Distribution of isoasparagine among different characean species

Thirteen species of Characeae were analyzed for their free amino acid contents. Large amounts of isoasparagine, accounting for 10 to 50% of the total free amino acids, were found in extracts fromChara (5 species including one unidentified),Nitellopsis (1 species), andLamprothamnium (1 species). In contrast, no isoasparagine was detected inNitella (5 species) andTolypella (1 species), except forN. flexilis in which as much as 40% of the free amino acids was isoasparagine. Other major amino acids found in the tested materials were Ala, Asp, Glu and Gln.     

Characean charasome-complex and plasmalemma vesicle development

Summary We report on an unusual phenomenon which occurs in some characean algae as a normal plasma membrane activity and also in association with charasome formation. The phenomenon of formation of coated invaginations of the plasma membrane was observed in twoChara and 6Nitella species. These invaginations are coated on their cytoplasmic surface, are 50–60 nm in diameter and rarely exceed 60 nm in length. They are abundant in the young cells ofChara andNitella and also occur in mature cells, but at a lower frequency.N. translucent is an exception in that coated invaginations were few in the young cells and absent in mature cells. Coated vesicles (50–60 nm diameter) were closely associated with these invaginations. Our observations suggest the vesicles may be derived from the invaginations by endocytosis.A close relationship was noted between the development of charasomes (plasmalemma modifications) and coated invaginations. Numerous coated invaginations are seen along the membranes of young charasomes; these invaginations appear to be associated with growth of the charasomes. Coated vesicles were not associated with the coated invaginations of the charasome membrane. The tubular network of cytoplasm and wall space seen in the mature charasome may be formed by fusion of coated invaginations of the developing charasomes, leaving cytoplasmic strands between the fused portions. Coated invaginations were not present along charasomes of the mature cells.     

Actin in living and fixed characean internodal cells: identification of a cortical array of fine actin strands and chloroplast actin rings

Summary We report on the novel features of the actin cytoskeleton and its development in characean internodal cells. Images obtained by confocal laser scanning microscopy after microinjection of living cells with fluorescent derivatives of F-actin-specific phallotoxins, and by modified immunofluorescence methods using fixed cells, were mutually confirmatory at all stages of internodal cell growth. The microinjection method allowed capture of 3-dimensional images of high quality even though photobleaching and apparent loss of the probes through degradation and uptake into the vacuole made it difficult to record phallotoxin-labelled actin over long periods of time. When injected at appropriate concentrations, phallotoxins affected neither the rate of cytoplasmic streaming nor the long-term viability of cells. Recently formed internodal cells have relatively disorganized actin bundles that become oriented in the subcortical cytoplasm approximately parallel to the newly established long axis and traverse the cell through transvacuolar strands. In older cells with central vacuoles not traversed by cytoplasmic strands, subcortical bundles are organized in parallel groups that associate closely with stationary chloroplasts, now in files. The parallel arrangement and continuity of actin bundles is maintained where they pass round nodal regions of the cell, even in the absence of chloroplast files. This study reports on two novel structural features of the characean internodal actin cytoskeleton: a distinct array of actin strands near the plasma membrane that is oriented transversely during cell growth and rings of actin around the chloroplasts bordering the neutral line, the zone that separates opposing flows of endoplasm.     

Variation in velocity of cytoplasmic streaming and gravity effect in characean internodal cells measured by laser-Doppler-velocimetry

Summary Velocities of cytoplasmic streaming were measured in internodal cells ofNitella flexilis L. andChara corallina Klein ex Willd. by laser-Doppler-velocimetry to investigate the possibility of non-statolith-based perception of gravity. This was recently proposed, based on a report of gravity-dependent polarity of cytoplasmic streaming. Our measurements revealed large spatial and temporal variation in streaming velocity within a cell, independent of the position of the cell with respect to the direction of gravity. In 58% of the horizontally positioned cells the velocities of acropetal and basipetal streaming, measured at opposite locations in the cell, differed significantly. In 45% of these, basipetal streaming was faster than acropetal streaming. In 60% of the vertically positioned cells however the difference was significant, downward streaming was faster in only 61% of these. When cell positions were changed from vertical to horizontal and vice versa the cells reacted variably. A significant difference between velocities in one direction, before and after the change, was observed in approx. 70% of the measurements, but the velocity was faster in the downward direction, as the second position, in only 70% of the significantly different. The ratio of basipetal to acropetal streaming velocities at opposite locations of a cell was quite variable within groups of cells with a particular orientation (horizontal, normal vertical, inverted vertical). On average, however, the ratio was close to 1.00 in the horizontal position and approx. 1.03 in the normal vertical position (basipetal streaming directed downwards), which indicates a small direct effect of gravity on streaming velocity. Individual cells, however, showed an increased, as well as a decreased, ratio when moved from the horizontal to the vertical position. No discernible effect of media (either Ca^{2 +}-buffered medium or 1.2% agar in distilled water) on the streaming velocities was observed. The above mentioned phenomenon of graviperception is not supported by our data.Abbreviations g gravitational acceleration (9.81 m/s²) - LDV laser-Doppler-velocimetry - VR velocity ratio Dedicated to Professor Peter Sitte on the occasion of his 65th birthday     

Wall appositions induced by ionophore A 23187, CaCl2, LaCl3, and nifedipine in characean cells

Summary The formation of wall appositions (plugs) by ionophore A 23187, CaCl₂, LaCl₃, and nifedipine was studied in mature internodal cells of characeaen algae. CaCl₂ at concentrations above 10^–2M induces thick fibrillar plugs without callose inNitella flexilis. InChara corallina andNitella flexilis ionophore A 23187 (1.25×10^–5 to 5×10^–5M) and LaCl₃ (7.5×10^–5 to 2.5×10^–4M) cause flat appositions which contain callose and have a more granular structure. Plug formation by ionophore A 23187, CaCl₂, and LaCl₃ is pH-dependent and occurs beneath the alkaline regions of the cell. Nifedipine (10^–4 to 10^–5M) induces plugs inNitella flexilis after previous injury. These callose-containing wall appositions consist of a heterogeneous granular core which is covered by a fibrillar layer. The results of this work are compared with previous studies on wound wall formation and chlortetracycline (CTC)-induced plug formation which reveal that abundant coated vesicles occur only when a thick fibrillar wall layer is formed. Neither LaCl₃ nor nifedipine inhibit the formation of CaCl₂- or CTC-plugs. The unusual effects of these substances, which normally act as Ca²⁺ antagonists and therefore should prevent and not induce plug formation, are discussed. It is suggested that La³⁺ mimicks the effects of calcium and that nifedipine binding to the Ca²⁺ channels is altered in the alkaline regions of characean internodes and allows an influx of Ca²⁺.Abbreviations AFW artificial fresh water - CTC chlortetracycline - DCMU dichlorphenyldimethylurea - DMSO dimethylsulfoxide - EGTA ethyleneglycoltetraacetic acid - MES 2-(N-morpholino) ethanesulfonic acid - HEPES N-2-hydroxyethylpiperazine-N-2-ethanesulfonic acid - TAPS N-tris[hydroxymethyl]methyl-3-aminopropanesulfonic acid     

Phycobionts of some Japanese species ofPertusaria (Pertusariaceae)

12 species ofPertusaria DC. were collected from various areas in Japan. Their phycobionts were isolated and studied taxonomically. The phycobionts of these species were indentified asTrebouxia potteri Ahmadjian exGärtner.Dedicated to Prof. DrLothar Geitler on the occasion of the 90th anniversary of his birthday.     

Measurement of motive force for cytoplasmic streaming in characean internodes

Summary With an attempt to measure the motive force responsible for cytoplasmic streaming in characean internodal cells, the difference between densities of cytoplasm and vacuolar sap was heightened by about 10 times (density of vacuolar sap was made larger than that of cytoplasm) by replacing the natural vacuolar sap ofChara corallina with an artificial one of higher density. Endoplasmic flow contiguous to the peripheral actin cables (peripheral flow of endoplasm) in the centrifugal direction was not influenced at all by the application of centrifugal acceleration up to 1400 g. We thus concluded that the motive force for the peripheral flow should be much larger than 12dyn/cm², a figure more than 10 times larger than that for bulk endop lasmic flow so far reported.Dedicated to Emeritus Professor Noburo Kamiya on the occasion of his 80th birthday     

Karyotaxonomic study of some species ofPeucedanum (Umbelliferae)

A study of morphometric karyotype characters was carried out in 14 species ofPeucedanum s. lat. (Umbelliferae — Apioideae) and in one species of the related genusLomatium. The differences of the species in their karyotype characters are correlated only to a limited degree with their similarities of dissimilarities in morphological (i. a. carpological) characters, and thus with their taxonomic position. The evolution of these two sets of characters seems to have proceeded not synchronously or even in different directions in the group of platycarpousUmbelliferae united in the genusPeucedanum. Therefore, it is unlikely that morphometric chromosome characters revealed by monochrome staining can be used appropriately in the taxonomic revision of the polymorphous genusPeucedanum.     

Inversion of extracellular current and axial voltage profile inChara andNitella

Summary Reducing the pH of the bathing solution from 8.2 to pH 6 can induce an inversion of the extracellular current pattern that develops at the surface ofChara corallina internodal cells. A similar result can be obtained on some cells by changing the medium to a pH value of 10. In noninvertingChara cells the currents were strongly reduced when the pH value of the medium was changed between 3 and 11. Simultaneous measurements of theChara transmembrane potential in the acid and alkaline regions revealed that a light-induced electrical potential gradient of approximately 24 mV was present in the axial (or longitudinal) direction. Correlated to the external current pattern inversion was an inversion of this internal longitudinal voltage gradient. Reillumination ofNitella cells, after a period of darkness, often resulted in a complete inversion of the extracellular current pattern. These results are discussed in terms of spatial and temporal control of membrane transport processes, and in particular the control of current loops that pass through these cells.     

Distribution of alkanes and fatty acids in theCondalia montana (Rhamnaceae) species complex

The alkane and fatty acid composition of the lipid extracts from leaves of 14 populations ofCondalia: C. microphylla f.xanthocarpa, C. microphylla f.erythrocarpa, C. microphylla f.melanocarpa, C. montana, andC. buxifolia, were determined by gas chromatography. Seventeen alkanes and 14 fatty acids were investigated and a predominance of nonacosane, hentriacontane, palmitic and linoleic acids in all species was found.     

Isozyme polymorphism in some yellow- and blue-floweredVigna species complexes (Fabaceae, Phaseoleae)

An electrophoretic comparison of variation at 28 isozyme loci was performed for 58Vigna accessions belonging to theV. luteola, V. ambacensis, andV. racemosa groups of species. In all three groups, strong divergence is noted between results and actual nomenclature.     

B-Chromosomes inAllium flavum (Liliaceae) and some related species

B-Chromosomes ofAllium flavum, A. stamineum andA. carinatum were C-banded. InA. flavum different types of B's were found, one of them possessing a nucleolus organizer. In the B's ofA. flavum andA. stamineum the banding patterns resemble those found in the standard chromosomes. The B inA. carinatum is only terminally banded. InA. flavum chiasma localization in the B's appears to be dependent on C-band location, just as is the case in the A-chromosomes. An increase in B's was found to cause a small (but significant) increase in chiasma frequency in the PMC's. This may result from an alteration in the nucleotype by the B's.     

The relationship of the charasome to chloride uptake in Chara corallina: physiological and histochemical investigations

A possible role of the charasome in terms of chloride transport into Chara corallina Klein ex. Willd., em. R.D.W. is examined. The branches of Chara contain the most charasome material and are shown to be very effective in acquiring Cl^- to support continued shoot growth. The early maturation of the branches, the rather large Cl^- fluxes into these cells, and their ability of translocate Cl^- to growing cells of the shoot indicate a special role of these branches in Cl^- accumulation. The structure of the charasome, with its extensive periplasmic space, appears especially suited as a site for H⁺–Cl^- cotransport (influx). We show, by histochemical assay, that the charasomes of mature cells contain ATPase activity; such activity is absent in growing charasomes of very young cells. ATPase activity is also associated with the plasmodesmata of C. corallina. Charasome ATPase activity and Cl^- uptake are both inhibited by p-chloromercuribenzenesulfonic acid (1 mM) or diethylstibestrol (40 M; 45 min). The anion transport inhibitor, 4,4-diisothiocyano-2,2-disulfonic acid stilbene (1 mM) had no effect on Cl^- transport and inhibited ATPase activity only when applied after chemical fixation of the cells. Results of an attempt to demonstrate the presence of Cl^- within the cytoplasmic tubules of the charasome, using a silver precipitation technique, proved difficult to interpret because of a reaction between the silver and a cellular substance produced in the light.Abbreviations CPW Chara pond water - DES diethylstilbestrol - DIDS 4,4-diisothiocyano-2,2-disulfonic acid stilbene - Mops 3-(N-morpholino)propanesulfonic acid - PCMBS p-chloromercuribenzenesulfonic acid     

Mechanism of plasmodesmata formation in characean algae in relation to evolution of intercellular communication in higher plants

It is generally accepted that higher plants evolved from ancestral forms of the modern charophytes. For this reason, we chose the characean alga, Chara corallina Klein ex Willd., em. R.D.W. (C. australis R. Br.), to determine whether this transition species produces plasmodesmata in a manner analogous to higher plants. As with higher plants and unlike most green algae, Chara utilizes a phragmoplast for cell division; however, in contrast with the situation in both lower and higher vascular plants, the developing cell plate and newly formed cell wall were found to be completely free of plasmodesmata. Only when the daughter cells had separated completely were plasmodesmata formed across the division wall. Presumably, highly localized activity of wall-degrading (or loosening) enzymes inserted into the plasma membrane play a central role in this process. In general appearance characean plasmodesmata are similar to those of higher plants with the notable exception that they lack an appressed endoplasmic reticulum. Further secondary modifications in plasmodesmal structure were found to occur as a function of cell development, giving rise to highly branched plasmodesmata in mature cell walls. These findings are discussed in terms of the evolution of the mechanism for plasmodesmata formation in algae and higher plants.This work was supported in part by National Foundation grant No. DCB-9016756 (W.J.L.). We thank the Electron Microscopy Center of Washington State University and the Zoology Department, University of California, Davis, for the use of their microscopy facilities.     

Morphological analysis of some cryptic species in the Acanthocyclops vernalis species complex from North America

Patterns of morphological variation and reproductive isolation were examined for several North American populations of copepods in the Acanthocyclops vernalis Fischer A., 1853 (Copepoda, Cyclopinae) species complex. The copepods were collected from six sites in Wisconsin, U.S.A. Morphological analysis of 120 adult females revealed that a character used previously to distinguish species in this group was unreliable because of phenotypic plasticity. Most of the morphological variance was due to environment (Laboratory vs. field) and to field site. Relatively little of the variation was due to measurement error or asymmetry. Multivariate ordination analysis produced poorly-defined clusters of individuals, suggesting that different biological species are difficult or impossible to distinguish using a set of easily-measurable morphological characters. In our study, morphological similarity was independent of geographic distance among sites, between 0.05 and 300 km. Isofemale lines within sites showed little or no reproductive isolation, but nearly complete isolation among sites. Reproductive isolation was also independent of morphology. These results suggest that the Acanthocyclops population at each site could be considered a distinct cryptic biological species. These copepods expressed morphological stasis – persistence of morphological uniformity despite reproductive isolation. Because of the effect of site and environment on morphology, we recommend using much larger collections (many sites), common garden experiments, and a multi-disciplinary approach (morphological, reproductive, chromosomal, and molecular) as the basis for future taxonomic research on putative copepod species.     

Oscillations of electric spatial patterns emerging from the homogeneous state in characean cells

Electric spatial patterns of bands formed along the cell wall of the characean internode were studied using a multi-electrode measuring system. The electric potential near the surface of the cell was measured by arranging about 25 electrodes along the cell at approximately 1.6 mm intervals. Since the time required for one scan over the cell length is only 1 s, the temporal change in the spatial pattern of surface electric potential can be readily observed. Oscillations were sometimes found as the electric pattern started to appear after the cell was illuminated. Fourier analysis shows that a single spatial mode arises gradually and then becomes stabilized in an oscillatory manner. A simple electric circuit model comprising three variables, i.e., a membrane potential, an electric current across the membrane and an electromotive force, can simulate well the oscillatory rise of bands. These results imply that the electric spatial pattern observed in characean internodes is a self-organized structure emerging far from equilibrium, known as a dissipative structure. Biophysical mechanisms of band formation are discussed.     

Differences in flavonoid constituents among the species ofFagonia sinaica complex (Zygophyllaceae)

Eight flavonol glycosides were detected in the three species of theFagonia sinaica complex. They were fully characterized as the 3-glucosides of kaempferol, quercetin and isorhamnetin, 3-rutinoside of quercetin and 3,7-diglucoside of quercetin and isorhamnetin. Two additional glycosides were partially characterized as a kaempferol 3,7-diglycoside and quercetin 3-diglycoside.     

A comparison of the pollen wall ultrastructure of aureoid and non-aureoidSenecio species (Asteraceae) in North America

The structure of the pollen wall as revealed by transmission electron microscopy is presented for 29 speciesSenecio including 19 aureoid and ten non-aureoid species. The previous report of a helianthoid wall structure (exine with internal foramina) is confirmed for all the aureoid species examined while all of the non-aureoid taxa showed a senecioid pattern. The data provide support for the treatment of the aureoid complex as a distinct taxon.     

Hybridisation in the Littorina saxatilis species complex (Prosobranchia : Mollusca)

This paper details attempts to breed members of the L. saxatilis species complex in the laboratory and to construct interspecific hybrids between them. Success in reciprocal crosses between animals from the type localities of the species of Littorina saxatilis (Olivi) and Littorina rudis (Maton) indicates that these two taxa are synonymous. Six of the twelve possible reciprocal hybridisation crosses between the four species (L. saxatilis, L. arcana Hannaford Ellis, L. nigrolineata Gray and L. neglecta Bean) have been attempted, with only one proving successful, that between male L. saxatilis and female L. arcana. This hybrid cross produces viable offspring, although at a lower frequency than either of the parental crosses in the laboratory, while the reciprocal cross has, as yet, proved unsuccessful. Limited work on the F1 hybrids shows them to be inter-fertile and also capable of backcrossing with male L. saxatilis, but not with female L. saxatilis or L. arcana. Details are also given of attempts to find natural hybrids in sympatric populations of the species of the complex, using gel electrophoresis.