New Taxa of Heracleum L. (Umbelliferae) from China

Three new species of the genus Heracleum L. (Umbelliferae) are described from Sichuan Province, China. They are H. wenchuanense; H.wolongense and H. xiaojinense.     

The cell wall-plasmalemma interface in sieve tubes of barley

Both thick- and thin-walled sieve tubes in leaf-blade veins of Hordeum vulgare L. exhibit a distinct, electron-opaque inner wall layer after fixation in glutaraldehyde-osmium tetroxide and staining with uranyl acetate and lead citrate. This inner wall layer is thickest at the sieve plates and lateral sieve areas where it is permeated by a labyrinth of tubules formed by the plasmalemma. Along the lateral walls between sieve areas the inner wall layer apparently is penetrated by numerous microvilli-like evaginations of the plasmalemma, giving the cell wall-plasmalemma interface the appearance of a brush border. It is suggested that a similar brush-border-like structure may occur at the cell wall-plasmalemma interface of sieve elements in a wide variety of vascular plants.Abbreviation ER endoplasmic reticulum     

中国独活属的分支分类学研究

研究运用分支分析方法研究了16种独活属植物的系统发育关系。采用最简约性分析的分支与界限法Branch—and—Bound(bandh)的严格一致树(thestrict consensus cladogram)和非加权配对算术平均法up weighted pair group method using arithmetic average method(UPGMA)对其进行分支分析。研究结果表明：两种方法均支持将独活属分为4个组,即多裂叶组Sect．Millefolia、多毛组Sect．Villosa、多管组Sect．Plurivittata、独活组Sect．Heracleum。《中国植物志》中少管组Sect．Wendia的法落海Heracleum apaense没有独立成组,它的系统位置还需要进一步研究。根据PAUP软件分析中的不同处理方法,探讨了中国独活属植物的系统进化及其起源问题。     

Transient gene expression in pine pollen tubes following particle bombardment

 A biolistic particle delivery system was used to genetically transform pollen tubes of three species of white pine (Pinus aristata, P. griffithii and P. monticola). The introduced plasmid DNA contained the GUS coding sequence flanked by the 35S CaMV promoter and NOS terminator sequences. Successful gene delivery was demonstrated by transient GUS expression as evaluated by standard histochemical assay. Distance of target specimens significantly influenced transient GUS expression in all three species of white pine. A target distance of 6 cm resulted in a significant number of transformed pollen tubes in P. aristata and P. griffithii, while distances of 6 and 9 cm resulted in a significant number of transformed pollen tubes in P. monticola. Generally, the number of pollen tubes expressing GUS activity was higher in P. aristata than in P. griffithii and P. monticola. The possibility of using GUS-transformed pollen tubes in conjunction with in vitro fertilization in conifers was examined. Gene expression in pollen tubes was also examined under electron microscopy where the X-glu reaction product occurred as large crystalline electron-dense precipitates in the cytoplasm. Received: 17 December 1998 / Revision received: 17 March 1999 / Accepted: 14 April 1999     

Karyotype and cytogeography of the genus Heracleum （Apiaceae） in the Hengduan Mountains

In the present study, the karyotypes of 34 populations belonging to 11 species and one variety of Heracleum from the Hengduan Mountains in China were examined. Chromosome numbers and the karyotypes of three species （H. souliei, H. la＇ngdoni, and H. wenchuanense） are reported for the first time, as are the karyotypes of H. moellendorffii and H. henryi （tetraploid）. Populations of H. candicans, H. franchetii, and H. kingdoni in the Hengduan Mountains were found to consist of a mixture of diploid and tetraploid plants. Except for four species of Heracleum, namely H. candicans, H. franchetii, H. henryi, and H.kingdoni, which have both diploid and tetraploid karyotypes, all other species of Heracleum are were found to be diploid. All karyotypes were found to belong to the 2A type of Stebbins, with the exception ofH. candicans var. obtusifolium, which belongs to 2B, and H. hemsleyanum and H. franchetii （Mt. Dujuan, Daocheng, Sichuan, China）, which belong to 1A. There was only a slight difference in the karyotype asymmetry index, which suggests a close kinship for species of Heracleum and that the entire phylogenetic development of Heracleum is relatively primitive. Species that exhibited advanced morphological features were also more advanced in karyotype structure, with the order ofkaryotype evolution being 1A→2A→2B. This phenomenon indicates that the species distributed in the Hengduan Mountains have not diverged completely and that the Hengduan Mountains are a relatively young and active area for the evolution of Heracleum. Polyploidization in Heracleum may be an important evolutionary mechanisms for some species, generating diversity. The biological attributes, distribution range, and the geological history of the genus have all played a part in accelerating the evolution through polyploidization or aneuploidization. It is known that as the distribution latitude of Heracleum decreases from north to south, the chromosome number, ploidy level, and asymmetry structure appear to increase. In the Hengd     

Karyotype and cytogeography of the genus Heracleum (Apiaceae) in the Hengduan Mountains

Abstract In the present study, the karyotypes of 34 populations belonging to 11 species and one variety of Heracleum from the Hengduan Mountains in China were examined. Chromosome numbers and the karyotypes of three species (H. souliei, H. kingdoni, and H. wenchuanense) are reported for the first time, as are the karyotypes of H. moellendorffii and H. henryi (tetraploid). Populations of H. candicans, H. franchetii, and H. kingdoni in the Hengduan Mountains were found to consist of a mixture of diploid and tetraploid plants. Except for four species of Heracleum, namely H. candicans, H. franchetii, H. henryi, and H. kingdoni, which have both diploid and tetraploid karyotypes, all other species of Heracleum are were found to be diploid. All karyotypes were found to belong to the 2A type of Stebbins, with the exception of H. candicans var. obtusifolium, which belongs to 2B, and H. hemsleyanum and H. franchetii (Mt. Dujuan, Daocheng, Sichuan, China), which belong to 1A. There was only a slight difference in the karyotype asymmetry index, which suggests a close kinship for species of Heracleum and that the entire phylogenetic development of Heracleum is relatively primitive. Species that exhibited advanced morphological features were also more advanced in karyotype structure, with the order of karyotype evolution being 1A→2A→2B. This phenomenon indicates that the species distributed in the Hengduan Mountains have not diverged completely and that the Hengduan Mountains are a relatively young and active area for the evolution of Heracleum. Polyploidization in Heracleum may be an important evolutionary mechanisms for some species, generating diversity. The biological attributes, distribution range, and the geological history of the genus have all played a part in accelerating the evolution through polyploidization or aneuploidization. It is known that as the distribution latitude of Heracleum decreases from north to south, the chromosome number, ploidy level, and asymmetry structure appear to increase. In the Hengduan Mountains, these tendencies are also evident. Finally, based on all the available cytogeographic data, we speculate that the more advanced tetraplont or aneuploid species of Heracleum in India may be derived from early diplont species that were distributed in the Caucasus region and Hengduan Mountains. The dispersal of Heracleum was from Eurasia to India, because this correlates with the emergence of the Himalayan Mountains through tectonic movement. Thus, the Hengduan Mountains are not only a center of diversity for Heracleum, but also a center of active speciation in modern times.     

中国独活属和当归属系统演化及区系地理的温带性

中国复杂的地质地貌和古老的地史,尤其在横断山区和四川地区,是伞形科独活属及其近邻属的演化和分化中心,区系古老且温带性质明显.从核型和花粉方面分析,独活属和当归属的系统演化表现出比较密切的亲缘关系.     

The microscopy of P-protein filaments in freeze-etched sieve pores

Intact vascular bundles from Nymphoides peltata (S.G. Gmel.) O. Kuntze, shown to have translocated carbon-14, were freeze-fractured and etched for electron microscopy. The interpretation of freezefractured and etched sieve pores and P-protein filaments seen in them is discussed. The entire widths of most of the sieve pores seen contained filaments separated by less than 100 nm. Their arrangement indicates too high a resistance to flow for pressure flow alone to drive translocation at known rates; pumps would be necessary at places along sieve tubes. However, calculations are presented to show that during the time taken to fix pores, by fast freezing or chemically, the filaments in them could rearrange and move further by Brownian and other motion than the distances between filaments which we need to measure. These calculations show that it is not possible, by microscopy alone, to answer the outstanding question “How are filaments arranged in translocating sieve pores?” with enough certainty to tell us whether pressure flow is adequate to explain translocation where filaments are present. The calculations are relevant also to microscopy of other cell structures which may move.     

Evidence for the presence and activity of a complete antioxidant defence system in mature sieve tubes

The phloem is the major route for the transport of solutes and nutrients from source to sink organs in plants. The functional transport phloem consists of parenchymal tissue, enucleate sieve elements, and the intimately connected companion cells. The general absence of a nucleus and functional ribosomes in sieve tubes poses problems especially for damage avoidance and repair of sieve element components. To examine how sieve tubes can remain functional during oxidative stress, we analysed phloem sap of cucumber and pumpkin plants with respect to the presence of antioxidant defence enzymes, their enzymatic activity, and activity changes after exposure to drought stress. Using 1D SDS-PAGE and nano ESI MS/MS, the presence of proteins such as cytosolic Cu/Zn superoxide dismutase, monodehydroascorbate reductase, and peroxidase could be shown. Moreover, activities for several antioxidant enzymes (superoxide dismutase, dehydroascorbate reductase, peroxidase) in phloem exudate could be demonstrated. The activity of these enzymes in phloem sap from cucumber and pumpkin plants increased in response to drought stress. The presented results together with earlier findings provide evidence supporting the presence of a complete machinery of antioxidant defence enzymes and detoxifying metabolites important for avoiding damage to essential components of the sieve elements due to oxidative stress.     

Transmission of Electric Signals in Sieve Tubes of Zucchini Plants

Membrane potentials of ?;160 to ?210 mV were recorded with microelectrodes inserted into meta-phloem sieve tubes of intact zucchini plants (Cucurbita pepo L. var. medullosa Alef.). The effects of darkness, white light and colored light on membrane potential were studied. Reference electrodes were in contact with the apoplast via fluid-filled cavities or “drinks”. Electrolyte solutions (100 mM) in the cavities could be quickly replaced by flushing with 100 mM solutions of sucrose, KCl, sorbitol, or EDTA without altering osmolarity. KCl and EDTA caused depolarization of the sieve tube membrane potential, while sucrose caused depolarization or hyperpolarization of the sieve tube membrane potential in mature or growing plant parts respectively. Recovery of the original voltage was recorded when rapid (sucrose) or slow (sorbitol) transients occurred. When two measuring circuits were installed, one in a growing fruit and the other in the petiole of the subtending mature leaf, the alteration of the sieve tube membrane potential at one site was accompanied by an alteration of the potential at the other site after a few seconds. The responses were opposite in the exporting leaf and importing fruit when sucrose was applied. The signal, transmitted via the sieve tubes, reached maximum velocities of 10 cm per second.     

Correlated motion and oscillation of neighboring cells in vitro

It has long been realized that fibroblastic and epithelial cells establish recognizable patterns in tissue culture. This behavior implies that neighboring cells interact with one another to produce organized populations. Interaction between cells that are separated by many intervening cells is also possible and is demonstrated here using a special configuration of a biosensor referred to as electric cell-substrate impedance sensing (ECIS). Normally the electrical impedance of a single electrode covered with a confluent cell layer is measured, and the morphological changes of the cells are reflected in the impedance. In this case the cells are cultured on two closely spaced electrodes whose impedances are measured independently as a function of time, and communication between the cell populations is revealed as a correlation between these two time series. We also report for the first time another striking manifestation of dynamic cell interaction, where confluent layers of Madin-Darby canine kidney cells (MDCK) on a single electrode are observed to oscillate in synchrony with a period of approximately 2.5 h.     

细菌显微追踪技术在生物被膜中的应用

细菌生物被膜是粘附于物体表面的由细菌细胞及其胞外物质组成的复杂膜样物聚集体,具有很强的耐药性和免疫逃逸能力。生物被膜内细菌的代谢活性、运动状态等与浮游细菌有明显区别。近年来,先进的显微成像技术结合新型图像处理方法,在研究细菌的运动、生理等方面发挥了重要作用。本文围绕生物被膜,概述了细菌显微追踪技术在其研究中的应用。主要从细菌的运动方式和生物被膜形成过程的调控两方面出发,介绍了在单细胞水平上利用该技术研究生物被膜的进展,包括细菌的游泳、蹭行、群集运动和多种信号通路调控下生物被膜的形成过程等,并展望了该技术在生物被膜其他相关研究领域的应用前景。     

Wall architecture with high porosity is established at the tip and maintained in growing pollen tubes of Nicotiana tabacum

A major question in pollen tube growth in planta remains: do the pollen tube walls form a barrier to interaction with the environment? Using cryo‐FESEM, we directly assessed the 3D construction and porosity of tobacco pollen tube walls. Fractured mature primary walls showed a 40–50 nm spaced lattice of continuous fibers interconnected by short rods in the primary wall. These observations agree with TEM observations of sectioned walls. In the secondary callose wall, for which no structure is visible using TEM, cryo‐FESEM also revealed a 50 nm lattice consisting of longer fibers, approximately 10–15 nm wide, with rod‐like, thinner interconnections at angles of approximately 90° with the longer fibers. Such architecture may reflect functional needs with respect to porosity and mechanical strength. The wall does not form a mechanical barrier to interaction with the environment and is gained at low cost. Cryo‐FESEM additionally revealed another special feature of the wall: the tubes were tiled with scales or rings that were highly conspicuous after pectin extraction with EDTA. These rings cause the typical banding patterns of pectin that are commonly seen in pollen tubes during oscillatory growth, as confirmed by staining with toluidine blue as well as by DIC microscopy. Growth analysis by VEC‐LM showed that the ring‐ or scale‐like structures of the primary wall consist of material deposited prior to the growth pulses. The alternating band pattern seen in the callose wall is probably imposed by constrictions resulting from the rings of the primary wall.     

Influence of thermal motion on 1H chemical shifts in proteins: the case of bovine pancreatic trypsin inhibitor.

The possible influence of thermal motion on 1H chemical shifts is discussed for a small stable protein, the bovine pancreatic Kunitz trypsin inhibitor (BPTI). The thermal effects on the aromatic side chains and on the backbone are treated separately. The thermal motion of the aromatic side chains is accounted for in terms of their rotation around the C(alpha)-C(beta) bond and the motion of each individual proton is interpreted as a ratio between the amount of ordered and quite disordered states. The influence of hydrogen bonds is introduced as an extra contribution to the chemical shifts of the bonded proton. Their contribution to the chemical shifts resulting from the polarization of the peptide bond is investigated, as is their influence on local flexibility. Finally, the relative importance of each contribution to the chemical shift information is compared.     

Perturbed turnover of microtubule-based nutritive tubes in ovarioles of virgin and precocene-treated Dysdercus fasciatus

During early oogenesis in Dysdercus fasciatus, anteriorly positioned nurse cells supply each oocyte with mRNA, ribosomes, and proteins via a microtubule-rich nutritive tube that lengthens as the oocyte is displaced backwards down an ovariole. Nurse cell-dependent development of an oocyte continues until the latter reaches a particular stage of oogenesis after which the nutritive tube supplying it becomes redundant and breaks down. The signal for nutritive tube breakdown is believed to derive from the oocyte, and to be developmental stage-specific. To explore this, nutritive tube turnover has been investigated following the experimental inhibition of oocyte maturation both by the prevention of mating, and also the topical application of precocene II. In each case, the nutritive tubes with their component microtubules continued to extend and failed to show normal tube redundancy, typified by microtubule rearrangement and then depolymerisation. This provided an in vivo demonstration that the dynamics of a large microtubule aggregate are influenced by the developmental state of the cytoplasm.     

Specific interaction of cytochalasins with muscle and platelet actin filaments in vitro

The cytochalasins (CE, CD, CB and H₂CB) inhibit numerous cellular processes which require the interaction of actin with other structural and contractile proteins. In this report we describe the effects of the cytochalasins on the viscosity and morphology of muscle and platelet actin. The cytochalasins decreased the viscosity of F-actin solutions. The effect of H₂CB, CB and CD on F-actin viscosity was maximal at concentrations of 20–50μM and did not increase with time. In contrast, CE caused a progressive decrease in the viscosity of F-actin solutions which was dependent upon the concentration of CE and the duration of incubation of the CE-actin mixture. After two hours of incubation of drug-actin mixtures, the relative effectiveness of the cytochalasins in reducing the viscosity of F-actin was CE > CD > CB = H₂CB. The effects of CD and CE were paralleled by morphologic changes in negatively stained actin filaments. The effects of the cytochalasins on the viscosity and morphology of muscle and platelet actin were the same whether the drugs were added before or after the polymerization of the protein. These studies show that the interaction of the cytochalasins with actin is highly specific. Because the relative potencies of these drugs for affecting motile processes and the relative affinities of the drugs for binding sites within a variety of cells are CE > CD > CB = H₂CB, the effects of cytochalasins on actin described here may contribute to some of the biological effects of the drugs on motile processes.     

Internal motion of a tryptophan residue in Streptomyces subtilisin inhibitor: deuterium nuclear magnetic resonance in solution

Deuterium NMR spectroscopy was used to study internal motions of a deuterium-labeled single tryptophan (Trp) residue (per subunit) of Streptomyces subtilisin inhibitor (SSI) in solution. The free inhibitor with the five ring protons of the Trp replaced with deuterons showed a narrow resonance component (56 Hz) of about one-quarter of the total intensity, in addition to the broad resonance component (about 600 Hz) at 25 degrees C, showing that it exits in an equilibrium mixture of two conformers, in one of which the tryptophan side chain is highly mobile. In analogy to the two structures of SSI found in the crystal, these two conformers were attributed to the one in which the contact between the alpha-lobe and the beta-lobe of the subunit is tight and the other in which the same contact is loose. When SSI forms a complex with subtilisin BPN', the broad component becomes invisibly broad, but the narrow component increases with even further narrowing, suggesting that the binding to the enzyme favors the "loose" conformer over the "tight" conformer.     

Capacitation of mammalian spermatozoa in vivo, with a specific focus on events in the Fallopian tubes

This essay argues strongly that for those sperm cells involved in fertilisation, the process of capacitation represents an active and specific coordination within succeeding regions of the female tract and one whose completion is synchronised with the events of ovulation. Observations on the time-course of capacitation when spermatozoa are first exposed to the uterus and then progress to the Fallopian tubes indicate a synergistic influence of these adjoining portions of the female tract on the rate of capacitation. Three concepts on the control of capacitation are introduced to emphasise the importance of integration in vivo, namely that (1) completion of capacitation is a peri-ovulatory event, (2) suppression of completion of capacitation is an essential storage strategy during a long pre-ovulatory interval, and (3) the process of capacitation comes under the influence of local and systemic ovarian control mechanisms, especially the secretion of progesterone from Graafian follicles soon to ovulate. The last would act to coordinate the final maturation and meeting of male and female gametes. Despite the preceding points, the requirement for such integrated in vivo programming of sperm cell maturation can clearly be overridden in systems of culture. The most reasonable interpretation here would be that a microdrop of culture medium containing eggs, follicular cells and components of follicular fluid would to a considerable extent represent a post-ovulatory environment. Within such a preparation, there would be leaching of the sperm surface among the relatively vast and heterogeneous population of cells, and a proportion of spermatozoa could then respond to 'post-ovulatory signals', not least to molecular influences of the zona pellucida and vitelline products for completion of capacitation. Nonetheless, a physiologically meaningful interpretation of capacitation calls for a stepwise analysis of the dynamic interactions between sperm cell and female tract at successive stages between the uterus and ampullary-isthmic junction.     

Analysis of cell locomotion on ligand gradient substrates

Directional cell motility plays a key role in many biological processes like morphogenesis, inflammation, wound repair, angiogenesis, immune response, and tumor metastasis. Cells respond to the gradient in surface ligand density by directed locomotion towards the direction of higher ligand density. Theoretical models which address the physical basis underlying the regulatory effect of ligand gradient on cell motility are highly desirable. Predictive models not only contribute to a better understanding of biological processes, but they also provide a quantitative interconnection between cell motility and biophysical properties of the extracellular matrix (ECM) for rational design of biomaterials as scaffolds in tissue engineering. In this work, we consider a one‐dimensional (1D) continuum viscoelastic model to predict the cell velocity in response to linearly increasing density of surface ligands on a substrate. The cell is considered as a 1D linear viscoelastic object with position dependent elasticity due to the variation in actin network density. The cell–substrate interaction is characterized by a frictional force, controlled by the density of ligand–receptor pairs. The generation of contractile stresses is described in terms of kinetic equations for the reactions between actins, myosins, and guanine nucleotide regulatory proteins. The model predictions show a reasonable agreement with experimentally measured cell speeds, considering biologically relevant values for the model parameters. The model predicts a biphasic relationship between cell speed and slope of gradient as well as a maximum limiting speed after a finite migration time. For a given slope of ligand gradient, the onset of the limiting speed appears at longer times for substrates with lower ligand gradients. The model can be applied to the design of biomaterials as scaffolds for guided tissue regeneration as it predicts an optimum range for the slope of ligand gradient. Biotechnol. Bioeng. 2009;103: 424–429. © 2009 Wiley Periodicals, Inc.