A glycine-rich protein that facilitates exine formation during tomato pollen development

Formation of the unique and highly diverse outer cell wall, or exine, of pollen is essential for normal pollen function and survival. However, little is known about the many contributing proteins and processes involved in the formation of this wall. The tomato gene LeGRP92 encodes for a glycine-rich protein produced specifically in the tapetum. LeGRP92 is found as four major forms that accumulate differentially in protein extracts from stamens at different developmental stages. The three largest molecular weight forms accumulated during early microspore development, while the smallest molecular weight form of LeGRP92 was present in protein extracts from stamens from early microsporogenesis through anther dehiscence, and was the only form present in dehisced pollen. Light microscopy immunolocalization experiments detected LeGRP92 at only two stages, late tetrad and early free microspore. However, we observed accumulation of the LeGRP92 at the early tetrad stage of development by removing the callose wall from tetrads, which allowed LeGRP92 detection. Transmission electron microscopy confirmed the LeGRP92 accumulation from microspore mother cells, tetrads through anther dehiscence. It was observed in the callose surrounding the microspore mother cells and tetrads, the exine of microspores and mature pollen, and orbicules. Plants expressing antisense RNA had reduced levels of LeGRP92 mRNA and protein, which correlated to pollen with altered exine formation and reduced pollen viability and germination. These data suggest that the LeGRP92 has a role in facilitating sporopollenin deposition and uniform exine formation and pollen viability.     

柴胡大、小孢子发生及雌、雄配子体发育

以柴胡(Bupleurum chinense)为研究对象, 运用石蜡切片技术对其大、小孢子发生及雌、雄配子体发育进行了研究。结果表明: 柴胡花药具4个药室, 花药壁由表皮、药室内壁、中层和绒毡层4层细胞构成, 花药壁发育为双子叶型, 腺质绒毡层。小孢子母细胞减数分裂的胞质分裂为同时型, 产生正四面体型小孢子。成熟花粉三细胞型。胚珠倒生型, 单珠被, 薄珠心。大孢子母细胞常为一个雌性孢原直接发育而成, 大孢子四分体呈线型或T型排列, 多数情况为合点端一个大孢子分化为功能大孢子, 由功能大孢子发育为蓼型成熟胚囊。在胚囊发育过程中, 珠被内表皮细胞特化成珠被绒毡层。同一朵花中, 雄蕊先熟, 记 录了花蕾大小及雌、雄配子体发育的对应关系。     

柴胡大、小孢子发生及雌、雄配子体发育

以柴胡（Bupleurum chinense）为研究对象,运用石蜡切片技术对其大、小孢子发生及雌、雄配子体发育进行了研究。结果表明：柴胡花药具4个药室,花药壁由表皮、药室内壁、中层和绒毡层4层细胞构成,花药壁发育为双子叶型,腺质绒毡层。小孢子母细胞减数分裂的胞质分裂为同时型,产生正四面体型小孢子。成熟花粉三细胞型。胚珠倒生型,单珠被,薄珠心。大孢子母细胞常为一个雌性孢原直接发育而成,大孢子四分体呈线型或T型排列,多数情况为合点端一个大孢子分化为功能大孢子,由功能大孢子发育为蓼型成熟胚囊。在胚囊发育过程中,珠被内表皮细胞特化成珠被绒毡层。同一朵花中,雄蕊先熟,记录了花蕾大小及雌、雄配子体发育的对应关系。     

一品红雄配子体发育研究

一品红花药来源于雄蕊原基,花药由表皮(1层)、药室内壁(1层)、中层(1层)、绒毡层(1层)及造胞细胞组成,花药四室,药壁发育为双子叶型。小孢子发生和雄配子体发育是经由小孢子母细胞减数分裂形成四分体,该四分体胞质分裂为同时型,四分体排列为四面体型,小孢子再经有丝分裂形成2-核花粉。花药壁层的变化是表皮在花药成熟期消失,中层在四分体时消失,药室内壁在花药成熟期形成柱状纤维层。绒毡层在单核小孢子期径向伸长,有双核或多核,另外有的绒毡层细胞形成横隔或类胎座;进入2-核花粉期,绒毡层细胞分泌颗粒物进入药室,为非典型腺质绒毡层;进入成熟期绒毡层消失。同时观察到花药发育异常现象。     

楸树大小孢子发生与雌雄配子体发育的研究

运用石蜡切片法和整体透明法对楸树(Catalpa bungei C.A.Meyer)大、小孢子发生及雌、雄配子体发育过程进行了研究.结果表明:楸树可育雄蕊2枚,花药4室,药壁发育属双子叶型,腺质异型绒毡层.小孢子母细胞减数分裂为同时型,四分体后小孢子不分离形成正四面体型四合花粉,偶有左右对称型和十字交叉型.成熟花粉为二细胞型,无萌发孔.子房上位,2室,中轴胎座,胚珠多数,倒生,单珠被,薄珠心,具珠被绒毡层.单孢原直接发育为大孢子母细胞,四分体线形排列,合点端大孢子发育为功能大孢子,胚囊发育为蓼型.雄蕊发育早于雌蕊,花开后雌、雄蕊趋于同熟.研究认为:虽然楸树雌、雄蕊发育过程中均存在一定比例的败育,但其花而不实"并非雄性或雌性不育所致.推测与其授粉受精和胚后发育有关.     

长春蒲公英雄性不育株形态学观察与败育细胞学研究

在长春蒲公英(Taraxacum junpeianum Kitam.)株群中发现雄性不育现象,为研究其败育机理及特点,探寻其不育基因,采用形态观察法、石蜡切片技术和染色体压片法,对长春蒲公英野生型及其雄性不育株的花药发育过程和花粉母细胞减数分裂过程进行了观察。结果表明:(1)长春蒲公英雄性不育株花药中部发红、干瘪、无花粉散出。与野生型比较,雄性不育株雄蕊更短,子房更窄,种子形态更加狭长;(2)长春蒲公英雄性不育株败育时期为四分体到单核小孢子前期,败育方式为小孢子自身异常发育,绒毡层异常分解,互相粘连败育;(3)长春蒲公英雄性不育株花粉母细胞减数分裂二分体时期出现落后微核,随后产生极少四分体,并且四分体产生大量染色体桥,小孢子营养物质流失,彻底败育。因此,长春蒲公英雄性不育株败育彻底、稳定,并且有种的特点。小孢子自身异常发育和绒毡层异常分解是导致败育的主要原因。     

雄性不育与可育楸树花发育的细胞学比较研究

楸树（Catalpa bungei C.A.Meyer.）属紫葳科(Bignoniaceae)梓树属(Catalpa),落叶乔木,是我国特有的珍贵优质用材树种。本文用石蜡切片法对可育株和雄性不育株楸树的大、小孢子发生及雌、雄配子体发育过程进行了详细地比较观察。结果表明：可育株和不育株楸树雌蕊的发育基本相同,胚珠倒生,薄珠心,单珠被,胚囊发育为蓼型。可育株雄蕊花药四室,药隔薄壁组织发达;异型绒粘层,由药壁绒粘层和药隔绒粘层组成;花药壁表皮细胞在小孢子母细胞减数分裂前后开始径向伸长加厚,直到花药开裂并不降解,这可能与花药开裂有关;成熟花粉为四合花粉。雄性不育株花药的早期发育到次生造胞细胞时期与可育雄蕊的相同,小孢子母细胞减数分裂前绒毡层发育不充分;四分体时期,绒毡层细胞高度液泡化,细胞质稀薄,已提前降解,小孢子四分体因绒毡层结构和功能异常而不能正常发育,因此楸树雄性不育为结构型雄性不育。     

Expression of ASK1-like genes in arrested stamens of female Silene latifolia plants

Silene latifolia is a dioecious plant in which sex is determined by heteromorphic sex chromosomes. In female plants, stamen development is arrested before microspore mother cells are formed. In this study, we isolated four cDNAs (SlSKP1-1 to 4) encoding ASK1-like protein as expression markers to reveal when expression levels are reduced in arrested stamens of female flowers. Expression patterns of the SlSKP1 genes were analyzed by in-situ hybridization. We use the flower development classification of Grant et al. (in Plant J 6:471–480, 1994). SlSKP1 genes were highly expressed in primary parietal cells and primary sporogenous cells that develop into microspore mother cells in male flowers. Expression levels started to be reduced in the external stamens of the female flowers when stamen development was arrested at stage 7. Although microspore mother cells could not be developed in female flowers and SlSKP1 expression may be unnecessary in arrested stamens, SlSKP1 genes were still expressed in sporogenous cells of degenerated stamens at stage 8. Parietal cells stopped differentiating earlier than sporogenous cells in arrested stamens. These results suggest that not all types of cell are arrested simultaneously at a particular stage of stamen development during stamen suppression of S. latifolia.     

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...     

紫斑牡丹的花药发育和小孢子发生

紫斑牡丹的花药是在花芽发育的第３个年周期中,从雄蕊原厚基发育而来,花药壁按特有方式发生,主要特点是绒毡层与次生造孢细胞同源。花药由４个花粉囊组成,绒毡层属分泌型,中层３－４层,其中１－３层与药室内壁同步发育出纤维素壁加厚,并在花药成熟时宿存。小包子母细胞减数分裂同时开始,但不同时结束,分裂过程高度不同步。胞质分裂为同时型,四分体中小孢子排列呈正四面体形,减数分裂前期Ⅰ通常有Ｂ－染色体和染色体桥形。     

Exine and tapetum development in Nymphaea capensis (Nymphaeaceae): a comparative study

The developmental events in microspore envelope and cytoplasm and in tapetum from premeiosis until late tetrad stage were studied in Nymphaea capensis. The exceptional feature of microspore development in this species is that post-meiosis cytokinesis is retarded until the late tetrad stage. Thus, the entire development of the exine becomes completed during the tetrad stage. As a consequence of the retarded cytokinesis, the proximal portion of the forming exine lags behind the distal one during the major part of the tetrad period, but eventually the proximal part of the exine overtakes the distal part in development. The significance of this retardation is discussed. This sequence of events differs sharply from corresponding sporoderm development in other Nymphaea species. Another important topic is the microspore surface activities during exine development. The surface coatings-glycocalyx-are very similar in microspores and in tapetum cells, but their functions are completely different; the roots for this difference are discussed. A noteworthy feature of the developing microspores is the presence of gigantic, deeply cup-like mitochondria; this property is also characteristic of the microspore cytoplasm of N colorata and N. mexicana. A functional significance of these organelles and their adaptive role is discussed.     

中国鹅掌楸小孢子发生的细胞化学研究

多糖、脂类和蛋白质的消长变化与花粉发育过程中营养供应存在着密切的关系。雄蕊特别是花药壁内的多糖在减数分裂过程中逐渐消耗殆尽。初期的小孢子母细胞内脂滴的含量丰富,无淀粉粒,也无颗粒状的蛋白质。进行减数分裂的小孢子母细胞内、中层及绒毡层细胞积累了大量的蛋白质与脂类;减数分裂完成后,绒毡层和四分体小孢子开始逐渐积累淀粉与脂类,但蛋白质的含量较少,至小孢子时期,小孢子壁及绒毡层细胞的径切向壁和内切向壁沉积了大量的脂类物质,而且小孢子核周围分布大量淀粉粒,未观察到蛋白质颗粒。     

彩色马蹄莲大小孢子发生和雌雄配子体发育的研究

选用石蜡切片法观察了彩色马蹄莲品种‘Majestic Red’的大小孢子发生及雌雄配子体发育的过程。研究结果表明：彩色马蹄莲的胚珠为倒生,具双珠被、厚珠心和珠被绒毡层。大孢子母细胞的减数分裂后形成的四分体为直线型或T型排列,合点端的大孢子发育成为功能大孢子,其余3个大孢子则退化,表明胚囊发育方式为单孢子发生的蓼型胚囊。观察到每个雄花花药多数,花粉囊呈蝶形,每侧有2个小孢子囊。花药壁由外到内分别为表皮、药室内壁、中层和绒毡层,其中绒毡层为变形绒毡层类型。在小孢子形成时,胞质分裂属于连续型,小孢子排列成十字形的四分体,成熟花粉则为二胞花粉粒。     

Ultrastructural expression of cytoplasmic male sterility in sugar beet (Beta vulgaris L.)

Summary The development of microspore mother cells (MMC) and tapetum in male-fertile and male-sterile anthers of Beta vulgaris L. was compared at the electron microscope level. These studies were complemented by morphometric analyses of mitochondria in both tissues through successive stages of microsporogenesis. The earliest irregularities in the ultrastructure of male-sterile anthers were noted within the tapetum at the tetrad stage. These disturbances were initially expressed by a slight reduction in mitochondrial size and the appearance of concentric configurations of endoplasmic reticulum. As development proceeded, a further decrease in mitochondrial size become more conspicuous and was accompanied by a reduction in ribosome population and a failure of the tapetum to produce Ubisch bodies. This failure to produce Ubisch bodies is reflected in the underdevelopment of sterile microspore exine.     

Expression of ogu cytoplasmic male sterility in cybrids of Brassica napus

Summary A light and electron microscopic investigation revealed that ogu cytoplasmic male sterility (CMS) in cybrids of Brassica napus is primarily a deficiency of the tapetum and clearly time and site specific. Three patterns of ogu CMS were found, and specific conclusions drawn. First, the partially male fertile cybrid 23 was highly variable. It sometimes produced heterogeneous stamens with an endothecium formed exclusively around the fertile locules, thus delineating each microsporangium as a functional unit. The second type, including cybrids 27, 58 and 85, on the contrary, was stable and completely male sterile. In the four locules of normal length, microspores were observed to die at the vacuolate polarized stage while the tapetum disappeared prematurely through excessive vacuolization by the end of meiosis followed by a rapid autolysis during the tetrad or early free microspore stage. The subepidermal layer of the locule wall failed to form characteristic thickenings. The male-sterile stamens were completely indehiscent. At the time of anthesis they contained only collapsed empty exines adhering to each other. These cybrids, 27, 58 and 85, were closest to the ogu CMS trait of radish and seemed to be the best suited for further use in plant breeding. The third pattern was found in cybrids 77 and 118, which besides showing abortion of the microsporangia also showed a feminization of the stamens. We suggest that this feminization might be due to an alloplasmic situation associating Brassica napus nuclear genes with the mitochondrial DNA of radish.     

星星草大、小孢子发生与雌、雄配子体发育的观察

利用常规石蜡制片法研究了星星草[Puccinellia tenutiflora(Griseb．)Scribn．et Merr．]大、小孢子发生及其雌、雄配子体的发育过程。主要结论是：(1)小孢子母细胞减数分裂过程中的胞质分裂为连续型,四分孢子为左右对称型;(2)成熟的花粉为三细胞型,具单萌发孔;(3)花药壁由4层结构组成,最外层为表皮,其内分别为药室内壁、中层、绒毡层,绒毡层为分泌型,花药壁的发育属于单子叶型;(4)星星草为单子房,单胚珠,双珠被,薄珠心,倒生型胚珠。大孢子母细胞经减数分裂形成线形排列的4个大孢子,合点端大孢子具功能;(5)胚囊发育属于蓼型,成熟胚囊形成时,反足细胞经无丝分裂形成4～6个反足细胞,反足细胞内可能存在多次DNA复制过程。     

Protein analysis during the ontogeny of normal and male sterile stamenless-2 mutant stamens of tomato (Lycopersicon esculentum Mill.)

The levels and synthesis of proteins during the ontogeny of normal and male sterile stamenless-2 (sl-2/sl-2) mutant stamens of tomato (Lycopersicon esculentum) were examined. The mutant stamens contained low levels of soluble protein which were related to reduction in protein synthesis. The mutant stamens, however, possessed many polypeptides similar to the normal and synthesized a 53-kd polypeptide at stages when there are abnormalities in tapetum development. The mutant stamens also possessed a 23-kd and some low molecular weight polypeptides that were considered as degradative proteins. Normal stamens exhibited the synthesis of many polypeptides not found in the mutant, from microspore mother cell to the preanthesis stages. In addition, at the time of pollen maturation there was a greater synthesis of several polypeptides, particularly those of 42 and 37 kd. Although the causative mechanisms of male sterility in the sl-2/sl-2 mutant are not known, the synthesis, and the lack, of specific polypeptides reported here appears to be associated with pollen degeneration.This work was supported by an operating grant from the Natural Sciences and Engineering Research Council of Canada to V.K.S.     

Cytochemistry of pollen development in Campsis radicans (L.) Seem. (Bignoniaceae)

In this study, cytochemical staining methods were used to follow the cytochemical modifications of microspore cytoplasm and sporoderm in Campsis radicans (L.) Seem. from tetrad stage to mature pollen. Flower buds were collected at different stages of development, and the anthers were fixed and embedded in Araldite. To make cytochemical observations under light microscope, semithin sections were cut and stained with different dyes. Cytochemical methods provided the opportunity to localize the reserve material in the microspore and pollen cytoplasm, to distinguish the different layers of the sporoderm, and to determine its chemical structure at different developmental stages. Microspore cytoplasm contains variable amounts of proteins, lipids, and insoluble carbohydrates at different stages of microsporogenesis. Sporoderm formation starts at tetrad stage by the formation of primexine and is completed at vacuolated microspore stage by the addition of sporopollenin from tapetum. During the vacuolization and enlargement of the microspores, the structure and the chemical composition of the exine are modified. The endexine becomes chemically different from the ectexine. The ectexine is composed of sporopollenin and a small amount of protein, whereas the endexine is composed of sporopollenin, proteins, and traces of polysaccharides.     

三叶木通大小孢子发生和雌雄配子体发育研究

三叶木通（Akebia trifoliata）属于木通科（Lardizabalaceae）木通属（Akebia）,为雌雄异花,雌雄同株的木质藤本植物,具有药食两用的经济价值。本文运用石蜡切片技术观察了三叶木通的大小孢子发生和雌雄配子体发育过程中胚胎学特征,以期了解三叶木通有性生殖过程,分析该物种自然条件下结实率低的生殖原因,为其后续进行杂交育种和新品种培育提供理论基础。结果表明：三叶木通雄蕊6枚,每枚花药具4个小孢子囊,花药壁完全分化时由外到内依次为1层表皮、1层药室内壁、2～3层中层及1层绒毡层,且绒毡层为分泌型绒毡层。小孢子母细胞减数分裂的胞质分裂为同时型,小孢子四分体为四面体型排列,从四分体分离出来的小孢子经过进一步发育形成2-核成熟花粉细胞。小孢子囊中的小孢子四分体时期存在少量败育现象。雌蕊具1室子房,为侧膜胎座,胚珠横生,多枚,双珠被,厚珠心。大孢子母细胞减数分裂后形成的大孢子四分体呈线型排列,靠珠孔端的3个大孢子退化,合点端的1个大孢子发育成为功能大孢子,经3次连续有丝分裂并进一步发育为七细胞八核的蓼型胚囊。雌花中的雄蕊早期发育正常,但小孢子发育至单核靠边期不在进一步发育,...