AM真菌摩西管柄囊霉对干旱胁迫下枳抗氧化酶基因表达的影响

测定分析了接种丛枝菌根（AM）真菌摩西管柄囊霉Funneliformis mosseae对正常供水与干旱处理的盆栽枳Poncirus trifoliata实生苗生长、活性氧代谢及抗氧化酶基因表达量的影响。结果表明,7周干旱处理显著降低了根系菌根侵染率。接种摩西管柄囊霉显著促进了干旱处理的枳植株生长,增加了根系体积和叶片相对含水量,显著降低了叶片脯氨酸含量,同时也上调了干旱处理的枳叶片精氨酸脱羧酶基因（PtADC1和PtADC2）和超氧化物歧化酶基因（PtFe-SOD和PtMn-SOD）、过氧化物酶基因（PtPOD）和过氧化氢酶基因（PtCAT1）的表达,因而维持了一个相对更低的活性氧水平（如过氧化氢）,有利于增强植株的抗旱性。     

丛枝菌根通过调节碳磷代谢相关基因的表达增强植物对低磷胁迫的适应性

丛枝菌根(AM)共生体系对于植物适应低磷胁迫具有重要作用。AM不仅直接调节宿主植物对低磷胁迫的响应,还可能通过分泌物影响相邻的非菌根植物。该研究采用分室培养系统,以玉米(Zea mays)和AM真菌Rhizophagus irregularis为试验材料,考察低磷(10 mg·kg~(–1))和高磷(100 mg·kg~(–1))条件下,菌根共生体系对植物生长、磷营养以及碳磷代谢相关基因表达的影响,以揭示AM调节植物低磷胁迫响应的生理机制。分室培养系统由0.45μm微孔滤膜分隔成供体室、缓冲室和受体室3个分室,以供体室菌根化植物为AM分泌物来源,通过微孔膜阻止菌根真菌对未接种受体植物的直接影响,但允许AM分泌物在分室间的扩散。采用实时荧光定量PCR技术分析玉米以及AM真菌自身碳磷代谢相关基因的表达情况。试验结果表明,低磷条件下接种AM真菌显著提高了供体植物干质量和磷浓度,上调了玉米碳磷代谢相关基因的表达。AM真菌磷转运蛋白基因和碳代谢相关基因在低磷条件下的表达水平显著高于高磷水平;对于受体植物而言,仅高磷处理显著提高了玉米植株干质量和磷含量,而接种处理显著上调了受体植物磷转运蛋白基因和碳代谢相关基因的表达水平。该研究表明,低磷胁迫下AM可能通过分泌物调控植物碳磷代谢相关基因的表达,进而调节植物对低磷胁迫的生理响应。     

丛枝菌根真菌对烟草香气相关物质代谢的影响

丛枝菌根(AM)真菌能够与植物共生,促进寄主植株营养元素的吸收,提高植株抗逆性,但鲜见其对香气物质代谢作用的报道。本试验研究了AM真菌摩西球囊霉对烟草叶片腺毛和香气相关物质代谢的影响。结果表明: 接种AM真菌能够增加烟草叶片腺毛的密度,并诱导烟草叶片腺体毛状体脂质分泌所必需的腺体特异性脂质转运蛋白基因(NtLTP1)表达量增加;提高香气相关物质绿原酸、茄尼醇和类胡萝卜素含量;同时促进了香气物质合成途径中关键酶苯丙氨酸解氨酶(PAL)和多酚氧化酶(PPO)活性;诱导香气相关物质代谢关键酶苯丙酰胺转氨酶和多酚氧化酶基因NtPAL和NtPPOE,以及黄酮醇合酶和角鲨烯合酶基因NtFLS和NtTSS表达上调。说明接种摩西球囊霉能够增加香气产生部位腺毛的数量和分泌活性,并促进烟草叶片香气物质的生物合成过程。     

水孔蛋白在细胞延长、盐胁迫和光合作用中的作用

水孔蛋白属于一个高度保守的、能够进行跨生物膜水分运输的通道蛋白MIP家族。水孔蛋白作为膜水通道,在控制细胞和组织的水含量中扮演重要角色。本研究的重点是属于PIP亚家族的GhPIP1;2和属于TIP亚家族的γTIP1在植物细胞延长中的作用。使用特异基因探针的Northern杂交和实时荧光PCR技术证明GhPIP1;2和GhγTIP1主要在棉花纤维延长过程中显著表达,且最高表达量在开花后5d。在细胞延长过程中,GhPIP1;2和GhγTIP1表达显著,表明它们在促使水流迅速进入液泡这一过程中扮演重要角色。而且也研究了盐胁迫植物中钙离子对水孔蛋白的影响。分别或一起用NaCl或CaCl2处理原生质体或细胞质膜。结果发现在盐胁迫条件下,水渗透率值在原生质体和质膜颗粒中都下降了,同时PIP1水孔蛋白的含量也下降了,表明NaCl对水孔蛋白的功能和含量有抑制作用。同时也观察了Ca2+的两种不同的作用。感知胁迫的胞质中游离钙离子浓度的增加可能导致水孔蛋白的关闭。而过剩的钙离子将导致水孔蛋白的上游调控。同时实验已经证明大麦的一类水孔蛋白-HvPIP2;1有更高的水和CO2转移率。本研究的目标是确定负责转运水和CO2的关键水孔蛋白...     

干旱对水稻生物钟基因和干旱胁迫响应基因每日节律性变化的影响

内源生物钟的节律运动不仅调控植物的生长发育,而且在调控植物响应和适应环境过程中发挥重要的作用。为了解水稻(Oryza sativa L.)干旱胁迫响应基因和生物钟基因在干旱条件下每日表达变化情况,本文利用实时荧光定量PCR方法研究旱稻品种IRAT109在干旱胁迫下相关基因的表达变化。结果表明,干旱胁迫导致早晨生物钟基因OsPRRs、OsLHY和OsZTL1的表达量显著下降,振幅减弱;同时导致夜晚生物钟基因OsTOC1、OsGI和OsELF3整体表达量升高,振幅增强,但对OsFKF1基因影响不大。同样,大部分水稻干旱胁迫响应基因在干旱胁迫后整体表达量显著升高,但OsDST基因表达量下降;同时大部分抗逆基因周期性表达被扰乱,但OsCIPK12、OsCDPK7和OsDREB1A依然保持24 h内震荡。本研究结果表明干旱胁迫能影响生物钟元件的基因表达,这种互相影响改变了部分基因每日的震荡变化。     

AM真菌对枳吸收磷和分泌磷酸酶的影响

研究0.1mmol/L P和1mmol/L P下接种丛枝菌根(AM)真菌Funneliformis mosseae对沙培枳Poncirus trifoliata实生苗P吸收、转运和分泌磷酸酶的影响。结果显示,在0.1mmol/L P和1mmol/L P下,接种AM真菌都增加了枳地上部和地下部干重、一级侧根和二级侧根数,但1mmol/L P下接种AM真菌显著降低了枳主根长。与不接种AM真菌相比,1mmol/L P下接种AM真菌显著降低了枳叶片酸性磷酸酶活性,但显著增加了根系酸性磷酸酶活性及根系分泌酸性磷酸酶基因Pt PAP1的表达水平。接种AM真菌显著增加了0.1mmol/L P和1mmol/L P下叶片磷含量,根系磷含量仅在1mmol/L P下接种AM真菌后显著增加。在0.1mmol/L P和1mmol/L P下接种AM真菌显著降低了枳叶片(PtPAP1和PtPAP3)和根系(PtPAP3)分泌磷酸酶基因以及叶片磷转运蛋白基因(PtaPT5和PtaPT6)的表达水平,显著增加了枳根系PtaPT3、PtaPT5和PtaPT6的表达量。因此,推断接种AM真菌能刺激枳侧根数增多和酸性磷酸酶分泌,增强磷转运蛋白基因的表达,从而改善枳对磷养分的吸收。     

盐穗木水通道蛋白基因的克隆与功能鉴定

非生物胁迫(如盐渍和干旱)会引起植物水分代谢的紊乱,导致植物细胞水分丧失,直接抑制植物的生长发育。研究水通道蛋白(aquaporins,AQPs)在非生物胁迫下对植物生理功能的影响十分重要。本研究利用RACE技术克隆获得一个盐穗木(Halostachys caspica)水通道蛋白家族亚类质膜内嵌蛋白(plasma intrinsic protein,PIPs)基因,命名为Hc PIP1。Hc PIP1基因全长序列为1 244 bp,包含858 bp的开放阅读框(open reading frame,ORF)序列,编码285个氨基酸,分子量大小约为30.6 k D。q RT-PCR检测表明Hc PIP1在盐穗木根部的表达量显著高于同化枝中,盐胁迫诱导其上调表达。在酿酒酵母INVSc1中Hc PIP1的异源过表达显著提高了重组菌的耐盐能力;和野生型植株相比过表达Hc PIP1的拟南芥能够显著缓解渗透胁迫和离子胁迫对生长的抑制,并且根长明显比野生型的长。结果表明Hc PIP1在胁迫时能够通过增加根的生长以抵御胁迫的影响。     

白花柽柳质膜水孔蛋白基因克隆及序列分析

植物水孔蛋白在植物体内形成水选择性运输通道,在植物种子萌发、细胞伸长、气孔运动、受精等过程中调节水分的快速跨膜运输。有的水孔蛋白还在干旱胁迫应答中起重要作用。本文根据白花柽柳的PEG6000胁迫处理构建的SSH消减文库的水孔蛋白基因表达序列标签(EST),设计基因特异性引物进行5′RACE,克隆出一个1 043 bp的核苷酸序列。应用生物信息学软件进行分析,预测该序列编码287个氨基酸,具有6个跨膜区,有MIP家族信号序列SGXHXNPAVT,高等植物PIP高度保守序列GGGANXXXXGY和TGI/TNPARSL/FGAAI/VI/VF/YN,这是质膜水孔蛋白基因的典型的结构特征。经NCBI比对,与Arabidopsis thaliana (MIP-C),同源性达到95%,预测该蛋白的相对分子量是30.9KD,理论等电点是8.84。     

添加物及植物组合对瞿麦幼苗铅吸收、生长及品质的影响

通过盆栽试验研究了接种丛枝菌根真菌(AM真菌)、种植共生植物(三叶草)、添加重金属螯合物EDTA和磷肥磷酸钙对铅污染下瞿麦生长和品质的影响,为科学种植中药材提供理论依据.结果表明: 接种AM真菌可以显著抑制铅的吸收(P＜0.05),促进根部发育,使根冠比增大,且活性成分累积最多,大黄素含量为6.5 mg·g^-1;与三叶草共生后,抑制铅吸收的效果不佳且药材品质下降,大黄素含量低于对照组,降至3.2 mg·g^-1.但在AM真菌共同参与后瞿麦的生长量和活性成分有所增加,铅含量达到最低,低至1.3 mg·g^-1;添加重金属螯合剂会使瞿麦生长量下降并促进根部对铅的吸收,铅含量最高可达340.0 mg·g^-1;磷酸钙可固定土壤中其他重金属元素,故更适宜在复合型污染时使用.综合考虑,在保护中药材安全、品质方面AM真菌具有良好的应用价值.     

葡萄NCED基因家族进化及表达分析

9-顺式-环氧类胡萝卜素双加氧酶(NCED)是植物体内ABA生物合成的关键限速酶, 参与植物对干旱、外源ABA和高盐的响应过程, 降低环境胁迫对植株的危害。基于全基因组鉴定分析葡萄(Vitis vinifera) NCED基因家族成员, 探讨各成员的物种进化关系及各个基因成员在不同组织中的时空表达模式及对干旱、ABA和高盐(NaCl)胁迫的响应, 为进一步揭示该基因家族成员的生物学功能奠定基础。在葡萄基因组中共发现12个NCED基因。其推测的编码蛋白质长度在510 (VvNCED2)-625 aa (VvNCED10)之间。VvNCED蛋白的分子量最大值是70.53 kDa (VvNCED10), 最小值是57.85 kDa (VvNCED2)。在从祖先基因分化之后, 葡萄NCED基因发生了5次复制事件, 同时有2次丢失事件。NCED1/2、NCED3/4、NCED6/7和NCED9/10基因对被认为是通过片段复制产生。上述4对复制基因复制时间分布在3.08-120.0百万年前, 晚于单双子叶植物分化的时间。与对照相比, VvNCED1在ABA处理48小时后显著上调(72.1%), 而VvNCED2显著下调(84.0%)。VvNCED6只在干旱处理14、21和28天的根系中表达量高于对照, 分别为对照的2.49、1.05和1.09倍。VvNCED7只在干旱处理14天的根系中表达量高于对照, 为对照的1.07倍。在ABA处理72小时后, VvNCED3表达量较对照显著下调(59.5%), 而VvNCED4较对照显著上调(169.9%)。VvNCED3/VvNCED4分别在NaCl处理24和48小时出现显著性峰值, 较对照分别上调219.2%和114.4%。保守结构域不同组成和不同胁迫处理下差异表达模式是NCED蛋白发生功能分化的基础。推测NCED在进化过程中发生的功能分化有利于复制事件的发生。     

Aquaporin genes GintAQPF1 and GintAQPF2 from Glomus intraradices contribute to plant drought tolerance

Arbuscular mycorrhizal (AM) symbiosis, established between AM fungi (AMF) and roots of higher plants, occurs in most terrestrial ecosystems. It has been well demonstrated that AM symbiosis can improve plant performance under various environmental stresses, including drought stress. However, the molecular basis for the direct involvement of AMF in plant drought tolerance has not yet been established. Most recently, we cloned two functional aquaporin genes, GintAQPF1 and GintAQPF2, from AM fungus Glomus intraradices. By heterologous gene expression in yeast, aquaporin localization, activities and water permeability were examined. Gene expressions during symbiosis in expose to drought stress were also analyzed. Our data strongly supported potential water transport via AMF to host plants. As a complement, here we adopted the monoxenic culture system for AMF, in which carrot roots transformed by Ri-T DNA were cultured with Glomus intraradices in two-compartment Petri dishes, to verify the aquaporin gene functions in assisting AMF survival under polyethylene glycol (PEG) treatment. Our results showed that 25% PEG significantly upregulated the expression of two aquaporin genes, which was in line with the gene functions examined in yeast. We therefore concluded that the aquaporins function similarly in AMF as in yeast subjected to osmotic stress. The study provided further evidence to the direct involvement of AMF in improving plant water relations under drought stresses.     

Impairment of NtAQP1 gene expression in tobacco plants does not affect root colonisation pattern by arbuscular mycorrhizal fungi but decreases their symbiotic efficiency under drought

We investigated in two tobacco (Nicotiana tabacum) plant lines (wildtype or antisense mutant) whether impairment in expression of the plasma membrane aquaporin gene (NtAQP1) affects the arbuscular mycorrhizal (AM) fungal colonisation pattern or the symbiotic efficiency of AM fungi. These two objectives were investigated under well-watered and drought stress conditions. Both plant lines had a similar pattern of root colonisation under well-watered and drought stress conditions. In contrast, under drought stress, AM wildtype plants grew faster than mycorrhizal antisense plants. Plant gas exchange also appeared to depend on the expression of NtAQP1 and parallelled the determined growth increments. The implications of enhanced symplastic water transport via NtAQP1 for the efficiency of the AM symbiosis under drought stress conditions are further discussed.     

How does arbuscular mycorrhizal symbiosis regulate root hydraulic properties and plasma membrane aquaporins in Phaseolus vulgaris under drought, cold or salinity stresses?

Here, we evaluated how the arbuscular mycorrhizal (AM) symbiosis regulates root hydraulic properties and root plasma membrane aquaporins (PIP) under different stresses sharing a common osmotic component. Phaseolus vulgaris plants were inoculated or not with the AM fungus Glomus intraradices, and subjected to drought, cold or salinity. Stress effects on root hydraulic conductance (L), PIP gene expression and protein abundance were evaluated. Under control conditions, L in AM plants was about half that in nonAM plants. However, L was decreased as a result of the three stresses in nonAM plants, while it was almost unchanged in AM plants. At the same time, PIP2 protein abundance and phosphorylation state presented the same trend as L. Finally, the expression of each PIP gene responded differently to each stress and was dependent on the AM fungal presence. Differential expression of the PIP genes studied under each stress depending on the AM fungal presence may indicate a specific function and regulation by the AM symbiosis of each gene under the specific conditions of each stress tested.     

Differential Effects of Pseudomonas mendocina and Glomus intraradices on Lettuce Plants Physiological Response and Aquaporin PIP2 Gene Expression Under Elevated Atmospheric CO2 and Drought

Arbuscular mycorrhizal (AM) symbiosis and plant-growth-promoting rhizobacterium (PGPR) can alleviate the effects of water stress in plants, but it is unknown whether these benefits can be maintained at elevated CO₂. Therefore, we carried out a study where seedlings of Lactuca sativa were inoculated with the AM fungus (AMF) Glomus intraradices N.C. Schenk & G.S. Sm. or the PGPR Pseudomonas mendocina Palleroni and subjected to two levels of watering and two levels of atmospheric CO₂ to ascertain their effects on plant physiological parameters and gene expression of one PIP aquaporin in roots. The inoculation with PGPR produced the greatest growth in lettuce plants under all assayed treatments as well as the highest foliar potassium concentration and leaf relative water content under elevated [CO₂] and drought. However, under such conditions, the PIP2 gene expression remained almost unchanged. G. intraradices increased significantly the AMF colonization, foliar phosphorus concentration and leaf relative water content in plants grown under drought and elevated [CO₂]. Under drought and elevated [CO₂], the plants inoculated with G. intraradices showed enhanced expression of the PIP2 gene as compared to P. mendocina or control plants. Our results suggest that both microbial inoculation treatments could help to alleviate drought at elevated [CO₂]. However, the PIP2 gene expression was increased only by the AMF but not by the PGPR under these conditions.     

干旱条件下AM真菌对植物生长和土壤水稳定性团聚体的影响

采用分室培养系统,模拟正常水分和干旱胁迫两种环境条件,探讨不同丛枝菌根真菌(arbuscular mycorrhizal fungi,AMF)对紫花苜蓿(Medicago sativa L.)生长和土壤水稳性团聚体的影响.试验条件下,Glomus intraradices对苜蓿根系的侵染率均显著高于Acaulospora scrobiculata和Diversispora spurcum接种处理.正常水分条件下,供试AM真菌均能显著提高植株生物量及磷浓度.干旱胁迫显著抑制了植株生长和菌根共生体发育,总体上菌根共生体对植株生长没有明显影响,接种D.spurcum甚至趋于降低植株生物量;同时,仅有G.intraradices显著提高了植株磷浓度.AM真菌主要影响到＞2mm的水稳性团聚体数量,以G.intraradices作用效果最为显著.在菌丝室中,G.intraradices显著提高了总球囊霉素含量.研究表明AM真菌对土壤大团聚体形成具有积极作用,而菌根效应因土壤水分条件和不同菌种而异,干旱胁迫下仅有G.intraradices对土壤结构和植物生长表现出显著积极作用.在应用菌根技术治理退化土壤时,需要选用抗逆性强共生效率高的菌株,对于不同AM真菌抗逆性差异的生物学与遗传学基础尚需进一步研究.     

Host- and virus-induced gene silencing of HOG1-MAPK cascade genes in Rhizophagus irregularis inhibit arbuscule development and reduce resistance of plants to drought stress

Arbuscular mycorrhizal (AM) fungi can form beneficial associations with the most terrestrial vascular plant species. AM fungi not only facilitate plant nutrient acquisition but also enhance plant tolerance to various environmental stresses such as drought stress. However, the molecular mechanisms by which AM fungal mitogen-activated protein kinase (MAPK) cascades mediate the host adaptation to drought stimulus remains to be investigated. Recently, many studies have shown that virus-induced gene silencing (VIGS) and host-induced gene silencing (HIGS) strategies are used for functional studies of AM fungi. Here, we identify the three HOG1 (High Osmolarity Glycerol 1)-MAPK cascade genes RiSte11, RiPbs2 and RiHog1 from Rhizophagus irregularis. The expression levels of the three HOG1-MAPK genes are significantly increased in mycorrhizal roots of the plant Astragalus sinicus under severe drought stress. RiHog1 protein was predominantly localized in the nucleus of yeast in response to 1 M sorbitol treatment, and RiPbs2 interacts with RiSte11 or RiHog1 directly by pull-down assay. Importantly, VIGS or HIGS of RiSte11, RiPbs2 or RiHog1 hampers arbuscule development and decreases relative water content in plants during AM symbiosis. Moreover, silencing of HOG1-MAPK cascade genes led to the decreased expression of drought-resistant genes (RiAQPs, RiTPSs, RiNTH1 and Ri14-3-3) in the AM fungal symbiont in response to drought stress. Taken together, this study demonstrates that VIGS or HIGS of AM fungal HOG1-MAPK cascade inhibits arbuscule development and expression of AM fungal drought-resistant genes under drought stress.     

Plant potassium content modifies the effects of arbuscular mycorrhizal symbiosis on root hydraulic properties in maize plants

It is well known that the arbuscular mycorrhizal (AM) symbiosis helps the host plant to overcome several abiotic stresses including drought. One of the mechanisms for this drought tolerance enhancement is the higher water uptake capacity of the mycorrhizal plants. However, the effects of the AM symbiosis on processes regulating root hydraulic properties of the host plant, such as root hydraulic conductivity and plasma membrane aquaporin gene expression, and protein abundance, are not well defined. Since it is known that K(+) status is modified by AM and that it regulates root hydraulic properties, it has been tested how plant K(+) status could modify the effects of the symbiosis on root hydraulic conductivity and plasma membrane aquaporin gene expression and protein abundance, using maize (Zea mays L.) plants and Glomus intraradices as a model. It was observed that the supply of extra K(+) increased root hydraulic conductivity only in AM plants. Also, the different pattern of plasma membrane aquaporin gene expression and protein abundance between AM and non-AM plants changed with the application of extra K(+). Thus, plant K(+) status could be one of the causes of the different observed effects of the AM symbiosis on root hydraulic properties. The present study also highlights the critical importance of AM fungal aquaporins in regulating root hydraulic properties of the host plant.     

Branched absorbing structures (BAS): a feature of the extraradical mycelium of symbiotic arbuscular mycorrhizal fungi

The present work describes the morphogenesis and cytological characteristics of 'branched absorbing structures' (BAS, formely named arbuscule-like structures, ALS), small groups of dichotomous hyphae formed by the extraradical mycelium of arbuscular mycorrhizal (AM) fungi. Monoxenic cultures of the AM fungus Glomus intraradices Smith & Schenck and tomato ( Lycopersicum esculentum Mill.) roots allowed the continuous, non-destructive study of BAS development. These structures were not observed in axenic cultures of the fungus under different nutritional conditions or in unsuccessful (asymbiotic) monoxenic cultures. However, extraradical mycelium of G. intraradices formed BAS immediately after fungal penetration of the host root and establishment of the symbiosis. The average BAS development time was 7 d under our culture conditions, after which they degenerated, becoming empty septate structures. Certain BAS were closely associated with spore formation, appearing at the spore's substending hypha. Branches of these spore-associated BAS (spore-BAS) usually formed spores. Electron microscopy studies revealed that BAS and arbuscules show several ultrastructural similarities. The possible role of BAS in nutrient uptake by the mycorrhizal plant is discussed.     

Cloning of cDNAs encoding SODs from lettuce plants which show differential regulation by arbuscular mycorrhizal symbiosis and by drought stress

In the present study three cDNA fragments were cloned using degenerate primers for Mn-sod genes and PCR: two showed a high degree of identity with Mn-sods from plants and the third with Fe-sod. Arbuscular mycorrhizal (AM) symbiosis down-regulated their expression pattern under well-watered conditions. In contrast, AM symbiosis in combination with drought stress considerably increased the expression of the Mn-sod II gene and this correlated well with plant tolerance to drought. These results would suggest that mycorrhizal protection against oxidative stress caused by drought may be an important mechanism by which AM fungi protect the host plant against drought.