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ACTA BIOLOGICA CRACOVIENSIA Series Botanica 45/1: 19–26 2003

PLANT SEXUAL : ASPECTS OF INTERACTION, HISTORY AND REGULATION

MICHIEL T. M. WILLEMSE*

Laboratory of , Wageningen University, Arboretumlaan 4, 6703 BD Wageningen, The Netherlands

Received July 16, 2002; revision accepted February 11, 2003

Sexual reproduction in angiosperms is an interactive process involving the , , and as well as the environment, aimed at achieving , fertilization and dispersal. This interaction occurs via an interface with nutrients and signals outside the cell and even outside the plant. has a history. In water, have different types of organs and , and in some cases the stays on the plant. The uses water movement and gravity for dispersal. Some algae have alternation of generations in the life cycle, and only the functions in sexual reproduction. On land, and inherited alternation of generations, with oogamy and zygote development on the gametophyte, with wind dispersal of the meiospore. In angiosperms, and the retention of the , megagametophyte and embryo on the sporophyte lead to a with gravity and biotic dispersal. The history of sexual reproduction is based on sex determination, due to cross-fertilization and recombination. Sex differentiation is manifested in the increasing complexity of interaction in the nutrient supply, the retention of the gametophyte or even the embryo, and the type of vector of dispersal. Regulation of sexual reproduction in angiosperms is governed mainly by the sporophyte, with the expression of new for biotic pollination and . In the heterotrophic gametophyte some expression is suppressed. The development of sexual reproduction is due to the communication between the and a dynamic environment.

Key words: Sex differentiation, dispersal, interaction, regulation, environment.

INTRODUCTION between the sporophyte and gametophytes during development, the sporophyte and the environment The organism represents a unit of form, function and during pollination, the male and female gameto- biotope. When a plant starts sexual reproduction, its phytes during fertilization, the sporophyte and em- flowering means the origin of new forms and func- bryo with endosperm after fertilization, and the tions in connection with the environment. sporophyte and the environment during seed disper- Sexual is a complex se- sal. Based on the sequence of ordered steps, the quence of different steps leading to a new organism. complex regulation of sexual reproduction will be The genome is renewed through recom- bination in presented. first and cross-fertilization. In the The process of sexual reproduction in angios- of angiosperms the anther and develop to perms can be compared with the process of sexual prepare the male and female gametophytes. In the reproduction in algae, ferns, seed ferns and gymnos- open flower, pollination and fertilization occur, and perms. Storage in a gamete or megaspore, retention then seed and development begins. of the gamete or the megaspore on the plant, and the This review presents sexual reproduction in means of dispersal, all determine the development angiosperms as an ordered and dynamic interaction of angiosperm sexual reproduction. Such a com-

*e-mail: [email protected]

PL ISSN 0001-5296 © Polish Academy of Sciences, Cracow 2003 20 Willemse parative approach will give insight into the origin of droplet or pellicula, followed by the nutrients pres- the interaction and into the regulation of sexual ent in the tube pathway. Nutrient supply reproduction. flows between the sporophyte and female gameto- phyte. The interface consists of degenerating nucel- lus cells, and in the megagametophyte are transfer INTERACTION cells and a filiform apparatus with attrac- tion products. The interaction of the male and female Interplay between involves interactions gametophyte is realized by attraction products and the taking place outside the organisms. Signaling sub- opening of the pollen tube in the synergid. stances and the production of special nutrients or In the fruit, the sporophyte interacts with the other substances require cooperation of organisms. embryo and endosperm by providing nutrients and The area of cooperation is called the interface and is space. In the beginning the interface has degenera- marked by special products in the functioning of ting nucellus cells and inner integument cells. Later, sexual reproduction outside the cell or even outside after development of the endosperm, depending on the organism (Willemse, 1996). Due to its spe- the type of seed, breakdown products supply an cialized function connected with reproduction, the interface for embryo development. Again there is diversity of products and their occurrence outside of interaction between the sporophyte and the environ- the organism, the interface is not considered as an ment, defined by seed or fruit dispersal via a vector. apoplast (Hong-Yuan Yang, 2001). Commonly the abiotic vector for seed dispersal is In the flower bud, the sporophyte of angios- gravity, but wind and water are also used. In biotic perms begins the interaction in the transport, the sporophyte elaborates a mode of ad- after meiosis. In the anther the develops vertisement, expressed in the odor, color and form into pollen, and in the ovule the megaspore develops of the fruit. The are exposed on the plant, and into a megagametophyte. The sporophyte provides the sporophyte adds nutrients to the fruit. Again, space for the developing gametophytes, attends to advertisement and nutrition represent interaction nutrient supply and retains the megagametophyte. with animals. Signals regulate the interaction between organisms. From flower bud to fruit there is continuous The interface between the sporophyte and develo- interaction, with the main role for the sporophyte ping male gametophyte consists of the callose wall, and a specialized role for the gametophytes. The the locular fluid, and the remnants of the tapetal interaction with the environment leading to pollen cells; these can remain as a coat around the pollen. and seed dispersal is marked by the open flower and Pollen can also export signal substances in the pol- the fruit. The interface has nutrients and signal len wall. The interface between the sporophyte and substances coming from the surrounding tissues but developing megaspore contains callose and degener- also consists in special cell walls and excretion and ating nucellus cells. degeneration products. In the open flower the interaction between spo- This interaction and the interface mark the rophyte and environment is expressed by way of flower as an of communication. This system is pollination. In abiotic pollination, pollen is adapted the result of the history of the process of sexual to wind, water or gravity; and in biotic pollination, reproduction in . to animals or to the plant’s selfing mechanism. Pol- len uptake occurs on the , and is adapted to the means of dispersal. In biotic pollination, the HISTORY sporophyte elaborates a mode of advertisement, through odor and color and the form of and/or The history of plant life is marked by the transition . Routing, commonly leading to cross-pollina- from water to land. tion, takes place in the open flower, with or without guiding and . WATER The interaction between the sporophyte and male gametophyte consists in the uptake of pollen, In water, unicellular algae have sexual reproduction including recognition as well as activation of the in the simplest form. Gametes are formed after pistil. After acceptance, the sporophyte guides the meiosis and prepared for flagellar movement, at- pollen tube towards the micropyle and supplies nu- tracting and fusing in water. The isogametes fuse to trients. The interface is the pollen coat and stigmatic form a zygote, the unit of dispersal. The vector of Plant sexual reproduction 21 dispersal is water, and then gravity plays a minor Ferns have alternation of generations, oogamy role. Recombination takes place during meiosis, and depending on water, and the development of the if cross-fertilization is promoted the zygote has a plantlet on the gametophyte. A terminal sporan- renewed genome. This renewal is the main goal of gium develops apically on the stem of primitive sexual reproduction. ferns, preparing the meiospores as units of disper- The origin of multicellular algae is the result of sal. More advanced ferns have sporangia on their incomplete cytokinesis, with contact, as in . The meiospores are transported by wind, and colonies, or wall and contact forming an land on the ground by gravity. organism with symplast and apoplast. Cells begin The next important change happens in the hete- independent development after meiosis or rosporic ferns. A functional (as in some horsetails) followed by complete cytokinesis. For the reproduc- or morphological difference in the meiospores devel- tive function, localized sex organs, the ops, mainly through an increase of storage in one of and oogonium, develop from the . Morpho- the , the megaspore. With this development logically dissimilar gametes are also formed. This is there begins an interaction, with the sporophyte due to enlargement of the cell with storage products. providing extra nutrition to the megaspore. Again, The enlarged gamete is called the female gamete; it this difference in meiospores has a consequence for can be mobile or immobile. The fusion of such mobile dispersal. are transported by cells is termed , whereas the fusion of an wind, and later gravity lets them fall downwards. immobile and a mobile male gamete is The megaspore has gravity dispersal and falls on the referred to as oogamy. Enlargement of the female ground. The use of different vectors presents a risk gamete entails greater dependence on gravity for to fertilization when the spores are far apart, but on dispersal. Because of this, the fertilized oocyte deter- the other hand it promotes cross-fertilization. mines the final position of the zygote and its settle- The extinct seed ferns are heterosporic, with ment on the substrate. prolonged retention and development of the mega- The introduction of alternation of generations spore in the megasporangium on the sporophyte. in the life cycle of algae is the next important step The sporangium develops from the epidermis and for plant sexual reproduction. The possibility to orig- hypodermis. This long interaction results in the inate an organism from each unicellular stage dur- formation of a seed, the unit of dispersal. Megaga- ing the life cycle results in a sequence of organisms. metophyte development, fertilization and seed de- Each unicellular cell stage is a totipotent cell ca- velopment all take place on the sporophyte, which pable of making a plant. The mitospore and the provides space and nutrients. The gametophytes zygote form a sporophyte, the meiospore forms the become heterotrophic. The seed is transported gametophyte, and a functional difference between mainly by gravity to the ground, as is still the case plants develops. Only one plant is able to produce with modern . In the microsporan- gametes, the gametophyte. Moreover, in a life cycle gium the microspores develop into microgameto- with alternation of generations there are two mo- phytes with a few cells having a simple male sex ments of dispersal, one by the meiospore and one by organ and producing immobile cells, or else the zygote. into pollen grains. The pollen grains are dispersed The gametophyte has sex organs and can have by wind and fall on the pollination droplet, which oogamy. In some algae the oocyte stays on the gameto- places them at the top of the nucellus. Pollen tubes phyte, and the zygote and young plant develop on the grow in the nucellus and the sperm cells are trans- gametophyte. With this retention an interaction takes ported in the pollen tube cytoplasm towards the place between the gametophyte and the zygote and megagametophyte. In the macrosporangium the even the young plant. In such a life cycle there is only megaspore develops a coenocytic liquid endosperm one moment of dispersal by the meiospore. surrounded by a megagametophyte wall. Cellulari- zation follows, and one or more sex organs develop LAND on the top of the cell mass, each with an isolated oocyte and venter cell, both surrounded by A life cycle with alternation of generations, with cells. Two or more neck cells form the entrance for oogamy and the development of the plant after fer- the pollen tubes. The pollen tube grows through the tilization on the gametophyte, characterizes the neck canal and one of the sperm cells fuses with the early land plants such as mosses and ferns. All these oocyte to form the zygote. After fertilization the seed characteristics were already developed in water. remains on the sporophyte. The mature seed will be 22 Willemse dispersed by gravity, or by wind in cases where the moments of dispersal are introduced. The meiospore seed is extended with a membranous wing. and zygote may use other vectors of dispersal, with In some gymnosperms such as Cycadales, high- the consequence that another biotope can be selected ly flagellated spermatozoids are released in water (Fig. 1c). present just above the neck cells of the megagameto- On land, the life cycle with oogamy, the devel- phyte, a last example of fertilization in water. In opment of the zygote on the gametophyte, and dis- Welwitschia and Gnetum, development of the mega- persal only by the meiospore is an inheritance from gametophyte is delayed and it remains coenocytic. aquatic plants. The heterosporic ferns develop a Pollen tube penetration in the coenocyte results in microspore and megaspore, as already happens in one or even two . Ephedra sp. has double some plants in water. Sporophyte-gametophyte in- fertilization: one sperm cell fuses with the cell teraction develops extra nutrition to the megaspore and another with the venter cell, and in Gnetum sp. by the sporophyte. The interaction again has a direct both sperm nuclei fuse, each with a megagameto- consequence for the means of spore dispersal. The phyte nuclleus (Herzfeld, 1922; Friedmann and Car- megaspore with storage tends to fall downwards but michel, 1996). the microspore is dispersed by wind (Fig. 1d). In angiosperms the cell number of gametophytes The next step in seed ferns and gymnosperms is very low and sex organs are absent. The two sperm is the prolonged retention of the megaspore on the cells are immobile and move in the cytoplasm of the sporophyte. This again means an interaction via an pollen tube. In the megagametophyte an is interface, with nutrition and signaling during spore selected to fuse with one of the sperm cells ; it is an egg and gametophyte development, and pollen uptake cell initial (Favre-Duchartre, 1984) bordered by two and guiding. Megaspore dispersal is blocked, and synergids. Before fertilization the development of the the prolonged retention results in seed after fertili- megagametophyte stops; it has antipodal cells and a zation (Fig. 1e). coenocytic central cell which needs fertilization to form In angiosperms the interaction between the spo- the endosperm. The seed is the unit of dispersal. Pollen rophyte and gametophytes becomes more intensi- and seed have abiotic dispersal but in most angio- fied. The effect is a gametophyte with a low number a biotic vector is used and animals become of specialized cells. The contribution of the sporo- involved in pollen and seed dispersal. The sporophyte phyte to biotic pollination (Fig. 1f) and seed disper- begins to function in advertisement and nutrition, and sal (Fig. 1g) is elaborated mainly in angiosperms. in the case of pollination it also provides routing. This This includes interaction and signaling with the development points to the interaction of the sporo- biotic environment. phyte with the environment. Sexual reproduction is connected with nutrition, signaling, interaction and the means of dispersal. Gamete storage and retention or prolonged retention Trends is a historical trend, and it evokes interaction via an In water the position of the sex organs, commonly interface. A consequence is gravity as the main vector developed from the epidermis, is directed to gamete for megaspore and seed dispersal. In cooperation with release. The storage in one of the gametes deter- dynamic changes in the environment, transport via mines the female gamete. This swelling of the biotic vectors on land becomes a characteristic of the gamete has a direct consequence in gravity disper- sporophyte and is realized by it. sal. Before and after fertilization, vector transport is influenced by gravity. This manner of formation of the oocyte points to the very strong connection REGULATION between sexual reproduction and dispersal (Fig. 1a). The retention of the isolated oocyte on the plant The history and trends leading to the process of or on the gametophyte means an interaction, which sexual reproduction in angiosperms are associated includes nutrition and signaling via an interface. If with changes in the regulation of reproduction and the zygote develops further on the plant or gameto- dispersal. In angiosperms many genes associated phyte the interaction continues (Fig. 1b). Both with sexual reproduction have been isolated, and the gamete storage and retention are effects of nutri- functions of some are well known (Raghavan, 1997). tion. The introduction of alternation of generations The expression of sex differentiation plays an im- means selection of one organism for sexual repro- portant role in the regulation of reproduction and duction: the gametophyte. In such a life cycle two dispersal. Plant sexual reproduction 23

Fig. 1. Scheme representing life cycles, arranged in order of the history of plants. First is the life cycle of unicellular algae followed by multicellular algae, algae with alternation of generations, ferns, heterosporic ferns, seed ferns along with gymnosperms and angiosperms. a–g – important steps during the history of sexual reproduction. a – storage of gamete with the consequence of gravity dispersal; b – retention of the gamete on the plant, with the consequence of nutrition; c – two moments of dispersal by meiospore and zygote, with a fit to different biotopes possible; d – storage of the megaspore, with the consequence of gravity dispersal; e – prolonged retention of the megaspore and megagametophyte, leading to seed dispersal; f – biotic pollination, involving a strong interaction between sporophyte and environment; g – biotic seed dispersal, involving strong interaction between sporophyte and environment. 24 Willemse

SEX DIFFERENTIATION are heterosporic sporangia, and are related to the terminal sporangium of a primitive , the The sex of an organism is determined through for meiospore dispersal on land. The role of differentiation. Each organism is totipotent and has the sporophyte in nutrition of heterotrophic gameto- genes for the expression of both , but during sex phytes results in more intensive interaction, which differentiation only one sex is developed. This is an effect includes other gene actions. of the male- or female-determining factors activating For pollination, the sporangia become surrounded the related genes. The simplest example of the process with petals and sepals for advertising and for routing of sexual reproduction is found in unicellular algae. pollen to be deposited on the stigma, and with nec- Gene expression with respect to sexual reproduction in taries for nutrition. In the sporophyte new genes unicellular plants involves at least meiosis and gamete become involved to develop this advertisement, rout- differentiation. Gamete differentiation includes cell iso- ing and nutrition, as well as the release and uptake of lation and preparation for mobility, attraction and fu- pollen. Genes are selected during flower induction and sion. The amount of storage and degree of immobility the formation of petals, sepals, and pistil, have consequences for the way gametes are supplied represented in the ABC model (Coen and Meyerowitz, with nutrition and dispersed. A gamete can be small, 1991) and its revisions (Gutierrez-Cortines and Davis, light, large or heavy as a consequence of nutrition; it can 2000; Maheswari et al., 2001; Soltis et al., 2002). If the be mobile or immobile, which has an effect on dispersal anther and ovule are sporangia their origin should be by water or gravity. Thus, sex differentiation includes expressed in the model. It is suggested that the ovule interaction with the dynamic environment. Another can be considered an independent sporangium (Co- expression of differentiation is the retention of the fe- lombo et al., 1995), but the same can be postulated for male gamete and even the fertilized oocyte on the plant. the anther. Regulation of the important nectar glands This retention involves interaction through nutrition commonly present in the flower also should be incor- and signaling via an interface. porated in the model. In life cycles with alternation of generations, sex differentiation is first expressed in the gametophyte only. On land, sex differentiation is also expressed in the Gametophytic gene actions before fertilization sporophyte in a specialized sporangium as the organ In the open flower the heterotrophic gametophytes are producing the microspores and , as in hete- partially developed. In the anther the two-celled rosporic ferns and seed plants. The interaction between microgametophyte (pollen grain) has no sex organ but sporophyte and gametophyte becomes more intensive. forms two immobile sperm cells, which are moved in Sex differentiation again relates to the means of disper- the cytoplasm of the pollen tube. In the ovule the sal by abiotic or biotic vectors, and this includes a further megaspore forms a simple female gametophyte, a interaction with the dynamic environment. coenocyte which is converting into a few highly dif- ferentiated cells. No sex organ develops, and there is SEX DIFFERENTIATION AND REGULATION no cell isolation but only an egg cell initial with some IN ANGIOSPERMS storage. The sister cells next to the egg cell initial attract the pollen tube. This means interaction be- In angiosperms the sporophyte is very involved in sex tween the male and female gametophyte. Fertilization differentiation. This has consequences for sex differen- of the egg cell initial is the beginning of the embryo, tiation in gametophytes. These get a reduced number and fertilization of the central cell is the beginning of of cells, but each with a higher level of differentiation. the endosperm; both depend on pollination and fertili- The sporophyte plays a dominant role in nutrition of zation. In the microgametophyte and the megaga- the gametes, embryo and endosperm. The sporophyte metophyte of angiosperms, gene expression becomes also governs pollen and seed dispersal. suppressed and the sporophyte dominates gameto- phyte development and the pollination mechanism. Sporophytic gene action in the flower bud During flower development, macro- and microspo- Sporophytic gene actions during pollination rangia, ovules and anthers develop. The composition and after fertilization of these sporangia is comparable, with a common structure having a multi-layered wall and spo- The sporophyte directs pollen uptake and guides the rogenic prepared for meiosis. Anthers and pollen tube to the ovules. This interaction at the Plant sexual reproduction 25 interface consists of nutrition and signaling, such as in sex determination. Sex determination involves recognition and pistil activation, and is regulated by the formation of new genes as well as the suppres- a complex of genes. Probably the whole system of sion or perhaps loss of genes. Sex is determined by nutrition and signals is oriented toward promotion sex differentiation, the formation of sex organs and of cross-pollination. gametes. Sex differentiation is connected with inter- can also be considered a action between organisms, nutrition via an inter- signal to start embryo and endosperm development. face, and the means of dispersal. Fertilization of the central cell is a condition for In water, sex differentiation is expressed in the endosperm development. Unlike autonomic endo- plant, or in the gametophyte in the case of alterna- sperm development as happens in gymnosperms, tion of generations. Here sex differentiation be- endosperm development in angiosperms depends comes expressed in storage of the gamete and its partly on a fertilization stimulus. In fact the primary retention on the organism, with consequences for endosperm is formed partly by the antipodal cells, the manner of dispersal. but the main part is organized as new tissue with a On land, to beome independent of the medium higher level from the fertilized central cell. of water and to use the vectors of dispersal, the However, autonomous endosperm development oc- sporophyte becomes an actor in sexual reproduction. curs after culture in vitro of young pollen (Chatelet Similarly to the events in water, sex differentiation et al., 1999), embryo sacs (Mol, 1999) and in some becomes expressed in storage of the meiospore and apomicts (Koltunow, 2000), and can be considered its retention on the organism, with consequences for gametophyte tissue, developed through some sup- the manner of dispersal: intensified interaction. In pression of gametophytic genes. The endosperm is angiosperms the sporophyte dominates the gameto- nutritive tissue for embryo development. Endo- phyte, and the sporophyte implements biotic disper- sperm decay will supply the interface with nutrients sal through pollen and seed. around the embryo. The sporophyte develops the The changes in the process of sexual reproduc- seed coat from the sporangium wall and the integu- tion and the variety of its expressions are the result ments, and the fruit from the surrounding carpels. of the creation of new organisms through recombi- New genes will become engaged in regulating this nation and cross-fertilization in a dynamic environ- great diversity of activities, which are directed to- ment. This development is possible because of the ward fulfilling the abiotic or biotic means of disper- unity of the organism and its environment. Sexual sal of the seed or the diaspore. In biotic dispersal of reproduction is an expression of dynamic communi- the diaspore a similar system of advertisement and cation between organisms and the environment. nutrition is involved. 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