Spore Mother Cells Research Articles

Chromatin reprogramming during the somatic-to-reproductive cell fate transition in plants

The life cycle of flowering plants is marked by several post-embryonic developmental transitions during which novel cell fates are established. Notably, the reproductive lineages are first formed during flower development. The differentiation of spore mother cells, which are destined for meiosis, marks the somatic-to-reproductive fate transition. Meiosis entails the formation of the haploid multicellular gametophytes, from which the gametes are derived, and during which epigenetic reprogramming takes place. Here we show that in the Arabidopsis female megaspore mother cell (MMC), cell fate transition is accompanied by large-scale chromatin reprogramming that is likely to establish an epigenetic and transcriptional status distinct from that of the surrounding somatic niche. Reprogramming is characterized by chromatin decondensation, reduction in heterochromatin, depletion of linker histones, changes in core histone variants and in histone modification landscapes. From the analysis of mutants in which the gametophyte fate is either expressed ectopically or compromised, we infer that chromatin reprogramming in the MMC is likely to contribute to establishing postmeiotic competence to the development of the pluripotent gametophyte. Thus, as in primordial germ cells of animals, the somatic-to-reproductive cell fate transition in plants entails large-scale epigenetic reprogramming.

Characterization of a Chitin-Binding Protein from Bacillus thuringiensis HD-1

Strains of Bacillus thuringiensis produce insecticidal proteins. These strains have been isolated from diverse ecological niches, such as soil, phylloplane, insect cadavers and grain dust. To effectively propagate, these strains produce a range of molecules that facilitate its multiplication in a competing environment. In this report, we have examined synthesis of a chitin-binding protein and evaluated its effect on fungi encountered in environment and its interaction with insecticidal proteins synthesized by B. thuringiensis. The gene encoding chitin-binding protein has been cloned and expressed. The purified protein has been demonstrated to interact with Cry insecticidal protein, Cry1Ac by Circular Dichrosim spectroscopy (CD) and in vitro pull down assays. The chitin-binding protein potentiates insecticidal activity of bacillar insecticidal protein, Cry1Ac. Further, chitin-binding protein was fungistatic against several soil fungi. The chitin binding protein is expressed in spore mother cell and deposited along with insecticidal protein, Cry1Ac. It interacts with Cry1Ac to potentiate its insecticidal activity and facilitate propagation of Bacillus strain in environment by inhibiting growth of certain fungi.

Observations onOphioglossum eliminatum(Pteridophyta: Ophioglossaceae) with comments on the lowest chromosome number in the genus

Population biology data collected during 1995–2009 from more than a dozen localities in Central India have revealed that both the interspecific as well as intraspecific chromosome variations are inherent in many species of Ophioglossum L. The lowest count of chromosome recorded for any plant of Ophioglossum eliminatum Khandelwal et Goswami reported to be n = 30 chromosomes has not been confirmed even in a single spore mother cell or in any mitotically dividing root tip cell. This is emphatically emphasised to be characterised by n = 86±2 chromosomes little lower than originally suggested 90±2 chromosomes. Therefore this is reemphasised that lowest chromosome count for the genus is n = 86, not n = 30 as claimed earlier. Somatic chromosomes from root tip squashes and microtome sections of root tip cells stained with haematoxylin clearly reveal polysomaty (chromosome variations in multiples within the same tissue) in O. eliminatum and O. costatum, which have been probably ignored by earlier workers. Additionally, typical epidermal feature of linear splitting of cells to form several openings in the mesophyll, inherent and expressed only by leaves of O. eliminatum has been found to be a diagnostic trait of the species collected from original localities in Shivpuri forest in Central India and far away in Rajasthan in Western India. As far as known, no living or a fossil leaf has ever shown such natural tearing of the mesophyll tissue.

The R2R3 MYB Transcription Factors FOUR LIPS and MYB88 Regulate Female Reproductive Development

Gamete formation is an important step in the life cycle of sexually reproducing organisms. In flowering plants, haploid spores are formed after the meiotic division of spore mother cells. These spores develop into male and female gametophytes containing gametes after undergoing mitotic divisions. In the female, the megaspore mother cell undergoes meiosis forming four megaspores, of which one is functional and three degenerate. The megaspore then undergoes three mitotic cycles thus generating an embryo sac with eight nuclei. The embryo sac undergoes cellularization to form the mature seven-celled female gametophyte. Entry into and progression through meiosis is essential for megasporogenesis and subsequent megagametogenesis, but control of this process is not well understood. FOUR LIPS (FLP) and its paralogue MYB88, encoding R2R3 MYB transcription factors, have been extensively studied for their role in limiting the terminal division in stomatal development by direct regulation of the expression of cell cycle genes. Here it is demonstrated that FLP and MYB88 also regulate female reproduction. Both FLP and MYB88 are expressed during ovule development and their loss significantly increases the number of ovules produced by the placenta. Despite the presence of excess ovules, single and double mutants exhibit reduced seed set due to reduced female fertility. The sterility results at least in part from defective meiotic entry and progression. Therefore, FLP and MYB88 are important regulators of entry into megasporogenesis, and probably act via the regulation of cell cycle genes.

Cryptosexuality and the genetic diversity paradox in coffee rust, Hemileia vastatrix.

BackgroundDespite the fact that coffee rust was first investigated scientifically more than a century ago, and that the disease is one of the major constraints to coffee production - constantly changing the socio-economic and historical landscape of the crop - critical aspects of the life cycle of the pathogen, Hemileia vastatrix, remain unclear. The asexual urediniospores are regarded as the only functional propagule: theoretically, making H. vastatrix a clonal species. However, the well-documented emergence of new rust pathotypes and the breakdown in genetic resistance of coffee cultivars, present a paradox.Methods and ResultsHere, using computer-assisted DNA image cytometry, following a modified nuclear stoichiometric staining technique with Feulgen, we show that meiosis occurs within the urediniospores. Stages of spore development were categorised based on morphology, from the spore-mother cell through to the germinating spore, and the relative nuclear DNA content was quantified statistically at each stage.ConclusionsHidden sexual reproduction disguised within the asexual spore (cryptosexuality) could explain why new physiological races have arisen so often and so quickly in Hemileia vastatrix. This could have considerable implications for coffee breeding strategies and may be a common event in rust fungi, especially in related genera occupying the same basal phylogenetic lineages.

Germline specification in plant reproduction

Three years ago, the journal Sexual Plant Reproduction featured a special issue on ‘‘Deciphering molecular mechanisms of fertilization in seed plants,’’ which captured our state of understanding of gamete function and double fertilization in flowering plants at that time. This issue received considerable attention, making it one of the journal’s most successful. Since then, the pace of research has accelerated further, and recently we have seen numerous scientific breakthroughs. We anticipate that in the near future we will have sufficient tools and knowledge of molecular reproductive mechanisms to allow unparalleled manipulation and genetic improvement of crop plants. Basic data on the molecular regulation of cell lineages that govern plant reproduction will need to be among the resources employed to meet the challenges posed by increasing population in a context of dwindling arable land, demands on fresh water supplies and the need to provide the genetic diversity to resist increasing global challenges in climate, energy resources and disease. With the current special issue, we address several enduring themes that relate to the fundamental biology of gametes and germ lineages, taking into account the most recent developments in the field of plant reproduction. In particular we examine the specification of the plant germline in reproduction and explore the general themes and molecular mechanisms that govern germ lineage and cell fate establishment, which expands upon the coverage in our previous special issue. In particular we address the designation and nature of germline identity in plants, from the simplest version of a germ cell-producing somatic cell, to the strongly reduced angiosperm germline, which derives two distinct germ lineages from spore mother cells that ultimately are the founding cells of the embryo and endosperm in flowering plants. In this issue, we include such aspects as crosstalk between the gametophyte, germ cells and the sporophyte, specification of gametes, the role of epigenesis in this process, and the establishment of polarity during gametogenesis and early embryogenesis. The selected papers address unifying concepts of critical importance to biology in general, focusing on flowering plants, which constitute the majority of crop plants upon which man relies. As before, we have asked leading scientists in the field to contribute to the special issue. We would like to thank Ueli Grossniklaus for his kind introduction to the special issue and to the authors, Alma Armenta-Medina, Stefano Bencivenga, Lucia Colombo, Edgar Demesa-Arevalo, Rita Gros-Hardt, Armin Hallmann, Thomas Laux, Simona Masiero, Stefanie Sprunck, David Twell, Jean-Philippe Vielle-Calzada, Zhongjuan Zhang, and also to our reviewers.

Cytological study of a fern Cystopteris fragilis in Mongolian Altai

Cystopteris fragilis collected in Mongolian Altai showed three cytotypes with chromosome numbers of tetraploid (2n=c.168), pentaploid (2n=c.210) and hexaploid (2n=c.252). Flow cytometry analysis also showed three different peaks of DNA contents of nuclei in fronds. Meiotic metaphase chromosomes of the spore mother cells in the tetraploid and hexaploid C. fragilis showed all bivalents with c.84II and c.126II, while those of the pentaploid C. fragilis showed many univalents and bivalents that produced mostly sterile spores. The pentaploid C. fragilis was suggested to be a hybrid between the tetraploid and the hexaploid C. fragilis.

Resistance of Helicoverpa armigera to Cry1Ac toxin from Bacillus thuringiensis is due to improper processing of the protoxin

The bacterium Bacillus thuringiensis produces ICPs (insecticidal crystal proteins) that are deposited in their spore mother cells. When susceptible lepidopteran larvae ingest these spore mother cells, the ICPs get solubilized in the alkaline gut environment. Of approx. 140 insecticidal proteins described thus far, insecticidal protein Cry1Ac has been applied extensively as the main ingredient of spray formulation as well as the principal ICP introduced into crops as transgene for agricultural crop protection. The 135 kDa Cry1Ac protein, upon ingestion by the insect, is processed successively at the N- and C-terminus by the insect midgut proteases to generate a 65 kDa bioactive core protein. The activated core protein interacts with specific receptors located at the midgut epithilium resulting in the lysis of cells and eventual death of the larvae. A laboratory-reared population of Helicoverpa armigera displayed 72-fold resistance to the B. thuringiensis insecticidal protein Cry1Ac. A careful zymogram analysis of Cry1Ac-resistant insects revealed an altered proteolytic profile. The altered protease profile resulted in improper processing of the insecticidal protein and as a consequence increased the LC50 concentrations of Cry1Ac. The 135 kDa protoxin-susceptible insect larval population processed the protein to the biologically active 65 kDa core protein, while the resistant insect larval population yielded a mixture of 95 kDa and 68 kDa Cry1Ac polypeptides. N-terminal sequencing of these 95 and 68 kDa polypeptides produced by gut juices of resistant insects revealed an intact N-terminus. Protease gene transcription profiling by semi-quantitative RT (reverse transcription)-PCR led to the identification of a down-regulated HaSP2 (H. armigera serine protease 2) in the Cry1Ac-resistant population. Protease HaSP2 was cloned, expressed and demonstrated to be responsible for proper processing of insecticidal protoxin. The larval population displaying resistance to Cry1Ac do not show an altered sensitivity against another insecticidal protein, Cry2Ab. The implications of these observations in the context of the possibility of development of resistance and its management in H. armigera to Cry1Ac through transgenic crop cultivation are discussed.

Male Sterile Lines of Zinnia elegans and Their Cytological Observations

In order to find out a new pathway for utilizing heterosis of Zinnia elegans and accelerate breeding process, the mechanism of anther development of a male sterile line was explored. Backcross, sibmating, selfing of fertile plants and testcross with inbred lines were analyzed and identified in the field, and cytology was observed. Recessive nucleus male sterile line AH209AB capable of being a maintainer was obtained by successive backcrosses with male sterile plants and fertile F1 plants as male parents. Cytological and anatomical studies indicated that: (1) The wall of normal anther was constituted of four layers of cells such as epidermis, powder chamber wall, middle level and tapetum cells. The process in meiosis of pollen mother cell in Zinnia elegans was normal and cytoplasm divided simultanously. Mature pollen grain was tricellular type. (2) The petal of male sterile plant degraded as a thread-like structure, the stamens were villiform in appearance and no pollens were formed. The result showed that the anther of male sterile plant no longer proceed to differentiate spore mother cell and the pollen sac after the formation of the tissue of sporogenous cells, there was no evident boundary between tapetum cell, middle lamella and inner wall of PMC, tapetal cells did not develop from the very beginning. So the abortion type was completely structural male sterility. The male sterile line belongs to non-sporange male sterile type and is of great use in F1 seeds production.

Ovule Development inGinkgo bilobaL., with Emphasis on the Collar and Nucellus

Ovule development in Ginkgo biloba is examined. Ovulate stalks are initiated in leaf axils, and each stalk primordium dichotomizes to produce two ovule primordia. Integument development appears circumferential and during micropyle formation becomes slightly lobed. Primary nucellar tissues are derived from periclinal divisions in the hypodermal layer, which is limited to several apically located cells of the nucellus and remains active through pollen chamber formation. One spore mother cell differentiates in the sporangial tissues; others differentiate as tapetum. The epidermal layer does not contribute directly to development of the primary nucellus but does contribute to secondary nucellar tissues. Secondary parietal tissues are divided into regions based on their growth patterns and location within the nucellus: the pollen chamber, including a distinct exothecium; the micropylar parietal tissue, including the distal and proximal tissues; the flanking parietal tissue; and the chalazal parietal tissue that lies between the flattened‐cell layer and the enlarging sporogenous tissue. The collar develops from periclinal divisions of cells in the hypodermal layer proximal to the ovule after the integument and nucellus have differentiated. The collar is interpreted as a unique structure within ginkgophytes. In some ovules, an organized flap of tissue develops between the collar and the ovule. There does not seem to be any homology of the flap with other structures in gymnosperms based on position or timing of growth, and it is interpreted as a structure sui generis.

Sporophyte Structure in the Neotropical Hornwort Phaeomegaceros fimbriatus: Implications for Phylogeny, Taxonomy, and Character Evolution

The first detailed structural description of the sporophyte of a Neotropical hornwort is presented herein. The morphology and ultrastructure of the placenta, basal meristem, and aerial sporophyte are described in populations of Phaeomegaceros fimbriatus, a poorly known but widespread species, from Panama, Venezuela, Costa Rica, and Ecuador. Gametophyte cells with wall ingrowths and haustorial sporophyte cells lacking ingrowths are closely intermingled in the placenta. Through anticlinal divisions, the basal meristem generates a protoderm, several rows of assimilative cells, and a single ring of archesporial cells surrounding a columella of 16 cells. Further periclinal division in the archesporium produces two or three rows of spore mother cells intermixed with elater initials, both of which are separated and suspended in mucilage as the sporophyte elongates. Spores are finely vermiculate with seven to 10 dimples on the distal surface. Spore walls comprise a perine‐like outer layer derived from the spore mother cell wall, a two‐layered exine, and a translucent intine. An aperture of thickened exine and sparse sporopollenin deposition defines the conspicuous trilete mark. The mature sporophyte consists of elongated, thick‐walled epidermal cells with stomata, sporogenous tissue, a central columella of up to 30 cells, and seven to 13 layers of assimilative cells with chloroplasts lacking pyrenoids. This study reveals a sharp morphological separation of plants from Venezuelan paramos from the rest of the populations, indicating recent evolutionary events. Superficial structural features such as foot shape, spore color, and stomata in the sporophyte align P. fimbriatus with Phaeoceros, while more elaborate development of histological regions, such as massive sporogenous tissue and the ultrastructure of plastids, suggests affinities with Megaceros. The unique mixture of Phaeoceros and Megaceros traits defines the genus Phaeomegaceros, including P. fimbriatus, and provides further evaluation of the taxonomy and evolutionary diversification of hornworts.

Sporogenesis in Haploid Sporophytes of Osmunda japonica (Osmundaceae)

Three out of four haploid sporophytes of Osmunda japonica (2n=x=22), produced artificially by induction of apogamy, developed fertile fronds and spores. The spores that germinated (up to 25% of the total green spores) were monoploid, determined by the nuclear DNA content and the somatic chromosome numbers of the gametophytes. The haploid 2 sporophyte produced more fertile spores than the two other haploids, providing an opportunity to observe the details of fertile spore development. In some spore mother cells of this plant, the random separation of 22 univalent chromosomes did not occur at the first meiotic division, but restitution nuclei were formed. The observation of chromosome behavior in the haploid 2 sporophyte showed that sporogenesis in haploid O. japonica resembles the Braithwaite scheme in apogamous ferns. Gametophytes, developed from spores of the three haploid sporophytes, produced a few sporophytes with diploid chromosome numbers. One of the new sporophytes was trisomic.

Approach to the early sporangial development in angiosperms considering meiosis control and cellular differentiation

Applying current data on cell differentia- tion and meiosis control to the early sporangial development in angiosperms, a strict relationship between cell lineage and its differentiation fate is rejected. An evaluation of cytological features indicative of a meiotic (sporogenous) fate discards the sterilization phenomenon and introduces the premeiotic cellular differentiation (PCD) concept. The early sporangial development comprises 5 basic steps and 4 cellular stages, where PCD and meiosis extension and gradient are related to mechanisms of spore mother cell selection. Concepts here discussed explain the exceptions to the normal early sporangial development and allow a precise definition of archesporium and archespore. PCD and meiotic extension and gradient recover more information of the early sporangial development, distinguishing different developmental patterns leading to the same final result and retaining slight developmental differences. However, there are no early developmental characteristics distinctive of andro- or gynosporangia. Therefore, the heterosporic condition is not related to early developmental changes.

Cryptospores: The Origin and Early Evolution of the Terrestrial Flora

The Cryptogamic, or spore-producing, plants of today are composed of three nonvascular, bryophyte groups (mosses, liverworts, and hornworts) and several vascular groups (ferns, club mosses, and horsetails). All of these plants produce abundant spores, which serve as propagules for dispersing and, to some extent, preserving plants through periods of ecological stress. Plant spores are typically formed as the end products of meiosis (reduction division) from a dividing sporocyte, or spore mother cell (smc). Because of this, they typically occur in groups of four, with each individual spore bearing a characteristic trilete, or Y-shaped mark on its common contact surface. Spore walls, composed of an inert, heterogeneous polymer called sporopollenin, are extremely resistant to the chemical vicissitudes of the terrestrial environment. This property of typical plant spores ultimately allows them to be quite abundant in fine grained clastic rocks. Although fossilized spores represent only a small part of the once-living plant, in many cases, they represent an important component of the plant fossil record, especially when the preservation of macroscopic tissues is lacking.

Reproductive biology of gametophytes of Cyathea podophylla (Hook.) Copel.

This study focused on characteristics related to the reproductive biology of Cyathea podophylla (Hook.) Copel. gametophytes, including growth habits, sexual expression, antheridiogen, ploidy, and genetic load. Antheridium formation was not promoted when young gametophytes of C. podophylla were treated with their own mature gametophytes or those of Pteridium aquilinum (L.) Kuhn, Lygodium japonicum (Thunb.) Sw., or Ceratopteris thalictroides (L.) Brongn. Their spores did not germinate at all in dark conditions. These results indicate that there is no antheridiogen (and possibly no germinin, either) in this species. The sexual progression of their gametophytes is from male to hermaphroditic. This sexual expression is not consistent with the model of antheridiogen action, either. The chromosome number of the spore mother cells is 69(subscript Ⅱ), which indicates that the sporophytes are diploid. Because of its high genetic load, C. podophylla reproduces mainly through intergametophytic mating, especially intergametophytic crossing. The facts that spores mature almost throughout the year may facilitate this type of mating behavior.

Changes in Plastid DNA Content during the Life Cycle of the Hornwort Anthoceros punctatus L.

The plastid DNA contents of egg, sperm, sporophyte, and gametophyte cells were measured to clarify the relative changes in plastid DNA content during the life cycle of the hornwort Anthoceros punctatus. Samples stained with DAPI were observed using epifluorescence microscopy and CHIAS. The plastid DNA content of unfertilized egg cells did not change before and the fertilization because of preferential digestion of organelle DNA in sperm cells. In the 2n-generation, plastids multiplied by binary fission, with DNA duplication. It is thought that plastid DNA content is halved in spore mother cells, but recovers before, or immediately after, spore germination. In the ngeneration, plastids multiplied by binary fission, with DNA duplication, and the plastid DNA content was maintained at the same level as that of the 2n-generation.

Meiotic cytokinetic apparatus in the formation of the linear spore tetrads of Conocephalum japonicum (Bryophyta)

Most bryophytes produce tetrahedral spore tetrads. However, linear spore tetrads have been reported to occur in Conocephalum japonicum (Thunb.) Grolle. In this study, the distribution of microtubules (MTs) during meiosis in C. japonicum was examined to determine the division pattern resulting in a linear tetrad. Spore mother cells in the pre-meiotic stage were cylindrical with randomly distributed cytoplasmic MTs. In the prophase-metaphase transition, spindle MTs replaced cytoplasmic MTs and a barrel-shaped spindle with two flattened poles developed. Cortical MT arrays were not detectable throughout meiosis. Although a phragmoplast appeared between sister nuclei in telophase-I, it disappeared without expanding to the parental cell wall. Metaphase-II spindles oriented parallel to the long axis of the cell and in tandem to each other resulted in a linear arrangement of telophase nuclei. Radial arrays of MTs developed from the nuclear surfaces and three phragmoplasts appeared among the four nuclei to produce four spores. Two phragmoplasts separating the paired sister nuclei appeared prior to the appearance of a phragmoplast between non-sister nuclei. The MT cycle is basically the same as that reported in meiosis of C. conicum, which produces non-linear tetrads. A morphometric study indicated that the difference in the division pattern between C. conicum and C. japonicum is due to a difference in the shape of spore mother cells. The cylindrical shape of sporocytes of C. japonicum restricts the orientation of spindles and phragmoplasts so that the four resultant spores are arranged linearly.

Studies on Plastid DNA Replication Cycle in Anthoceros punctatus

We have investigated the plastid DNA replication cycle of Anthoceros Punctatus. In mitotic cycles of this species, plastids divided into two daughter plastids with DNA synthesis. By contrast, in spore mother cells, the DNA content of plastids which had completed their division was halved as well as the DNA content of cell nuclei. The plastid DNA content of unfbrtilized egg cells did not change before and the fertilization because of prefbrential digestion of organelle DNA in sperm cells. In the 2n-generatio, plastids multiplied by binary fission, with DNA duplication. It is thought that plastid DNA content is halved in spore mother cells but recovers before, or immediately after, spore germination. In the n-generation, Plastids multiplied by binary fission, with DNA duplication, and the plastid DNA content was maintained at the same level as that of the 2ngeneration.

Generating autotetraploid sporophytes and their use in analyzing mutations affecting gametophyte development in the fern Ceratopteris.

The haploid gametophytes of the fern Ceratopteris richardii are autotrophic and develop independently of the diploid sporophyte plant. While haploid genetics is useful for screening and characterizing mutations affecting gametophyte development in Ceratopteris, it is difficult to assess whether a gametophytic mutation is dominant or recessive or to determine allelism by complementation analysis in a haploid organism. This report describes how apospory can be used to produce genetically marked polyploid sporophytes whose gametophyte progeny are heterozygous for mutations affecting sex determination in the gametophyte and a known recessive mutation affecting the phenotype of both the gametophyte and sporophyte. The segregation ratios of wild-type to mutant phenotypes in the gametophyte progeny of polyploid sporophyte plants indicate that all of the mutations examined are recessive. The presence of many multivalents and few univalents in meiotic chromosome preparations of spore mother cells confirm that the sporophyte plants assayed are polyploid. The DNA content of the sperm of their progeny gametophytes was also found to be approximately twice that of sperm from wild-type haploid gametophytes.

Cytochemical Characterization of the Synaptic Poles in Equisetum Spore Mother Cells in the Synaptic Stage of Meiosis.

Two kinds of poles, the PP and the MP, are manifested just prior to and during the synaptic stage of meiosis in the SMCs of Equisetum. A spectroscopic and ultracytochemical characterization of these poles has been carried out to elucidate their functions. The former pole not only works as the intracellular center of carbohydrate metabolism in this stage, but also as a possible supply center of intermediates for protein and fat biosyntheses. In contrast, the latter pole functions as the intracellular center of ATP production in this stage. This synaptic pole formation is causally related to the “chromosome and molecular pairing” of meiosis.