Localization of Galactolipid Biosynthesis in Etioplasts Isolated from Dark-Grown Wheat (Triticum aestivum L.)

Etioplasts were isolated from leaves of dark-grown wheat (Triticum aestivum L. var Starke II). Galactolipid biosynthesis was assayed in an envelope-rich fraction and in the fraction containing the rest of the etioplast membranes by measuring incorporation of ¹⁴C from uridine-diphospho[¹⁴C]galactose into monogalactosyl diacylglycerol and digalactosyl diacylglycerol. More than half of the galactolipid biosynthetic capability was found in the fraction of inner etioplast membranes. This fraction was subfractioned into fractions enriched in prolamellar bodies and membrane vesicles (prothylakoids), respectively. All membrane fractions obtained from etioplasts were able to carry out galactolipid biosynthesis, although the activity was very low in prolamellar body-enriched fractions. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis revealed markedly different polypeptide patterns between the different fractions. It is concluded that the capability of galactolipid biosynthesis of etioplasts probably is not restricted to the envelope, but is also present in the inner membranes of this plastid.     

Isolation of Protoplasts and Chloroplasts from Flag Leaves of Triticum aestivum L

Intact protoplasts and chloroplasts have been isolated from mature flag leaves of wheat (Triticum aestivum L.). Both showed high rates of photosynthesis, the best of which equaled those observed in the parent tissue (greater than 150 micromoles O₂ per milligram chlorophyll per hour). The presence of ethylenediaminetetraacetate and an alkaline medium (pH 8.4) were required in the isolation and assay for the achievement of maximum rates of photosynthesis by chloroplasts. Photosynthesis by isolated chloroplasts was inhibited at very low concentrations of external orthophosphate.     

Isolation and Partial Characterization of DNA Topoisomerase I from the Nucleoids of White Mustard Chloroplasts

DNA topoisomerase was isolated for the first time from nucleoids of white mustard (Sinapis alba L.) chloroplasts. The enzyme had a molecular weight of 70 kDa; was ATP-independent, required the presence of mono- (K⁺) and bivalent (Mg²⁺) cations, and was capable of relaxing both negatively and positively supercoiled DNA. These results suggest that the enzyme isolated belongs to the type IB DNA topoisomerases.     

Changes in the Endoplasmic Reticulum During Sieve Element Differentiation in Triticum aestivum

The changes in structure of the endoplasmic reticulum (ER) andits associations with other cell components have been studiedin differentiating protophloem sieve elements of root tips ofTriticum aestivum. In the young sieve elements single ER cisternaebearing ribosomes are dispersed in the cytoplasm. As differentiationprogresses ER increases in amount while a small proportion ofit aggregates into stacks or becomes associated with the nuclearenvelope and the mitochondria. These modifications occur inthe last two sieve elements containing ribosomes and coincidewith most dramatic changes in the degenerating nucleus. Stacksconsist of relatively few ER cisternae and may be encounteredfree in the cytoplasm or applied to the nuclear envelope. Electron-densematerial accumulates between the contiguous cisternae of thestacks. ER-attached ribosomes persist even in nearly maturesieve elements, but their pattern of arrangement becomes changed.The structural evidence indicates that only a few highly degradedER elements are retained in fully mature sieve elements. Triticum aestivum, root protophloem, sieve elements, endoplasmic reticulum, differentiation     

Expression of Green Fluorescent Protein from Bacterial and Plastid Promoters in Tobacco Chloroplasts

The expression of the green fluorescent protein reporter gene (gfp) from the bacterial trc and plastid rrn and psbA promoters has been compared in transplastomic tobacco plants produced by microprojectile bombardment. The homoplasmic nature of the regenerated plants was confirmed by Southern blot analysis. Northern blot analysis indicated that plants expressing gfp from the rrn promoter contained 3-fold more gfp RNA than plants containing the psbA promoter and 12-fold more than plants with the trc promoter. Immunoblot analysis and fluorescence spectroscopy indicated that plants expressing gfp from the rrn promoter contained approximately 90-fold more green fluorescent protein (GFP) than plants containing the psbA or trc promoters. This study demonstrates that the bacterial trc promoter is significantly weaker than the plastid rrn promoter for expression of gfp in tobacco chloroplasts.     

Cytological Basis of Plastid Inheritance in Phaseolus vulgaris-Investigations of Plastids,Mitochondria and Their DNA Nucleoids During Pollen Development

Electron microscopic and DNA fluorescence microscopic observations of the plastids, mitochondria and their DNA in the developing pollen of Phaseolus vulgaris L. have demonstrated that the male plastids were excluded during microspore mitosis. The formed generative cell was free of plastids because of regional localization of plastids in early developing microspore and the extremely unequal distribution during division. The fluorescence observations of DNA showed that cytoplasmic (plastid and mitochondria) nucleoids degenerated and disappeared during the development of microspore/pollen, and were never presented in the generative cell at different development stages. These results provided precise cytological evidence of maternal plastid inheritance in Phaseolus vulgaris, which was not in accord with the biparental plastid inheritance identified from early genetic analysis. Based on authors' previous observations in a variety of common bean that the organelle DNA of male gamete was completely degenerated, the early genetic finding of the biparental plastid inheritance was unlikely to be effected by genotypic difference. Thus those biparental plastid inheritance might be caused by occational male plastid transmission, and plastid uniparental maternal inheritance was the species character of Phaseolus vulgaris.     

Plastid development in germinating wheat (Triticum aestivum) is enhanced by gibberellic acid and delayed by gabaculine

Etioplast development and protochlorophyllide (Pchlide) accumulation was studied in wheat seedlings ( Triticum aestivum L. cv. Walde, Weibull) grown in darkness on gibberellic acid (GA₃), gabaculine (3-amino-2,3-dihydrobenzoic acid), or on a combination of the two. The results were compared with the features of seedlings grown on water only. GA₃ enhanced shoot growth and promoted etioplast development. A correlation was observed between the appearance of prolamellar bodies (PLBs) and of phototransformable Pchlide. Gabaculine, a known tetrapyrrole biosynthesis inhibitor, delayed growth, slowed down the rate of PLB formation and caused structural alterations of the etioplasts up to 48 h of germination. Gabaculine also delayed the formation of phototransformable Pchlide as well as overall Pchlide biosynthesis, as determined by low-temperature fluorescence emission in vivo. The spectral blue-shift of newly formed chlorophyllide (Chlide) was delayed in irradiated dark-grown gabaculine-grown seedlings, indicating an inhibited dissociation of Chlide and NADPH-Pchlide oxidoreductase (Pchlide reductase: EC 1.3.1.33). Thus there is a close correlation between accumulation of Pchlide and etioplast development, also under conditions when development is enhanced or delayed.     

Plant regeneration from isolated microspores of Triticum aestivum

Summary Wheat microspores were isolated, without prior anther culture, from a range of genotypes and cultured to regenerate self-fertile plants. Microspores were isolated using a microblender and competent microspores were enriched by gradient centrifugation. The use of maltose as the sole carbohydrate in the culture medium and co-culture of microspores with wheat or barley ovaries were critical for development of microspore-derived embryos. Results were also improved when spikes were pretreated by emersion of the basal ends of detached heads in water at 25°C for 2d. This procedure leads to highly reproducible production of plants.     

Changes in the Number and Composition of Chloroplasts during Senescence of Mesophyll Cells of Attached and Detached Primary Leaves of Wheat (Triticum aestivum L.)

Changes in the number and composition of chloroplasts of mesophyll cells were followed during senescence of the primary leaf of wheat (Triticum aestivum L.). Senescence was due to the natural pattern of leaf ontogeny or was either induced by leaf detachment and incubation in darkness, or incubation of attached leaves in the dark. In each case discrete sections (1 centimeter) of the leaf, representing mesophyll cells of the basal, middle, and tip regions, were examined. For all treatments, senescence was characterized by a loss of chlorophyll and the protein ribulose 1,5-bisphosphate carboxylase (RuBPCase). Chloroplast number per mesophyll cell remained essentially constant during senescence. It was not until more than 80% of the plastid chlorophyll and RuBPCase was degraded that some reduction (22%) in chloroplast number per mesophyll cell was recorded and this was invariably in the mesophyll cells of the leaf tip. We conclude that these data are consistent with the idea that degradation occurs within the chloroplast and that all chloroplasts in a mesophyll cell senesce with a high degree of synchrony rather than each chloroplast senescing sequentially.     

Structure-function relationships during metaphloem sieve elements d evelopment in Triticum aestivum

The differentiation of metaphloem sieve element (MSEs) in the developing caryopsis of wheat (Triticum aestivum L.) was a programmed cell semi-death process. We studied the changes of microtubules and polysaccharide contents during MSEs development. Some significant features are presented in MSEs, such as cell wall non-uniform thickening, chromatin condensation and so on. During the period of MSEs differentiation, numerous microtubules are distributed in the vicinity of the cell wall, but finally they vanished in mature MSEs. Large glycoconjugates in the cell wall and small glycoconjugates in the Golgi apparatus were observed in the developing MSEs. Programmed cell death (PCD) ceased in the mature MSEs after 6 d after flowering and higher aggregation of glycoconjugates appeared in the cytoplasm. All of these processes were in tight contact with the cell wall non-uniform thickening during PCD.     

Floret Initiation and Development in Spring Wheat (Triticum aestivum L.)

The number and developmental stages of florets were determinedin each spikelet of the spike in the main shoots of spring wheat.Samples were taken frequently from plants grown in a phytotronand in a nitrogen application field-test. Ten stages of development,from floret initiation until anthesis, were recognized and described. Inter-spikelet variation in the total number of initiated floretswas rather small. However, the number of florets at advancedstages of development, as well as the number of grains, washighest in the central spikelets in which florets initiatedfirst. Floret initiation did not proceed beyond spike emergence,whereafter the distal florets and the spikelet apex degenerated.Grain-set was restricted to florets which had developed at leastto the stage of visible anther lobes at spike emergence. Thenumber of these florets was increased significantly by nitrogenapplication. Wheat, Triticum aestivum L., spikelet, floret, grain set, nitrogen     

Structures of adenylosuccinate synthetase from Triticum aestivum and Arabidopsis thaliana

Catalyzing the first step in the de novo synthesis of adenylmonophosphate, adenylosuccinate synthetase (AdSS) is a known target for herbicides and antibiotics. We have purified and crystallized recombinant AdSS from Arabidopsis thaliana and Tritium aestivum, expressed in Escherichia coli. The structures of A. thaliana and T. aestivum AdSS in complex with GDP were solved at 2.9 A and 3.0 A resolution, respectively. Comparison with the known structures from E. coli reveals that the overall fold is very similar to that of the E. coli protein. The longer N terminus in the plant sequences is at the same place as the longer C terminus of the E. coli sequence in the 3D structure. The GDP-binding sites have one additional hydrogen-bonding partner, which is a plausible explanation for the lower K(m) value. Due to its special position, this partner may also enable GTP to initiate a conformational change, which was, in E. coli AdSS, exclusively activated by ligands at the IMP-binding site. The dimer interfaces show up to six hydrogen bonds and six salt-bridges more than in the E. coli structure, although the contact areas have approximately the same size.     

Sucrose metabolism during endosperm development in wheat (Triticum aestivum)

This work reports changes in sucrose synthase and invertase activities throughout endosperm development in wheat, together with the associated substrates and metabolites, sucrose, UDP, glucose, fructose and UDP-glucose. Throughout endosperm development, sucrose synthase had consistently higher activity than invertase and indeed invertase activity did not change appreciably. The observed variation in pattern and amounts of glucose and fructose present during the mid- and late stages of endosperm development confirmed the suggestion that invertase was not the preferred pathway of sucrose catabolism. Kinetic parameters for sucrose synthase were determined in crude extracts. Estimates of UDP and sucrose concentrations suggest that sucrose synthase is unlikely to achieve its potential maximum velocity. This limitation may however be overcome in part by the apparent excess catalytic activity measured during endosperm development.     

Enzymatic Activities modified during Multiplication and Differentiation of Neuroblastoma Cells

THE neuroblastoma cell system of Augusti-Tocco and Sato¹ is useful for studying neurone differentiation and function. The cells multiply rapidly in vitro and have low oxygen consumption². When serum is withdrawn from the culture medium, cellular expansion³, increase of acetylcholinesterase activity⁴ and increase of oxygen consumption² occur. The differences in respiratory metabolism of neuroblastoma cells, depending on whether they are proliferating or differentiating, may suggest major modifications of intermediary metabolism.     

Aminoacyl-tRNA Synthetases in Triticum aestivum L. During Seed Development and Germination

Changes in the level of total and individual aminoacyl-tRNAsynthetase activity in the various tissues of wheat (Triticumaestivum L.) were followed by the ATP-pyrophosphate (ATP-PP₁)exchange procedure throughout seed maturation and germination. During seed development the total synthetase activity in theendosperm increased up to the 5th week after fertilization andthereafter decreased rapidly. Over the same period, synthetaseactivity in the testa-pencarp decreased markedly, whilst theactivity in the developing embryo increased. Many of the individualsynthetases conformed with this general pattern although therewere several exceptions. The total synthetase activity of both the coleoptile and coleorhiza(root) increased rapidly during the first 2 d of germinationwhilst the total activity of these enzymes in the scutellumremained constant. After an initial increase on germination,aminoacyl-tRNA synthetase activity in the testa-aleurone layerremained almost constant until most of the endosperm had beendigested. With a few exceptions the relative levels of individualsynthetases in the various tissues did not change significantlyduring seed maturation or germination.     

Fertility of Triticum aestivum plants derived from anther culture

The fertility of plants from wheat anther culture was studied. It was found that one half of the 36 plants with diploid root tips didn't set seeds at all, and that 41 of the 42 plants with haploid root tips were completely sterile. It was surprising that sterility was so widely distributed even among the plants with diploid root tips.     

红盖鳞毛蕨孢子囊早期发育与质体分化的研究

利用光学显微镜和透射电子显微镜观察了红盖鳞毛蕨(Dryopteris erythrosora(Eaton)O.Ktze.)孢子囊的发育及在此期间质体的分化过程。研究表明:(1)红盖鳞毛蕨孢子囊的发育类型属于薄囊蕨型;(2)绒毡层为混合型,即内层绒毡层为原生质团型,外层绒毡层为腺质型;(3)孢子囊原始细胞中的质体通过3条路径分化,其一,原始细胞中含淀粉粒的质体通过分裂分配到下方细胞,继而进入孢子囊柄;其二,原始细胞分裂产生的新生质体被分配到上方细胞,进而被分配到除顶细胞外的原基细胞中,顶细胞将含淀粉粒的质体通过分裂分配到外套层原始细胞中;其三,顶细胞也将具淀粉粒的质体通过分裂分配到内部细胞,使分裂产生的孢原细胞和绒毡层原始细胞具新生质体;造孢细胞和孢子母细胞的质体具淀粉粒,孢子母细胞还具油体,新生孢子中具造粉体和油体;两层绒毡层具新生质体,随着退化外层绒毡层出现造粉体,内层绒毡层出现油体;(4)红盖鳞毛蕨与少数被子植物小孢子发育阶段质体分化模式类似,由前质体分化为造粉体再到油体。研究结果为蕨类植物质体在孢子囊发育过程不同组织细胞中的差异分化提供了新观察资料,为蕨类植物发育生物学和系统演化研究提供科学依据。     

Changes in the DNA,RNA and Protein Contents During the Root-Tip Cell Differentiation of Triticum aestivum

One to two cm long root tips of Triticum aestivum L., after three days' germination at 25 ℃, each were cut into three regions--the meristem region, elongation region and mature region. The cells in different regions were stained with Hochest 33258, Pymnin G and FITC respectively. Nuclear DNA was measured by micmfluorometry with VIDAS digital image analysis system. The relative contents of RNA and protein in the cells were measured with a MPV-Ⅲ microspectrofluorometer. It was shown that the nuclear DNA content increased during root tip cell differentiation, being maximum in the mature region. The relative content of RNA was maximum in the meristem region, but decreased continuously during the growth of tissue. The relative content of protein was maximum in the elongation region, but minimum in the meristem region. The relationships among DNA, RNA, protein and cell differentiation were discussed.