玉米C4光合叶不同部位解剖结构和光抑制特性的比较

以玉米第54i全展叶（C4光合叶）为材料,分别测定基部、中部和顶部的光合速率后,将叶片置于强光（2000μmol·m^2·s^-1）下处理3h和暗中恢复3h,再测定这3个部位在处理期间的叶绿素荧光参数变化;然后分别从叶片的基部、中部和顶部取样观察显微结构和超微结构,测定叶绿素含量。结果表明,3个部位光合速率和叶绿素含量的大小依次为：中部〉顶部〉基部。基部的维管束鞘细胞叶绿体数量少,体积小,排列无规律,类囊体膜有部分垛叠;中部和顶部维管束鞘细胞叶绿体数量多,体积大,大部分围绕维管束呈离心排列,类囊体膜垛叠消失。在强光下,基部、中部和顶部均发生光抑制,但光抑制程度不同,根据严重度依次为：基部〉顶部〉中部,3个部位在暗中的光抑制恢复能力依次为：中部〉顶部〉基部。与叶基部相比,叶中部在强光下能维持较高的电子传递效率（φEo）和较低的热耗散比率（φDo）。这表明,C4光合循环是保持较高电子传递效率、减轻光抑制的重要因子。     

莲子光下萌发过程中光合膜超分子结构与27kD多肽变化

冰冻撕裂电镜观察及膜多肽组分的研究结果表明,随着莲子在光下萌发时间的延长,莲(Nelumbonucifera Gaertn.)胚芽叶的叶绿体光合膜的超分子结构发育与膜多肽组分中的27kD多肽含量变化具有明显的相关性:1.萌发2d后,胚芽叶的叶绿体巨基位变成解垛叠状态,其光合膜的超分子结构只呈现解垛叠类囊体区外质膜撕裂面(EF)和解垛叠类囊体的原生质膜撕裂面(PF)两个面;膜组分中主要是30kD多肽,而27kD多肽含量甚微。2.萌发4d后,光合膜从解垛叠开始转变成小基粒垛,垛叠区类囊体外质膜撕裂面(EFs)和垛叠类囊体的原生质膜撕裂面(PFs)开始发育;27kD多肽含量开始增加,30kD多肽含量开始减少。3.萌发6～8d后,光合膜明显分化出非垛叠膜区,非垛叠类囊体的外质膜撕裂面(EFu)和非垛叠类囊体的原生质膜撕裂面(PFu)开始呈现,EFs和PFs功能蛋白颗粒逐渐增多;27kD多肽逐渐增加,30kD多肽逐渐减少。4.萌发10～12d后,光合膜垛叠和非垛叠膜区分化完善,排列有序,EFs、PFs、EFu和PFu面功能蛋白颗粒的密度、大小、分布等超分子构象发育正常;27kD多肽更加增多,30kD多肽几乎消失。表明其超分子结构的发育动态既与其超微结构变化相一致,又与27kD多肽含量变化相吻合,却与一般高等植物的叶绿体发育相反,可为显示莲在被子植物系统演化中的独特地位提     

不同光质下生长的满江红叶绿体和共生者满江红鱼腥藻的吸收光谱和荧光光谱的研究

测定了不同光质下培养的满江红的叶绿体和满江红鱼腥藻的吸收光谱和荧光光谱。吸收光谱表明:对于前者叶绿体,红光和蓝光比白光和绿光更有利于Chl a的形成。对于后者光对其色素的影响比叶绿体更为敏感。在白光下其胆藻素含量最高,在红光下P_(330)含量最高,依次为蓝光、白光、绿光;而类胡萝卜素相对稳定。荧光发射光谱表明:红光下培养的满江红的叶绿体PS Ⅱ荧光发射最强,蓝光、白光、绿光依次减弱。这与我们在电镜下观察到叶绿体膜垛叠的结果是一致的。而在满江红鱼腥藻中,不同光质下的差异与叶绿体不同。从荧光光谱和类囊体膜垛叠的分析表明,我们从另一方面支持了Arntzen等(1977)关于系统Ⅱ颗粒主要分布在基粒膜上的观点。     

玉米C4光合叶不同部位解剖结构和光抑制特性的比较

以玉米第5位全展叶(C4光合叶)为材料,分别测定基部、中部和顶部的光合速率后,将叶片置于强光(2000μmol·m-2·s-1)下处理3h和暗中恢复3h,再测定这3个部位在处理期间的叶绿素荧光参数变化;然后分别从叶片的基部、中部和顶部取样观察显微结构和超微结构,测定叶绿素含量。结果表明,3个部位光合速率和叶绿素含量的大小依次为:中部>顶部>基部。基部的维管束鞘细胞叶绿体数量少,体积小,排列无规律,类囊体膜有部分垛叠;中部和顶部维管束鞘细胞叶绿体数量多,体积大,大部分围绕维管束呈离心排列,类囊体膜垛叠消失。在强光下,基部、中部和顶部均发生光抑制,但光抑制程度不同,根据严重度依次为:基部>顶部>中部,3个部位在暗中的光抑制恢复能力依次为:中部>顶部>基部。与叶基部相比,叶中部在强光下能维持较高的电子传递效率(φEo)和较低的热耗散比率(φDo)。这表明,C4光合循环是保持较高电子传递效率、减轻光抑制的重要因子。     

光周期对光敏核不育水稻叶绿体类囊体膜超分子结构的影响

通过冰冻撕裂叶绿体类囊体膜电镜术的研究,在超分子水平上揭示出长日照（ＬＤ）与短日照（ＳＤ）对突变型光敏核雄性不育水稻农垦５８Ｓ及其野生型农垦５８ 倒二叶叶绿体类囊体膜的形成有不同的影响。农垦５８－ＳＤ 和农垦５８－ＬＤ差别不明显;而农垦５８Ｓ－ＳＤ和农垦５８Ｓ－ＬＤ之间出现较明显的差异：（１）农垦５８Ｓ－ＳＤ 叶绿体类囊体膜的垛叠和非垛叠膜区的４ 个冰冻撕裂膜小面呈现的功能蛋白粒的密度、大小及构象分布与其对照农垦５８－ＳＤ 和农垦５８－ＬＤ 类似,均属正常的超分子结构类型。（２）农垦５８Ｓ－ＳＤ叶绿体垛叠类囊体膜区的外质撕裂面（ＥＦｓ）功能蛋白粒的密度比５８Ｓ－ＬＤ的大,且在有的垛叠膜区呈现出类晶格状的规则排列。（３）农垦５８Ｓ－ＬＤ 叶绿体非垛叠膜区的原生质撕裂面（ＰＦｕ）和外质撕裂面（ＥＦｕ）出现频率较低,往往在基粒的边缘膜或末端中能找到,且分布于膜上的功能蛋白粒的密度较小。有的叶绿体ＥＦｕ、ＰＦｕ、ＥＦｓ和垛叠膜区原生质撕裂面（ＰＦｓ）功能蛋白粒极少或丧失,表明类囊体膜受到严重损伤     

定位于叶绿体的玉米ZmFDR3参与铁转运

铁是植物代谢中一种必需的营养元素,植物缺铁会表现出失绿等症状.ZmFDR3（Zea maize Fe-deficiency-related）是从缺铁诱导的玉米根cDNA文库中筛选到的铁转运相关基因.玉米根中,缺铁胁迫下ZmFDR3加强表达.异源互补实验表明,ZmFDR3与铁转运有关.序列分析表明,ZmFDR3蛋白与细菌Ⅲ型分泌系统的FliN有同源结构域,并预测定位在叶绿体类囊体中.通过转基因烟草的荧光免疫细胞定位,ZmFDR3主要存在于根、茎、叶的质体,尤其是保卫细胞的叶绿体中;转基因烟草的光合指标高于野生型;转基因烟草类囊体的基质片层垛叠较野生型的紧密;测定叶片、种子铁锌含量发现转基因烟草的铁含量高于野生型.因此,推测ZmFDR3定位在叶绿体中,参与叶绿体的铁转运。     

低浓度尼日利亚菌素或氯化铵促进ATP形成机理的再探

在低盐介质中,含有垛叠类囊体（基粒）与不垛叠类囊体（间质片层）结构的叶绿体进行的光系统Ｉ电子传递与磷酸化反应（ＰＳＰ）都为低浓度的尼日利亚菌素所促进,低浓度的氯化铵对基粒结构叶绿体的系统Ｉ与包括两个系统的电子传递以及与之相偶联的磷酸化反应有促进效应,而对时间质片偶联的磷酸反应有促进效应,尼日利亚菌素或氯化铵对ＰＳＰ的促进效应在高盐介质中消失,且它们对高能态（Ｚ）形成的抑制效应在低盐介质中较高盐介质     

类囊体膜的垛叠、松散与它的功能关系

菠菜完整叶绿体置于4mM MgCl_2或20 mM KCl低浓度介质中低渗10秒钟后,得到由Mg~(++)或K~+离子诱导的类囊体垛叠膜和松散膜。它们在功能上表现出明显的差异。垛叠膜有较高的毫秒级延迟光发射(ms-DLE),松散膜显著降低DLE的快相,垛叠膜比松散膜的9-AA荧光猝灭快,并保持稳定;而松散膜有H~+渗漏。在非循环或Fd催化的循环光合磷酸化中,垛叠膜比松散膜活力高。但是,若在同样的低渗介质中低渗1分钟以上,Mg~(++)离子诱导的垛叠膜,在显微结构上不同于低渗过10秒钟的垛叠膜,它垛叠较松,而且在磷酸化活力上也与松散膜差别不大。揭示了H~+传递速度受二个光系统、电子载体间的距离及偶联程度的限制。新鲜制备的垛叠或松散膜,在NADP~+还原系统中,具有相同的电子传递放O_2速度,说明电子传递速度在一定范围内不受膜间的距离和偶联程度的影响。但是松散膜不稳定,随着膜的老化而解联,牛血清白蛋白(BSA)能稳定松散膜的电子传递。     

Hg2+对菠菜离体类囊体膜光化学活性和多肽组分的影响

重金属Ｈｇ＾２＋对菠菜（Ｓｐｉｎａｃｉａ ｏｌｅｒａｃｅａ Ｌ．）离体类囊体膜的光合电子传递活性、室温吸收光谱、室温荧光发射光谱以及多肽组分影响的研究结果表明：Ｈｇ＾２＋对两个光系统的电子传递活性都有抑制作用,且Ｈｇ＾２＋对ＰＳＩ的抑制作用较ＰＳⅡ大;Ｈｇ＾２＋处理使类囊体膜的室温吸收光谱峰及室温荧光发射峰降低,但未使类囊体膜的多肽组分发生改变。     

墨兰幼叶和成熟叶不同部位叶绿体超微结构和光合作用

墨兰试管苗植株成熟叶片叶绿体基粒较发达,类囊本膜垛叠较紧密。幼叶叶绿体中少有亲锇颗粒,成熟叶的叶绿体中往往既有亲锇颗粒又有淀粉粒。幼叶中基粒数目比成熟叶的少,叶绿体也比成熟叶的小。幼叶的光合放氧速率比成熟叶的低。幼叶中叶尖部叶绿体最大而叶基部最小,但叶尖部的光合放氧速率比叶基部小。成熟叶中叶绿体大小及光合放氧速率区别不明显。通过对各部位叶绿素含量的测定发现,叶绿素含量与光合放氧速率之间没有正相关性     

Differential changes in degradation of chlorophyll-protein complexes of photosystem I and photosystem II during flag leaf senescence of rice.

The stability of chlorophyll-protein complexes of photosystem I (PSI) and photosystem II (PSII) was investigated by chlorophyll (Chl) fluorescence spectroscopy, absorption spectra and native green gel separation system during flag leaf senescence of two rice varieties (IIyou 129 and Shanyou 63) grown under outdoor conditions. During leaf senescence, photosynthetic CO(2) assimilation rate, carboxylase activity of Rubisco, chlorophyll and carotenoids contents, and the chlorophyll a/b ratio decreased significantly. The 77 K Chl fluorescence emission spectra of thylakoid membranes from mature leaves had two peaks at around 685 and 735 nm emitting mainly from PSII and PSI, respectively. The total Chl fluorescence yields of PSI and PSII decreased significantly with senescence progressing. However, the decrease in the Chl fluorescence yield of PSI was greater than in the yield of PSII, suggesting that the rate of degradation in chlorophyll-protein complexes of PSI was greater than in chlorophyll-protein complexes of PSII. The fluorescence yields for all chlorophyll-protein complexes decreased significantly with leaf senescence in two rice varieties but the extents of their decrease were significantly different. The greatest decrease in the Chl fluorescence yield was in PSI core, followed by LHCI, CP47, CP43, and LHCII. These results indicate that the rate of degradation for each chlorophyll-protein complex was different and the order for the stability of chlorophyll-protein complexes during leaf senescence was: LHCII>CP43>CP47>LHCI>PSI core, which was partly supported by the green gel electrophoresis of the chlorophyll-protein complexes.     

Heat stress induces an inhibition of excitation energy transfer from phycobilisomes to photosystem II but not to photosystem I in a cyanobacterium Spirulina platensis.

The effects of high temperature (30-52.5 degrees C) on excitation energy transfer from phycobilisomes (PBS) to photosystem I (PSI) and photosystem II (PSII) in a cyanobacterium Spirulina platensis grown at 30 degrees C were studied by measuring 77 K chlorophyll (Chl) fluorescence emission spectra. Heat stress had a significant effect on 77 K Chl fluorescence emission spectra excited either at 436 or 580 nm. In order to reveal what parts of the photosynthetic apparatus were responsible for the changes in the related Chl fluorescence emission peaks, we fitted the emission spectra by Gaussian components according to the assignments of emission bands to different components of the photosynthetic apparatus. The 643 and 664 nm emissions originate from C-phycocyanin (CPC) and allophycocyanin (APC), respectively. The 685 and 695 nm emissions originate mainly from the core antenna complexes of PSII, CP43 and CP47, respectively. The 725 and 751 nm band is most effectively produced by PSI. There was no significant change in F725 and F751 during heat stress, suggesting that heat stress had no effects on excitation energy transfer from PBS to PSI. On the other hand, heat stress induced an increase in the ratio of Chl fluorescence yield of PBS to PSII, indicating that heat stress inhibits excitation energy transfer from PBS to PSII. However, this inhibition was not associated with an inhibition of excitation energy transfer from CPC to APC since no significant changes in F643 occurred at high temperatures. A dramatic enhancement of F664 occurring at 52.5 degrees C indicates that excitation energy transfer from APC to the PSII core complexes is suppressed at this temperature, possibly due to the structural changes within the PBS core but not to a detachment of PBS from PSII, resulting in an inhibition of excitation energy transfer from APC to PSII core complexes (CP47 + CP43). A decrease in F685 and F695 in heat-stressed cells with excitation at 436 nm seems to suggest that heat stress did not inhibit excitation energy transfer from the Chl a binding proteins CP47 and CP43 to the PSII reaction center and the decreased Chl fluorescence yields from CP43 and CP47 could be explained by the inhibition of the energy transfer from APC to PSII core complexes (CP47 + CP43).     

Light-dependent reversal of dark-chilling induced changes in chloroplast structure and arrangement of chlorophyll-protein complexes in bean thylakoid membranes

Changes in chloroplast structure and rearrangement of chlorophyll-protein (CP) complexes were investigated in detached leaves of bean (Phaseolus vulgaris L. cv. Eureka), a chilling-sensitive plant, during 5-day dark-chilling at 1 °C and subsequent 3-h photoactivation under white light (200 μmol photons m⁻² s⁻¹) at 22 °C. Although, no change in chlorophyll (Chl) content and Chl a/b ratio in all samples was observed, overall fluorescence intensity of fluorescence emission and excitation spectra of thylakoid membranes isolated from dark-chilled leaves decreased to about 50%, and remained after photoactivation at 70% of that of the control sample. Concomitantly, the ratio between fluorescence intensities of PSI and PSII (F736/F681) at 120 K increased 1.5-fold upon chilling, and was fully reversed after photoactivation. Moreover, chilling stress seems to induce a decrease of the relative contribution of LHCII fluorescence to the thylakoid emission spectra at 120 K, and an increase of that from LHCI and PSI, correlated with a decrease of stability of LHCI-PSI and LHCII trimers, shown by mild-denaturing electrophoresis. These effects were reversed to a large extent after photoactivation, with the exception of LHCII, which remained partly in the aggregated form. In view of these data, it is likely that dark-chilling stress induces partial disassembly of CP complexes, not completely restorable upon photoactivation. These data are further supported by confocal laser scanning fluorescence microscopy, which showed that regular grana arrangement observed in chloroplasts isolated from control leaves was destroyed by dark-chilling stress, and was partially reconstructed after photoactivation. In line with this, Chl a fluorescence spectra of leaf discs demonstrated that dark-chilling caused a decrease of the quantum yield PSII photochemistry (F_v/F_m) by almost 40% in 5 days. Complete restoration of the photochemical activity of PSII required 9 h post-chilling photoactivation, while only 3 h were needed to reconstruct thylakoid membrane organization and chloroplast structure. The latter demonstrated that the long-term dark-chilled bean leaves started to suffer from photoinhibition after transfer to moderate irradiance and temperature conditions, delaying the recovery of PSII photochemistry, independently of photo-induced reconstruction of PSII complexes.     

Immunological characterization of chlorophyll a/b-binding proteins of barley thylakoids

Monoclonal antibodies have been raised against the light-harvesting chlorophyll a/b-binding proteins of photosystem I (LHCI) using a photosystem (PS) I preparation (PSI-200) wild-type from barley (Hordeum vulgare L. cv. Svaløf's Bonus) as the antigen. These antibodies cross-reacted with a minor light-harvesting chlorophyll a/b-protein of PSII (Chla/b-P1=CP29), but not with the major one, LHCII (=Chla/b-P2^**). Similarly, a monoclonal antibody to Chla/b-P1, elicited by a PSII preparation as the antigen, cross-reacted with LHCI, but not LHCII. This explains why an antigen consisting of LHCII, free of LHCI, but contaminated with Chla/b-P1, can elicit antibodies which cross-react with LHCI. Immunoblot assays showed that LHCI and Chla/b-P1 have at least two epitopes in common. Immunogold labelling of thin-sectioned wild-type thylakoids confirmed a preferential localisation of Chla/b-P1 in grana partition membranes and LHCI in stroma lamellae. The presence of LHCI was demonstrated in barley mutants lacking the PSI reaction centre (viridis-zb ⁶³) and chlorophyll b (chlorina-f2), and was correlated with the presence of long-wavelength (730 nm) fluorescence emission at 77 K. The mutant viridis-k ²³, which has a 77 K long-wavelength fluorescence peak at 720 nm, was shown by immune-blot assay to lack LHCI, although Chla/b-P1 was present.Abbreviations Chl-P chlorophyll-protein - CM Carlsberg Monoclonal - Da dalton - LHC light-harvesting complex - PAGE polyacrylamide gel electrophoresis - PSI, II photosystem I, II - PSI-200 PSI containing LHCI polypeptides - SDS sodium dodecyl sulphate     

Interaction of phycobilisomes with photosystem II dimers and photosystem I monomers and trimers in the cyanobacterium Spirulina platensis.

Distribution of phycobilisomes between photosystem I (PSI) and photosystem II (PSII) complexes in the cyanobacterium Spirulina platensis has been studied by analysis of the action spectra of H2 and O2 photoevolution and by analysis of the 77 K fluorescence excitation and emission spectra of the photosystems. PSI monomers and trimers were spectrally discriminated in the cell by the unique 760 nm low-temperature fluorescence, emitted by the trimers under reductive conditions. The phycobilisome-specific 625 nm peak was observed in the action spectra of both PSI and PSII, as well as in the 77 K fluorescence excitation spectra for chlorophyll emission at 695 nm (PSII), 730 nm (PSI monomers), and 760 nm (PSI trimers). The contributions of phycobilisomes to the absorption, action, and excitation spectra were derived from the in vivo absorption coefficients of phycobiliproteins and of chlorophyll. Analyzing the sum of PSI and PSII action spectra against the absorption spectrum and estimating the P700:P680 reaction center ratio of 5.7 in Spirulina, we calculated that PSII contained only 5% of the total chlorophyll, while PSI carried the greatest part, about 95%. Quantitative analysis of the obtained data showed that about 20% of phycobilisomes in Spirulina cells are bound to PSII, while 60% of phycobilisomes transfer the energy to PSI trimers, and the remaining 20% are associated with PSI monomers. A relevant model of organization of phycobilisomes and chlorophyll pigment-protein complexes in Spirulina is proposed. It is suggested that phycobilisomes are connected with PSII dimers, PSI trimers, and coupled PSI monomers.     

Resolution of the Photosystem I and Photosystem II contributions to chlorophyll fluorescence of intact leaves at room temperature

Green leaves illuminated with photosynthetically active light emit red fluorescence, whose time-dependent intensity variations reflect photosynthetic electron transport (the Kautsky effect). Usually, fluorescence variations are discussed by considering only the contribution of PSII-associated chlorophyll a, although it is known that the fluorescence of PSI-associated chlorophyll a also contributes to the total fluorescence [Aust. J. Plant Physiol. 22 (1995) 131]. Because the fluorescence emitted by each photosystem cannot be measured separately by selecting the emission wavelength in in vivo conditions, the contribution of PSI to total fluorescence at room temperature is still in ambiguity. By using a diode array detector, we measured fluorescence emission spectra corresponding to the minimal (F(O)) and maximal (F(M)) fluorescence states. We showed that the different shapes of these spectra were mainly due to a higher contribution of PSI chlorophylls in the F(O) spectrum. By exciting PSI preferentially, we recorded a reference PSI emission spectrum in the near far-red region. From the F(O) and F(M) spectra and from this PSI reference spectrum, we derived specific PSI and PSII emission spectra in both the F(O) and F(M) states. This enables to estimate true value of the relative variable fluorescence of PSII, which was underestimated in previous works. Accurate separation of PSI-PSII fluorescence emission spectra will also enable further investigations of the distribution of excitation energy between PSI and PSII under in vivo conditions.     

Fluorescence changes accompanying short-term light adaptations in photosystem I and photosystem II of the cyanobacterium <Emphasis Type="Italic">Synechocystis</Emphasis> sp. PCC 6803 and phycobiliprotein-impaired mutants: State 1/State 2 transitions and carotenoid-induced quenching of phycobilisomes

The features of the two types of short-term light-adaptations of photosynthetic apparatus, State 1/State 2 transitions, and non-photochemical fluorescence quenching of phycobilisomes (PBS) by orange carotene-protein (OCP) were compared in the cyanobacterium Synechocystis sp. PCC 6803 wild type, CK pigment mutant lacking phycocyanin, and PAL mutant totally devoid of phycobiliproteins. The permanent presence of PBS-specific peaks in the in situ action spectra of photosystem I (PSI) and photosystem II (PSII), as well as in the 77 K fluorescence excitation spectra for chlorophyll emission at 690 nm (PSII) and 725 nm (PSI) showed that PBS are constitutive antenna complexes of both photosystems. The mutant strains compensated the lack of phycobiliproteins by higher PSII content and by intensification of photosynthetic linear electron transfer. The detectable changes of energy migration from PBS to the PSI and PSII in the Synechocystis wild type and the CK mutant in State 1 and State 2 according to the fluorescence excitation spectra measurements were not registered. The constant level of fluorescence emission of PSI during State 1/State 2 transitions and simultaneous increase of chlorophyll fluorescence emission of PSII in State 1 in Synechocystis PAL mutant allowed to propose that spillover is an unlikely mechanism of state transitions. Blue–green light absorbed by OCP diminished the rout of energy from PBS to PSI while energy migration from PBS to PSII was less influenced. Therefore, the main role of OCP-induced quenching of PBS is the limitation of PSI activity and cyclic electron transport under relatively high light conditions.     

Nuclear pea mutants deficient in chlorophyll b and the major polypeptide of the light-harvesting chlorophyll a/b protein of photosystem II

Eight chlorophyll b deficient nuclear mutants of pea (Pisum sativum L.) have been characterized by low temperature fluorescence emission spectra of their leaves and by the ultrastructure, photochemical activities and polypeptide compositions of the thylakoid membranes. The room temperature fluorescence induction kinetics of leaves and isolated thylakoids have also been recorded. In addition, the effects of Mg²⁺ on the fluorescence kinetics of the membranes have been investigated. The mutants are all deficient in the major polypeptide of the light-harvesting chlorophyll a/b protein of photosystem II. The low temperature fluorescence emission spectra of aurea-5106, xantha-5371 and –5820 show little or no fluorescence around 730 nm (photosystem I fluorescence), but possess maxima at 685 and 695 nm (photosystem II fluorescence). These three mutants have low photosystem II activities, but significant photosystem I activities. The long-wavelength fluorescence maximum is reduced for three other mutants. The Mg²⁺ effect on the variable component of the room temperature fluorescence (685 nm) induction kinetics is reduced in all mutants, and completely absent in aurea-5106 and xantha-5820. The thylakoid membranes of these 2 mutants are appressed pairwise in 2-disc grana of large diameter. Chlorotica-1-206A and–130A have significant long-wavelength maxima in the fluorescence spectra and show the largest Mg²⁺ enhancement of the variable part of the fluorescence kinetics. These two mutants have rather normally structured chloroplast membranes, though the stroma regions are reduced. The four remaining mutants are in several respects of an intermediate type.Abbreviations Chl chlorophyll - CPI Chi-protein complex I, F_o, F_v - F_m parameters of room temperature chlorophyll fluorescence induction kinetics - F685, F695 and F-1 components of low temperature chlorophyll emission with maximum at 685, 695 and ca 735 nm, respectively - PSI photosystem I - PSII photosystem II - LHCI and LHCII light-harvesting chlorophyll a/b complexes associated with PSI and PSII, respectively - SDS sodium dodecyl sulfate     

Light-dependent reversal of dark-chilling induced changes in chloroplast structure and arrangement of chlorophyll-protein complexes in bean thylakoid membranes

Changes in chloroplast structure and rearrangement of chlorophyll-protein (CP) complexes were investigated in detached leaves of bean (Phaseolus vulgaris L. cv. Eureka), a chilling-sensitive plant, during 5-day dark-chilling at 1 degrees C and subsequent 3-h photoactivation under white light (200 mumol photons m(-2) s(-1)) at 22 degrees C. Although, no change in chlorophyll (Chl) content and Chl a/b ratio in all samples was observed, overall fluorescence intensity of fluorescence emission and excitation spectra of thylakoid membranes isolated from dark-chilled leaves decreased to about 50%, and remained after photoactivation at 70% of that of the control sample. Concomitantly, the ratio between fluorescence intensities of PSI and PSII (F736/F681) at 120 K increased 1.5-fold upon chilling, and was fully reversed after photoactivation. Moreover, chilling stress seems to induce a decrease of the relative contribution of LHCII fluorescence to the thylakoid emission spectra at 120 K, and an increase of that from LHCI and PSI, correlated with a decrease of stability of LHCI-PSI and LHCII trimers, shown by mild-denaturing electrophoresis. These effects were reversed to a large extent after photoactivation, with the exception of LHCII, which remained partly in the aggregated form. In view of these data, it is likely that dark-chilling stress induces partial disassembly of CP complexes, not completely restorable upon photoactivation. These data are further supported by confocal laser scanning fluorescence microscopy, which showed that regular grana arrangement observed in chloroplasts isolated from control leaves was destroyed by dark-chilling stress, and was partially reconstructed after photoactivation. In line with this, Chl a fluorescence spectra of leaf discs demonstrated that dark-chilling caused a decrease of the quantum yield PSII photochemistry (F(v)/F(m)) by almost 40% in 5 days. Complete restoration of the photochemical activity of PSII required 9 h post-chilling photoactivation, while only 3 h were needed to reconstruct thylakoid membrane organization and chloroplast structure. The latter demonstrated that the long-term dark-chilled bean leaves started to suffer from photoinhibition after transfer to moderate irradiance and temperature conditions, delaying the recovery of PSII photochemistry, independently of photo-induced reconstruction of PSII complexes.     

A retrieval algorithm to evaluate the Photosystem I and Photosystem II spectral contributions to leaf chlorophyll fluorescence at physiological temperatures

A new computational procedure to resolve the contribution of Photosystem I (PSI) and Photosystem II (PSII) to the leaf chlorophyll fluorescence emission spectra at room temperature has been developed. It is based on the Principal Component Analysis (PCA) of the leaf fluorescence emission spectra measured during the OI photochemical phase of fluorescence induction kinetics. During this phase, we can assume that only two spectral components are present, one of which is constant (PSI) and the other variable in intensity (PSII). Application of the PCA method to the measured fluorescence emission spectra of Ficus benjamina L. evidences that the temporal variation in the spectra can be ascribed to a single spectral component (the first principal component extracted by PCA), which can be considered to be a good approximation of the PSII fluorescence emission spectrum. The PSI fluorescence emission spectrum was deduced by difference between measured spectra and the first principal component. A single-band spectrum for the PSI fluorescence emission, peaked at about 735?nm, and a 2-band spectrum with maxima at 685 and 740?nm for the PSII were obtained. A linear combination of only these two spectral shapes produced a good fit for any measured emission spectrum of the leaf under investigation and can be used to obtain the fluorescence emission contributions of photosystems under different conditions. With the use of our approach, the dynamics of energy distribution between the two photosystems, such as state transition, can be monitored in vivo, directly at physiological temperatures. Separation of the PSI and PSII emission components can improve the understanding of the fluorescence signal changes induced by environmental factors or stress conditions on plants.