Monoclonal antibody against the glutaraldehyde-conjugated polyamine,spermine

We developed a mouse monoclonal antibody (ASPM-29, mAb) against spermine (Spm) conjugated to human serum albumin (HSA) using glutaraldehyde-sodium borohydride, for applications in immunocytochemistry (ICC). The antibody specificity was evaluated by an enzyme-linked immunosorbent assay (ELISA) binding test, simulating the ICC of tissue sections. ASPM-29 showed an almost equal immunoreactivity to Spm and spermidine (Spd) but no reactivity to any of the other polyamine (PA)-related compounds tested. By use of this antibody, indirect immunoperoxidase staining was observed in different tissues fixed with glutaraldehyde in combination with borohydride reduction. In contrast, immunoreactivity was quite low in tissues fixed only with glutaraldehyde. Absorption controls indicated that the immunostaining could be completely inhibited by 50 g/ml of Spm or Spd and partially inhibited by N-acetylspermine (Ac-Spm), N¹-acetylspermidine (N¹-Ac-Spd), or N⁸-acetylspermidine (N⁸-Ac-Spd), but was hardly inhibited at all by other PA-related compounds or amino acids. The reactivity of the antibody with Spm conjugated on wells in an ELISA plate was inhibited by micromolar concentrations of Spm, Spd, Ac-Spm, N¹-Ac-Spd, or N⁸-Ac-Spd, in decreasing order, but not by other small molecules. Dense ICC staining was observed in the paranuclear and basal cytoplasm of acinar cells of rat pancreas, submandibular gland and paratid gland, these results being in complete agreement with our recent ICC methods using other mAbs produced against N-(-male-imidobutyryloxy) succinimide-conjugated Spm.     

A monoclonal antibody against the glutaraldehyde-conjugated polyamine, putrescine: application to immunocytochemistry

We developed a mouse monoclonal antibody (mAb; APUT-32, IgG1 subisotype mAb) against putrescine (Put) conjugated to bovine serum albumin using a glutaraldehyde (GA)-sodium borohydride procedure, for applications in immunocytochemistry (ICC). The antibody specificity was evaluated by an ELISA binding test, simulating the ICC of tissue sections. APUT-32 mAb was highly specific to Put, and distinguished alterations in the chemical structure of other polyamine (PA) analogs, showing 3.8% crossreaction with cadaverine, 3.3% with spermidine (Spd), and 2.3% with 1,3-diaminopropane. Comparable results in immunoreactivity of APUT-32 mAb were obtained with the ELISA inhibition test. By the indirect immunoperoxidase method using the APUT-32 mAb, Put-like immunoreactivities were observed in the cytoplasm of HeLa and MCF-7 cell lines fixed with GA in combination with NaBH4 reduction, but almost no immunoreaction was seen in the cytoplasm of the human melanoma BD cell line. On the other hand, the same method but using a previously prepared ASPM-29 mAb, specific for spermine (Spm) and Spd, produced intense immunostaining in the cytoplasm of all the three cell types. The Put-like immunoreaction was completely abolished by absorption of the APUT-32 mAb with 10 microg/ml Put-human serum albumin conjugate prepared using GA and NaBH4. HPLC analysis was also performed for the levels of each of the PAs in the three types of cell, showing that the levels of Put detected were much lower than those of Spm and Spd, and were strikingly different in the three cell lines among which the human melanoma BD cell line contained the lowest levels of Put. These results strongly suggest that APUT-32 mAb reacts specifically with Put in the tumor cells and therefore has the potential as a new tool for elucidating the biological roles of Put in cells and tissues.     

An evaluation of polyamine immunocytochemistry using immunocytochemical model systems

Polyamine (PA) immunocytochemistry (ICC) was evaluated by a recently developed enzyme-linked immunosorbent assay (ELISA) binding test (ELISABT) using an anti-spermine (Spm) serum raised against Spm conjugated via the cross-linker N-(-maleimidobutyryloxy)succinimide (GMBS) with bovine serum albumin. In the test the antiserum showed strong immunoreactivity with N ¹-acetylspermine (Ac-Spm) and acetylspermidine (N ¹-Ac-Spd and N ⁸-Ac-Spd), and low immunoreactivity with Spm and Spd, which was, however, markedly enhanced after reaction with GMBS, acetic anhydride or glutaraldehyde. Complete agreement with results of immunoblot analysis was observed. PA-like immunoreactivity observed in the present PA ICC in cells in the foveolae and isthmus of rat gastric glands was completely abolished by absorption of the serum with N ¹,N ¹²-diacetyl-Spm, Ac-Spm or N ¹-Ac-Spd, but not by Spm or Spd. This absorption test was then simulated by an ELISA inhibition test (ELISAIT) with a solid phase conjugated with Ac-Spm or Spm, and by a PA ICC model system using Sepharose gel beads conjugated with each of several PAs. The results strongly suggest that the immunostaining in the gastric mucosa was mainly due to antibody species in the serum specific to acylated Spm and Spd, but not to Spm or Spd. Acetyl PAs exist at such low concentrations in animal tissues that they are virtually undetectable by current ICC methods. Therefore Spm and Spd are likely candidates for those detected, after having been converted by fixation into such PA derivatives as become reactive with the antiserum.     

Production and characterization of monoclonal antibodies against the polyamine,spermine: immunocytochemical localization in rat tissues

Two monoclonal antibodies of types IgG_2b and IgG_2a, anti-spermine-(Spm)-1 (ASPM-1) and anti-Spm-2 (ASPM-2) respectively were found among five clones of murine monoclonal antibodies, which were raised against Spm conjugated with bovine serum albumin via the cross-linker N-(-maleimidobutyryloxy) succinimide (GMBS). Antibody specificity was evaluated by a recently developed ELISA binding test, and led to the study of tissue sections by immunocytochemistry (ICC). ASPM-1 showed exclusive immunoreactivity with Spm, with the exception of a negligible cross-reactivity (2.0%) with spermidine (Spd). ASPM-2, on the other hand, reacted almost equally with acetylspermine (Ac-Spm) and N ¹-acetylspermidine (N¹-Ac-Spd) but with none of the other polyamine-related compounds tested. Complete agreement was obtained with the results of immunoblot analysis. Furthermore, results for antibody specificity obtained with the ELISA inhibition test and ICC model experiments using Sepharose gel beads strongly suggested that ASPM-1 recognizes the Spm molecule possessing at least a free terminal primary amino group, while ASPM-2 recognizes the Spm molecule acylated at both the terminal primary amino groups. An ICC method using ASPM-2 produced strong staining for polyamines (PAs) in the cytoplasm (but very few in the nuclei) of two different tumor cell lines and protein- or peptide-secreting cell systems, including exocrine and endocrine cell types; ASPM-1 showed immunoreactivity only with the tumor cell lines. These results strongly suggest that ASPM-2 may be useful for studies on actively proliferating and neoplastic cells, supporting our previously proposed idea that in immunocytochemistry PAs were converted to a variety of PA derivatives during the fixation process.     

A new enzyme-linked immunosorbent assay (ELISA) for studying immunocytochemical procedures using an antiserum produced against spermidine as a model

Antiserum was produced in rabbits against the polyamine spermidine (Spd) conjugated to bovine serum albumin (BSA). The reactivity of the serum to Spd and a variety of structurally related compounds was quantified by a new immunocytochemical model system incorporating an enzyme-linked immunosorbent assay (ELISA) binding test. This is based on the principle of coupling these compounds to the wells of microtiter plate activated with poly-l-lysine and glutaraldehyde and incubating the wells by the indirect immunoperoxidase method. The antiserum showed a 25% cross reaction with spermine (Spm), putrescine (Put), and cadaverine (Cad), and a 1% cross reaction with 1,3-diaminopropane (Dap), but no cross reaction with monoacetyl polyamines and amino acids. The antibody binding was inhibited most effectively by absorption of the antiserum with N ¹-acetylspermidine and Spd in the ELISA inhibition test. Also, immunoblot analysis of the antiserum with nitrocellulose paper gave completely identical results to the ELISA binding tests. Spd-like immunoreactivities in human melanoma BD and neuroblastoma IMR 32 cell lines are presented as examples of the staining pattern obtained with the antiserum. Absorption of the serum with N ¹-acetylspermidine and Spd was demonstrated to abolish the immunostaining reaction. The immunohistochemical model is simple: amines and amino acids are bound in the same way as in aldehyde-fixed tissues and, in comparison to immunoblot analysis, the immunoreactivity can be more easily and accurately quantified by assay with the antibody. The model should prove useful in assessing the specificity of other antisera.     

水稻细胞质雄性不育系幼穗发育过程中多胺与乙烯的关系初探(英文 )

利用ＨＰＬＣ和ＧＣ分别测定了水稻细胞质雄性不育系及其保持系幼穗多胺（ 腐胺,亚精胺和精胺） 含量和乙烯释放速率,并研究了外施多胺合成抑制剂ＭＧＢＧ 和乙烯前体ＡＣＣ生成抑制剂ＡＶＧ 对两系幼穗多胺含量和乙烯释放速率以及花粉育性的影响。结果表明, 不育系幼穗乙烯释放速率显著高于其保持系幼穗, 外施ＡＶＧ 引起两系幼穗乙烯释放速率下降,并使不育系花粉育性得以部分恢复; 不育系幼穗多胺含量显著低于保持系幼穗, 外施ＭＧＢＧ 使两系幼穗Ｓｐｄ 和Ｓｐｍ 含量下降, 并使保持系花粉育性降低。外施ＡＶＧ 抑制乙烯释放,促进多胺合成;而外施ＭＧＢＧ 抑制Ｓｐｄ和Ｓｐｍ 合成, 却促进乙烯的释放; 而且,乙烯释放速率与多胺（精胺和亚精胺） 含量呈显著负相关。提示在水稻ＣＭＳ 系及其保持系幼穗发育过程中乙烯与多胺（ 精胺和亚精胺） 的生物合成竞争ＳＡＭ。     

Bridging the gap between plant and mammalian polyamine catabolism: a novel peroxisomal polyamine oxidase responsible for a full back-conversion pathway in Arabidopsis

In contrast to animals, where polyamine (PA) catabolism efficiently converts spermine (Spm) to putrescine (Put), plants have been considered to possess a PA catabolic pathway producing 1,3-diaminopropane, Delta(1)-pyrroline, the corresponding aldehyde, and hydrogen peroxide but unable to back-convert Spm to Put. Arabidopsis (Arabidopsis thaliana) genome contains at least five putative PA oxidase (PAO) members with yet-unknown localization and physiological role(s). AtPAO1 was recently identified as an enzyme similar to the mammalian Spm oxidase, which converts Spm to spermidine (Spd). In this work, we have performed in silico analysis of the five Arabidopsis genes and have identified PAO3 (AtPAO3) as a nontypical PAO, in terms of homology, compared to other known PAOs. We have expressed the gene AtPAO3 and have purified a protein corresponding to it using the inducible heterologous expression system of Escherichia coli. AtPAO3 catalyzed the sequential conversion/oxidation of Spm to Spd, and of Spd to Put, thus exhibiting functional homology to the mammalian PAOs. The best substrate for this pathway was Spd, whereas the N(1)-acetyl-derivatives of Spm and Spd were oxidized less efficiently. On the other hand, no activity was detected when diamines (agmatine, cadaverine, and Put) were used as substrates. Moreover, although AtPAO3 does not exhibit significant similarity to the other known PAOs, it is efficiently inhibited by guazatine, a potent PAO inhibitor. AtPAO3 contains a peroxisomal targeting motif at the C terminus, and it targets green fluorescence protein to peroxisomes when fused at the N terminus but not at the C terminus. These results reveal that AtPAO3 is a peroxisomal protein and that the C terminus of the protein contains the sorting information. The overall data reinforce the view that plants and mammals possess a similar PA oxidation system, concerning both the subcellular localization and the mode of its action.     

渗透胁迫对小麦胚芽鞘内多胺的种类、形态和含量的影响

用高压液相色谱法研究了豫麦18（抗旱性较强）和扬麦9号（抗旱性较弱）小麦胚芽鞘中三种不同形态的多胺（polyamine,PA）：游离态多胺（PA）、高氯酸可溶性结合态多胺（ps结合态PA）和高氯酸不溶性结合态多胺（PIS结合态PA）与渗透胁迫的关系。结果发现：渗透胁迫2d,豫麦18胚芽鞘中的游离态Spd和游离态Spm的含量明显上升,而扬麦9号的游离态Put的上升明显。S-腺苷蛋氨酸脱羧酶（S—AMDC）的抑制剂——甲基乙二醛-双（鸟嘌呤腙）（MGBG）处理豫麦18,明显抑制了渗透胁迫诱导的游离态Spd和游离态Spm的增加,并且加重了渗透胁迫伤害,外源Spd处理扬麦9号明显促进了渗透胁迫诱导的游离态Spd和游离态Spm的增加,并且减缓了渗透胁迫的伤害。渗透胁迫下,豫麦18胚芽鞘中的PS结合态PA和PIS结合态PA的上升幅度都明显大于扬麦9号。菲咯啉（o—Phen）抑制渗透胁迫下PIS结合态PA的合成并加重了渗透胁迫对胚芽鞘的伤害。这些结果表明：小麦胚芽鞘中的游离态Spd、游离态Spm、PS结合态PA和PIS结合态PA的升高有利于增强渗透胁迫抗性。     

Polyamine metabolism of potato seed-tubers during long-term storage and early sprout development

Growth potential of potato (Solanum tuberosum L.) plants is influenced by seed-tuber age. After 24 days of growth, single-eye seedcores from 7-month-old seed-tubers produced 64% more foliar dry matter than those from 19-month-old seed-tubers, reflecting a higher growth rate. This study was initiated to determine if differences in polyamine (PA) metabolism are associated with aging and age-reduced vigor of potato seed-tubers. As tubers aged in storage, putrescine (Put) increased 2.2-fold, while spermidine (Spd) and spermine (Spm) decreased 33% and 38%, respectively. Ethylene content of the tuber tissue also increased with advancing age, suggesting that during the aging process S-adenosylmethionine was directed toward ethylene biosynthesis at the expense of the PAs. Single-eye cores from 7- and 19-month-old tubers were sown and PA levels in core and shoot tissues were monitored during plant development. Put titer of younger cores increased 8.8-fold by 12 days. In contrast, the increase in Put over the initial titer in older cores was 2.9-fold. The reduced ability of older cores to synthesize Put during plant establishment is probably due to a 45% decline in ornithine decarboxylase activity between 12 and 16 days after planting. Lack of available Put substrate limited the biosynthesis of Spd and Spm, and thus their concentrations remained lower in older cores than in younger cores. Lower PA titer in older cores during plant establishment is thus coincident with reduced growth potential. Concentrations of Put and Spd were higher in shoots developing from older cores throughout the study, but there was no age-related difference in Spm content. In contrast, activities of arginine and S-adenosylmethionine decarboxylases were higher in shoots from younger cores during establishment. The results indicate that aging affects PA metabolism in both tuber and developing plant tissues, and this may relate to loss of growth potential with advancing seed-tuber age.     

Promoter strength influences polyamine metabolism and morphogenic capacity in transgenic rice tissues expressing the oat adc cDNA constitutively

We analyzed molecularly and biochemically a series of transgenic rice lines expressing the oat adc (arginine decarboxylase) cDNA under the control of the constitutive maize ubiquitin 1 promoter. We established baseline biochemical parameters to elucidate the role of polyamines (PAs) during morphogenesis. We measured mRNA levels, ADC enzyme activity and cellular PAs in dedifferentiated callus. Polyamine levels were also quantified in two subsequent developmental stages – regenerating tissue and differentiated shoots. We observed significant (P<0.05) differences in the levels of individual PAs at the three developmental stages. The amounts of putrescine (Put) and spermidine (Spd) in dedifferentiated transgenic callus were lower than those in the wild type or in hpt (hygromycin resistant)-controls, whereas the amount of spermine (Spm) was increased up to two-fold. In regenerating tissue, this trend was reversed, with significantly higher levels of Put and Spd (P<0.05), and lower levels of Spm (P<0.05) compared to non-transformed or hpt-control tissues at the same developmental stage. In differentiated shoots, there was a general increase in PA levels, with significant increases in Put, Spd, and Spm (P<0.05); on occasion reaching six times the level observed in wild type and hpt-control tissues. These results contrast those we reported previously using the weaker CaMV 35S promoter driving adc expression. mRNA measurements and ADC enzyme activity were consistently higher (P<0.01) in all tissues expressing pUbiadcs compared to equivalent tissues engineered with 35Sadc. Our findings are consistent with a threshold model which postulates that high adc expression leading to production of Put above a basal level is necessary to generate a big enough metabolic pool to trigger PA flux through the pathway leading to an increase in the concentration of Spd and Spm. This can be best accomplished by a strong constitutive promoter driving adc. We discuss our results in the context of flux through the PA pathway and its impact on morphogenesis.     

The Relationship between Polyamines and Hormones in the Regulation of Wheat Grain Filling

The grain weight of wheat is strongly influenced by filling. Polyamines (PA) are involved in regulating plant growth. However, the effects of PA on wheat grain filling and its mechanism of action are unclear. The objective of the present study was to investigate the relationship between PAs and hormones in the regulation of wheat grain filling. Three PAs, spermidine (Spd), spermine (Spm), and putrescine (Put), were exogenously applied, and the grain filling characteristics and changes in endogenous PA and hormones, i.e., indole-3-acetic acid (IAA), zeatin (Z) + zeatin riboside (ZR), abscisic acid (ABA), ethylene (ETH) and gibberellin 1+4 (GAs), were quantified during wheat grain filling. Exogenous applications of Spd and Spm significantly increased the grain filling rate and weight, but exogenous Put had no significant effects on these measures. Exogenous Spd and Spm significantly increased the endogenous Spd, Spm, Z+ZR, ABA, and IAA contents and significantly decreased ETH evolution in grains. The endogenous Spd, Spm and Z+ZR contents were positively and significantly correlated with the grain filling rate and weight of wheat, and the endogenous ETH evolution was negatively and significantly correlated with the wheat grain filling rate and weight. Based upon these results, we concluded that PAs were involved in the balance of hormones that regulated the grain filling of wheat.     

Glutaraldehyde (GA)-hapten adducts, but without a carrier protein, for use in a specificity study on an antibody against a GA-conjugated hapten compound: histamine monoclonal antibody (AHA-2) as a model

In our recent study on monoclonal antibodies (mAbs AHA-1-5) against glutaraldehyde (GA)-conjugated histamine (HA), we identified one mAb (AHA-2) which can detect neuronal HA in the rat brain with an immunocytochemistry method (ICC) [Fujiwara et al. (1999) J. Biochem. 126, 503-509]. In the present study the specificity of AHA-2 mAb for use for ICC has been examined by means of competitive experiments involving HA and analogs, all of which had been allowed to react with GA followed by sodium borohydride, but not allowed to couple with the carrier protein. It was demonstrated that the antibody distinguished alterations in the chemical structure of the molecule, showing decreased immunoreactivity with all the GA-adducts of (R)-(-)-alpha-methylhistamine, 1- and 3-methylhistamine, L-histidine, and 1- and 3-methyl-L-histidine. On the other hand, AHA-1 mAb only reacted with GA-adducts of 3-MeHA (3-MeHA-GA) and HA (HA-GA), to almost the same degree, in relatively high concentration ranges. AHA-3, 4, and 5 mAbs reacted about 10-times more strongly with 1-MeHA-GA than with HA-GA, but reacted very little or not at all with the other analogs. These results may suggest that AHA-2 mAb recognized both the non-substituted imidazole and alpha-methine groups of a HA molecule in addition to the conjugation site of GA including the part(s) reduced with NaBH(4), and especially the imidazole group more strictly than the other mAbs. This may partly explain why AHA-2, among the five AHA mAbs, can detect neuronal HA with an ICC method. The present ELISA method for GA-hapten adducts should be applicable to other antibodies against GA-conjugated biologically active amines or amino acids, thus allowing the study of antibody specificity for ICC more easily and accurately than was previously possible with hapten-protein conjugates as antigens.     

Involvement of polyamines in the post-anthesis development of inferior and superior spikelets in rice

Early-flowered superior spikelets usually exhibit a faster grain filling rate and heavier grain weight than late-flowered inferior spikelets in rice (Oryza sativa L.). But the intrinsic factors responsible for the variations between the two types of spikelets are unclear. This study investigated whether and how polyamines (PAs) are involved in regulating post-anthesis development of rice spikelets. Six rice genotypes differing in grain filling rate were field grown, and PA levels and activities of the enzymes involved in PA biosynthesis were measured in both superior and inferior spikelets. The results showed that superior spikelets exhibited higher levels of free spermidine (Spd) and free spermine (Spm) and higher activities of arginine decarboxylase (ADC, EC 4.1.1.19), S-adenosylmethionine decarboxylase (SAMDC, EC 4.1.1.50) and Spd synthase (EC 2.5.1.16) than inferior spikelets at the early endosperm cell division and grain filling stage. The maximum concentrations of free Spd and free Spm and the maximum activities of ADC, SAMDC and Spd synthase were significantly correlated with the maximum cell division and grain filling rates, maximum cell number and grain weight. Application of Spd and Spm to panicles resulted in significantly higher rates of endosperm cell division and grain filling in inferior spikelets along with the activities of sucrose synthase (EC 2.4.1.13), ADP glucose pyrophosphorylase (EC 2.7.7.27) and soluble starch synthase (EC 2.4.1.21), suggesting that these PAs are involved in the sucrose-starch metabolic pathway. The results indicate that the poor development of inferior spikelets is attributed, at least partly, to the low PA level and its low biosynthetic activity.     

干旱胁迫下小麦幼苗根、叶多胺含量和多胺氧化酶活性的变化

 以抗旱性不同的两个小麦品种（‘晋麦33’和‘温麦8’）（Triticum aestivum cv. Jinmai 33 and Wenmai 8）为材料,研究了干旱胁迫下多胺含量和多胺氧化酶活性的变化。结果表明：旱过程中,幼苗根、叶中腐胺（Put）、亚精胺（Spd）、精胺（Spm）3种多胺含量和多胺氧化酶（PAO）活性先迅速升高,而后下降。与抗旱性弱的‘晋麦33’相比,抗旱性强的品种‘温麦8’幼苗根、叶中Spd、Spm 含量初期升高幅度大,之后下降速率减慢;PAO活性的变化与之相反,‘晋麦33’的PAO活性提高的幅度大于‘温麦8号’。多胺含量和PAO活性在小麦幼苗的根与叶之间呈极显著正相关。干旱初期,小麦根、叶中多胺迅速积累可能是干旱胁迫反应的一个信号,随后较高的Spd、Spm 水平有利于增强小麦幼苗的抗旱性。     

Involvement of polyamines in the drought resistance of rice

This study investigated whether and how polyamines (PAs) in rice (Oryza sativa L.) plants are involved in drought resistance. Six rice cultivars differing in drought resistance were used and subjected to well-watered and water-stressed treatments during their reproductive period. The activities of arginine decarboxylase, S-adenosyl-L-methionine decarboxylase, and spermidine (Spd) synthase in the leaves were significantly enhanced by water stress, in good agreement with the increase in putrescine (Put), Spd, and spermine (Spm) contents there. The increased contents of free Spd, free Spm, and insoluble-conjugated Put under water stress were significantly correlated with the yield maintenance ratio (the ratio of grain yield under water-stressed conditions to grain yield under well-watered conditions) of the cultivars. Free Put at an early stage of water stress positively, whereas at a later stage negatively, correlated with the yield maintenance ratio. No significant differences were observed in soluble-conjugated PAs and insoluble-conjugated Spd and Spm among the cultivars. Free PAs showed significant accumulation when leaf water potentials reached -0.51 MPa to -0.62 MPa for the drought-resistant cultivars and -0.70 MPa to -0.84 MPa for the drought-susceptible ones. The results suggest that rice has a large capacity to enhance PA biosynthesis in leaves in response to water stress. The role of PAs in plant defence to water stress varies with PA forms and stress stages. In adapting to drought it would be good for rice to have the physiological traits of higher levels of free Spd/free Spm and insoluble-conjugated Put, as well as early accumulation of free PAs, under water stress.     

UV-B 辐射对番茄花粉生活力的影响与内源激素和多胺的关系

以成熟期不同的两个番茄品种同辉(早熟型)和霞光(晚熟型)为试材, 模拟兰州地区12%和20%臭氧层减薄时增强的UVB辐射(分别为T1=2.54 KJ.m-2.d-1和T2=4.25 KJ.m-2.d-1), 研究了大田条件下增强UV-B辐射对其花粉生活力的影响以及雄蕊中4种内源激素(IAA、GAs、ZR和ABA)和多胺(Put、Spm和Spd)及脯氨酸的变化。结果表明: 辐射抑制了同辉的花粉萌发和花粉管伸长, 但只降低了霞光的花粉萌发率; 两种辐射明显降低了两品种番茄雄蕊中的GAs含量, 同时造成同辉雄蕊中Put和Spd含量明显增加, Spm和Put/Spd+Spm比值显著降低; 霞光中3种多胺含量都显著减少, 导致高辐射时Put/Spd+Spm比值上升; 同辉番茄雄蕊的脯氨酸含量不受影响, 但高辐射使霞光番茄雄蕊的脯氨酸含量降低。实验表明, 两品种番茄花粉生活力的变化与增强UV-B 辐射下雄蕊中GAs水平、Spm含量以及脯氨酸含量的减少有关。雄蕊中多胺和脯氨酸含量变化对UVB辐射的响应说明霞光品种对UV-B辐射更敏感。     

Do exogenous polyamines have an impact on the response of a salt-sensitive rice cultivar to NaCl?

In order to analyze the putative impact of polyamines (PAs) on the plant response to salt, seedlings from the salt-sensitive rice cultivar I Kong Pao (IKP) were exposed for 5, 12 and 19 days to 0, 50 or 100 mM NaCl in the absence, or in the presence of exogenous PAs (putrescine (Put), spermidine (Spd) or spermine (Spm) 1mM) or inhibitors of PA synthesis (methylglyoxalbis-guanyl hydrazone (MGBG) 1mM, cyclohexylammonium (CHA) 5mM and D-arginine (D-Arg) 5mM). The addition of PAs in nutritive solution reduced plant growth in the absence of NaCl and did not afford protection in the presence of salt. PA-treated plants exhibited a higher K+/Na+ ratio in the shoots, suggesting an improved discrimination among monovalent cations at the root level, especially at the sites of xylem loading. The diamine Put induced a decrease in the shoot water content in the presence of NaCl, while Spd and Spm had no effects on the plant water status. In contrast to Spd, Spm was efficiently translocated to the shoots. Both PAs (Spd and Spm) induced a decrease in cell membrane stability as suggested by a strong increase in malondialdehyde content of PA-treated plants exposed to NaCl. These results are discussed in relation to the putative functions of PAs in stressed plant metabolism.     

Stress-dependent accumulation of spermidine and spermine in the halophyte Mesembryanthemum crystallinum under salinity conditions

We studied the effects of chloride salinity (300 and 500 mM NaCl) on the content of free polyamines (PAs) from putrescine (Put) family in Mesembryanthemum crystallinum L. leaves and roots. The contents of Put and spermidine (Spd) in leaves increased temporarily, achieving the highest values on the third day of salinity treatment; thereafter (by days 7–14), they dropped sharply. The content of spermine (Spm) increased gradually, and its high level was maintained until the end of experiment. The dynamics of Spm accumulation in leaves under salinity conditions resembled that of phosphoenolpyruvate carboxylase (PEPC), a key enzyme of the water-saving CAM pathway of photosynthesis. This indicates indirectly the involvement of Spm in the common ice plant adaptation to salinity. A decrease in the molar ratios of Spd to Spm in the leaves under salinity conditions could point to the acceleration of Spm biosynthesis (accumulation) during plant adaptation, whereas the levels of Spm precursors, Put and Spd, did not increase. This phenomenon could be explained by an accelerated conversion of Spd into Spm, an active liberation of free Spm from its conjugates, or changes in the rates of studied PA biosynthesis and degradation under salinity. At the same time, the intracellular concentration of ethylene rose under these conditions. It was supposed and then demonstrated, that the pathway of ethylene biosynthesis and that of the synthesis of Put family PAs compete under severe salinity conditions. This competition might be based on the disturbances in sulfur metabolism and a decrease in the methionine content, an immediate precursor of S-adenosyl-L-methionine.     

节瓜茎尖多胺含量和比值与花性别分化的关系

以4个不同基因型的节瓜为材料,通过两个发育时期（10、19片叶展平）茎尖取样,研究了多胺（PA）含量和比值与植株花性别分化的关系。结果表明,节瓜茎尖4种多胺含量差异显著,两个取样时期都是亚精胺（Spd）〉腐胺（Put）〉尸胺（Cad）〉精胺（Spm）。10片叶展平时期多胺含量与节瓜花性别分化之间没有明确的相关性;19片叶展平时期,节瓜茎尖Put、Spd和多胺总量与植株雌花分化比例呈极显著的正相关,而Cad则与雌花分化比例呈极显著的负相关。在两个取样时期,复合指标Spd/PA都与植株雌花分化比例呈显著的正相关,而（Put＋Cad）/（Spd＋Spm）均与之呈显著的负相关,可以较好地预测节瓜的花性别分化状况。     

Immunocytochemical localization of polyamines in the gastrointestinal tracts of rats and mice

An immunocytochemical method using a recently produced monoclonal antibody (ASPM-29) with an antibody specificity to spermine (Spm) and spermidine (Spd) fixed in situ, was used to demonstrate an immunocytochemical localization of polyamine (PA) pools in the gastrointestinal tracts of rats and mice. High PA immunoreactivity was always found in the cytoplasm of cells not only at the cell proliferative zone or the precursor cell zone but also at the neighboring non-proliferative premature cell zone of the epithelium, and a gradient of decreasing PA levels was noticed from these cells to the fully mature differentiated gastric surface mucous cells and absorptive cells of the small and large intestines. Also, strong staining for PAs was seen in the cytoplasm of fully differentiated gastric chief cells and neurons of both the myenteric and submucous plexuses, whereas the nuclei of the cells remained virtually unstained. These results may suggest that PAs are closely associated with the high biosynthetic activity in the cells of the gastrointestinal mucosa of normal rats and mice. This seems to be consistent with the PA imunocytochemical results previously obtained for neoplastic cells and active protein- or peptide-secreting cells, including exocrine or endocrine cell types.