SECRETORY CELLS OF LILY PISTILS. I. FINE STRUCTURE AND FUNCTION

The fine structure of the cells which line the canal of Lilium longiflorum pistils confirms the secretory function which has been ascribed to them. The cells differ in structure from the secretory cells which cover the stigma surface and are therefore referred to as canal cells rather than stigmatoid cells. Their most striking feature is an elaborate wall, 8–14 μ in maximum depth, on the side facing the canal, which with associated structures, we term the secretion zone. Pollination, which triggers chemotropic activity in the style and secretory activity in the canal cells, is not correlated with marked changes in the fine structure of the canal cells. The canal cells appear to fit well into that category which Gunning and Pate have termed “transfer cells.”     

Incorporation of Label into Pollen Tube Walls from Myoinositol-labeled Lilium longiflorum Pistils

Compatible and incompatible pollen tubes growing on detached Lilium longiflorum pistils which had been prelabeled with myoinositol-U-(14)C take up a portion of the label and utilize it for biosynthesis of tube wall substance. The label is transferred from pistil to pollen tubes apparently via the secretion products (exudate) of the pistil. The exudate thus appears to have a major nutritional role in pollen tube growth in vivo.     

SECRETORY CELLS OF LILY PISTILS. II. ELECTRON MICROSCOPE CYTOCHEMISTRY OF CANAL CELLS

The secretory cells which line the canal of Lilium longiflorum pistils possess, on the side facing the canal, an elaborate wall which, with associated structures, Rosen and Thomas (1970) termed the “secretion zone.” We examined the secretion zone in the electron microscope following treatment of excised pistil slices with extraction procedures which remove pectin, hemicellulose, cellulose, lipid, or protein. The outer, fibrillar wall (layer 1) of the secretion zone contains protein, pectin, and cellulose. Internal to layer 1 is a granular-fibrillar wall (layer 2) several microns thick. It consists of outer and inner regions which can be distinguished from each other cytochemically. The granular component is composed of pectin which is not esterified with methyl groups and which may be complexed with protein. The short, randomly dispersed microfibrils of layer 2 were sensitive to procedures which dissolve cellulose. The extraction procedures did not reveal the chemical nature of the “osmiophilic islands” of layer 2. Paramural body membranes appear to be composed of glycoprotein and may function in secretion by serving as sites of pinocytic interchange at the plasmalemma. The origin of stigmatic exudate and the release of canal cell secretion product are discussed.     

The Nutritional Role of Pistil Exudate in Pollen Tube Wall Formation in Lilium longiflorum: II. Production and Utilization of Exudate from Stigma and Stylar Canal

Detached pistils of Lilium longiflorum were labeled with d-glucose-U-¹⁴C 24 hours after anthesis and then sampled for the next 6 days to determine the appearance of label into exudate from the stylar canal and the stigmatic surface of the pistil. Results were obtained with unpollinated cv. Ace pistils and pollinated cv. Ace pistils, selfed or crossed (cv. Croft pollen). Limited data were also obtained on cv. Croft pistils, selfed or crossed (cv. Ace pollen).     

CHEMOTROPIC ACTIVITY AND THE PATHWAY OF THE POLLEN TUBE IN LILY,

A study of the pollen tube pathway in Lilium leucanthum var. centifolium and in L. regale reveals that the entire pathway from stigma to ovule is lined with cytologically unique stigmatoid cells. Assays for chemotropic activity of tissues and exudates along the pathway of pollinated or unpollinated pistils showed that onset of chemotropic activity progressed basipetally (and, when pollinated, in advance of the pollen tubes), commencing at the stigma 3-5 days before anthesis and appearing in the ovules 1-2 days after anthesis. Activity persists about 10 days in ovules of pollinated pistils and for 14-16 days in ovules of non-pollinated pistils. Attempts to localize the source of the chemotropic factor showed that gynoecial tissues bearing stigmatoid cells are chemo-tropically active while slices of style or ovary wall lacking stigmatoid cells are inactive. When ovules were sliced transversely and the micropylar and chalazal halves assayed, only the micropylar half showed activity. We suggest that the ovules and the stigmatoid tissue along the pollen tube pathway are the sources of the chemotropic factor responsible for the directional growth of the pollen tube.     

Stigma development in Nicotiana tabacum. Cell death in transgenic plants as a marker to follow cell fate at high resolution

Pistil development was studied in transgenic tobacco plants in which the stigma is ablated by expression of a stigma-specific cytotoxic gene. These plants offer a tool to investigate the process of differentiation of the secretory zone, in that cell death caused by barnase activity provides a marker to follow cell fate at high resolution. After fusion of the carpel walls in the region most distal from the ovary, the epidermal cells begin to divide in both wild-type and stigmaless plants. Divisions of the L1 layer of the pistil are immediately followed by the morphogenetic events that lead to three different cell types: rounded-angular cells showing an equal number of anti- and periclinal divisions, cells that are more oblong forming the transition zone, and the square cells of the transmitting tissue dividing mostly anticlinally with respect to the original carpel wall. In the stigmaless plants, cell death caused by the expression ofSTIG 1-barnase begins at stage –1 and proceeds gradually, but is always associated with round epidermal cells and with angular-rounded cells underneath them. Studies at the ultrastructural level show that cell death caused by barnase activity occurs first in solitary cells and gradually extends to groups of cells.In situ hybridizations using the STIG 1 RNA probe in wild-type pistils confirm these results. Most likely, the cells in whichSTIG 1 is expressed are those that have just differentiated into the secretory cell type. Our results indicate that the transition zone or neck is autonomously differentiated from the secretory zone and the transmitting tissue. Furthermore, our results indicate that in both wild-type and stigmaless pistils secretion of lipids most likely occurs through the plasmodesmata. This observation suggests that bulk transport can occur via plasmodesmata.     

湖北黄精大小孢子发生及雌雄配子体发育

【目的】研究湖北黄精花部形态结构特征和大小孢子发生及雌雄配子体发育过程,以丰富黄精属植物的生殖生物学理论,为进一步开展湖北黄精的品种选育提供依据。【方法】以不同发育时期的湖北黄精花芽为试验材料,用显微观察法观察花部形态结构特征,石蜡切片技术对单花雌雄蕊进行切片观察。【结果】湖北黄精的花被为白色或淡黄绿色,花被筒近喉部稍缢缩;具6枚雄蕊,花丝下端与花被合生,花药开裂方式为纵裂;雌蕊子房上位,3心皮,花柱与子房等长。湖北黄精花药壁由4层细胞组成,成熟的绒毡层具多核,绒毡层发育类型为分泌型;小孢子母细胞减数分裂为连续型,四分体呈左右对称型排列,成熟花粉粒为2-细胞型;存在小孢子发育不同步的现象。雌蕊胚珠具双珠被、厚珠心;四分体呈直线型排列,胚囊发育类型为蓼型;存在双胚囊胚珠现象。在雄蕊的花药壁和雌蕊的子房壁都观察到有束状草酸钙针晶。【结论】湖北黄精雌雄蕊具有较原始的发育特征,虽然在发育过程中都存在异常现象,但雄蕊最终能形成正常的雄配子体,雌蕊低频率的双胚囊现象对总体受精结果影响很小。湖北黄精杂交育种可以选择花药开裂前一时期的花粉,花药壁和子房壁观察到的束状草酸钙针晶无法作为湖北黄精物种鉴定的...     

INFLUENCE OF THE PISTIL ON POLLEN TUBE KINETICS IN PEACH (PRUNUS PERSICA)

Pollen tube growth has been studied in peach and has been related to changes in the pistil structures which the pollen tube has to traverse in its way from the stigma down to the ovule. Growth of the pollen tubes along the pistil is not continuous. While pollen tubes reach the base of the style 7 days after pollination, fertilization does not take place until 12 days later. Pollen tubes stop for 5 days at the top of the obturator and they further stop for 3 days before entering the ovule. The pollen tube growth is heterotrophic; starch, present all along the pistilar tract at anthesis, vanishes as the pollen tubes pass by. Discontinuous pollen tube growth appears to be controlled by the pistil. At anthesis the pistil is not fully matured. Maturation of the pistil implies a number of secretory processes that occur in a basipetal way starting from the stigma down to the style and ending in the ovule. Some of these secretions at the stigma and the style are triggered by pollination; others appear to be a maturative stage of the pistil and are produced in a discrete way. The fact that the pollen tube depends on these secretions together with the fact that these secretions are not continuously produced confer upon the pistil a role of controlling pollen tube kinetics and point out that, for a successful fertilization, male gametophyte development and pistil maturation need to by synchronized.     

Fine structure of postgonopodial glands of a myriapod Glomeris marginata (Villers)

The postgonopodial gland of the myriapod Glomeris marginata (Villers), which produces a pheromone, is an integumentary gland comprising numerous functional secretory units. Each secretory unit consists of two proximal secretory cells, an intermediary cell lacking secretory characteristics and a canal cell surrounding the canal, which is secretory in nature. Secretory proximal cells exhibit a zone of small channels originating from invaginations of the plasma membrane and through which secreted material is released. Apposing each invagination of these cells is a corresponding invagination of the intermediary cell: the two units in the centre of the intermediary cell join another which communicates with the canal. Secretion produced by the latter passes through the canal wall and blends with secretion of the two proximal cells. The most striking feature of all these cells is the abundance of tubules and fibrils in the small canal zone in the proximal cells, which also exhibits a centriole; in the intermediary cell around cytoplasmic membrane invaginations where a diplosome is present, and in almost the entire canal cell.     

Characterization of the pollen growth transition in self-incompatible <Emphasis Type="Italic">Petunia inflata</Emphasis>

In Petunia inflata, as in other species that shed bicellular pollen, early pollen tube growth in the pistil is slow, then increases 2- to 5-fold depending on the genotype of the female parent. We refer to the time point at which pollen tubes enter the accelerated phase of growth as the pollen growth transition (PGT). Here, we present evidence that pre-PGT and post-PGT growth are quantitatively and qualitatively different, and that the PGT is triggered when pollen tubes reach the transition zone (TZ) below the stigma. The capacity of various pistil zones to precipitate the PGT was tested through 'stump' pollinations: varying lengths of the pistil apex were excised, the cut surface of the remaining pistil (the stump) coated with stigmatic exudates then dusted with compatible pollen. Pollen applied to TZ tissues entered the PGT earlier than pollen growing in intact control pistils; the PGT was delayed in stylar stumps, largely because of delayed germination and reduced pre-PGT growth. In immature pistils, the PGT was delayed by several hours relative to its onset in mature pistils. The PGT fails to occur in pollen cultured in vitro. Collectively, the data suggest that pollen tubes become competent to enter the PGT when they reach a critical size, but the physicochemical environment of the transmitting tissue is necessary for triggering the cellular changes that result in accelerated growth. An analysis of the distribution of pollen tube tips before and after the PGT suggests that pollen competition is most intense during the pre-PGT phase.     

THE EFFECTS OF GIBBERELLIC ACID AND A GROWTH RETARDANT ON OVULE FORMATION AND GROWTH OF EXCISED PISTILS OF NIGELLA RANUNCULACEAE

The effects of gibberellic acid (GA₃) and the growth retardant AMO-1618 on ovule formation in excised pistils of Nigella sativa L. were studied by sterile culture techniques. Gibberellic acid promoted pistil growth and inhibited ovule formation. The role of endogenous gibberellins in ovule formation and pistil growth was investigated by adding AMO to the basal medium. Both pistil lengths and ovule formation were reduced significantly with increasing concentrations of AMO. The addition of low concentrations of GA₃ to the medium restored pistil growth but did not reverse the inhibitory effect of AMO on ovule formation. The addition of kinetin or indoleacetic acid (IAA) to the medium containing AMO had no effect on pistil lengths. However, with the addition of 10⁻⁷ m kinetin, the number of ovules in pistils was increased but not to the levels found in pistils grown in the absence of AMO.     

The Nutritional Role of Pistil Exudate in Pollen Tube Wall Formation in Lilium longiflorum: I. Utilization of Injected Stigmatic Exudate

A quantity of labeled stigmatic exudate, collected from detached Lilium longiflorum (cv. Ace) pistils labeled with d-glucose-1-¹⁴C, was fractionated on Sephadex G-100 and the polysaccharide component, G-100-I, was injected into the hollow styles of unlabeled detached pistils (cv. Ace) which had been removed on the day after anthesis from the plant. Injected pistils were immediately cross-pollinated with L. longiflorum (cv. No. 44) pollen. Eighty-four hours later, pistils were dissected to recover the pollen tubes, expended exudate, and labeled tissues of the stigma and style. Distribution of label revealed that at least 25% of the carbohydrate substance in excised pollen tubes was derived from G-100-I. The composition of expended exudate adhering to pollen tubes, of pollen tube cytoplasm, and of pollen tube walls suggests that utilization of exudate by growing pollen tubes involves uptake and incorporation into pollen tube cytoplasm of exudate polysaccharide fragments followed by extensive metabolism of at least a portion of the incorporated carbohydrate prior to its utilization for pollen tube wall biosynthesis. Results suggest the presence of at least two polysaccharide components in G-100-I, one which resists major degradation following injection into the style and another which undergoes measurable degradation both before and after entry into the pollen tube.     

Ultrastructural and physiological differences between buds and mature flowers of Petunia hybrida prior to and following pollination

The structural events accompanying the maturation of the pistil of Petunia hybrida have been studied in detail, together with the changes in the protein spectrum of the transmitting tissue that occur over this period. These events have been considered in terms of the acquisition of the self-incompatibility response, which occurs while the pistil is enclosed in the bud. Apart from several minor differences, the young pistils differ only from the mature in that their transmitting tissue cells fail to respond to pollination by undergoing characteristic ultrastructural changes. Data from electrofocusing indicates that several proteins, mobilised in the mature transmitting tissue some three hours after pollination, are absent from bud pistils. It is proposed that the pollination-stimulated release of certain polypeptides, believed to be involved in the self-incompatibility response, does not take place in young pistils. These observations are considered with reference to currently-accepted models of the operation of the self-incompatibility mechanism in Petunia.     

倒地铃花的形态分化与花药结构研究

利用体视显微镜、半薄切片和超薄切片法对倒地铃(Cardiospermum halicacabum Linn.)雄花和假两性花开花过程及花药发育过程进行了观察和比较研究。结果显示:(1)花蕾发育早期,倒地铃雄花和假两性花的花蕾形态没有区别;花蕾发育后期,雄花雌蕊退化,假两性花雌蕊继续发育,花蕾外部形态出现差异;开花时雄花花药开裂,假两性花花药不开裂。(2)倒地铃雄花和假两性花均具四室花药,呈蝶形;花药壁细胞从外到内依次是表皮、药室内壁、中层(2层)和绒毡层;花药壁发育为基本型,绒毡层为单核分泌型,四分体为四面体型,花粉粒两核;开花时雄花和假两性花中层都有残留;小孢子液泡化时,绒毡层开始降解,两核花粉粒时,假两性花绒毡层降解较快。(3)雄花药室内壁次生加厚完全,裂口区发育,连接同侧花粉囊的连接组织降解,花药开裂;假两性花药室内壁次生加厚不完全,具唇形细胞,药隔细胞壁未降解,同侧花粉囊未连通,花药四室,不开裂;假两性花成熟花粉粒细胞质稀少,内壁不完整。本研究结果表明,倒地铃的雄花是由两性花在发育早期雌蕊停止发育形成的,假两性花则由两性花在发育晚期雄蕊功能退化造成的。     

Inositol Metabolism in Plants: VII. Distribution and Utilization of Label from Myoinositol-U-14C and -2-3H by Detached Flowers and Pistils of Lilium longiflorum1

When detached flowers or isolated pistils of Lilium longiflorum are given myoinositol-U-(14)C or -2-(3)H as dilute solution through the severed pedicel, label is quickly distributed by the vascular system. In the case of pistils, a pattern of labeling in ovary, style, and stigma is obtained which indicates that products of myoinositol metabolism are utilized in the biosynthesis of exudate (secretion product) of the stigma and style as well as for components of pistil cell walls. Pollination had no discernible effect on labeling pattern.     

溶液流动对等电溶菌酶晶体生长的影响

本文对等温自由生长和强制性溶液生长的等电溶菌酶的晶体形态进行了研究,发现这些形态变化与溶液相的流动密切相关,指出生物晶体生长停止是由于生长晶体周围的溶质贫乏造成的;通过某些手段减薄或消除这一溶质贫乏区,就可以保证晶体的持续生长。本文的研究对改善大尺寸晶体的生长提供了一条途径。     

Citrus Tissue Culture : AUXINS IN RELATION TO ABSCISSION IN EXCISED PISTILS

An in vitro bioassay for chemicals that affect Citrus abscission was used to identify three inhibitors of stylar abscission in lemon pistil explants incubated on defined nutrient media. The three inhibitors (picloram, 4-chlorophenoxyacetic acid, and 3,5,6-trichloropyridine-2-oxyacetic acid) are all auxins, and the most potent of them (i.e. picloram) was found to be at least 10 times more active in the bioassay than 2,4-dichlorophenoxyacetic acid. Picloram (2 micromolar) also was shown to be effective in inhibiting stylar abscission in pistil explants from other Citrus cultivars such as mandarin, Valencia, and Washington navel oranges and grapefruit. To study the physiology of auxins active as abscission inhibitors versus inactive auxins in lemon pistils, the transport and metabolism of [1-¹⁴C]-2,4-dichlorophenoxyacetic acid was compared with that of [2-¹⁴C]indole-3-acetic acid, which is without effect in the bioassay over the range from 0.1-100 micromolar. Insignificant quantities of labeled indole-3-acetic acid and/or labeled derivatives were found to reach the presumptive zone of stylar abscission under the test conditions. Labeled 2,4-dichlorophenoxyacetic acid and/or labeled derivatives also were transported slowly through pistils, but some radioactivity could be detected in the stylar abscission zone as early as 24 hours after the start of incubation. Extensive conversion of [2-¹⁴C]indole-3-acetic acid to labeled compounds tentatively considered to be glycoside and cellulosic glucan derivatives was found with the use of solvent extraction methodology. A significantly smaller percentage of the radioactivity in pistils incubated on [1-¹⁴C]-2,4-dichlorophenoxyacetic acid was found in fractions corresponding to these derivatives. Both transport and metabolism appear to be important factors affecting the activity of auxins as abscission inhibitors in the bioassay.     

SECRETORY CAVITY DEVELOPMENT IN GLANDULAR TRICHOMES OF CANNABIS SATIVA L. (CANNABACEAE)

Development of the secretory cavity and formation of the subcuticular wall of glandular trichomes in Cannabis sativa L. was examined by transmission electron microscopy. The secretory cavity originated at the wall-cuticle interface in the peripheral wall of the discoid secretory cells. During the presecretory phase in development of the glandular trichome, the peripheral wall of the disc cells became laminated into a dense inner zone adjacent to the plasma membrane and a less dense outer zone subjacent to the cuticle. Loosening of wall matrix in the outer zone initiated a secretory cavity among fibrous wall materials. Membrane-bound hyaline areas, compressed in shape, arose in the wall matrix. They appeared first in the outer and subsequently in the inner zone of the wall. The membrane of the vesicles, and associated dense particles attached to the membrane, arose from the wall matrix. Hyaline areas, often with a conspicuous electron-dense content, were released into the secretory cavity where they formed rounded secretory vesicles. Fibrous wall material released from the surface of the disc cells became distributed throughout the secretory cavity among the numerous secretory vesicles. This wall material was incorporated into the developing subcuticular wall that increased five-fold in thickness during enlargement of the secretory cavity. The presence of a subcuticular wall in the cavity of Cannabis trichomes, as contrasted to the absence of this wall in described trichomes of other plants, supports a polyphyletic interpretation of the evolution of the secretory cavity in glandular trichomes among angiosperms.     

NUTRITIONAL REQUIREMENTS FOR OVULE FORMATION IN EXCISED PISTILS OF NIGELLA

The nutritional requirements for ovule formation in Nigella saliva L. were investigated by growing excised pistils on defined media. Pistils grown on a medium containing the minerals of Murashige and Skoog produced significantly more ovules than on a medium containing the minerals of Bilderback. When the nitrogen, sulfate, and phosphate of the Bilderback medium were adjusted to levels comparable to those of the Murashige and Skoog medium, a similar number of ovules was formed. The effect of different forms and concentrations of nitrogen on ovule formation and pistil growth was investigated. High concentrations of nitrate (40 mil) favored pistil growth and ovule formation, but comparable levels of ammonium were toxic. When ammonium at concentrations above 10 mM was added to nitrate media, ovule formation was inhibited. A medium containing low concentrations of ammonium (10 mM) and nitrate (5 mM) supported more ovule formation and pistil growth in young pistils than a low-nitrate (5 mM) medium without ammonium. However, ovule formation on a medium containing 10 mM ammonium and 5 mM nitrate was significantly less than on a medium containing only 15 mM nitrate. Low concentrations of organic nitrogen in the form of α-alanine (1 mM) and γ-aminobutyric acid (5 mM) supported ovule formation and pistil growth similar to a high nitrate medium.