芜菁花叶病毒外壳蛋白在寄主植物叶绿体中的积累及其对光系统Ⅱ活性的影响

分别以接种感染芜菁花叶病毒(TuMV)和健康时照的青菜苏州青品种(Brassica chinensis L. cv.Suzhou)和芥菜温州芥菜品种(B.juncea L.cv.Wenzhou)叶片为材料提取完整叶绿体,用胰蛋白酶消除其表面蛋白后,抽提总蛋白,经SDS-PAGE电泳和Weste rn blot检测,发现TuMV的外壳蛋白(CP)存在于感病寄主的叶绿体中.免疫金标记电镜实验显示TuMV-CP定位在感病青菜和芥菜的细胞质和叶绿体中.对两种寄主植物叶片的叶绿素荧光动力学参数测定结果显示,青菜、芥菜的Fv/F.、Fv/Fm、φPSII、qp值都有不同程度的降低,qN值增大.实验结果表明TuMV侵染后在寄主细胞叶绿体中积累的CP,抑制了光系统Ⅱ (PS Ⅱ)的光化学活性,这可能是影响寄主植物光合作用的一个重要原因.     

芜菁花叶病毒外壳蛋白在寄主植物叶绿体中的积累及其对光系统II活性的影响

分别以接种感染芜菁花叶病毒(TuMV)和健康对照的青菜苏州青品种(Brassica chinensis L. cv.Suzhou)和芥菜温州芥菜品种(B.juncea L. cv.Wenzhou)叶片为材料提取完整叶绿体,用胰蛋白酶消除其表面蛋白后,抽提总蛋白,经SDS-PAGE电泳和Western blot检测,发现TuMV的外壳蛋白(CP)存在于感病寄主的叶绿体中。免疫金标记电镜实验显示TuMV-CP定位在感病青菜和芥菜的细胞质和叶绿体中。对两种寄主植物叶片的叶绿素荧光动力学参数测定结果显示,青菜、芥菜的Fv/Fo、Fv/Fm、φPSII、qp值都有不同程度的降低,qN值增大。实验结果表明TuMV侵染后在寄主细胞叶绿体中积累的CP,抑制了光系统II(PS II)的光化学活性,这可能是影响寄主植物光合作用的一个重要原因。     

Rubisco和RCA在青菜叶绿体中的分布及病毒侵染对其细胞定位的影响

运用免疫金标电镜术观察了青菜叶细胞中光合作用关键酶Rubisco和Rubisco活化酶(RCA)的细胞化学定位,结果显示Rubisco和RCA免疫金颗粒主要分布于薄壁组织叶绿体的间质中,在基粒片层上很少,表皮的气孔保卫细胞和维管束薄壁细胞叶绿体内也有分布,在细胞质及线粒体等细胞器中无特异性分布。同时比较观察了感染芜菁花叶病毒(TuMV)的青菜叶绿体Rubisco和RCA免疫金标记结果,发现病组织中结构尚完整的叶绿体Rubisco和RCA标记率略有下降,而结构严重破坏的叶绿体中两种酶标记率分别仅为正常叶绿体的58．44％和64．67％,表明病毒侵染可导致Rubisco和RCA含量下降,影响寄主植物的光合作用。     

Rubisco和RCA在青菜叶绿体中的分布及病毒侵染对其细胞定位的影响

运用免疫金标电镜术观察了青菜叶细胞中光合作用关键酶Rubisco和Rubisco活化酶(RCA)的细胞化学定位,结果显示Rubisco和RCA免疫金颗粒主要分布于薄壁组织叶绿体的间质中,在基粒片层上很少,表皮的气孔保卫细胞和维管束薄壁细胞叶绿体内也有分布,在细胞质及线粒体等细胞器中无特异性分布。同时比较观察了感染芜菁花叶病毒(TuMV)的青菜叶绿体Rubisco和RCA免疫金标记结果,发现病组织中结构尚完整的叶绿体Rubisco和RCA标记率略有下降,而结构严重破坏的叶绿体中两种酶标记率分别仅为正常叶绿体的58.44%和64.67%,表明病毒侵染可导致Rubisco和RCA含量下降,影响寄主植物的光合作用。     

水杨酸对高温强光下小麦叶绿体蛋白酶Deg5和PSⅡ功能的调节作用

以小麦(Triticum aestivum)矮抗58为材料,采用0.1mmol/L的外源水杨酸(SA)处理小麦叶片,以清水为对照,通过Western blotting蛋白质印记技术和叶绿素荧光分析,研究了高温强光胁迫(38℃和1600μmol m-2s-1)对小麦叶绿体Deg5蛋白酶、D1蛋白和叶绿素荧光参数的影响及SA的调节作用。结果表明,高温强光胁迫导致小麦叶绿体Deg5蛋白酶、D1蛋白含量和PSⅡ最大光能转化效率(Fv/Fm)降低,原初荧光(Fo)升高。和对照相比,外源SA处理可维持较高的Deg5蛋白酶、D1蛋白、Fv/Fm水平和较低的Fo。说明外源水杨酸可减轻高温强光对Deg5蛋白酶和D1蛋白的损伤,维持较强的PSⅡ功能。     

寄主抗性与黄龙病寄主植物次生化合物相关性初步探讨

本文通过使用甲醇浸泡浸提和气相色谱-质谱连用(GC-MS)方法,对黄龙病易感寄主椪柑Citrus reticulata Blanco cv. Ponkan的健叶、病叶以及隐症寄主九里香Murraya paniculata(L.) Jack健叶中部分次生化合物的种类及相对含量进行了差异分析。结果显示:椪柑的健康叶片鉴定出32种次生化合物,感病叶片有31种,九里香健康叶片有21种。其中脂肪酸类和简单芳香族类化合物在3样本之间未显示差异;烷烃类化合物在椪柑健叶中含量较高,椪柑病叶和九里香健叶中相对较少;萜类、黄酮类和香豆素类化合物在椪柑健病间的含量相差无几,其中萜类和黄酮类显著多于九里香,而香豆素类则相反。萜类化合物α-石竹烯在椪柑健叶中未检出,但在其病叶和九里香健叶之间无显著差异,此外九里香中特有的萜类有α-姜黄烯(0.15%)、左旋-姜烯(1.00%)、β-倍半水芹烯(0.26%);特有且含量较高的香豆素类化合物主要为蛇床子素(13.97%);而黄酮类化合物在九里香中未检出。两种寄主植物之间以及同种植物在感病与不感病的状态下,次生化合物的种类及相对含量均有显著差异,并初步探讨了其与寄主抗性的相关性。     

高温强光对小麦叶绿体Deg1蛋白酶和D1蛋白的影响及水杨酸的调节作用

以小麦品种矮抗58为材料,采用0.3 mmol/L水杨酸(SA)溶液预处理灌浆期小麦叶片,以水预处理为对照,进行3种不同的光温处理:适宜温度中等光强(25℃,600 μmol m-2 s-1)2h、高温强光(38℃,1600μmol m-2 s-1)2h、高温强光2h后置于适宜温度中等光强下恢复3h.测定不同光温条件下,小麦叶绿体的Deg1蛋白酶、D1蛋白和PSⅡ功能的变化及SA的调节效应.结果表明,高温强光胁迫导致Deg1蛋白酶和D1蛋白降解,PSⅡ功能发生可逆损伤.与对照相比,水杨酸预处理不仅能够抑制高温强光下小麦叶绿体Deg1蛋白酶和D1蛋白的降解,维持较高的PSⅡ原初光化学效率(Fv/ Fm)、实际光化学效率(φPSⅡ)、电子传递速率和净光合速率(Pn),而且加快回到非逆境下PSⅡ功能的恢复.     

外源多胺对盐胁迫下玉米叶绿体结合态多胺水平和光合作用的影响

在100 mm o l/L N aC l胁迫下,研究了外源多胺-腐胺(Pu t)、尸胺(C ad)、亚精胺(Spd)和精胺(Spm)对玉米幼苗生长、光合速率和PSⅡ光化学效率(Fv/Fm)、叶绿体结合多胺和叶片丙二醛(M DA)含量以及抗氧化酶活性的影响.结果表明,叶面喷施1 mm o l/L的Pu t、Spd和Spm可显著增加盐胁迫下玉米幼苗干物质重、叶绿体内结合态多胺的含量、叶片净光合速率和PSⅡ光化学效率(Fv/Fm),并降低了叶片中M DA含量.外源Spd和Spm明显增加盐胁迫下玉米幼苗的抗坏血酸过氧化物酶(APX)、过氧化物酶(POD)和过氧化氢酶(CAT)活性;外源Pu t可增强APX和POD活性,对CAT活性的影响不明显.这些结果表明多胺对玉米盐害的缓解作用可能是由于提高了叶绿体中结合态多胺的含量和叶片抗氧化酶的活性,从而增强了盐胁迫下的玉米光合能力.     

高温胁迫对柑橘光合速率和光系统Ⅱ活性的影响

用红外CO2分析仪和叶绿素荧光仪测定了温州蜜柑和脐橙叶片的净光合速率(Pn)、初始荧光(Fo)、最大光能转换效率(Fv／Fm)及电子传递速率(ETR)．结果表明,与常温(25℃)相比,高温胁迫(38～40℃)使温州蜜柑和脐橙叶片的Pn、Fv／Fm及ETR下降,Fo升高．胁迫25d后温州蜜柑和脐橙叶片的Pn分别下降55．6％和39．8％．Fv／Fm下降22．0％和6．7％,ETR下降55．0％和41．5％,Fo分别上升了113．8％和14．9％．柑橘经高温胁迫后,在25℃下处理10d,叶片的Pn、Fv／Fm、Fo及ETR恢复明显．这些结果说明柑橘的光合速率下降与PSⅡ反应中心失活有关．     

外源硅对纹枯病菌(Rhizoctonia solani)侵染下水稻叶片光合功能的改善

为了进一步探讨外源加硅增强水稻对纹枯病的抗性作用,以抗病品种91SP和感病品种Lemont为材料,研究了人工接种纹枯病菌条件下外源硅对水稻叶片叶绿素含量、光合作用、叶绿素荧光特性和MDA含量的影响。结果表明：（1）外源加硅能降低抗病品种91SP的纹枯病病级和病情指数,显著降低感病品种Lemont的病级和病情指数;（2）接种纹枯病菌后,水稻叶片叶绿素含量、净光合速率（Pn）、气孔导度（Gs）均明显降低,胞间CO2浓度（Ci）增大,而加硅处理的水稻叶片叶绿素含量、Pn、Gs不同程度增加,Ci有所降低;（3）接种纹枯病菌后,两个品种PSⅡ最大光化学效率（Fv／Fm）、PSⅡ有效光化学效率（Fv＇／Fm＇）、PSⅡ实际光化学效率（ФPSⅡ）、光化学猝灭系数（qP）和表观光合电子传递速率（ETR）均降低,非光化学猝灭系数（qNP）增大,而对于加硅处理的水稻叶片,上述荧光参数在纹枯病菌侵染条件下的变化均受到不同程度的抑制。（4）外源硅可不同程度地减缓纹枯病菌侵染引起的丙二醛（MDA）含量的增加,对感病品种Lemont的缓解作用要大于抗病品种91SP。可见,外源硅处理可以不同程度地缓解纹枯病菌侵染条件下非气孔因素引起的水稻叶片光合速率的下降以及对光合机构的破坏作用,提高光化学效率,改善叶片的光合功能,减轻叶片膜脂过氧化程度,增强水稻对纹枯病的抗性。     

蚕豆萎蔫病毒2号分离物侵染对蚕豆叶片光合活性和叶绿体超微结构的影响

研究了受蚕豆萎蔫病毒2号（Broad bean wilt virus 2,BBWV2）中国分离物B935和欧洲分离物PV131侵染的蚕豆（Vicica faba）叶片光合特性、叶绿素荧光诱导动力学参数和叶绿体超微结构变化。感病蚕豆叶绿素含量减少,叶绿素a／b比逐步降低;光合气体交换参数Pn值和Gs值降低,Ci值升高;叶绿素荧光诱导动力学参数Fv/Fm、FV＇/Fm＇、ΦPSII、qP值均有不同程度降低,NPQ值升高;光合器结构遭到不同程度的破坏,B935侵染后叶绿体发育不良,片层结构疏松,PVl31侵染后叶绿体肿胀变圆,片层结构疏松瓦解。与B935相比,PV131侵染对以上各参数的变化有更大影响,且对叶绿体的破坏更为严重。实验结果表明BBWV2不同分离物对光系统II（PSII）的抑制作用与光合器受损程度相关。     

Drought-induced changes in photosynthetic membranes of two wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis> L.) cultivars

Two wheat (Triticum aestivum L.) cultivars contrasting in architectonics and differing in drought resistance, Azamatli-95 (short stature, vertically oriented small leaves, drought-tolerant) and Giymatli-2/17 (short stature, broad and drooping leaves, drought-sensitive), were studied. It was found that the content of CP I (115 kDa) and 63-kDa apoprotein P700 and also LHC II polypeptides increases slightly in the drought-resistant cv. Azamatli-95 under extreme water supply limitation, while their content decreases in drought-sensitive cv. Giymatli-2/17. The intensity of synthesis of α- and β-subunits of CF₁ (55 and 53.5 kDa) and 33–30.5 kDa proteins also decreases in the sensitive genotype. The intensity of short wavelength peaks at 687 and 695 nm sharply increases in the fluorescence spectra (77K) of chloroplasts from Giymatli-2/17 under water deficiency, and there is a stimulation of the ratio of fluorescence band intensity F687/F740. After exposure to drought, cv. Giymatli-2/17 shows a larger reduction in the actual PS II photochemical efficiency of chloroplasts than cv. Azamatli-95. Published in Russian in Biokhimiya, 2009, Vol. 74, No. 8, pp. 1109–1116.     

Rapid immunofluorescent determination of cells in the S phase in pea root meristems: An alternative to autoradiography

Photosystem II (PS II) activity and the localization of ribulose-l,5-bisphosphate (RuBP) carboxylase (EC 4.1.1.39) were studied in primary leaves of young maize plants ( Zea mays L. cv. Fronica) by tetra-nitro-blue-tetrazoliumchloride reduction and immunolocalization, respectively. In tissue of 3-day-old plants all chloroplasts were structurally identical. From day 4 they developed into their typical appearance of mesophyll and bundle sheath chloroplasts. First PS II-activity was present in both types of chloroplasts. From day 4 it disappeared in bundle sheath chloroplasts concomitant with the loss of grana. RuBP carboxylase on the other hand was only present in bundle sheath chloroplasts at all stages of development. Thus, the control of the development of the photosystems and the Calvin cycle enzymes seem to differ.     

Structural-functional state of thylakoid membranes of wheat genotypes under water stress

Plants were grown in field conditions in the wide area under normal water supply and severe water deficit. Two wheat (Triticum aestivum L.) genotypes contrasting by architectonics and differing in drought-resistance were used: Giymatli-2/17, short stature, with broad and drooping leaves, drought-sensitive, and Azamatli-95, short stature, with vertically oriented small leaves, drought-tolerant). It was found out that Giymatli-2/17 was characterized by relatively low content of Chl a-protein of PS I (CP I) and beta-subunit of ATP-synthase complex, the high content of proteins in the 33-30.5 kDa region and LHC polypeptides (28-24.5 kDa), the intensive fluorescence at 740 nm and more high photochemical activity of PS II under normal irrigation compared with Azamatli-95. However, the content of CP I (M(r) 115 kDa) and apoprotein of P700 with M(r) 63 kDa insignificantly increases in the drought-resistant genotype Azamatli-95 under extreme water supply condition while their content decreases in drought-sensitive cv Giymatli-2/17. Intensity of synthesis alpha- and beta-subunits of CF(1) (55 and 53.5 kDa) also decreases in Giymatli-2/17. The levels of the core antenna polypeptides of FS II with M(r) 46 and 44.5 kDa (CP47 and CP43) remains stable both in normal, and stressful conditions. At the same time the significant reduction is observed in the content of polypeptides in the 33-30.5 kDa region in the more sensitive genotype Giymatli-2/17. There is an increase in the LHC II polypeptides level in tolerant genotype Azamatli-95 in contrast to Giymatli-2/17 (where the content of these subunits is observed decreasing). The intensity of short wavelength peaks at 687 and 695 nm sharply increases in the fluorescence spectra (77 K) of chloroplasts from sensitive genotype Giymatli-2/17 under water deficiency and there is a stimulation of the ratio of fluorescence band intensity F687/F740. After exposure to drought, cv Giymatli-2/17 shows a larger reduction in the actual PS II photochemical efficiency of chloroplasts than cv Azamatli-95.     

The reactions of swede (Brassica napus) to infection by turnip mosaic virus

Turnip mosiac virus (TuMV) and cucumber mosaic virus (CMV) were the only viruses commonly isolated from naturally diseased swedes (Brassica napus) showing leaf mosaic and leaf and root necrosis. Only TuMV caused these symptoms when re-inoculated to swede. TuMV-infected plants showed a severe loss in leaf (55%) and root (63%) fresh weight after 140 days. Systemic leaf symptoms in infected swede plants varied greatly, but were predominantly necrotic (N) or mosaic (M). Plants were classified into one of seven reaction classes ranging from slight or severe necrosis, and mosaics with and without slight veinal necrosis. Swede cultivars differed markedly in their reaction to TuMV and contained different proportions of N- or M-reacting plants. The reactions of progeny of four resistant cv. Bangholm plants were separately inherited; progeny of two plants reacting either symptomlessly or necrotically and those of the other two plants developing mosaic symptoms only. Five isolates of TuMV from swede crops in different regions caused similar reactions but differed in virulence in the progeny of a self-pollinated resistant swede plant.     

绿斑病藻寄生对夏橙叶片光合作用特性的影响

以盆栽的2年生奥灵达夏橙为试材,研究了绿斑病藻寄生对夏橙叶片光合作用特性的影响.结果表明,轻度病叶对叶绿素总量（Chl a+b）、类胡萝卜素含量（Car）、净光合速率（P_n）、胞间CO₂浓度（C_i）、原初光能转换效率（F_v/F_m）、光合电子传递量子效率（ΦPS Ⅱ）和光化学猝灭系数（qP）无显著影响;中度病叶和重度病叶的Chla+b、Car、P_n、F_v/F_m、ΦPS Ⅱ和qP较对照分别下降了23.85%、26.49%、43.3%、4.5%、35.1%、22.5%和37.61%、44.04%、64.5%、8.6%、63.6%、40.1%,与对照差异显著,而C_i较对照显著上升.绿斑病藻的大量寄生减弱了夏橙叶片的光合作用,而净光合速率的下降主要是由非气孔因素限制引起.     

Protective action of salicylic acid against bean yellow mosaic virus infection in Vicia faba leaves

In this study, morphological, ultrastructural and physiological modifications of faba bean (Vicia faba cv Giza 461) leaves in response to bean yellow mosaic virus (BYMV) infection and salicylic acid (SA) treatments were examined. Under BYMV stress, leaves showed symptoms including severe mosaic, mottling, crinkling, size reduction and deformations. Three weeks after virus inoculation, photosynthetic rate, pigment contents and transpiration rate were significantly reduced in response to BYMV infection.

Ultrastructural investigations of BYMV-infected leaves demonstrated that most chloroplasts with increased stromal area became spherical in shape and some lost their envelopes, either partially or totally. The internal structures of chloroplast, grana and thylakoids were dilated. Two kinds of inclusions were detected in BYMV-infected leaves: straight or slightly curved bands sometimes coiled or looped at the end, and electron opaque crystals with varied shapes. BYMV-infected cells showed lower chloroplast number in comparison to the control.

Spraying of SA on faba bean leaves helped to reduce or prevent the harmful effects produced after virus infection. Application of 100 μM SA three days before inoculation restored the metabolism of infected leaves to the levels of healthy controls. SA treatment improved plant health by increasing the photosynthesis rates, pigment contents and levels of other parameters studied similar to control values.

Moreover, SA treatment increased plant resistance against BYMV. This was observed through induction of chloroplast number, reduction in percentage of infected plants, decrease in disease severity and virus concentration of plants treated with SA prior to BYMV inoculation. Cells of SA-treated samples showed well-developed chloroplasts with many starch grains and well-organized cell organelles.

The present results provide an overview of the negative effects on faba bean leaves due to BYMV infection from physiological and subcellular perspectives. Also, a role of SA involved in induction of resistance against BYMV infection in bean plants is discussed.