Embryology ofCymbidium sinense:the Microtubule Organization of Early Embryos

InCymbidium sinense, the pattern of embryo development is unusualin that oblique cell divisions result in the formation of severalsuspensor cells prior to the development of the embryo proper.Characteristic changes in microtubular distribution can be foundwithin the zygote and the proembryo during their development.After fertilization, the ellipsoid-shaped zygote has randomlydistributed microtubules within its cytoplasm. As the zygotetakes on a more rounded appearance, microtubules organize intoa dense meshwork. Furthermore, microtubule bundles appear atthe chalazal region of the cell prior to the first mitotic divisionof the zygote. At the preprophase stage of mitosis, a preprophaseband of microtubules appears in the cytoplasm of the zygote.The zygote divides obliquely and unequally and gives rise toan apical cell and a slightly larger basal cell. Many randomly-alignedmicrotubules can be found in the cortex of the basal cell. Theincrease in the abundance of microtubules coincides with theisotropic expansion of the basal cell. The early division ofthe basal cell and subsequent division of the apical cell resultsin the formation of a four-celled embryo, of which three cellsnear the micropylar pole develop as suspensor cells. In thesuspensor cells, the microtubules tend to orient in the samedirection as the long axis of the cell. In addition, prominentmicrotubules can also be found near the adjoining cell wallsof the four-celled embryo. The terminal cell is highly cytoplasmicwith abundant microtubules within the cell. Subsequent divisionsof the terminal cell give rise to additional suspensor cellsand the embryo proper. In the mature embryo, five suspensorcells are usually present; one eventually grows through themicropyle of the inner integument and four grow towards thechalazal pole. The cortical microtubules of suspensor cellsredistribute from a longitudinal to a transverse direction asthey grow towards their respective poles.Copyright 1998 Annalsof Botany Company Embryogenesis, endosperm, microtubules, preprophase band, suspensor cells,Cymbidium sinense(Andr.) Willd.     

Aplanogamous sexual reproduction in Carteria eugametos (Volvocales,Chlorophyta)

Morphological details of sexual reproduction in Carteria eugametos (Volvocales, Chlorophyta) were studied under controlled laboratory conditions. Protoplasts of the two pairing, flagellate cells were released from cell walls to become isogametes. Such gametes were nonmotile and soon fused to form a completely immobile zygote. The zygote then secreted a cell wall to enter the dormant period. After dark treatment, the zygote produced four, eight or 16 quadriflagellate germ cells, and a transparent vesicle enclosing all the germ cells was released from the zygote wall. This type of zygote germination and aplanogamy in C. eugametos is unique and may be related to its peculiar phylogenetic position within the Volvocales (Chlamydomonadales).     

Suppression of nucleic acid synthesis in chromatin of Spirogyra during conjugation process

Marked changes in RNA- and DNA-synthesizing activities of thechromatin fraction were found to be connected with the conjugationprocess of Spirogyra sp. RNA-synthesizing activity of the chromatin fraction was muchlower in conjugating and zygote cells than in vegetatively growingcells. No significant differences were observed in the templateactivities of purified DNA prepared from vegetatively growing,conjugating, and zygote cells. The activities of DNA purifiedfrom vegetatively growing and zygote cells and calf thymus werestrongly inhibited at different levels by binding with histonesprepared from vegetatively growing and zygote cells. DNA-synthesizingactivity of the chromatin fraction of conjugating or zygotecells was much lower than that of vegetatively growing cells. (Received December 24, 1971; )     

Reproduction in Halopteris (Sphacelariales)

Oogamy is recorded in five southern species of Halopteris. Ineach the gametangia and asexual sporangia occur in similar sori,although situated on separate plants. The plurilocular antheridiaand the unilocular oogonia occur within one and the aame sorus.In both the dehiscence is apical. In H. congesta fertilization has been observed in cultures.The polarity of the zygote is established by the direction ofthe first-formed wall. Early divisions convert it into a nodulecomposed of small cells. First rhizoids appear and then cylindricalshoots arise from the opposite pole. Sporelings of five NewZealand species of Halopteris have been raised in culture. Theconditions in the culture greatly affected the character ofthe sporelings, the simplest of which at the end of 2 months'growth were only uniseriate filaments. The best developed sporelingshad a compact basal disc with up to four upright shoots showingthe beinnings of branching.     

Ultrastructural Changes of the Egg Apparatus Associated with Fertilization and Proembryo Development of Soybean, Glycine max (Fabaceae)

As part of a study involving pod retention in soybean, Glycinemax (L.) Merr., we investigated changes occurring in the eggapparatus of non-abscised flowers from the time immediatelypreceding fertilization through early embryogeny. Prior to theentry of the pollen tube into the embryo sac, one of the synergidsbegins to degenerate as evidenced by increased electron densityand a loss of volume. This cell serves as the site of entryfor the pollen tube. The cytoplasm of the second, or persistentsynergid, remains unaltered until after fertilization. Bothsynergids contain, in addition to a filiform apparatus, a singleunidentified inclusion of flocculent material located in thechalazal portion of each cell. The zygote can be distinguishedfrom the egg by its consistently narrow wall; and it dividesto form a proembryo, a mass of cells not yet differentiatedinto embryo proper and suspensor. The basal cells of the proembryoare more vacuolate than the apical ones, characteristic of thebasal vacuolation of both egg and zygote. Cells of the proembryoare connected to one another via plasmodesmata, and with theexception of the basal-most cell, are isolated symplasticallyfrom the surrounding endosperm. Wall ingrowths frequently occurin certain cells of the proembryo, notably those cells in contactwith the degenerate synergid and embryo sac wall. At a laterstage of ontogeny, by which time the globular embryo properhas become distinct from the suspensor, the wall ingrowths areconcentrated in the suspensor. Glycine max, soybean, embryogeny, synergids     

Complementation and Preliminary Linkage Analysis of Zygote Maturation Mutants of the Homothallic Alga, CHLAMYDOMONAS MONOICA

Sexual reproduction in Chlamydomonas monoica is homothallic: pair formation and cell fusion occur in clonal culture and give rise to a heavily walled diploid zygospore. During maturation of the young zygote, a distinctive "primary zygote wall" is released before the development of the highly reticulate zygospore wall. Using ethyl methanesulfonate and ultraviolet irradiation as mutagens, we have isolated 19 maturation-defective (zym ) mutant strains which upon self-mating produce inviable zygotes. These zygotes fail to release a primary zygote wall, fail to develop the normal zygospore wall, and eventually undergo spontaneous lysis. In nearly all cases, the mutations appear to be expressed only in the diploid zygote; pleiotropic effects on vegetative cell growth or morphology are not evident.—Complementation testing performed on 17 of these mutants indicates that all are recessive and that they define seven distinct complementation groups. Preliminary tetrad analysis of two-factor and multifactor zym crosses provides no evidence for physical clustering of the maturation genes, and instead suggests that they are widely distributed throughout the nuclear genome.     

SEXUAL REPRODUCTION OF PERIDINIUM WILLEI (DINOPHYCEAE)1

Sexual reproduction was induced in the dinoflagellate Peridinium willei Huitfeld-Kass when exponentially growing cells were inoculated into nitrogen deficient medium. Small, naked vegetative cells produced by division of thecate cells acted as gametes. The zygote remained motile 13–14 days, during which time it enlarged and the theca formed became warty. Fourteen to 15 days following plasmogamy the zygote was nonmotile with the protoplast contracted. A large red oil droplet appeared and the wall thickened, becoming chitinized. Hypnozygotes with 4 nuclei were observed 7–8 wk following formation. Meiosis was inferred. The hypnozygote germinated, within 8 wk producing one post-zygotic cell retaining the red oil droplet. This cell divided within 24 h into 2 daughter cells each with a prominent red oil droplet. These daughter cells divided after 2 to 3 days into ordinary vegetative cells. Attempts to induce sexual reproduction by inoculation of cells into media deficient in a number of basic elements were unsuccessful.     

SEXUAL REPRODUCTION OF PERIDINUM CINCTUM F. OVOPLANUM (DINOPHYCEAE)1,2

Sexual reproduction is induced in the dinoflagelate Peridinium cinctum f. ovoplanum Lindemann when exponentially growing cells are inoculated into nitrogen deficient medium. Small, naked vegetative cells, produced by division of the thecate cells, then act as gametes. The zygote remains motile for 12–13 days during which time it enlarges and the theca which it forms becomes warty. Thirteen to 14 day s following plasmogamy the zygote is nonmotile, the protoplast contracts, a large red oil droplet appears, the wall thickens and becomes chitinized. This hypnozygote germinates within 7–8 weeks at 20 c producing 1 post-zygotic cell retaining the large red oil droplet. The presence of 4 nuclei in these post-zygotic cells may be demonstrated by staining them with acetocarmine. Two of these nuclei are smaller than the other two and probably abort. One may infer that meiosis occurs immediately prior to or at the germinartion of the hypnozygote. This post-zygotic cell divides within 24 h into 2 daughter cells each with a promment red oil droplet. These daughter cells divide after 2–3 days into ordinary vegetative cells. Attempts to induce sexual reproduction by changes in temperature or light and by inoculation of cells into media deficient in a number of basic elements were unsuccessful.     

Light dependency of the mating process in Closterium acerosum

Sexual cell division and activation of gametangial cells forconjugation in Closterium acerosum were induced by light. L₂₀₀cells conjugated at maximum level under the following conditions;(i) a light intensity higher than 1,000 lux in a 16-hr lightand 8-hr dark regime and (ii) an illumination time longer than12 hr at 3,000 lux. L₂₀₀ cells also conjugated under continuousillumination at 3,000 lux. The action spectrum for the activation of gametangial cellshad peaks around 450, 611 and 665 nm. 3-(4'-Chlorophenyl)-l,l-dimethylurea (CMU) inhibited the accumulationof carbohydrates and sexual cell division at 10^–5 M andthe activation of gametangial cells for conjugation at 10^–4M. (Received August 15, 1977; )     

Effect of concanavalin A on the mating reaction in Saccharomyces cerevisiae

The effect of concanavalin A (Con A) on the process of massmating of Saccharomyces cereoisiae was studied. Sexual agglutinationwas repressed by Con A at concentrations of 400 µg/mland 500 µg/ml, but zygote formation was little affectedat these concentrations. The action of Con A was antagonizedspecifically by -methyl-D-mannoside. We compared the mode ofinhibitory action on sexual agglutination of Con A with thatof agglutination substances, cell surface glycoproteins responsiblefor sexual agglutination. The agglutination substances inhibitedthe formation of small cell aggregates consisting of less thanfifty cells thought to be necessary for the formation of zygotes.Con A, on the contrary did not inhibit the formation of smallaggregates, but inhibited the formation of large cell aggregatesconsisting of more than hundred cells by interfering with thefusion of small cell aggregates. Univalent Con A inhibited isoagglutination caused by high concentrationsof native Con A. Specific binding of Con A to the cell surfacewas observed by using fluorescent Con A. A procedure to prepareunivalent Con A using Enzygel, a trypsin-Sepharose conjugantis described. ¹ On leave from Osaka City University. ² Present address: Department of Physiology, Japan Women's University,Bunkyo-ku, Tokyo 112, Japan. (Received March 28, 1978; )     

Sperm morphology and storage in the female reproductive tract of the fat-tailed dunnart,Sminthopsis crassicaudata (Marsupialia: Dasyuridae)

A study of spermatozoa in the isthmus of the oviduct and of the surrounding epithelial cells in the dasyurid marsupial, Sminthopsis crassicaudata, was carried out. At least 10% of the ejaculated spermatozoa probably populate the isthmus region, where many come to reside in crypts until around the time of ovulation. Ultrastructural observations of spermatozoa in this region indicated that they had intact acrosomes and were identical in their morphology with those in the cauda epididymidis. After ovulation spermatozoa rapidly disappeared, some of which may be phagocytosed by the cells lining the crypts. These epithelial cells were also found to have many large, electron-dense granules at the time of sperm storage, but the contents did not appear to be released until the zygotes passed along the tract. The secretory activity of these cells may thus relate more to the production of the shell membrane that comes to surround the zygote than to the cells performing a nutritive or protective function for the spermatozoa during their period of storage within the female reproductive tract.     

SEXUAL REPRODUCTION OF PERIDINIUM GATUNENSE (DINOPHYCEAE)1

Sexual reproduction was induced in the dinoflagellate Peridinium gatunense Nygaard when exponentially growing cells were inoculated into nitrogen deficient medium. Small thecate cells produced by division of vegetative cells then acted as gametes. Thecae of fusing gametes broke in the girdle region and were lost. Zygotes thus formed remained motile 3–5 days during which time they enlarged slightly with the newly formed theca becoming warty. Three to 5 days following plasmogamy the zygote became nonmotile, the protoplast contracted, and the cell wall thickened. Hypnozygotes with 4 nuclei were observed ca. 10–12 h following formation. Meiosis was inferred. Hypnozygotes germinated within 12 h of formation producing 2 vegetative cells which divided within a 24 h period. Attempts to induce sexual reproduction by inoculation of cells into media deficient in a number of basic elements other than N were unsuccessful.     

Sexual selection for large male size in a polyandrous butterfly: the effect of body size on male versus female reproductive success in Pieris napi

Sexual selection theory predicts that the larger sex shouldbe that for which fitness increases at the faster rate withsize. In butterflies, as in most invertebrates, females areusually the larger sex, but previous comparative analysis hasshown that relative male size increases with female polyandryamong butterflies. In agreement with this pattern, males arelarger than females in the strongly polyandrous green-veinedwhite butterfly, Pieris napi L., and in this article we assessthe size dependence of reproductive success in both sexes. Inan experiment where virgin males and females were released inthe field, we found no strong association between size and malemating success. However, laboratory experiments showed thatthere was a strong correlation between size and the ejaculatethat the male delivered to the female at mating and that largeejaculates delayed female remating for a longer time comparedto small ejaculates. Moreover, female P. napi utilize male-derivednutrients received at mating to increase their fecundity. Hence,large males sire more offspring both by way of donating morenutrients to female egg production and by way of delaying femaleremating (given that the last male to mate with the female willfather most of the offspring). Laboratory experiments showedthat the association between size and fecundity was low, ornonexistent, among P. napi females allowed to mate only once.However, weak size dependence was found for polyandrous females.We hypothesize that size dependence of female fecundity maybe especially weak among polyandrous butterflies because a fundamentalsource of variation in fecundity relates to their ability tofind nutrient giving males, an ability which may be unrelatedto female size. According to this hypothesis there is a causalassociation between weak size dependence of female fecundityand polyandry, and a strong size dependence of male reproductivesuccess that may underlie the comparative pattern of positivecorrelation between relative male size and polyandry.     

Observations on the Cytology of Halidrys siliquosa (L.) Lyngb.

A study of the cytology of Halidrys siliquosa suggests thatit has a haploid thallus with reduction division immediatelyfollowing fertilization. There appear to be eight chromosomesin vegetative cells and developing antheridial and oogonialcells. Typical ‘bouquet’ and diakinesis stages suggestinga reduction division have been seen on germination of the zygote.     

Detection and Evaluation of an Inducer of a Diffusible Mating Pheromone of the Heterothallic Closterium peracerosum-strigosum-littorale Complex

When mating-type plus cells of the Closterium peracerosum-strigosum-littoralecomplex were incubated in nitrogen-deficient medium obtainedfrom a 24-h-old mixed culture of mating-type plus and mating-typeminus cells, protoplast-release-inducing activity specific formating-type minus cells was detected in the medium. When mating-typeplus cells were incubated in the medium from a culture of exclusivelymating-type minus cells, protoplast-release-inducing activitywas also detected. These results suggested the existence ofa substance, released from mating-type minus cells, that hasthe ability to make mating-type plus cells release protoplast-release-inducingprotein (PR-IP). We designated it PR-IP Inducer. The PR-IP Inducerwas constitutively released from mt^– cells in the light.The PR-IP Inducer was heat-labile and had a relative molecularweight of 10,000 on gel filtration. We suggest that the PR-IPInducer is also a pheromonal substance that plays a role inthe initial events in the sexual communication of this Closteriumcomplex. (Received April 26, 1993; Accepted July 15, 1993)     

Studies on the Hill reaction of membrane fragments of blue-green algae III. Fluorescence characteristics of membrane fragments of Anabaena variabilis and Anabaena cylindrica

Fluorescence spectra of the pigment system at –196°Cin membrane fragments of Anabaena variabilis and A. cylindricawere investigated. The fluorescence spectra of membrane fragments having four emissionbands at 645–655, 685, 695 and 725 nm were basically similarto those reported for intact cells of blue-green algae, thoughthe emission from phycocyanin (645–655 nm) was far strongerwith membrane fragments than with intact algal cells. Incubation of membrane fragments of A. variabilis in a dilutebuffer (10^–2M, pH 7.5) caused an increase in the 645 nmfluorescence and slight decreases in the 685 and 695 nm fluorescences,but had no influence on the 725 nm fluorescence. The decreasein the 685 and 695 nm fluorescences of A. cylindrica was moremarked and had the same kinetics as the inactivation of photosystemII reaction measured by DPIP-photoreduction. When membrane fragments of A. cylindrica were incubated in thebuffer solution at room temperature or in the presence of MgCl₂(10^–3M) at 0°C; phycobilin aggregates, which emittedthe 655 and 685 nm fluorescence, were solubilized. This solubilizationwas not observed with membrane fragments of A. variabilis. (Received August 31, 1972; )     

Changes in the Chromosome Complements of Cells of Vicia faba Roots following Irradiation

1. Primary roots of Vicia faba were irradiate after one day'sgermination. The X-ray dose was 620 r 2. Cells with changed chromosome complements appeared in lateralroots. It is inferred that these cells are descended from cellsdamaged by irradiation; chromosome breakage was seen in thefirst mitoses in the irradiated roots but was not scored. 3. Six types of chromosomal rearrangements were found. One typehad been reported previously in primary roots of V. faba afterX-raying. 4. The new complements appear to be stable because they occurseveral cells of a lateral. Affected laterals could containup to five new complements. The primordium of such a lateralmust have consisted of five changed and at least one normalcell. It had previously been suggested that a root of Viciahad three or four initial cells. 5. The survival of the cells with changed complements is notdue solely to successful competition with, not to support by,normal cells, otherwise other types of changes should also havebeen found. 6. It is suggest that the changes found were perpetuated becausethey were induced in initial cells of the primary root, andthat cells in which they were present beca,e part of the primordialand continued as initial cells in apical meristems of lateralroots. 7. Apical cells appear to have a lower sensitivity than othermeristematic cells, so providing, after treatment, a reservoirof normal cells from which regeneration can occur.     

The role of temperature in the seed germination of two species of the Solanum nigrum complex

The germination behaviour of S. nigrum L. and S. physalifoliumRusby var. nitidibacatum (Bitter) Edmonds is compared, basedon temperature requirements during imbibition. Three seed lotsof S. nigrum had base temperatures (T_b) between 7.5C and 10C,showing a lower T_b when the period of freezing days, duringwhich each population was collected, was reduced. S. physalifoliumhas a higher value for T_b at constant temperatures (21C) thatcan be interpreted as a dormancy constraint. This constraintis released by alternating temperatures at amplitudes exceeding5C and with the high temperature above 21C by apparently reducingT_b to 12.5C. This implies that for S. physaiifolium temperature has a dualeffect on germination. It is the driving force for changes indormancy, but germination also depends on the temperature. Therole of temperature for S. nigrum is simpler: each populationcollected showed differences in the thermal time required forgermination that could be related to the temperature regimenof the original environment. Key words: Dormancy, S. nigrum, S. physalifolium, thermal time     

Sexual process in eight-spored ascus-forming yeast, Schizosaccharomyces japonicus I. Culture medium for the synchronous sexual process

The sexual process of Schizosaccharomyces japonicus consistsof sexual flocculation, zygote formation, eight-spored ascusformation and liberation of spores from the ascus. The culturemedium in which this sexual process took place synchronouslywas prepared. For the completion of the sexual process, glucosewas essential and inorganic salts and vitamins were also required.Elimination of the nitrogen source stimulated the rate of sporeformation. The temporal relationship among the sexual eventswas also elucidated: sexual flocculation, zygote formation,ascus formation and spore liberation occurred at 4, 7, 13 and20 hr, respectively, after transfer to the medium for the synchronoussexual process. (Received May 11, 1978; )     

Sexual process in eight-spored ascus-forming yeast, Schizosaccharomyces japonicus II. Dependency of the sexual process on the carbon source

The carbon-source dependency of the sexual process in Schizosaccharomycesjaponicus was studied. Schiz. japonicus grew well in vegetativemedia containing glucose, sucrose, fructose or raffinose, anddid poorly in one containing mannose. On the other hand, itssexual process proceeded well in sporulation media containingglucose, sucrose or mannose, and was markedly delayed in thosecontaining fructose or raffinose. Neither vegetative growthnor sexual process occurred when non-fermentable carbon sources,such as glycerol, were used. The amount of glucose in the sporulationmedium sufficient for completion of the sexual process varieddepending on the cell-population density. Glucose was requiredfor both zygote and ascus formation but not for spore liberation.Cells were committed to sporulation shortly after the stageof zygote formation. (Received August 3, 1978; )