculated (AM) and non-inoculated (non-AM) with Rhizophagus irregularis and subjected to drought, cold, or salinity. Stomatal conductance, photosynthetic efficiency, biomass production, hydrogen peroxide accumulation, lipid peroxidation, antioxidants enzymes activities, and jasmonate levels were determined. Stomatal conductance and photosynthetic efficiency decreased in AM and non-AM plants under all stress conditions. However, AM plants subjected to drought, salinity, or non-stress conditions showed significantly higher stomatal conductance values. AM plants subjected to drought or non-stress conditions increased their shoot/root biomass ratios, whereas salinity and cold caused a decrease in these ratios. Hydrogen peroxide accumulation, which was high in non-AM plant roots under all treatments, increased significantly in non-AM plant shoots under cold stress and in AM plants under non-stress and drought conditions. Lipid peroxidation increased in the roots of all plants under drought conditions. In shoots, although lipid peroxidation decreased in AM plants under non-stress and cold conditions, it increased under drought and salinity. AM plants consistently showed high catalase (CAT) and ascorbate peroxidase (APX) activity under all treatments. By contrast, the glutathione reductase (GR) and superoxide dismutase (SOD) activity of AM roots was lower than that of non-AM plants and increased in shoots. The endogenous levels of cis-12-oxophytodienoc acid (OPDA), jasmonic acid (JA), and 12-OH-JA showed a significant increase in AM plants as compared to non-AM plants. 11-OH-JA content only increased in AM plants subjected to drought. Results show that D. eriantha is sensitive to drought, salinity, and cold stresses and that inoculation with AM fungi regulates its physiology and performance under such conditions, with antioxidants and jasmonates being involved in this process.     

Regulation of root morphogenesis in arbuscular mycorrhizae: what role do fungal exudates,phosphate, sugars and hormones play in lateral root formation?

Background

Arbuscular mycorrhizae (AMs) form a widespread root–fungus symbiosis that improves plant phosphate (Pi) acquisition and modifies the physiology and development of host plants. Increased branching is recognized as a general feature of AM roots, and has been interpreted as a means of increasing suitable sites for colonization. Fungal exudates, which are involved in the dialogue between AM fungi and their host during the pre-colonization phase, play a well-documented role in lateral root (LR) formation. In addition, the increased Pi content of AM plants, in relation to Pi-starved controls, as well as changes in the delivery of carbohydrates to the roots and modulation of phytohormone concentration, transport and sensitivity, are probably involved in increasing root system branching.

Scope

This review discusses the possible causes of increased branching in AM plants. The differential root responses to Pi, sugars and hormones of potential AM host species are also highlighted and discussed in comparison with those of the non-host Arabidopsis thaliana.

Conclusions

Fungal exudates are probably the main compounds regulating AM root morphogenesis during the first colonization steps, while a complex network of interactions governs root development in established AMs. Colonization and high Pi act synergistically to increase root branching, and sugar transport towards the arbusculated cells may contribute to LR formation. In addition, AM colonization and high Pi generally increase auxin and cytokinin and decrease ethylene and strigolactone levels. With the exception of cytokinins, which seem to regulate mainly the root:shoot biomass ratio, these hormones play a leading role in governing root morphogenesis, with strigolactones and ethylene blocking LR formation in the non-colonized, Pi-starved plants, and auxin inducing them in colonized plants, or in plants grown under high Pi conditions.     

Expression of a Soybean Hydroxyproline-Rich Glycoprotein Gene Is Correlated with Maturation of Roots

A novel extensin gene has been identified in soybean (Glycine max L.) that encodes a hydroxyproline-rich glycoprotein (SbHRGP3) with two different domains. In this study expression of SbHRGP3 was investigated during soybean root development. SbHRGP was expressed in roots of mature plants, as well as seedlings, and showed a distinct pattern of expression during root development. The expression of SbHRGP3 increased gradually during root development of seedlings and reached a maximum while the secondary roots were maturing. The maximum expression level was contributed mainly by the secondary roots rather than by the primary root. Furthermore, expression of SbHRGP3 was preferentially detected in the regions undergoing maturation of the primary and secondary roots. These results imply that the expression of SbHRGP3 is regulated in an organ- and development-specific manner and that in soybean SbHRGP3 expression may be required for root maturation, presumably for the cessation of root elongation. Wounding and sucrose in combination enhanced expression of SbHRGP3 in roots, whereas both wounding and sucrose were required for the expression of SbHRGP3 in leaves.     

Aboveground resource allocation in response to root herbivory as affected by the arbuscular mycorrhizal symbiosis

Aims

Arbuscular mycorrhizal (AM) fungi associate with the majority of terrestrial plants, influencing their growth, nutrient uptake and defence chemistry. Consequently, AM fungi can significantly impact plant-herbivore interactions, yet surprisingly few studies have investigated how AM fungi affect plant responses to root herbivores. This study aimed to investigate how AM fungi affect plant tolerance mechanisms to belowground herbivory.

Methods

We examined how AM fungi affect plant (Saccharum spp. hybrid) growth, nutrient dynamics and secondary chemistry (phenolics) in response to attack from a root-feeding insect (Dermolepida albohirtum).

Results

Root herbivory reduced root mass by almost 27%. In response, plants augmented investment in aboveground biomass by 25%, as well as increasing carbon concentrations. The AM fungi increased aboveground biomass, phosphorus and carbon. Meanwhile, root herbivory increased foliar phenolics by 31% in mycorrhizal plants, and increased arbuscular colonisation of roots by 75% overall. AM fungi also decreased herbivore performance, potentially via increasing root silicon concentrations.

Conclusions

Our results suggest that AM fungi may be able to augment plant tolerance to root herbivory via resource allocation aboveground and, at the same time, enhance plant root resistance by increasing root silicon. The ability of AM fungi to facilitate resource allocation aboveground in this way may be a more widespread strategy for plants to cope with belowground herbivory.

     

AM真菌和磷对小马安羊蹄甲幼苗生长的影响

丛枝菌根(arbuscular mycorrhizal,AM)真菌在退化生态系统恢复与重建实践中具有重要作用。采用盆栽模拟方法,重点分析不同土壤磷条件下小马鞍羊蹄甲(Bauhinia faberi)幼苗接种AM真菌后,幼苗的形态、生物量积累、菌根侵染率和菌根效应(mycorrhizal growth response, MGR)在一个生长季内的动态变化。结果表明,Glomus mosseae和 Glomus coronatum能较好地侵染幼苗,两种AM真菌显著地增加幼苗根系、叶片数和生物量;接种AM真菌显著影响幼苗的生物量分配,而土壤磷对幼苗的生物量分配影响不明显,AM真菌和土壤磷对幼苗生长的交互作用显著;G. mosseae是小马鞍羊蹄甲的优势AM菌,其接种的幼苗根长、叶片数、生物量、侵染率和菌根效应都显著高于G. coronatum处理的幼苗;菌根效应显著,接种AM真菌能有缓解土壤磷素缺乏的限制作用,且随着苗龄增大促生作用表现更为明显。不同AM菌种对小马鞍羊蹄甲幼苗生长的促生作用表现出的差异,提示在多元资源限制的干旱贫瘠环境中进行生物修复须为目标恢复物种筛选出高效的优势AM真菌。     

Effects of arsenate on tobacco hairy root and seedling growth,and its removal

Arsenic (As) is a highly toxic environmental contaminant to which most living organisms are exposed. Plants have evolved several mechanisms to cope with this toxic metalloid; however, these mechanisms are only partially understood. The response of plants to As phytotoxicity is highly complex, with considerable variation among species. In this study, arsenate (As⁺⁵) effects on germination and early root development of tobacco (Nicotiana tabacum) seedlings were investigated. Also, As⁺⁵ tolerance and removal efficiency of tobacco hairy roots (HRs) and seedlings were assessed and compared. Total seed germination capacity was not affected by 10 to 200 μM As⁺⁵, while primary root length and root branching were reduced by As⁺⁵ concentrations that were at or above 100 μM. Both systems were able to tolerate As⁺⁵ concentrations of 10 μM since no growth inhibition was detected. For higher As⁺⁵ concentrations, phytotoxicity increased, but it was mitigated by higher phosphate (Pi) availability. Under the studied conditions, As⁺⁵ removal efficiency of HRs greatly exceeded that of seedlings. Further, tobacco HRs were able to accumulate As in their tissues. These results justify further investigations on As tolerance and detoxification mechanisms in tobacco, an easy-to-transform crop species with high biomass, which could allow evaluation of the possible application of wild type or alternatively transgenic tobacco plants for As phytoextraction.     

Jasmonate response of the Nicotiana tabacum agglutinin promoter in Arabidopsis thaliana

NICTABA is a carbohydrate-binding protein (also called lectin) that is expressed in several Nicotiana species after treatment with jasmonates and insect herbivory. Analyses with tobacco lines overexpressing the NICTABA gene as well as lines with reduced lectin expression have shown the entomotoxic effect of NICTABA against Lepidopteran larvae, suggesting a role of the lectin in plant defense. Until now, little is known with respect to the upstream regulatory mechanisms that are controlling the expression of inducible plant lectins. Using Arabidopsis thaliana plants stably expressing a promoter-β-glucuronidase (GUS) fusion construct, it was shown that jasmonate treatment influenced the NICTABA promoter activity. A strong GUS staining pattern was detected in very young tissues (the apical and root meristems, the cotyledons and the first true leaves), but the promoter activity decreased when plants were getting older. NICTABA was also expressed at low concentrations in tobacco roots and expression levels increased after cold treatment. The data presented confirm a jasmonate-dependent response of the promoter sequence of the tobacco lectin gene in Arabidopsis. These new jasmonate-responsive tobacco promoter sequences can be used as new tools in the study of jasmonate signalling related to plant development and defense.