On Dark-Germination and Antheridium Formation in Anemia phyllitidis

Schraudolf's finding is confirmed that gibberellic acid induces antheridia in Anemia phyllitidis and Lygodium japonicum. The activity spectra of gibberellic acid and the native antheridiogen of Anemia phyllitidis are similar: Both induce antheridia in the tested species of the family Schizaeaceae but are inactive towards the tested representatives of other fern families. However, the native antheridiogens of two schizaeaceous species are more species-specific in their action than is gibberellic acid. Anemia medium cancels the light requirement for spore germination as is the case with gibberellic acid. Chromatographic studies indicate that the dark-germination-inducing factor is identical with, or very similar to, the earlier demonstrated antheridiogen. The specificities of the active factors towards dark-germination in A. phyllitidis and L. japonicum are similar to those encountered with antheridium formation. Anemia medium induces dark-germination and induces antheridia on the dark-grown protonemata to a concentration ca. 30 times lower than it induces antheridia on light-grown prothalli. The studies indicate that this differential activity results from a differential competence of the prothalli to react to the active substance in light and darkness.     

The Spontaneous Formation of Antheridia in Onoclea is not Blocked by Light

Onoclea sensibilis gametophytes were grown from spores on ashedsoil and agar to determine if the spontaneous formation of antheridiacan be blocked by light. Under most conditions, dark-grown gametophytesformed antheridia later than or at the same time as gametophytesgrown in the light. Under no circumstances was there a rapidonset of maleness in the dark. These results contradict thehypothesis that, in Onoclea, antheridiogen is required to inducemaleness because light inhibits the formation of antheridia.In the light, antheridia formed on heart-shaped thalli. In darkness,antheridia formed on filamentous gametophytes. The timing ofonset of maleness was affected by temperature and the presenceof sucrose. The effect of sucrose on the comparison betweenlight and dark treatments depended on both substrate and temperature Onoclea sensibilis, L., sensitive fern, fern gametophytes, sexuality, light-induced block     

On the physiology and chemistry of fern antheridiogens

Döpp has demonstrated an antheridium-inducing hormone (antheridiogen) in P.aquilinum. This antheridiogen (abbr. Apt.) is active in many, if not all, species of the family Polypodiaceae. Among responsive species, the minimally effective concentration varies widely. Apt was assayed againstOnoclea sensibilis because this species fails to form antheridia spontaneously under the prevailing conditions of culture and because none of the many species tested responds to a lower concentration A_Pt is inactive toward the investigated species of the fern families Osmundaceae, Cyatheaceae and Schizaeaceae. The two schizaeaceous speciesAnema phyllitidis andLygodium japonicum also elaborate antheridiogens (abbr. A_An and A_Ly). Both these antheridiogens are inactive in 0.sensibilis, the species used to assay for A_Pt. A_An, A_Ly, A_Pt and A_On (the antheridiogen of O.sensibilis) are distinct entities based both on the criteria of cross-testing and of Chromatographic separation. Cross-testing led to the conclusion that the antheridiogen ofCeratopteris thalictroides differ from A_Pt and A_An. Gibberellins have antheridiogenic properties in schizaeaceous species but, like A_An and A_Ly, they fail to hasten antheridium formation in the species used to assay for A_Pt. The native antheridiogens of schizaeaceous species are more species-selective in their action than is GA₃. A_An has recently been isolated. Its structure is similar to, if not identical with, that of gibberellins. A_An behaves like a weak gibberellin in several higher plant assay systems. The prothalli ofP. aquilinum andO. sensibilis become insensitive to Apt as they attain heart shape or shortly thereafter. Prothalli ofP. aquilinum do not begin to synthesize A_Pt and secrete it into the medium, until after they have become insensitive to it. It is in consequence of this that the most rapidly growing and developing individuals attain the archegonial phase without a prior antheridial phase. Various mechanisms and developmental characteristics are described, which strongly favor cross-fertilization inP. aquilinum without, however, eliminating an opportunity for self-fertilization. The cells of abortive antheridium initials, and of “green antheridia”, exhibit certain characteristics of green vegetative cells. These atypical structures appear to arise when early antheridial stages are overtaken by conditions unfavorable to antheridium differentiation. The observations suggest that A_Pt may be required beyond an initial inductive event. The investigations led to the conclusion that A_Pt functions by cancelling a light-dependent block to antheridium formation and suggest that in darkness this block decays without the intervention of A_Pt. InPolypodium crassifolium, the light-effect on antheridium formation is mediated by phytochrome. Other subject matters discussed include: The cellular location of antheridium initials; the relationship of antheridiogen to antheridiogen structure; the existence of a switching mechanism in the sexual development ofO. sensibilis; the retrieval of genetic information in the induction and differentiation of antheridia; the tempero-spatial pattern of competence to antheridiogen in schizaeaceous species and the inducibility of a physiological state antagonistic to antheridium formation in A.phyllitidis.     

Reproductive and competitive interactions among gametophytes of the allotetraploid fern Dryopteris corleyi and its two diploid parents

Background and Aims

Several models predict that the establishment of polyploids within diploid populations is enhanced by non-random mating (i.e. selfing and assortative mating) of cytotypes and by a higher relative fitness of polyploids. This report assesses the role that antheridiogens (i.e. maleness-inducing pheromones) and intercytotype differences in growth rate have on polyploid performance.

Methods

Three buckler-fern species were studied: the allotetraploid Dryopteris corleyi and its diploid parents, D. aemula and D. oreades. In one experiment, gametophytes of these species were cultured under rich growth conditions to compare the timing of gametangia production. The substrata on which these gametophytes had grown were used as antheridiogen sources in a second experiment. The three species were combined as source and target of antheridiogen (i.e. nine species pairs). Timing of antheridia production and gametophyte size were determined after those antheridiogen treatments.

Key Results

Under rich growth conditions the allotetraploid produced archegonia earlier than those of diploid parents. Female gametophytes of the three species produced antheridiogens that inhibited growth and favoured maleness both within and among species. Gametophyte size was similar in the three species but antheridia formed earlier in the allotetraploid.

Conclusions

Unisexuality, promoted by non-specific antheridiogens, enhances random mating both within and among species. The resulting hybridization can favour the reproductive exclusion of the allopolyploid in sites where it is outnumbered by diploids. However, the earlier production of gametangia in the allotetraploid favours assortative mating and may thus counterbalance reproductive exclusion.Key words: Allopolyploidy, antheridiogen, assortative mating, Dryopteris aemula, Dryopteris corleyi, Dryopteris oreades, gametophytes, gender expression, minority cytotype exclusion     

Significance of gametophyte form in long-distance colonization by tropical, epiphytic ferns

Gametophyte morphology of tropical epiphytic ferns may confer an advantage for establishment on islands. Most tropical, epiphytic ferns belong to five families: Hymenophyllaceae, Grammitidaceae, Vittariaceae, Polypodiaceae, and Elaphoglossaceae. Gametophytes of these families are long-lived and clone-forming. In addition, most Hymenophyllaceae, Grammitidaceae, and Vittariaceae produce dispersible gemmae. Each of these characteristics increases opportunity for outbreeding, and when island floras are statistically compared with floras of adjacent mainlands, island floras are found to be rich in epiphytic species possessing gemmae (Hymenophyllaceae, Grammitidaceae, and Vittariaceae), and depauperate in epiphytic species lacking gemmae (Polypodiaceae and Elaphoglossaceae). We propose that gametophytic gemmae significantly aid long-distance colonization of outbreeding species because gemmae 1) allow gametophytes to exploit available niches through dispersal of gemmae, and through clonal expansion and persistence of the resulting gametophyte, and 2) facilitate sexual reproduction by providing the opportunity for sperm and antheridiogen transfer when gametophytes are distant, and by providing a new source of tissue for antheridia formation.     

Antheridiogen,dark spore germination,and outcrossing mechanisms in Bommeria (Adiantaceae)

Isozymic analyses of the patterns of genetic variability in sporophyte populations have demonstrated that most fern species have outcrossing breeding systems. However, because fertilization takes place during the ephemeral, diminutive gametophyte generation, direct observation of breeding systems in nature has not been possible. Recent discoveries of soil-bound spore banks suggested that genetic diversity could be stored beneath the surface and subsequently released by appropriate chemical cues. Previous studies demonstrated that Bommeria sporophytes are the product of outcrossing, that their gametophytes carry high levels of genetic load, and that they produce and respond to antheridiogen. Research reported here demonstrated that Bommeria spores can survive long-term storage but will not germinate in the dark. Antheridiogen, however, will release spores from this light requirement and stimulate germination. Higher concentrations of antheridiogen result in higher germination rates. Gametophytes grown in the dark on antheridiogen-enriched agar form antheridia and release actively swimming sperms. Thus, spores housed beneath the soil surface could remain dormant until stimulated to germinate by antheridiogen secreted by surface-dwelling, archegoniate gametophytes. Sperm released from these subterranean gametophytes could fertilize eggs on the surface. Because spores housed in the soil are likely to be genetically different than those at the surface, heterozygous sporophytes would be more likely to result. Discovering that Bommeria species contain all of the prerequisites for this proposed outcrossing mechanism provides an explanation for the maintenance of genetic diversity in some fern populations.     

蕨类植物性别分化对环境的响应

蕨类植物是维管植物中唯一的孢子体和配子体都能独立生活的类群.同型孢子蕨类配子体的性别分化受到激素和环境因子的影响.生理学研究表明,成精子囊素与赤霉素能诱导雄配子体发育,抑制雌配子体发育;脱落酸阻止成精子囊素诱导的精子器形成;乙烯合成前体ACC促进赤霉素诱导的精子器形成,而乙烯合成抑制因子AOA通过抑制细胞分化来抑制精子器形成.光照对不同种类蕨类配子体分化的影响存在差异.糖类能够促进雄配子体形成,并可加速成熟雌配子体向两性分化.钙离子、钴离子和甲硫氨酸等分别参与了蓝光和赤霉素对配子体性别分化的调控过程.培养密度影响配子体生长及性别表达,高密度下雄性和无性配子体居多,而低密度下两性和雌性配子体居多.近年来的突变体表型分析与分子生物学研究表明,成精子囊素通过影响ANI1、HER、TRA、FEM和MAN等基因的表达调控配子体性别分化.综述了蕨类植物性别分化对环境响应的研究进展.     

狭眼凤尾蕨配子体发育及其成精子囊素对水蕨配子体发育的影响

对狭眼凤尾蕨（Pteris biaurita）配子体发育特征及其外源成精子囊素对模式植物水蕨（Ceratopteris thalictroides）在黑暗和光照条件下孢子萌发和配子体发育的影响进行了研究。结果表明：（1）狭眼凤尾蕨孢子深褐色,三裂缝,孢子萌发为书带蕨型,原叶体发育为水蕨型,无毛状体产生;培养发现,其配子体能产生精子器,但不产生颈卵器,当接种密度适中时,可进行无配子生殖。（2）在光照和黑暗条件下狭眼凤尾蕨成精子囊素有促进和抑制水蕨孢子萌发的作用,但效果均不显著。（3）在光照条件下,狭眼凤尾蕨成精子囊素可以延迟水蕨心脏形配子体分生组织缺刻的形成,但对其配子体形态和性别分化无明显影响;而在黑暗条件下狭眼凤尾蕨成精囊素对水蕨长条形配子体的形态发育具有一定影响,与对照组相比其顶端分生组织发达,整体呈长楔形,对性别分化影响不显著。可见,狭眼凤尾蕨和水蕨不具有同种成精子囊素系统。     

On separation and identity of fern antheridiogens

Chromatographic studies show that the hormones controlling antheridiuminduction in the fern species Pteridium aquilinum (Polypodiaceae),Anemia phyllitidis and Lygodium japonicum (Schizaeaceae) aredifferent molecular entities. SCHRAUDOLF's report that gibberellic acid induces antheridiain A. phyllitidis and L. japonicum was confirmed. The activityspectrum of GAs towards species of different fern families stronglyresembles that of the native Anemia antheridiogen. However,the native antheridiogens of A. phyllitidis, and of Lygodiumjaponicum, are more species-selective in their action than isGA₃. Preliminary studies have yielded no conclusive evidenceon whether the native antheridiogens are gibberellins. (Received August 21, 1967; )     

Purification,characterization, molecular cloning,and expression of novel members of jacalin-related lectins from rhizomes of the true fern Phlebodium aureum (L) J. Smith (Polypodiaceae)

A lectin was purified from rhizomes of the fern Phlebodium aureum by affinity chromatography on mannose-Sepharose. The lectin, designated P. aureum lectin (PAL), is composed of two identical subunits of approximately 15 kDa associated by noncovalent bonds. From a cDNA library and synthetic oligonucleotide probes based on a partial amino acid sequence, 5'- and 3'-rapid amplification of cDNA ends allowed the generation of two similar full-length cDNAs, termed PALa and PALb, each of which had an open reading frame of 438 bp encoding 146 amino acid residues. The two proteins share 88% sequence identity and showed structural similarity to jacalin-related lectins. PALa contained peptide sequences exactly matching those found in the isolated lectin. PALa and PALb were expressed in Escherichia coli using pET-22b(+) vector and purified by one-step affinity chromatography. Native and recombinant forms of PAL agglutinated rabbit erythrocytes and precipitated with yeast mannan, dextran, and the high mannose-containing glycoprotein invertase. The detailed carbohydrate-binding properties of the native and recombinant lectins were elucidated by agglutination inhibition assay, and native lectin was also studied by isothermal titration calorimetry. Based on the results of these assays, we conclude that this primitive vascular plant, like many higher plants, contains significant quantities of a mannose/glucose-binding protein in its storage tissue, whose binding specificity differs in detail from either legume mannose/glucose-binding lectins or monocot mannose-specific lectins. The identification of a jacalin-related lectin in a true fern reveals for the first time the widespread distribution and molecular evolution of this lectin family in the plant kingdom.     

Gibberellins and antheridiogen on sex in Blechnum spicant L.

The role of gibberellins (GAs) in determining sex in the gametophyte of the fern Blechnum spicant L. was studied through (a) the effect of exogenous GA₄₊₇ and GA₃ (b) quantitation of the endogenous levels of GA_1, GA₃, GA₄, GA₇, GA₉, and GA₂₀ in male and female gametophytes, and (c) the effect of flurprimidol, a GAs biosynthesis inhibitor of the steps of oxidation of ent-kaureno to ent-kaurenoic acid. Our results show that GA₄₊₇ had a slight effect of inducing either male or female sexual organs, antheridia and archegonia, respectively. The endogenous GAs content was not significantly different between sexes, but the GA₄, GA₇, and GA₂₀ levels were raised above those of the other GAs in both sexes. Neither antheridiogen biosynthesis nor antheridia formation was inhibited by flurprimidol. Gametophytes regenerated from homogenized mature gametophytes (HG) show a different physiological behavior than spore-derived gametophytes. In the first case, gametophytes are males and synthesize antheridiogen before they attain maturity, in contrast to what occurs in spore-derived gametophytes which are females and synthesize antheridiogen when mature.     

GENETIC ANALYSIS OF ANTHERIDIOGEN SENSITIVITY IN CERATOPTERIS RICHARDII

The pheromone antheridiogen promoted the production of males in gametophyte populations of the fern Ceratopteris richardii. Two homozygous strains of this species exhibited markedly different sensitivities to the pheromone, resulting in populations with either high or low frequencies of males. The two strains were characterized by dose responses to antheridiogen and by gametophyte population density studies to investigate the effect of antheridiogen produced within cultures. Analyses of Fl hybrids and an F2 generation indicated that differences in the antheridiogen responses of the two parental strains are associated with either one or two nuclear genes.     

Purification and Characterization of a Novel (R)-Mandelonitrile Lyase from the Fern Phlebodium aureum

Using high-performance liquid chromatography and nuclear magnetic resonance we identified vicianin as the cyanogenic compound of Phlebodium aureum. The (R)-hydroxynitrile lyase involved during cyanogenesis in the catabolism of the aglycon ([R]-mandelonitrile) was purified to apparent homogeneity. The purified holoenzyme is a homomultimer with subunits of Mr = 20,000. At least three isoforms of the enzyme exist. In contrast to other hydroxynitrile lyases, mandelonitrile lyase (MDL) from P. aureum was not inhibited by sulfhydryl- or hydroxyl-modifying reagents, suggesting a different catalytic mechanism. The enzyme is active over a broad temperature range, with maximum activity between 35 and 50[deg]C, and a pH optimum at 6.5. In contrast to (R)-MDLs isolated from several species of the Rosaceae family, (R)-MDL from P. aureum is not a flavoprotein. The substrate specificity was investigated using immobilized enzyme and diisopropyl ether as solvent. The addition of cyanide to aromatic and heterocyclic carbonyls is catalyzed by this (R)-MDL, whereas aliphatic carbonyls are poorly converted.     

Effects of tree crown structure on biomass of the epiphytic fern Polypodium scouleri (Polypodiaceae) in redwood forests

Redwood forests contain some of the largest and most structurally complex trees on Earth. The most abundant vascular epiphyte in these forests is the fern Polypodium scouleri (Polypodiaceae). We measured dimensions of all 765 P. scouleri mats on 32 trees (27 Sequoia sempervirens, 5 Picea sitchensis). Eighteen P. scouleri mats from 11 trees were randomly selected for removal and dissection in the laboratory. The total fern mat mass consisted of live fronds (3.3%), dead fronds (2.4%), live rhizomes (4.2%), dead rhizomes (8.9%), roots (34.4%), humus (28.0%), and debris (18.8%). We used multiple regression analysis to develop equations for estimation of fern masses, and we applied these equations to undisturbed fern mats on the 32 trees. Individual trees supported up to 742 kg dry mass of P. scouleri mats. These are the highest whole-tree epiphyte masses ever reported. We also quantified crown structure and counted the number of vascular plant species occurring as epiphytes on each tree. Very large, complex trees had more fern mat mass and higher vascular epiphyte species richness than smaller, simpler trees. Desiccation-sensitive organisms dependent on water stored in fern mats may be unable to survive in managed redwood forests lacking large, complex trees with abundant P. scouleri.     

Characterization and genetic analysis of antheridiogen-insensitive mutants in the fern Ceratopteris

WARNE, T. R., HICKOK, L. G. & SCOTT, R. S., 1988. Characterization and genetic analysis of antheridiogen-insensitive mutants in the fern Ceratopteris . The pheromone antheridiogen mediates the differentiation of male gametophytes in the fern Ceratopteris . Mutants insensitive to antheridiogen were isolated using an in vitro selection procedure. Antheridiogen-insensitive mutants exhibited partial or complete insensitivity to antheridiogen, but were normal in all other respects. Two mutants were completely insensitive to antheridiogen, whereas, another mutant was insensitive to supplemented antheridiogen, but produced male gametophytes in multispore cultures. Genetic analysis suggested a single gene basis for each mutant.     

Ant protection of the nectaried fern Polypodium plebeium in central Mexico

Nectaries on fronds of Polypodium spp. have been studied previously only in cultivated specimens. We conducted field observations in middle-elevation forests in Mexico and found five ant species associated with nectaries of Polypodium plebeium and P. lepidotrichum. To investigate whether nectaries promote protection against herbivores, we performed ant-exclusion experiments with nectary-bearing ferns (P. plebeium) and other ferns without nectaries (Polypodium plesiosorum, P. furfuraceum, and Phlebodium pseudoaureum). When ants were excluded from the developing fronds of Polypodium plebeium, damage from foliage-feeding sawfly and lepidopteran caterpillars was significantly greater than in control fronds. Ferns without nectaries did not show a difference in damage between ant-excluded and control fronds. Our results demonstrate that fern nectaries can support ant defense of the plant body as do the extrafloral nectaries of many angiosperms.     

The reproductive biology of the invasive ferns Lygodium microphyllum and L. japonicum (Schizaeaceae): implications for invasive potential

The effect of culture system and population source on sexual expression and sporophyte production was examined for two invasive fern species in Florida, USA, Lygodium microphyllum and L. japonicum (Schizaeaceae). Both species are currently spreading through Florida. Long-distance dispersal of ferns is thought to rely on successful intragametophytic selfing. Given the rate of spread observed in both Lygodium species, we hypothesized that both species are capable of intragametophytic selfing. To test this hypothesis, gametophytes of both species were grown in vitro as isolates, pairs, and groups. Both species were capable of intragametophytic selfing; 78% of L. microphyllum isolates produced sporophytes and over 90% of the L. japonicum isolates produced sporophytes. Lygodium microphyllum also displayed the ability to reproduce via intergametophytic crossing, facilitated by an antheridiogen pheromone. Sporophyte production was rapid across mating systems for both species, an advantage in Florida's wet and dry seasonal cycles. The high intragametophytic selfing rate achieved by both species has likely facilitated their ability to colonize and spread through Florida. The mixed mating system observed in L. microphyllum appears to give this species the ability to invade distant habitats and then adapt to local conditions.     

14种剑蕨属植物的孢子形态特征及其系统分类学意义

利用扫描电子显微镜对水龙骨科(Polypodiaceae)剑蕨属(Loxogramme)14种植物的孢子形态和表面纹饰进行了观察。结果显示:剑蕨属的孢子具有单裂缝和三裂缝两种类型,个别物种具有中间过渡类型。剑蕨属的孢子纹饰为蠕虫状或浅波纹状。经比较发现,具三裂缝孢子的物种通常植株较小,而具单裂缝的种类则通常植株较大。从进化的角度来看,三裂缝孢子为原始类型,单裂缝孢子为进化类型,因而从孢子形态特征的角度支持剑蕨属为水龙骨科基部类群。     

The Transformer Genes of the Fern Ceratopteris Simultaneously Promote Meristem and Archegonia Development and Repress Antheridia Development in the Developing Gametophyte

The sex of the haploid gametophyte of the fern Ceratopteris is determined by the presence or absence of the pheromone antheridiogen, which, when present, promotes male development and represses female development of the gametophyte. Several genes involved in sex determination in Ceratopteris have been identified by mutation. In this study, the epistatic interactions among new and previously described sex-determining mutants have been characterized. These results show that sex expression is regulated by two sets of genes defined by the FEM1 and TRA loci. Each promotes the expression of either male or female traits and simultaneously represses the expression of the other. A model describing how antheridiogen regulates the expression of these genes and the sex of the gametophyte is described. The observation that some gametophytic sex-determining mutants have phenotypic effects on the sporophyte plant indicates that sex determination in the Ceratopteris gametophyte is regulated by a mechanism that also regulates sporophyte development.     

Genetic diversity and breeding system in a group of neotropical epiphytic ferns (Pleopeltis; Polypodiaceae)

Epiphytes are ecologically important components of tropical forests worldwide and yet they have been underrepresented in studies of reproductive biology. Given the presumed ephemeral nature of their substrates, and the importance of dispersal and colonization, epiphytes might be expected to undergo substantial inbreeding to ensure reproductive success, as in weedy terrestrial plants. While there is some evidence for inbreeding in epiphytic angiosperms, the only previous studies of fern epiphytes indicate that they are predominantly outcrossing. The present study reports on the genetic diversity and breeding system of six members of the Neotropical epiphytic fern genus Pleopeltis (Polypodiaceae). A survey of isozyme variability using starch gel electrophoresis revealed high population levels of polymorphism (P = 0.62), allelic diversity (A = 2.3), and individual heterozygosity (Ho = 0.181), but little differentiation among conspecific populations (I 3 0.98; Gst = 0.048), and high interpopulational gene flow rates (Nm > 1). In addition, there was no indication of homozygote excess within populations that might indicate a history of selfing in these ferns: populations generally conformed to Hardy-Weinberg expected genotype frequencies, and both Wright's inbreeding coefficient (Fis) and Holsinger's intragametophytic selfing rates approached zero. Possible mechanisms limiting inbreeding in these ferns include antheridiogen activity and high levels of genetic load that would lead to inbreeding depression upon selfing.