Sporogenous Cells Research Articles

Arabidopsis SOMATIC EMBRYOGENESIS RECEPTOR KINASES1 and 2 Are Essential for Tapetum Development and Microspore Maturation

Among the >200 members of the leucine-rich repeat receptor kinase family in Arabidopsis thaliana, only a few have been functionally characterized. Here, we report a critical function in anther development for the SOMATIC EMBRYOGENESIS RECEPTOR KINASE1 (SERK1) and SERK2 genes. Both SERK1 and SERK2 are expressed widely in locules until stage 6 anthers and are more concentrated in the tapetal cell layer later. Whereas serk1 and serk2 single insertion mutants did not show developmental phenotypes, serk1 serk2 double mutants were not able to produce seeds because of a lack of pollen development in mutant anthers. In young buds, double mutant anthers developed normally, but serk1 serk2 microsporangia produced more sporogenous cells that were unable to develop beyond meiosis. Furthermore, serk1 serk2 double mutants developed only three cell layers surrounding the sporogenous cell mass, whereas wild-type anthers developed four cell layers. Further confocal microscopic and molecular analyses showed that serk1 serk2 double mutant anthers lack development of the tapetal cell layer, which accounts for the microspore abortion and male sterility. Taken together, these findings demonstrate that the SERK1 and SERK2 receptor kinases function redundantly as an important control point for sporophytic development controlling male gametophyte production.

Cellular organisation in meiotic and early post-meiotic rice anthers

We have used fluorescent, confocal laser and transmission electron microscopy (TEM) to examine cellular organisations, including callose (1,3-beta-glucan) behaviour, in meiotic and early post-meiotic rice anthers. These features are critical for pollen formation and provide information to better understand pollen sterility caused by abiotic stress in rice and other monocotyledonous species. Among organelles during meiosis, abundant plastids, mitochondria and nuclei of the anther cells show distinctive features. Chloroplasts in the endothecium store starch and indicate a potential for photosynthetic activity. During meiosis, the middle layer cells are markedly compressed and at the tetrad stage are either vacuolated or filled with degenerating electron-opaque organelles. Viable mitochondria, stained with Rhodamine 123, are seen in the endothecium and tapetum, but the mitochondria in the middle layer are not stained during meiosis. The radial walls of the tapetum are disorganised and degenerating, indicating the formation of a syncytium; pro-orbicules are located at the locular walls at the tetrad stage. Immunohistochemical studies show that the sporogenous cells are entirely enveloped by a thick callosic layer at early meiosis. Cell plate callose was assembled in a plane between the dyad cells. In the tetrads, however, callose formed only at the centre, showing that the tetrad microspores are not enveloped but separated by callose walls. Thick, undulating electron-opaque walls around the tetrads indicate the beginning of exinous microspore wall differentiation.

Microsporogenesis inAmelanchier alnifolia: sporogenous cells, microsporocytes, and tetrads

As part of an overall program aimed at increasing our knowledge of the male reproductive system of Amelanchier alnifolia Nutt., this study documents structural and developmental changes that occur in the sporogenous cells, microsporocytes, and tetrads of microspores during microsporogenesis using general cytochemical techniques in conjunction with bright field, fluorescence, and transmission electron microscopy. The sporogenous cells are thin walled and stain positively for β-1,4-glucans, pectic acids, and cellulose, but not callose. At the microsporocyte and tetrad stages of microsporogenesis, thick walls develop and stain positively for β-1,4-glucans (hemicelluloses but not cellulose), pectic acids, and callose. Thus, the eventual release of maturing microspores from the tetrads requires the digestion of all three of these carbohydrate wall materials. Postmeiotic cytokinesis is of the simultaneous type and is initiated when Golgi vesicles aggregate simultaneously into a network of cylindrical tubules in both central and peripheral cell locations of the coenocytic tetrad. Eventually, this network fuses to form the new cell walls within the microspore tetrad.

Cellular organisation and differentiation of organelles in pre-meiotic rice anthers

Pre-meiotic cellular organisation of rice anthers has a great significance in pollen formation. We have used a combination of confocal laser and transmission electron microscopy (TEM) to characterise and differentiate organelles in pre-meiotic rice anthers. Along with the characteristic organelles in the cytoplasm the epidermal cells of the pre-meiotic rice anther are coated on their outer surface by a conspicuous bi-lamellate cuticle. Chloroplasts of the endothecium contain immature grana, thylakoids and also starch granules. These plastids clearly contain photosynthetic pigments as shown by autofluorescence in confocal microscope studies. Both confocal and TEM studies reveal clusters of mitochondria in the middle layer. The tapetum contains electron opaque ribosomes, bundles of mitochondria and plastids. The nuclei of the tapetum occupy a large volume of the cytoplasm indicating the onset of mitotic prophase. Intense Rhodamine 123 staining reveals that a major portion of the structurally indistinguishable organelles that were seen throughout the densely ribosomic cytoplasm of sporogenous cells are mitochondria.

The effects of nitrite on the survival of Clostridium sporogenes and the autoxidation properties of the Kavurma

Kavurma is a traditional cooked (fried) meat product manufactured to preserve meat. Some bacterial genera, e.g., clostridia are important in kavurma. The objective of this study was to determine the influence of nitrite and the traditional cooking process on the survival and proliferation of Clostridium botulinum and the autoxidation properties of the kavurma. For this purpose, Clostridium sporogenes having similar characteristics to C. botulinum was used, and the samples were inoculated with 10 6 CFU/g C. sporogenes cells before the traditional cooking. The final products were packaged and stored under refrigeration for 6 months, and then the oxidation parameters (TBA, peroxide and free fatty acid values) and C. sporogenes counts of samples were determined. It was observed that C. sporogenes could survive during the traditional cooking process and storage. However, counts decreased during storage; for example, nitrite containing samples initially had 3.21 log CFU/g C. sporogenes and 2.73 log CFU/g at the end of storage. While nitrite had a slight antimicrobial effect on clostridia, it significantly reduced the TBA, peroxide and FFA values of the samples. In conclusion, it is suggested that addition of 100 ppm of nitrite might be useful in kavurma processing because of its role in limiting oxidation as well as its antimicrobial effect.

Overexpression ofTAPETUM DETERMINANT1Alters the Cell Fates in the Arabidopsis Carpel and Tapetum via Genetic Interaction withEXCESS MICROSPOROCYTES1/EXTRA SPOROGENOUS CELLS

Previously, we reported that the TAPETUM DETERMINANT1 (TPD1) gene is required for specialization of tapetal cells in the Arabidopsis (Arabidopsis thaliana) anther. The tpd1 mutant is phenotypically identical to the excess microsporocytes1 (ems1)/extra sporogenous cells (exs) mutant. The TPD1 and EMS1/EXS genes may function in the same developmental pathway in the Arabidopsis anther. Here, we further report that overexpression of TPD1 alters the cell fates in the Arabidopsis carpel and tapetum. When TPD1 was expressed ectopically in the wild-type Arabidopsis carpel, the number of cells in the carpel increased significantly, showing that the ectopic expression of TPD1 protein could activate the cell division in the carpel. Furthermore, the genetic analysis showed that the activation of cell division in the transgenic carpel by TPD1 was dependent on EMS1/EXS, as it did not happen in the ems1/exs mutant. This result further suggests that TPD1 regulates cell fates in coordination with EMS1/EXS. Moreover, overexpression of TPD1 in tapetal cells also delayed the degeneration of tapetum. The TPD1 may function not only in the specialization of tapetal cells but also in the maintenance of tapetal cell fate.

The Study of Pollen Development in Nine Cultivars of Hazelnut (Corylusavellana)

Pollen development is an important event in plant reproduction. Hazelnut (Corylus avellana) male flower differentiation starts in summer and pollen shed is in the winter. Hazelnut pollen shed can vary up to 3 months between early to late flowering genotypes. Microsporogenesis and microgametogenesis of hazelnut is not well understood. Pollen development and differentiation of nine genotypes, representing early to late blooming cultivars from the National Clonal Germplasm Repository in Corvallis, Ore., were studied. Catkins were collected weekly from Aug. to Nov. 2002. Tissue sections were examined under the light microscope. Microsporogenesis was divided into five stages: archesporial cells, sporogenous cells and parietal layers, pollen mother cells (PMC), tetrads, and microspores. Microgametogenesis was distinguished between young pollen grains (uninucleate) and mature pollen grains (binucleate). On 4 Aug., cultivars were at different developmental stages of microsporogenesis. Early blooming cultivars had PMCs present. Later-blooming cultivars only contained archesporial cells. PMCs were present in all cultivars by 22 Aug. Microspores were observed on 26 Sept. in all cultivars. This study contributes to a better understanding of male gametophyte development in hazelnut, which has increased our ability to correlate hazelnut pollen development with bloom phenology.

Ultrastructure of Urediniospore Development in Coleosporium ipomoeae as Revealed by High‐Pressure Freezing Followed by Freeze Substitution

Transmission electron microscopy of high‐pressure frozen/freeze‐substituted samples was used to study urediniospore development in Coleosporium ipomoeae in leaves of Ipomoea coccinea. The urediniospores of this species are morphologically identical to aeciospores. Our study showed that these spores developed in the same fashion as aeciospores. Spores formed in a basipetal fashion along with intercalary cells from a layer of binucleate sporogenous cells lining the base of each uredinium. High‐pressure freezing/freeze substitution yielded excellent preservation of ultrastructural details of the spore formation process until the point of secondary wall formation in maturing spores. When viewed with scanning electron microscopy, mature urediniospores were verrucose in appearance, except for a slightly reticulate area at the base of each spore. Both the reticulations and the small wartlike ornaments present on spore surfaces arose from tiny electron‐transparent deposits that formed on the outer surface of the ...

Temporal and spatial distribution of pectin epitopes in differentiating anthers and microspores of fertile and sterile sugar beet.

We studied the possible involvement of several pectin epitopes in anther differentiation and microsporogenesis in fertile and cytoplasmically male sterile sugar beets. The spatial and temporal distribution of five structural motifs were traced with a panel of monoclonal antibodies in six stages: premeiosis, meiotic prophase, young and mature tetrads, young and expanding microspores. The composition of the walls of sporogenous cells and meiocytes differed than that in the tapetum, as evidenced by the presence of alpha-Fuc(1-->2)-beta-Gal and alpha-(1-->5)-L-Ara epitopes binding CCRC-M1 and LM6 antibodies. At meiotic prophase, the meiocyte walls were additionally marked by the appearance of poorly methyl-esterified domains of homogalacturonan and of (1-->4)-beta-Gal residues, detected by JIM5 and LM5. Some constituents of the meiocyte wall which reacted with JIM5 and JIM7 persisted on the surface of the special callose sheath during tetrad development. In newly formed primexine and exine layers of tetrads and microspores, epitopes that were bound by JIM5, JIM7 and LM5 were abundant. No differences in the deposition or relative abundance of pectins were found between fertile and sterile anthers until microspore release from the callose. Later, at the time of abortion, sterile microspores had much larger amounts of epitopes detected by JIM5 than their fertile counterparts.

Stamen Structure and Function

Stamens are the male reproductive organs of flowering plants. They consist of an anther, the site of pollen development, and in most species a stalk-like filament, which transmits water and nutrients to the anther and positions it to aid pollen dispersal. Within the anther, male sporogenous cells

Monoplastidic cell in lower land plants

Summary: Many lower land plants have cells that contain a single plastid. A monoplastidic condition is observed in dividing vegetative cells of hornworts, the moss Takakia, and some microphyllophytes(Isoetes and Selaginella). Sporogenous cells and sporocytes are monoplastidic in all mosses, hornworts, Isoetes and Selaginella, as well as some species of liverworts, Lycopodium and Angiopteris. Additionally, spermatocytes in the taxa mentioned above are typically monoplastidic. In monoplastidic cells, the single plastid divides before nuclear division and divided plastids seem to serve as microtubule organizing centers of mitotic apparatus. Such a monoplastidic condition is considered to be a critical symplesiomorphy among bryophytes, lycopsid pteridophytes, and the coleochaetalean algae. In this paper, we summarize the results of recent investigations on monoplastidic cells of lower land plants.

Tapetum determinant1 is required for cell specialization in the Arabidopsis anther.

In flowering plants, pollen formation depends on the differentiation and interaction of two cell types in the anther: the reproductive cells, called microsporocytes, and somatic cells that form the tapetum. The microsporocytes generate microspores, whereas the tapetal cells support the development of microspores into mature pollen grains. Despite their importance to plant reproduction, little is known about the underlying genetic mechanisms that regulate the differentiation and interaction of these highly specialized cells in the anther. Here, we report the identification and characterization of a novel tapetum determinant1 (TPD1) gene that is required for the specialization of tapetal cells in the Arabidopsis anther. Analysis of the male-sterile mutant, tpd1, showed that functional interruption of TPD1 caused the precursors of tapetal cells to differentiate and develop into microsporocytes instead of tapetum. As a results, extra microsporocytes were formed and tapetum was absent in developing tpd1 anthers. Molecular cloning of TPD1 revealed that it encodes a small protein of 176 amino acids. In addition, tpd1 was phenotypically similar to excess microsporocytes1/extra sporogenous cells (ems1/exs) single and tpd1 ems1/exs double mutants. These data suggest that the TPD1 product plays an important role in the differentiation of tapetal cells, possibly in coordination with the EMS1/EXS gene product, a Leu-rich repeat receptor protein kinase.

Morphological development of anthers induced by the dimorphic smut fungus Microbotryum violaceum in female flowers of the dioecious plant Silene latifolia.

When inoculated with the dimorphic smut fungus Microbotryum violaceum (Pers.) G. Deml and Oberwinkler, the female flower of the dioecious plant Silene latifolia (Miller) E.H.L. Krause develops anther-like structures filled with spores instead of pollen grains. Using natural scanning electron microscopy, Nomarski interference microscopy, and fluorescence microscopy, we investigated the morphological modifications of the host plant resulting from this parasitism and the localization of smut hyphae in the flower bud. Flowers of infected plants lasted significantly longer than those of healthy plants, probably because the infection strengthened floral organs, such as the flower base and the anther filaments. Smut hyphae were observed throughout all organs of the young flower buds of infected plants, including sepals, petals, stamens, and pistil primordia. In healthy female flowers, anthers initiated sporogenous cell formation, but lacked parietal cell layers. By contrast, the parietal cell layers of infected female flowers differentiated into tapetal tissue, middle cell layers, and endothecial layers, as in the anthers of healthy male flowers. Smut spore formation in the infected anther was initiated in intercellular regions between the sporogenous cells, resulting in degeneration of premature sporogenous cells, tapetal tissue, and middle cell layers. The development of the endothecial layers and epidermis in the infected anther were morphologically normal.

Comparative development of aseptate and septate anthers of Annonaceae

We compared anther development in 13 genera and 15 species of Annonaceae to document the nature and development of anther septa. In aseptate anthers, all sporogenous initials proceed to sporogenesis and meiosis. In septate anthers, a small number of sporogenous initials, in a discontinuous distribution pattern, differentiate into sporogenous cells; the remaining initials become sterile and form cellular septa that partition each anther lobe into multiple sporangial chambers. In species where the septum is 1-2 cell layers thick, the entire septum becomes tapetal (T-type septa) and breaks down before anther dehiscence. In species in which the septum is three or more cell layers thick, only the layer in direct contact with the sporogenous cells becomes tapetal, and the remaining cells become parenchymatous (P-type septa). These thicker P-type septa are sometimes visible in dehisced anthers. Both types are homologous in ontogeny and are highly associated with the production of compound pollen. We propose that the evolution of anther septation in Annonaceae was mainly driven by the requirement for highly efficient nutrient and physical support to the development of large, compound pollen units.

Expression of female sterility in alfalfa (Medicago sativa L.)

Female sterility associated with the presence of callose in the nucellus at anthesis was studied in an F1 progeny of two alfalfa plants displaying 5 and 81% ovule sterility. Transgressive segregation was observed and 100% sterile plants were obtained. Two of the sterile plants were used for cytological analyses on sectioned and stain-cleared whole ovules, in comparison to a 100% fertile full sib plant. The first sign of sterility was callose deposition in the nucellus cell walls surrounding the sporogenous cells of the young ovules. At the same stage, no trace of callose was present in ovule primordia of the fertile plant. Megaspore mother cells differentiated in both fertile and sterile ovules and meiosis was initiated, as indicated by chromatin patterning typical of a zygotene stage. However, meiosis was never completed in the sterile plants. In the control, callose was deposited around the meiocyte and as sects between the cells of the dyads and tetrads during meiosis, and disappeared after the completion of meiosis; an embryo sac developed and female fertility was normal. In the sterile ovules, some nucellus cells enlarged and callose accumulation continued forming thick deposits. At anthesis, the sterile ovules lacked an embryo sac and showed massive callose accumulation in the nucellus. Male fertility was normal in female-sterile plants, thus a female-specific arrest of sporogenesis appears to be the cause of sterility. Pistil development was aberrant in some sterile genotypes, even with arrested pistil growth in early flower buds.

Synthesis and Deposition of Callose in Anthers and Ovules of Meiotic Mutants of Maize (Zea mays)

The pattern of callose deposition was studied in anthers and ovules of three meiotic mutants of Zea mays L. The synthesis of the callose wall in sporogenous cells was related to their transfer to meiotic division.

Molecular and biochemical characterization of a calcium/calmodulin-binding protein kinase from rice.

A Ca2+/calmodulin (CaM)-binding protein kinase from rice ( Oryza sativa ), OsCBK, has been characterized that lacks Ca2+-binding EF hands and has Ca2+/CaM-independent autophosphorylation and substrate-phosphorylation activity. OsCBK has all 11 subdomains of a kinase catalytic domain and a putative CaM-binding domain, and shares high identity with Ca2+-dependent-protein-kinase ('CDPK')-related protein kinases in plants. OsCBK bound CaM in a Ca2+-dependent manner as previously reported for Ca2+/calmodulin-dependent protein kinases in animals, but autophosphorylation and phosphorylation of histone IIIs were Ca2+/CaM-independent. Surface plasmon resonance analysis showed that OsCBK specifically bound CaM with high affinity ( K (D)=30 nM). Capillary electrophoresis showed that phosphorylation of OsCBK occurred on serine and threonine residues. These data show that OsCBK is a serine/threonine protein kinase that binds Ca2+/CaM, but whose enzymic activity is independent of Ca2+/CaM. In situ hybridization showed that OsCBK is expressed in reproductive and vegetative tissues of rice and shows temporal and spatial changes during plant growth and development. OsCBK is highly expressed in zones of cell division and it is particularly abundant in sporogenous cells of the anther at meiosis.

EXS, a Putative LRR Receptor Kinase, Regulates Male Germline Cell Number and Tapetal Identity and Promotes Seed Development in Arabidopsis

Background: Plant germlines arise late in development from archesporial initials in the L2 layer of the anther and ovule primordia. These cells generate a radially symmetrical array of tissues that, in the Arabidopsis anther, comprises a core of sporogenous cells (meiocytes) and the enveloping tapetum, middle cell, and endothecium layers. The putative transcription factor NZZ/SPL[1, 2] is required for the specification of archesporial cells, but nothing is known of how their number is regulated, or what controls cell fate in the lineages they generate. Here, we report detailed characterization of extra sporogenous cells (exs), a male sterile mutant that generates extra meiocytes but lacks tapetal and middle cell layers [3, 4].Results: We identified the EXS locus by map-based cloning and found it to encode a putative LRR receptor kinase. In the anther, an increased number of L2 layer cells assume an archesporial fate and divide to generate a larger number of sporogenous cells. In seeds, the exs mutation results in smaller embryonic cells, delayed embryo development, and smaller mature embryos. Consistent with the observed phenotype, EXS is expressed in the inflorescence meristem, floral apices, anthers, and in developing seeds.Conclusions:EXS regulates the number of cells that divide in the L2 layer of the anther, and thus the number of functional male archesporial initials. In the young seed, EXS affects cell size in the embryo and the rate at which it develops. The apparently contrasting roles of EXS in the anther and embryo suggest that signaling through the EXS receptor kinase is a feature of a number of regulatory pathways in Arabidopsis.

Embryology of Zippelia begoniaefolia (Piperaceae) and its systematic relationships

Microsporogenesis and embryology of the monotypic Zippelia (Z. begoniaefolia) Blume (Piperaceae) is described for the first time to assess its systematic relationships. The formation of the anther wall is of Basic Type such that the anther wall, consisting of an endothecium with fibrous thickenings, two middle layers, and a glandular septum with 2-nucleate cells, is derived from a primary parietal layer. Simultaneous cytokinesis follows meiosis of the microspore mother cell thence forming a tetrahedral tetrad of microspores. The single basal ovule is orthotropous, crassinucellate and bitegmic but only the inner integument forms the micropyle. The sporogenous cell of the nucellus functions directly as a megaspore mother cell. A coenocyte with four nuclei forms after meiosis of the megaspore mother cell. The formation of the embryo sac is tetrasporic ab initio and is of, or similar to, the Drusa Type of embryo sac in which the nuclei of the coenocyte undergo two successive mitoses and forms a 16-celled or 16-nucleate embryo sac that is ovoid in shape. The embryo sac has an egg apparatus consisting of an egg cell and two synergids (but one of the latter is less discernable). Two polar cells occur just beneath the egg apparatus and 11 antipodal cells or nuclei are arranged along the lower part of the inner wall of the embryo sac. They are linked by threads of cytoplasm. The two polar cells are separated or fused before fertilization. A large primary endosperm nucleus with many nucleoli, which resulted from the fertilized polar cells and with the participation of antipodal cells, divides into a free nuclei stage. The free nuclei are arranged along the lower part of the inner wall of the embryo sac or rarely assemble at the central part. The development of endosperm is thus of the Nuclear Type. The zygote remains undivided and fails to develop even when the seed is nearly mature. Frequently, the zygote and the endosperm abort later and leave an empty chamber in the top part of the seed. Most of the seed content is starchy perisperm. Only the inner integument forms the seed coat and the pericarp develops glochidiate hairs (anchor-like hairs) when the endosperm begins to develop. By comparison with the other piperaceous taxa using embryological and botanical features, Zippelia is referred to as a basal taxon and a more isolated evolutionary line or a blind branch in the Piperaceae.

Anatomical and Histochemical Studies on Pollen Degeneration in CMS Helianthus petiolaris Nutt.

The purpose of this microscopic histochemical study was to establish the developmental sequence leading to sterility in a cytoplasmic male sterile Helianthus petiolaris line. Histochemical studies showed that CMS anthers, in contrast to fertile anthers, lacked carbohydrate storage at sporogenous stage. CMS sporogenous cells were thick-walled and ascorbic acid-rich. Subsequent developmental pattern was not uniform in CMS anthers. The variations were found in different anthers and in different locules of the same anther. Two categories of CMS anthers lacked callose deposition and showed premature disintegration of tapetal cell layer, which lead to precocious formation of periplasmodium. In one type, CMS anthers contained naked microspores. In another type, coenocytic microspores were formed. The microspores lacked reserve metabolites. In the third category of CMS anthers normal callose deposition and tetrad formation were observed. In this category callose deposition and hypertrophied tapetal cell layer persisted until mature stage of the anther. Results in all 3 categories of CMS anthers link tapetal abnormality to male sterility.