Desaturation of Oleic and Linoleic Acids by Leaves of Dark- and Light-grown Maize Seedlings

Oleate and linoleate desaturation in leaves of maize seedlings was largely independent of previous light treatment of the seedlings; there was no evidence of light-induced desaturase activities. These results are in sharp contrast to those observed with developing cucumber cotyledons in which pronounced increase in desaturation occurs after exposure of tissue to light. The rates of desaturation of oleate were about four times those of linoleate in both etiolated and 16-hour greened maize leaves. In both etiolated and greened tissues, about two-thirds of the label from oleate was esterified after 4 hours, half of which was in phosphatidylcholine. Phosphatidylcholine and diglyceride contained large proportions of [¹⁴C]linoleate formed from [¹⁴C]oleate but not [¹⁴C]linolenate. In monogalactolipid, about two-thirds of the labeled fatty acids were linolenate. In vivo desaturase activity was present in tissue of widely different levels of differentiation and chlorophyll content obtained from light-grown maize seedlings.     

An Evaluation of Various Fixing and Staining Procedures for Mitochondria in Plant Root Tissues

Mitochondria were stained intensely by a Regaud iron-hematoxylin procedure for roots fixed in formalin-sublimate or in Helly's fluid. Formalin-sublimate fixation required iodization during the staining sequence, but roots fixed in Helly's fluid were best iodized to remove mercurial precipitates before embedding in paraffin. Both methods required treatment with 1% KOH before immersion in the staining solution to remove RNA and produce pale cytoplasm. A third successful method was to postosmicate methacrylate-embedded roots after fixation in Hermann's fluid. Blackened mitochondria were produced by the postosmication and further staining was unnecessary. Fixation in Regaud's fluid did not give successful stains in any of the three methods tested. A prefixation treatment in quinone did not aid in obtaining sharply stained mitochondria of roots fixed in Bouin's fluid and stained with Heiden-hain's iron-hematoxylin.     

铬在南瓜体内的分布与积累

采用^51Cr同位素标记法检测富铬南瓜品种‘永安2号’在不同发育时期对^51Cr的吸收和分配的结果表明：苗期（8片叶）,南瓜地上各部分^51Cr放射性强度的相对比例分别为：叶64．4％,茎30．2％,花5．4％,主要分布在距根部较近的叶片和茎中,幼叶中含量很低;幼果期分别为：果肉51．1％,叶片41．6％,茎4．8％,花2.4％,主要分布在果肉和中部叶片中。     

Sequence comparison of the 63-, 61-, and 59-kDa calmodulin-dependent cyclic nucleotide phosphodiesterases.

Partial protein sequences from the 59-kDa bovine heart and the 63-kDa bovine brain calmodulin-dependent phosphodiesterases (CaM-PDEs) were determined and compared to the sequence of the 61-kDa isozyme reported by Charbonneau et al. [Charbonneau, H., Kumar, S., Novack, J. P., Blumenthal, D. K., Griffin, P. R., Shabanowitz, J., Hunt, D. F., Beavo, J. A. & Walsh, K. A. (1991) Biochemistry (preceding paper in this issue)]. Only a single segment (34 residues) at the N-terminus of the 59-kDa isozyme lacks identity with the 61-kDa isozyme; all other assigned sequence is identical in the two isozymes. Peptides from the 59-kDa isozyme that correspond to residues 23-41 of the 61-kDa protein bind calmodulin with high affinity. The C-terminal halves of these calmodulin-binding peptides are identical to the corresponding 59-kDa sequence; the N-terminal halves differ. The localization of sequence differences within this single segment suggests that the 61- and 59-kDa isozymes are generated from a single gene by tissue-specific alternative RNA splicing. In contrast, partial sequence from the 63-kDa bovine brain CaM-PDE isozyme displays only 67% identity with the 61-kDa isozyme. The differences are dispersed throughout the sequence, suggesting that the 63- and 61-kDa isozymes are encoded by separate but homologous genes.     

Non-Enzymatic Reduction of Nitrite by Means of Ascorbic Acid in Citrus and Other Higher Plant Tissues

It has been proved that the nitrite reduction in the leaves and other plant tissues of citrus and other green plants is partly or mainly a non-enzymatic chemical process, and a heat-stable factor present in these tissues is responsible for this reduction. It is suggested that ascorbic acid plays a major role in this chemical reaction since the reduction is inhibited by ascorbic acid oxidase. A significant association was also found between the ascorbic acid content and the nitrite reduction capacity of citrus leaves. Evidence has been presented that this non-enzymatic chemical reduction of nitrite occurs also in vivo as undetached citrus leaves on branches placed in NaNO₂ solution have shown diminution of their ascorbic acid content along with the absorption of nitrite. Stronger accumulation of nitrite in these leaf tissues was observed under dark conditions, apparently due to the inhibition of the biosynthesis of the ascorbic acid.     

Effect of Salicylhydroxamic Acid on Respiration, Photosynthesis, and Peroxidase Activity in Various Plant Tissues

Earlier reports from our laboratory described salicylhydroxamicacid (SHAM) stimulation of O₂ uptake by expanded soybean leavesor older green cotyledons. This stimulation could not be interpretedin terms of engagement or capacity of the cytochrome and alternativerespiratory pathways. In this report, we tested the possibilitythat a soluble peroxidase, which can be easily eluted from soybeanleaves and cotyledons, might be responsible for SHAM stimulationin whole tissue. The peroxidase catalyzes oxidation of NADHby O₂, is strongly stimulated by SHAM and benzhydroxamic acid(BHAM) and inhibited by KCN, propyl gallate and gentisic acid.This peroxidase, however, does not seem to be responsible forSHAM-stimulated O₂ uptake in whole, green tissue. In our earlier work reporting SHAM-stimulated respiration ingreen tissue, the samples had not been shielded from room light(10–20 µmol photons m^–2.s^–1). In thisreport, we show that O²-uptake rates of controls measured indarkness were always greater than those measured in room light.SHAM stimulation was not observed in the dark or in tissue withoutchlorophyll. We also found that CO₂ uptake of whole leafletsin saturating light was completely inhibited by SHAM fed throughthe transpiration stream. SHAM, therefore, is a potent inhibitorof photosynthesis. We conclude that the SHAM-stimulated respirationof green tissues we reported earlier likely was due to verylow rates of photosynthesis occurring under room light. ³Present address: SANDOZ Ltd., Agrobiological Research Station,4108 Witterswil, Switzerland ⁴Present address: WTC 1A3, Weyerhaeuser Co., Tacoma, WA 98477,U.S.A. (Received June 23, 1989; Accepted October 20, 1989)     

黑线姬鼠与大林姬鼠组织中超氧化物酶和过氧化物酶的分布与活性比较

以黑线姬鼠(Apodemus agrarius)和大林姬鼠(A. peninsulae)为研究对象,采用聚丙烯酰胺凝胶电泳(PAGE)不连续体系的方法,比较分析了心、肝、肾、肌肉、脑、肺6种器官和组织中超氧化物酶(SOD)和过氧化物酶(POD)活性,并建立了2种酶的电泳图谱。结果显示,上述2种酶在黑线姬鼠和大林姬鼠的6种器官和组织中均有表达并表现出明显的特异性,其中,2种鼠中超氧化物酶共分离出迁移率由0.15～0.66的9条电泳谱带,过氧化物酶共分离出迁移率由0.09～0.83的20条电泳谱带。在肝和肺中酶的活性最强,黑线姬鼠6种器官和组织中超氧化物酶活性均强于大林姬鼠,2种鼠组织中过氧化物酶的活性和分布相似,但在同一物种不同器官和组织间过氧化物酶的活性及分布存在明显差异。     

Molecular cloning and characterization of the Cl(-) pump-associated 55-kDa protein in rat brain.

The Cl(-)-ATPase/pump in the plasma membrane of the rat brain is a candidate for active outwardly directed Cl(-) translocating systems. We recently isolated a Cl(-) pump, 520- or 580-kDa protein complex, which consisted of 51-, 55-, 60-, and 62-kDa proteins. In this study, we cloned a cDNA encoding a 55-kDa glycoprotein, designated as ClP55, which contained an open reading frame of 1512 base pairs encoding a protein of 504 amino acids including a signal peptide of 28 amino acids. Northern and Western blot analyses demonstrated expression of ClP55 mainly in the cerebrum. Application of antisense phosphorothioate oligonucleotides to cultured neurons resulted in a marked increase in the intracellular Cl(-) concentration ([Cl(-)](i)). Immunohistochemical analysis indicated that ClP55 was localized to the plasma membranes of neurons such as hippocampal pyramidal neurons and cerebellar Purkinje cells. Taken together, these results suggest that ClP55 is one of the Cl(-) pump subunits responsible for Cl(-) pump activity.     

Differential Accumulation of Salicylic Acid and Salicylic Acid-Sensitive Catalase in Different Rice Tissues

We previously proposed that salicylic acid (SA)-sensitive catalases serve as biological targets of SA in plant defense responses. To further examine the role of SA-sensitive catalases, we have analyzed the relationship between SA levels and SA sensitivity of catalases in different rice (Oryza sativa) tissues. We show here that, whereas rice shoots contain extremely high levels of free SA, as previously reported (I. Raskin, H. Skubatz, W. Tang, B.J.D. Meeuse [1990] Ann Bot 66: 369-373; P. Silverman, M. Seskar, D. Kanter, P. Schweizer, J.-P. Metraux, I. Raskin [1995] Plant Physiol 108: 633-639), rice roots and cell-suspension cultures have very low SA levels. Catalases from different rice tissues also exhibit differences in sensitivity to SA. Catalase from rice shoots is insensitive to SA, but roots and cell-suspension cultures contain SA-sensitive catalase. The difference in SA sensitivity of catalases from these different tissues correlates with the tissue-specific expression of two catalase genes, CatA and CatB, which encode highly distinctive catalase proteins. CatA, which encodes a catalase with relatively low sequence homology to the tobacco SA-sensitive catalases, is expressed at high levels exclusively in the shoots. On the other hand, in roots and cell-suspension cultures, with northern analysis we detected expression of only the CatB gene, which encodes a catalase with higher sequence homology to tobacco catalases. The role of catalases in mediating some of the SA-induced responses is discussed in light of these results and the recently defined mechanisms of catalase inhibition by SA.     

Free Sugars and Organic Acids in the Leaves of Various Plant Species and their Compartmentation Between the Tissues4

The distribution of free sugars and organic acids between theepidermis and mesophyll of Tulipa gesneriana L., Vicia fabaL., and Commelina communis L. leaves was studied using mainlygas-liquid chromatography. Fructose, glucose, sucrose, and myo-inositol were found in theepidermis and mesophyll of all three species. In T. geenerianaleaf tissues arabinose (trace levels), stachyose, tuliposidesA and B (mainly in the mesophyll), and xylose (trace levelsalso in V. faba tissues) were also detected. The acids were more difficult to detect and identify, beingat considerably lower concentrations than the sugars in bothtissues. Fumaric, citric, malic, ascorbic (trace levels), andan unidentified acid were common to the epidermis and mesophyllof all three species. Of special interest was the detectionof large amounts of glyceric acid in the epidermis and mesophyllof V. faba; this acid was not detected in the tissues from theother species. Fumaric acid was also very abundant in the epidermisof V.faba. A special study was made of the compartmentation of acids andsugars between the epidermis and mesophyll of T. geenerianaleaves after light and dark treatments. No changes in free acidor sugar levels were detected in the epidermis or mesophyllafter these treatments. Except for suceinic acid (P < 0·05),there were no statistically significant differences in acidlevels between the epidermis and mesophyll but for most of thesugars (myo-inositol, arabinose, and xylose being exceptions)differences were highly significant (P < 0·001), highestlevels occurring in the mesophyll. The differences in sugarlevels and the similarity in acid levels between epidermis andmesophyll of tulip leaves were considered to be essentiallydue to the different CO₂ fixing mechanisms and capacities ofthe two tissues. The energy source for the essentially non-greenepidermal tissue was discussed.     

The production of 53-55-kDa isoforms is not required for rat L-histidine decarboxylase activity

Post-translational processing of the histamine-producing enzyme, L-histidine decarboxylase (HDC), leads to the formation of multiple carboxyl-truncated isoforms. Nevertheless, it has been widely reported that the mature catalytically active dimer is dependent specifically on the production of carboxyl-truncated 53-55-kDa monomers. Here we use transiently transfected COS-7 cells to study the properties of carboxyl-truncated rat HDC isoforms in the 52-58-kDa size range. Amino acid sequences important for the production of a 55-kDa HDC isoform were identified by successive truncations through amino acids 502, 503, and 504. Mutating this sequence in the full-length protein prevented the production of 55-kDa HDC but did not affect enzymatic activity. Further truncations to amino acid 472 generated an inactive 53-kDa HDC isoform that was degraded by the proteasome pathway. These results suggested that processed isoforms, apart from 53-55-kDa ones, contribute toward histamine biosynthesis in vivo. This was confirmed in physiological studies where regulated increases in HDC activity were associated with the expression of isoforms that were greater than 55 kDa in size. We provide evidence to show that regulation of HDC expression can be achieved by the differential production or differential stabilization of multiple enzyme isoforms.     

大鼠肾脏和其它组织中磷酸酪氨酸蛋白的研究

本文采用P-tyr-BSA为免疫原免疫家无得抗血清。将纯化的IgG与HRP偶联,建立了P-tyr-Pr的ELISA法,并测定了正常大鼠肾脏等组织中P-tyr-Pr含量,其分布规律如下:上清中P-tyr-Pr含量高者,其颗粒部分则低,反之亦然;其中肾脏上清中含量远比其它组织(脾、肺、肝等)高。在此基础上,又研究了膜性肾炎大鼠肾脏P-tyr-Pr含量,发现其上清中的含量远远高于正常大鼠肾脏中的含量。     

Cholinesterases from Plant Tissues: III. Distribution and Subcellular Localization in Phaseolus aureus Roxb

The distribution and localization of cholinesterase in Phaseolus aureus, Glycine max, and Pisum sativum is described. The enzyme is present in roots, leaves, stems, root callus tissue, root cells suspension cultures, and root nodules. Cholinesterase in roots is found primarily in the cell wall. In cell fractionation experiments, at least 95% of the cholinesterase activity is associated with cell wall material. The enzyme can be solubilized by salt solutions, whereas Triton X-100 and sodium deoxycholate solubilize relatively small amounts of the enzyme. Cytochemical techniques have been employed to show the presence of cholinesterase activity at the cell surface and in the cell wall of certain cells of the root.     

Metabolic Stability of 3-O-Methyl-d-Glucose in Brain and Other Tissues

3-O-Methyl-D-glucose (methylglucose) is often used to study blood-brain barrier transport and the distribution spaces of hexoses in brain. A critical requirement of this application is that it not be chemically converted in the tissues. Recent reports of phosphorylation of methylglucose by yeast and heart hexokinase have raised questions about its metabolic stability in brain. Therefore, we have re-examined this question by studying the metabolism of methylglucose by yeast hexokinase and rat brain homogenates in vitro and rat brain, heart, and liver in vivo. Commercial preparations of yeast hexokinase did convert methylglucose to acidic products, but only when the enzyme was present in very large amounts. Methylglucose was not phosphorylated by brain homogenates under conditions that converted 97% of [U-14C]glucose to ionic derivatives. When [14C]methylglucose, labeled in either the methyl or glucose moiety, was administered to rats by an intravenous pulse or a programmed infusion that maintained the arterial concentration constant and total 14C was extracted from the tissues 60 min later, 97-100% of the 14C in brain, greater than 99% of the 14C in plasma, and greater than 90% of that in heart and liver were recovered as unmetabolized [14C]methylglucose. Small amounts of 14C in brain (1-3%), heart (3-6%), and liver (4-7%) were recovered in acidic products. Plasma glucose levels ranging from hypoglycemia to hyperglycemia had little influence on the degree of this conversion. The distribution spaces for methylglucose were found to be 0.52 in brain and heart and 0.75 in liver.     

hSef基因在不同人组织和细胞系的表达分布

将人源Sef蛋白胞内区编码序列与GST融合构建原核表达质粒进行重组蛋白的表达与纯化并制备多抗 .在COS 7细胞中转染表达hSef显示 ,其分子量分别为 80kD ,10 0kD ,比体外翻译的分子量偏大 ,提示可能有糖基化存在 .Northern印迹的结果表明 ,hSefmRNA主要分布在人肾和睾丸组织 .RT PCR检测到hSefmRNA在众多细胞系有广泛存在 .免疫组化的结果显示 ,hSef蛋白在人肾和睾丸及相应癌组织表达水平较高 .     

Distribution of Protein Phosphatase Inhibitor-1 in Brain and Peripheral Tissues of Various Species: Comparison with DARPP-32

The distribution of inhibitor-1, a cyclic AMP-regulated inhibitor of protein phosphatase-1, was analyzed in various brain regions and peripheral tissues of various species by immunolabeling of sodium dodecyl sulfate-poly-acrylamide gel transfers using specific antibodies. The distribution of inhibitor-1 was directly compared to that of DARPP-32, a structurally related cyclic AMP-regulated inhibitor of protein phosphatase-1. In rat CNS, a single immunoreactive protein of M(r) 30,000, identified as inhibitor-1, was widely distributed. In contrast, DARPP-32 was highly concentrated in the basal ganglia. Inhibitor-1 was detected in brain tissue from frog (M(r) 27,000), turtle (M(r) 29,000/33,000), canary (M(r) 26,000), pigeon (M(r) 28,000), mouse (M(r) 30,500), rabbit (M(r) 26,500), cow (M(r) 27,000), and monkey (M(r) 27,500), but not from goldfish. Inhibitor-1 was detected at various levels in most peripheral tissues of the species studied; however, it was not detectable in certain tissues of particular species (e.g., rat and cow liver). DARPP-32 was detected in brain tissue of all the species tested except frog and goldfish, but was not detectable in most peripheral tissues. Both inhibitor-1 and DARPP-32 were concentrated in the cytosol and synaptosomal cytosol of rat striatum. The developmental expressions of inhibitor-1 and DARPP-32 in rat striatum differed: the level of inhibitor-1 peaked in the first postnatal week and then declined by the third postnatal week, whereas the level of DARPP-32 increased to a peak level by the third postnatal week and remained elevated thereafter.(ABSTRACT TRUNCATED AT 250 WORDS)