外源水杨酸对盐胁迫下小桐子幼苗脯氨酸代谢的影响

以小桐子幼苗为材料,设置盐胁迫(200mmol·L-1 NaCl)和外源水杨酸处理(0~2.0mmol·L-1 SA)水培试验,通过检测幼苗叶片脯氨酸含量、脯氨酸代谢关键酶活性及相关代谢酶基因的表达水平,研究了外源水杨酸对盐胁迫下小桐子幼苗脯氨酸代谢机理的影响。结果显示:(1)外源0.9mmol·L-1 SA处理可显著提高盐胁迫下小桐子幼苗的脯氨酸含量,上调脯氨酸合成关键酶Δ1-吡咯琳-5-羧酸合成酶(P5CS)和鸟氨酸转氨酶(OAT)活性,以及上调JcP5CS和JcOAT基因的表达水平。(2)SA也显著抑制了脯氨酸降解酶ProDH的活性及JcProDH基因的表达水平。(3)SA处理还显著提高了盐胁迫下小桐子幼苗的组织活力,降低了叶片电解质渗漏率和丙二醛(MDA)含量。研究发现,外源SA可通过活化脯氨酸合成的谷氨酸途径和鸟氨酸途径,以及抑制脯氨酸的降解途径来促进盐胁迫下小桐子幼苗脯氨酸的积累;外源SA处理也可提高小桐子幼苗的耐盐性,且这种提高可能与SA诱导脯氨酸的积累密切相关。     

外源ABA对低温胁迫下小桐子幼苗脯氨酸积累及其代谢途径的影响

本文以小桐子为材料,研究了外源脱落酸（ABA）对低温胁迫下小桐子幼苗脯氨酸积累的影响。结果表明,低温胁迫（5℃）可促进小桐子幼苗体内脯氨酸的积累,上调脯氨酸合成关键酶Δ¹-吡咯琳-5-羧酸合成酶（P5CS）和鸟氨酸转氨酶（OAT）的活性,上调P5CS基因（JcP5CS）的表达,及抑制脯氨酸降解酶脯氨酸脱氢酶（Pro DH）的活性。用外源ABA处理低温胁迫下的小桐子幼苗,发现150μmol·L^-1的ABA可显著提高其低温胁迫下的脯氨酸含量,上调P5CS的活性和JcP5CS的表达水平,及抑制ProDH的活性。表明外源ABA可通过活化脯氨酸合成的谷氨酸途径和抑制脯氨酸的降解途径来促进低温胁迫下小桐子幼苗脯氨酸的积累。     

外源茉莉酸甲酯对盐胁迫下小桐子幼苗渗透调节和脯氨酸代谢的影响

以小桐子幼苗为材料,采用人工气候箱内水培试验,设置不同浓度(0、20、40、60、80、100μmol·L^(-1))茉莉酸甲酯(MeJA)和150 mmol·L^(-1)NaCl胁迫处理,分析不同处理条件下小桐子幼苗叶片的组织活力、MDA含量、水势、含水量和叶片渗透调节物质脯氨酸、甜菜碱和可溶性糖的含量,以及脯氨酸代谢关键酶P5CS、OAT和甜菜碱合成关键酶BADH活性和相关基因表达水平,探讨外源茉莉酸甲酯对盐胁迫下小桐子幼苗渗透调节能力的影响及其机制。结果表明:(1)外源MeJA处理可提高盐胁迫下小桐子幼苗叶片的组织活力和叶片含水量,降低小桐子幼苗叶片的MDA含量和水势,且60μmol·L^(-1)浓度处理效果最佳。(2)外源MeJA处理可提高盐胁迫下小桐子幼苗叶片的脯氨酸、甜菜碱和可溶性糖的含量,且60μmol·L^(-1)MeJA处理显著提高了盐胁迫下小桐子幼苗内源茉莉酸、脯氨酸和甜菜碱的含量。(3)60μmol·L^(-1)MeJA也提高了盐胁迫下小桐子BADH、P5CS和OAT的活性,并上调了JcBADH、JcP5CS、JcOAT基因的表达水平,但MeJA降低了脯氨酸降解酶ProDH的活性,下调了JcProDH基因的表达。研究发现,在盐胁迫条件下,外源MeJA通过活化脯氨酸生物合成的谷氨酸和鸟氨酸途径,尤其是鸟氨酸途径,以及抑制脯氨酸降解途径来促进小桐子幼苗脯氨酸的积累,同时MeJA也激活了幼苗体内甜菜碱的生物合成过程,从而强化了盐胁迫下幼苗的渗透调节作用和耐盐性,表明MeJA诱导的渗透调节在小桐子耐盐性形成过程中发挥着重要作用。     

外源H_2O_2对混合盐碱胁迫下燕麦幼苗叶片脯氨酸积累和代谢途径的影响

为探讨H_2O_2对盐碱胁迫下植物脯氨酸代谢的调控机理,以燕麦新品种‘定莜6号’幼苗为材料,采用水培法研究了外源H_2O_2对混合盐碱胁迫下燕麦脯氨酸积累和代谢途径的影响。结果表明,75 mmol·L-1混合盐碱(Na Cl∶Na_2SO_4∶Na HCO_3∶Na_2CO_3=12∶8∶9∶1)胁迫可促进燕麦幼苗叶片脯氨酸的积累,提高脯氨酸合成的鸟氨酸途径关键酶鸟氨酸δ-氨基转移酶(δ-OAT)活性,抑制脯氨酸合成的谷氨酸途径关键酶Δ1-吡咯啉-5-羧酸合成酶(P5CS)及脯氨酸降解限速酶脯氨酸脱氢酶(Pro DH)活性。在75 mmol·L-1混合盐碱胁迫下添加0.01~1 000μmol·L-1H_2O_2可显著提高燕麦幼苗叶片的脯氨酸含量,其中10μmol·L-1H_2O_2的作用最明显;10μmol·L-1H_2O_2上调了75 mmol·L-1混合盐碱胁迫下燕麦幼苗叶片的P5CS和δ-OAT活性,降低了Pro DH活性。此外,10μmol·L-1H_2O_2使75 mmol·L-1混合盐碱胁迫下燕麦幼苗叶片内源性H_2O_2含量急剧升高后迅速降低。表明外源H_2O_2能够提高混合盐碱胁迫下燕麦幼苗内源H_2O_2的含量,并通过活化脯氨酸合成的谷氨酸途径和鸟氨酸途径,抑制脯氨酸的降解,促进混合盐碱胁迫下燕麦幼苗脯氨酸的积累。     

一氧化氮参与调节盐胁迫下脱落酸诱导的小麦幼苗叶片脯氨酸的累积

不同浓度(0.01～5.00mmol/L)的外源一氧化氮(NO)供体硝普钠(SNP)以浓度依赖性的性式诱导150mmol/L NaCl胁迫下小麦(Triticum aestivum L.cv.Yangmai 158)幼苗叶片脯氨酸的累积。其中0.1 mmol/L的SNP效果最明显,而结合采用NO清除剂c-PTIO和血红蛋白的处理均分别逆转了该效应。研究结果还发现:0.1 mmol/L SNP诱导的脯氨酸累积还可能有利于盐胁迫下小麦幼苗的保水性;0.1 mmol/L的SNP显著激活了内源ABA的合成,而结合血红蛋白的处理则证实,在外源ABA诱导脯氨酸累积的过程中NO可能作用于ABA信号分子的下游,但NO和ABA信号分子在此诱导反应中不存在累积效应。进一步研究脯氨酸合成和降解的酶促反应途径,发现外源NO处理前4天内可能主要是通过提高Δ~1-吡咯啉-5-羧酸合成酶(P5CS)的活性来促进脯氨酸的合成,以后直至第8天主要是通过抑制脯氨酸脱氢酶(ProDH)的活性来抑制脯氨酸的降解;ABA对于P5CS和ProDH活性的调节能力弱于NO。此外,Ca~(2 )在NO诱导的盐胁迫下小麦叶片脯氨酸累积的信号分子途径中起重要的介导作用。     

ABA和NaCl对碱蓬多胺和脯氨酸代谢的影响

比较了 5 0 μmol/L的ABA和 4 0 0mmol/L的NaCl对碱蓬植株内脯氨酸 (Pro)和多胺 (PAs)含量和代谢关键酶活性的影响。结果表明 :ABA处理能诱导碱蓬植株吡咯啉 5 羧酸合成酶 (P5CS)、鸟氨酸脱羧酶(ODC)、多胺氧化酶 (PAO)和脯氨酸脱氢酶 (ProDH)活性上升 ,Pro和游离多胺含量增加。ABA NaCl处理促进碱蓬植株体内Pro和PA的积累。与NaCl处理相比 ,ABA NaCl处理后碱蓬植株P5CS活性显著上升 ,ProDH活性变化不明显 ,ODC活性极显著上升 ,PAO和转谷酰胺酶 (TGase)活性上升 ,从而使Pro和游离多胺含量均显著上升 ,游离态多胺 (Spd PAx) /Put提高 ,结合态多胺总量上升。ABA NaCl处理对碱蓬植株体内Pro和PAs合成的促进与其对生长速率的促进效应相一致     

Cd~（2＋）胁迫诱导烟草悬浮细胞脯氨酸积累的生化途径及外源脯氨酸对Cd~（2＋）胁迫下H_2O_2产生的抑制作用

Cd~（2＋）可提高烟草悬浮细胞脯氨酸的含量,顺序上调脯氨酸合成关键酶鸟氨酸转氨酶（OAT）、精氨酸酶、△~1-吡咯啉-5-羧酸合成酶（P5CS）和谷氨酸脱氢酶（GDH）的活性,降低脯氨酸降解关键酶脯氨酸脱氢酶（ProDH）的活性,表明Cd~（2＋）胁迫诱导烟草细胞脯氨酸的积累是脯氨酸合成的鸟氨酸途径和谷氨酸途径顺序激活、而脯氨酸降解途径显著抑制的综合结果。此外,Cd~（2＋）能导致烟草细胞H_2O_2的快速产生及H_2O_2产生相关酶（质膜NADPH氧化酶、细胞壁多胺氧化酶及共价结合与离子结合细胞壁过氧化物酶）活性升高和脂质过氧化产物丙二醛（MDA）增加,导致烟草细胞的氧化胁迫。外源脯氨酸预处理显著抑制了Cd~（2＋）诱导的烟草细胞H_2O_2的产生与MDA的增加,减轻了Cd~（2＋）诱导的氧化胁迫。而脯氨酸抑制Cd~（2＋）诱导的H_2O_2产生可能是由于脯氨酸抑制了H_2O_2产生相关酶的活性所致。     

脯氨酸对不结球白菜（Brassica chinensis）镉积累的影响

采用溶液培养的方法,研究了过量Cd对不结球白菜（Brassica chinensis）幼苗内源脯氨酸含量的影响及外源脯氨酸在不结球白菜Cd积累中的作用。100μmol？L-1 Cd处理明显增加不结球白菜叶片脯氨酸（Pro）的含量,并且随着Cd胁迫时间的延长,其含量也显著上升。100μmol？L-1 Cd处理明显降低脯氨酸脱氢酶（PDH）活性,而增加δ1-吡咯啉-5-羧酸合成酶（P5CS）的活性。1~5 mmol？L-1外源Pro处理显著降低不结球白菜幼苗地上部Cd含量,对根系Cd含量无显著影响,但外源Pro处理并不能缓解过量Cd对不结球白菜幼苗生物量的抑制作用。外源Pro处理下,不结球白菜叶片中Pro含量与其地上部干重呈显著的负相关关系。     

盐胁迫激活大麦幼苗脯氨酸合成的鸟氨酸途径

２００mmol/L NaCl处理结合^14C-Ｇlu和^14C-Arg叶面饲喂６d龄大麦（Ｈordeum vulgareL.)幼苗,结果证明６d龄大麦幼苗体内普遍存在Ａrg（谷氨酸）→Ｏｍ（鸟氨酸）→Ｐro(脯氨酸）途径。直 迫明显激活了Ａrg→Ｏｍ→Ｐro途径,胁迫处理７d,大麦幼苗叶片和根系中该途径对Pro含量上升的贡献是Glu→Pro途径的１．０－１．５倍。而盐的“鉴４”品种Ａrg→Ｏｍ→Ｐro途径对Pro含量上升的贡献是对盐敏感的“科品７号”的１．７－２．０倍,结果表明盐胁迫下Ａrg→Ｏｍ→Ｐro途径的激活对大麦幼苗耐盐性的提高具有重要意义。     

一氧化氮参与调节盐胁迫下脱落酸诱导的小麦幼苗叶片脯氨酸的累积

不同浓度(0.01～5.00mmol/L)的外源一氧化氮(NO)供体硝普钠(SNP)以浓度依赖性的性式诱导150mmol/LNaCl胁迫下小麦(Triticum aestivum L.cv.Yangmai 158)幼苗叶片脯氨酸的累积.其中0.1 mmol/L的SNP效果最明显,而结合采用NO清除剂c-PTIO和血红蛋白的处理均分别逆转了该效应.研究结果还发现:0.1 mmol/L SNP诱导的脯氨酸累积还可能有利于盐胁迫下小麦幼苗的保水性;0.1 mmol/L的SNP显著激活了内源ABA的合成,而结合血红蛋白的处理则证实,在外源ABA诱导脯氨酸累积的过程中NO可能作用于ABA信号分子的下游,但NO和ABA信号分子在此诱导反应中不存在累积效应.进一步研究脯氨酸合成和降解的酶促反应途径,发现外源NO处理前4天内可能主要是通过提高△'-吡咯啉-5-羧酸合成酶(P5CS)的活性来促进脯氨酸的合成,以后直至第8天主要是通过抑制脯氨酸脱氢酶(ProDH)的活性来抑制脯氨酸的降解;ABA对于P5CS和ProDH活性的调节能力弱于NO.此外,Ca2 在NO诱导的盐胁迫下小麦叶片脯氨酸累积的信号分子途径中起重要的介导作用.     

Proline peptide isomerization and protein folding

The unfolding-refolding of proteins is a cooperative process and, as judged by equilibrium properties, occurs in one step involving the native,N, and the unfoldedU, conformational states. Kinetic studies have shown that the denatured protein exists as a mixture of slow-(U)_Sand fast-(U)_Frefolding forms produced by proline peptidecis-trans isomerization. Proline residues inU _Fare in the same configuration as in the native protein while they are in non-native configuration inU _S. For protein folding to occur quicklyU _Smust be converted intoU _F. The fact that the equilibrium and kinetic properties of are the same as those found for prolinecis-trans isomerization taken together with the absence of slow phase in the kinetics of refolding of a protein devoid of proline, support this view. However, the absence of a linear correlation between half-time of reactivation of denatured enzymes and their proline-contents, as well as the dissimilarities in the kinetic properties of in unfolding and refolding experiments are not consistent with the model. Conformational energy calculation and experimental results on refolding of proteins suggest that some proline residues are non-essential. They will not block protein folding even in wrong isomeric form. The native-like folded structure with incorrect proline isomers will serve as intermediate state(s) in which these prolines will more readily isomerize to the correct isomeric form. The picture becomes more complex when one considers the consequence ofcis-trans isomerism of non-proline residues on protein folding.     

The Relation between Accumulation of Abscisic Acid and Proline in Detached Rice Leaves

The relation between abscisic acid (ABA) and proline accumulation was investigated in detached rice (Oryza sativa L.) leaves. In darkness, proline content increased about 2-, 2,5- and 6-fold after 24, 48 and 72 h. ABA content reached maximum after 48 h. In the light, proline content remained almost unchanged until 48 h and subsequently increased slightly. ABA content in the light was lower than in darkness, but the maximum was also after 48 h. During 12-h exposure to decreased air humidity, proline content gradually increased, but ABA content increased about 25-fold after 4 h and declined thereafter. Exogenous application of ABA resulted in an increase in proline content in detached rice leaves under both light and darkness.     

Effect of NaCl and Proline on Bean Seedlings Cultured in vitro

Effects of NaCl (150 mM), proline (10 mM) and their combination on growth and contents of chlorophyll, proline and protein of bean (Phaseolus vulgaris cv. Kizilhaç) seedlings in vitro were investigated. NaCl decreased seedling growth. Proline added to control seedlings did not change seedling growth but decreased chlorophyll and increased protein contents. When proline added to NaCl-treated seedlings growth was increased in comparison with NaCl-treated only. Thus, proline alleviated salinity stress in bean seedlings.     

Proline porters effect the utilization of proline as nutrient or osmoprotectant for bacteria

Proline is utilized by all organisms as a protein constituent. It may also serve as a source of carbon, energy and nitrogen for growth or as an osmoprotectant. The molecular characteristics of the proline transport systems which mediate the multiple functions of proline in the Gram negative enteric bacteria, Escherichia coli and Salmonella typhimurium, are now becoming apparent. Recent research on those organisms has provided both protocols for the genetic and biochemical characterization of the enzymes mediating proline transport and molecular probes with which the degree of homology among the proline transport systems of archaebacteria, eubacteria and eukaryotes can be assessed. This review has provided a detailed summary of recent research on proline transport in E. coli and S. typhimurium; the properties of other organisms are cited primarily to illustrate the generality of those observations and to show where homologous proline transport systems might be expected to occur. The characteristics of proline transport in eukaryotic microorganisms have recently been reviewed (Horak, 1986).     

Effects of NaCl and Proline on Polyphenol Oxidase Activity in Bean Seedlings

In this work, the effects of NaCl (0, 50, 100, and 150 mM), proline (0, 5 and 10 mM) and NaCl + proline in combinations on activity of polyphenol oxidase (PPO; E.C. 1.10.3.1) and soluble protein content have been investigated in the root, stem and leaf tissues of bean (Phaseolus vulgaris L.) seedlings grown in embryo culture. PPO activities were higher in all the tissues treated with NaCl, proline and NaCl + proline combinations those that of the control tissues. The protein content was very high in tissues exposed to proline and NaCl + proline combination, but NaCl alone decreased protein contents in root and leaf tissues. The results suggest that proline may play a role as an enzyme-stabilizing agent in salt stress.     

Transport of Proline and Hydroxyproline by the Neutral Amino-acid Exchanger ASCT1

ASCT1 is a member of the glutamate transporter superfamily cloned from human brain and characterized as a Na⁺-dependent neutral amino-acid exchanger, which displays substrate-induced chloride-channel activity and mediates concentrative transport of alanine. Initial studies in ASCT1-expressing Xenopus laevis oocytes showed that proline did not elicit measurable currents, in contrast to what occurred with alanine, serine or cysteine, suggesting that proline was not an ASCT1 substrate, although it induced the release of alanine from preloaded oocytes. Here, we have studied the uptake of proline and hydroxyproline by ASCT1-expressing oocytes in order to investigate the ability of ASCT1 to translocate these imino acids. The results demonstrate ASCT1-mediated proline transport that is Na⁺-dependent, saturable, inhibited by the reported ASCT1 substrates as well as by hydroxyproline and can drive the imino acid against its concentration gradient. The apparent kinetic constants for the transport of alanine and the imino acids, obtained with oocytes from the same batch, showed maximal transport rate for proline and hydroxyproline to be half of that for alanine. However, K _0.5 for proline was 704 ± 86 µM, about three times higher than alanine K _0.5 (203.3 ± 36.4 µM), whereas hydroxyproline K _0.5 was 33.2 ± 4.3 µM, indicating that the hydroxylation on carbon 4 of proline strongly increases the affinity of ASCT1 for this proline derivative. In summary, the present work demonstrates for the first time the ability of ASCT1 to transport proline and hydroxyproline.     

Light-Dark Changes in Proline Content of Barley Leaves under Salt Stress

Proline accumulation in leaves of barley (Hordeum vulgare L. cv. Alfa) seedlings treated with 150 mM NaCl was promoted in the light and suppressed in the dark. The light/dark changes of proline content was enhanced with each 12 h light/12 h dark cycle and the proline content increased steadily. Root and shoot concentrations of Na⁺ and Cl^– in salt treated plants increased about 10 to 25 times as compared to the control. The content of these ions and the content of malondialdehyde were higher in the shoot of seedlings exposed to salt stress for 4 d in the light in comparison with the seedlings exposed to NaCl for 4 d in darkness. Light stimulated both ions and proline accumulation in the leaves and has no effect in the roots. Oxygen uptake was higher in the seedlings kept 4 d in the light which have higher endogenous free proline content. Chlorophyll fluorescence measurements showed that the photochemical activity of PS 2 slightly decreased as a result of salt stress and was not influenced by light regimes during plant growth.     

Proline accumulation in a barley mutant resistant to trans-4-hydroxy-L-proline

Five proline analogues were tested for inhibition of the growth of mature barley (Hordeum vulgare L.) embryos in sterile culture. Inhibition by all analogues was relieved by proline. Inhibition by trans-4-hydroxy-L-proline was relieved by low amounts of proline. Twenty thousand mature embryos were dissected from M₂ seeds after sodium azide mutagenesis. Four plants (Rothamsted 5201, 6102, 6901, 6902) were selected with good growth on 4 mM trans-4-hydroxyproline. Properties of mutant R5201 were studied in detail. Selfed progeny of R5201 were all resistant to trans-4-hydroxyproline and also to L-thiazolidine-4-carboxylic acid and trans-3-hydroxy-L-proline but not L-azetidine-2-carboxylic acid. The content of soluble proline in progeny of R5201 was higher in leaves by a factor of up to six-fold. Proline content was measured in the soluble fraction of the terminal 20 mm of 4 d old plants subjected to severe water stress in 40% w/v polyethylene glycol. Leaves of the mutant contained more proline initially and accumulated proline morer rapidly than the parental leaves. As mutant leaves were larger and lost water more rapidly the greater increase in proline may have been caused by more severe water stress. Resistance to trans-4-hydroxyproline in R5201 was due to a single partially dominant nuclear gene.Abbreviations AZC L-azetidine-2-carboxylic acid - HYP trans-4-hydroxy-L-proline - ORN L-ornithine - CIT L-citrulline     

Proline betaine is a highly effective osmoprotectant for Staphylococcus aureus

Proline betaine is an osmoprotectant that is at least as effective as glycine betaine, and more effective than L-proline, for various strains of Staphylococcus aureus, and Staphylococcus epidermidis and Staphylococcus saprophyticus. ¹³C NMR studies revealed that proline betaine accumulated to high levels in osmotically stressed S. aureus, but was also detected in organisms grown in its presence in the absence of osmotic stress. Competition experiments indicated that proline betaine was taken up by the proline transport systems of S. aureus, but not by the high affinity glycine betaine transport system.Abbreviations PYK Peptode - Yeast extract K₂HPO₄     

Proline, Ornithine, Arginine and Glutamic Acid Contents in Detached Rice Leaves

The effects of water stress on the contents of proline, ornithine, arginine and glutamic acid in detached rice leaves were examined. In water stressed leaves, the content of proline was elevated to a content approximately 8-, 14- and 17-fold higher than in control leaves after 4, 8 and 12 h, respectively. We also observed that omithine and arginine contents were much higher under water stress than in control leaves. However, the content of glutamic acid in water stressed leaves was higher after 4 and 8 h and lower after 12 h than that in control leaves.