Cell signaling involved in the germination of the pollen tube
- Pollen tube germination
- Structure of the pollen tube wall
- Pollen interaction with receptive stigma
- Polarity of the pollen tube
- Ovular orientation mechanisms
- Pollinic germination in orchids
- Growth of the pollen tube in Orchids
Cell signaling involved in the germination of the pollen tube case family Orchidaceae
Abstract: The interest in understanding the processes involved in plant sexual reproduction has allowed us to identify the molecules involved in the processes of fertilization and therefore germination and elongation of the pollen tube, key aspects to achieve reproductive success , obtaining fruits and perpetuating the species. Currently, these processes have been widely described in Arabidopsis thaliana, the biological model par excellence, where advances have led to clarify mechanisms and networks Cells that control cell shape, growth, wall modifications and cellular signaling mediated by second messengers. In this review, the factors and mechanisms of cellular activation that allow the communication and interaction of male and female gametes in the stigmatic cavity are analyzed in a general manner . Likewise, it is described how fertilization is carried out in the orchids, since these species have modifications at the level of the reproductive organs and development of pollen aggregates (pollinia), which make the disposition and delivery of the male gametes, unfold mechanisms that are particular to this group of monocotyledons. This contrast is interesting to the extent that cellular strategiesand interactions are compared to achieve reproductive success.
Although plants exhibit different forms of propagation, in sexual reproduction the process of fertilization depends to a large extent on the growth and development of the pollen tube, since its rupture at the tip or “tip zone” facilitates the release of male gametes which they have as their purpose the fusion with the ovules in the flower. Product of this union occurs the formation of the zygote allowing the formation of a new individual .
At the moment when the flower opening stage begins, the arrival of the pollen from other flowers to the compatible and mature stigma is propitious. The process of release and delivery of pollen requires coordination between the different participating tissues , with the end of guiding the cells sperm towards the ovules (Soto, 2009). This process begins with the pollination stage in which the male gametophyte is transported from the anther to the stigmatic surface. In this place the adhesion, hydration and germination of the pollen is carried out, and then the pollen tube emerges penetrating the tissues of the pistil until reaching the ovary, crossing the micropyle and finally encountering the female gametophyte, to achieve fertilization (Suarez , 2009).
The main objective was to describe the cellular signaling processes that are activated to achieve the liberation of the pollen tube ( vegetative cell ) in the process of fertilization, allowing the delivery of sperm cells (generative) in the ovarian cavity. At the same time, reference is made to the germinative process in orchids, in order to clarify whether the mechanisms that drive pollen germination are similar processes or, on the contrary, they differ from those described for dicotyledons.
Pollen tube germination
Many authors agree, that once the process of germination of the pollen tube begins, its growth rates are very fast this in order to ensure the effective delivery of male gametes towards receptive stigma. Within the mechanisms used by pollen to achieve its purpose, is the extension of the cell wall of the tube and of course the regulation of it. Next, we will give a general overview of the enzymes and cell interactions that allow their development and viability.
Krichevsky et al., 2007 attribute the lengthening of the pollen tube to two biochemical processes:
2.1. An accentuated calcium gradient in the tip of the tube: which drives and guides the growth in the apical zone, in the same way it has been evidenced the increase of the concentration of calcium exclusively in the apical or tip zone, while in the rest of the tube its values remain normal. Li et al, 1996; Pierson et al, 1994; Rathore et al, 1991, who are cited by Krichevsky et al., 2007, point out that an inhibition of this factor, either by blockage of their channels or the induction of passive thermal shock, prevent the growth of the tube.
2.2 . An elongation favored by actin microfilaments: this cytoskeletal protein forms networks of actin under the plasma membrane, providing mechanical support, cellular structure and facilitating the movement of the cell surface. It also empowers the cell in activities of migration , division and absorption of particles (Cooper and Hausman, 2011). For the case of the pollen tube the longitudinal filaments of actin together with the motors of myocin, are key in the growth of the tip. Since when drugs were used inhibitory effects were observed in the lengthening of the pollen tube, by preventing the polymerization of the actin. However, this action can not be generalized because (Anderhag et al., 2000) obtained short tubes of Picea abies when it inhibited the microfilamnets with colchicine.
Dresselhaus & Franklin 2013, alludes to a series of sporophytic cells that support the growth of the tube as it is directed towards the ovular surface, especially the proteins belonging to the subclasses of small proteins rich in cysteine (CRPS), reactive oxygen species (ROS) / NO signaling, and the second Ca2 + messenger, which play a central role in most of these processes.
Soto (2009), describes several aspects related to the growth of the pollen tube, where an adequate coordination will be vital to achieve its elongation, such as:
2.3. A gradient and flow of ions : in the process of growth of the pollen tube an oscillation caused by the entrance of the ions is generated in the apical region thereof of the tube. The calcium ions and protons are the most important depending on the elongation, followed by the chlorine and potassium ions. The entrance of calcium and protons occurs at the tip of the pollen tube, in the latter there is a lateral outlet flow in the subapical zone, which causes changes in pH , in regions of the apical end (Holdaway-Clarke et al., 2003; Campanoni and Blatt, 2007).
2.4. Vesicular traffic: is a communication process mediated by the Golgi apparatus, under vesicular production , which have as a fate the plasma membrane for the release of its content to the cell wall (Lisboa et al., 2007). On the other hand, the process of displacement of organelles allows the growth of the tube in the measure in which they are correctly distributed and the accumulation of secretory vesicles in the apex of the tube.
2.5. Signaling: the type of signaling with greater participation in the processes of elongation of the tube, vesicular traffic, regulation of the cytoskeleton and channels of Ca2 +, is the signaling measured by phospholipids and inositol phosphate which involves kinase-like receptors associated with the membrane and Activation of signaling cascades (Monteiro et al., 2005 a and Malho et al., 2006). On the other hand, vesicular traffic and regulation of actin microfilaments is controlled by the signaling of small G proteins (Krichevsky et al., 2007). The alteration of any of these elements prevents maintaining fidelity in the fertility process, which results in a series of phenotypic aberrations (Cheung et al., 2004, Myers et al., 2009).
Structure of the pollen tube wall
The wall of the tube is characterized by a cylindrical structure with an exclusive growth of its apical end. The primary wall that forms at the apex of the tube is composed of esterified pectins that are released into vesicles from the Golgi and the secondary wall that is composed of callose and is contiguous to the plasma membrane. Among the multiple functions of the cell wall, is the protection of the generative cell by mechanical stress , physical control in the structure of the tube (Derksen 1996, Helpler et al., 2001 cited by Suarez, 2009).
The germination of the tube requires the activation of important factors to achieve its configuration and regulation. In the process of maturation of the wall, the growth of the tube displaces the peptinas that are in the apex towards the subapical zone, where they suffer a process of de-esterification mediated by enzymes peptina metilesterasas (PME). Tian et al., 2006 point out that the role of SMEs is crucial for the growth of the pollen tube wall, these authors identified a new specific PME of Arabidopsis pollen, the ATP ME1, when using genetic Inverse (alteration of DNA by mutations), demonstrating that the participation of this enzyme, is determinant in the formation of the tube and the rhythm of its lengthening. In parallel, the activation of the callose synthase brings about the formation of a layer of callose under the fibrous layer of pectins. At the end of the process, other proteins such as AGPs and extensins are joined to the wall of the pollen tube (Bosch and Hepler 2005, Chen and Ye 2007 cited by Suarez, 2009).
Pollen interaction with receptive stigma
The adhesion of the pollen grain in the stigma florar is a cellular interaction mediated by two factors, the presence of proteins that are part of the pollen cover and the capacity of the stigma to rehydrate the pollen grain after its adhesion. At the time of rehydration, the stigma acts as a water source that induces pollen germination. Problems in the rehydration of pollen are attributed mainly to dry stigmas, which are usually common in the Brassicaceae family . Pollen hydration is also related to expression acuapurinas in stigma and its influence on the transport of water fast and controlled manner, to the grains (Dixit et al., 2001).
Samuel et al., 2009, points out that the pollen-pistil interactions in the hydration stage are regulated by a pistil protein called Exo70A1. More precisely, Exo70A1 acts as a factor of stigmatic compatibility and is also believed to have a mechanical function, being implicit in the immobilization of post-Golgi secretory vesicles to the plasma membrane, and on the other hand it would participate in the immobilization of secretory vesicles. the papillary stigmatic membrane at the contact site of the grain, allowing delivery of the stigmatic components required for pollen hydration.
At the level of the pistil there is also the expression of key factors for the recognition of pollen. In the specific case of Arabidopsis, sulfinylated azadecalins were identified as a small molecule that stimulates pollen germination at an in vitro level . Pollen bioassays with sulfinylated azadecalins synthesized suggest that these molecules act in a specific way to ensure rapid fertilization when pollen from appropriate species interacts with the pistil (Qin et al., 2011).
Polarity of the pollen tube
The polarity and the direction of the pollen tube are mediated by a series of chemo attractants, including chemocyanins and plantacianins. Dresselhaus and Franklin 2013, consider that these proteins have a high redox potential, so that reactive oxygen species such as (ROS) could be responsible for the orientation of the pollen tube. Likewise, the extracellular pistil matrix (ECM) is recognized for providing signals and nutrients that guide the pollen tube and favor its growth, several components of the (ECM) have been identified with chemotropic activity in the direction of the pollen tube, which is destined for the ovules. Among them stand out, the protein rich in cysteine called adhesin stigma / style rich in cysteine (SCA), who is responsible for regulating the lily polar growth of the tip of the pollen tube. Park et al., 2000 sees the interaction of SCA with peptinas as a crucial factor in the orientation of the pollen tube, since this enables the formation of the adhesive matrix as the pollen tube grows, which influences the transmission way. Tissue-Specific Transmitting Tissue-Specific Protein (TTS) with chemo-attractant function was identified in the piston ECM of tobacco and for the specific case of A. thaliana it was found that the neurotransmitter (GABA) contributes to the orientation of the pollen tube.
Ovular orientation mechanisms
The ovular orientation, refers to the guide of the pollen tube to and within the ovule. This includes two orientation mechanisms, particularly identified in A. thaliana .
- Fenicular Orientation: it is understood as the opening of the transmission routes of the pollen tubes with growth on the surface of the septum and funicles towards the ovule, due to the ovular signals. In Arabidopsis this action is controlled by both the sporophyte and the gametophyte.
- Micropillary orientation: represents the passage of the tubes to the ovular apparatus, through the micropyle. And it is regulated by the female gametophyte that interacts through the secretion and attraction of attraction and repulsion signals (Higashiyama and Hamamura, 2008).
Three molecules have been fully identified in relation to the signaling of the pollen tube and its interaction with the pistil and the ovular tissues, in this way under the criteria of molecules of fenicular orientation are highlighted: the non-protein amino acid GABA which has been found which exhibits high concentrations at the moment of the micropillary opening of the ovule. The D-serine that regulates the expression of Ca2 + in the CNGC18 channel of the pollen tube and the NO gaseous molecule that in in vitro tests of A. thaliana proved to be required for the orientation of the micropyle (Palanivel et al., 2003; Michard et al. al., 2011; Lu et al., 2011)
Pollinic germination in orchids
Orchidaceae is the family with the largest number of species at a general level, with an estimated of more than 25,000 species, it is a cosmopolitan family that reaches its greatest diversity in tropical regions, being Tropical America the region with the largest number of species. In recent years, the family has been the subject of numerous research studies that have helped to clarify their phytogenetic relationships and to propose a new classification (García et al., 2004).
In many groups there is aggregation of pollen grains and in the subfamilies Epidendroideae and Orchidiodeae it forms well-defined structures , known as pollinia. Pollinia can be provided with accessory structures for the removal and disposition of pollen, which has allowed a greater success in pollen dispersal and colonization of different ecosystems (Garcia et al., 2004). Regarding the germination of pollen in orchids Sanfor et al., (1964) indicates the scarce availability of information of this process, difficulty associated with differences in the initial germination requirements in orchids compared to other plants. This author also developed a preliminary study of orchid pollen germination in (1964), finding a relatively low percentage of germination in 29 species evaluated at in vitro level . This essay provides convincing results, affirming that germination in orchids generally begins in the periphery of the pollinia, a postulate that has great acceptance at present. This author points out in particular the case of the Phataenopsis and Vanda hybridswho obtained a percentage of (20%) of germination under controlled conditions. Indicating a possible sterility in the pollen grains of the Hybrid Rodricidium “Tahiti”, due to the non germination of its pollen cells.
In 2013 Moraes et al., Evaluated the pollen viability and germination of the pollen tube in interploid hybrids of Epidendrum fulgens x E. puniceoluteum(Epidendroideae, Orchidaceae), finding that except for crosses related to E. fulgens and hybrid flowers , the pistils exhibited a growth of the pollen tube with different intensity, 12 days after the manual pollination at all the crosses. Noting further, that the use of hybrids as pollen donors negatively affects the germination of the tube, making it weaker.
On the other hand Sanfor et al., (1994) recognizes a high sensitivity of orchid pollen to calcium, and its influence to stimulate the growth of pollen, ratifying the affirmations of Brewbaker and Kwack (1963) who worked with a great variety of pollen. (except for Orchidaceae), reported that the presence of calcium ions in yeast extracts and extracts with different parts of the plant, stimulated the development of pollen.
Growth of the pollen tube in Orchids
Cocucci in (1973) in his search to understand the processes of pollen tube growth in orchids and taking as model species Epidendrum scutella Lind, I can determine that before the germination of the pollen grain, there is an association between structures known as bodies of dense electrons and the tonoplast of the cell. This union is a precursor of the concentric intravacuolar membranous bodies (ICMB). Apparently the formation of these structures occurs in two different stages that are part of the same process which culminates with the organization of a membrane reservoir. In this way, at the beginning of the germination of the pollen tube the ICMB will serve to increase the surface area of the tonoplast and the plasmalemma, avoiding the loss of energy and reducing the time when installing the tube in its ideal medium, that is to say the stigma
The orchids are characterized by having several reproductive peculiarities, which possibly originated in function of the increase in the number of grains contained in the units of pollinic dispersion (PDU), that is, the way in which the mature pollen grain is presented for its dispersion , since in particular in orchids the pollen can be delivered to the flower as a single grain, in mass or as a compound pollen. Monel pollen is the initial unit that allows the transition from monada to pollen in tetrad, which finally by cytological mechanisms will be combined generating different types of PDU, which can host about 4,000,000 pollen grains in poly (Schill et al., 1992).
Pacini 1990, describes a typical process of this Angiosperm family that presents a dispersion of pollen in monada, known as hermomegatia.
8.1. Hermomegatia: refers to the loss and gain of water from the pollen plant cell . Pollen as a measure of protection to environmental conditions, slows down its metabolism generating a loss of water before its dispersion to finally restore its metabolism completely after rehydration in the stigma.
This process is unnoticed by the pollen units that disperse thousands of pollen grains. This is because the devices used for hermomegatia (furrows that adjust to changes in water content) are absent due to the compaction of the tetrads. In more complex PDUs the absence of a pore or groove in mature pollen indicates that the site for the appearance of the pollen tube is determined only after the pollen has contact with the stigmatic surface Pandolfi and Pacini, 1995).
According to (Yeung, 1987) in more complex pollen organizations where the tetrads show a higher level of compaction, the shape of these tetrads (decussate, square, rhomboidal, T-shaped and linear) is usually a relevant character (Davis, 1966), since differences in their morphology allow a greater or lesser degree of union in the polynite, which in turn affects the obtaining of intertected spaces. Without the formation of these spaces it would be practically impossible to germinate the pollen tube.
In orchids the degree of pollen compaction is determinant in processes such as pollen tube germination. The substance that generally holds the pollen grains together inside the pollinium is known as elastoviscin, a type of elastic material made up of lipids that lead to the formation of a polymer Blackman and Yeung (1983). In the case of the masses where the tetrads that form it usually remain attached until deposited in a receptive stigma, it could be the case that a strong adhesion between these tetrads is unfavorable for the pollen tube emissions, due to the lack of intertetrated spaces (Pandolfi and Pacini, 1995). The cohesion of pollen grains is very variable within Orchidaceae, so this aspect should be considered when evaluating germinative processes.
Some authors affirm that in the receptive stigma of orchids there is a sugary substance that receives the pollinia and allows the pollen grains to germinate, but most of these studies are unaware of the properties and biological composition of said substance. What is clear is that stigmatic surfaces have evolved simultaneously to adapt to different units of pollen dispersion, for example it is normal to find flat or slightly convex stigmatic surfaces in species with sectile pollen, where this receptive organ has an adhesive cover that facilitates pollen trawling (Nilsson 1983).
In general, the pollen germination process of orchids begins once pollination occurs, in this way the pollen germinates and a pollen tube grows that grows downwards inside the pistil column towards the ovary, stimulating the development of the ovules, which will be receptive by the time the pollen tube reaches its proximity, fertilization occurs and the development of the embryo begins which will result in the development of the seed (Ackerman and Del Castillo, 1992). However Von Kirchner 1922 cited by Sanfor et al., (1994) reported difficulties for in vitro germinationof orchid pollen. Von Kirchner also highlights the importance of investigating in a more profound way about the conditions that promote the germination of pollen in orchids. But until now, not enough literature has been found to support whether the communication and cell signaling networks, described in species such as A. thaliana, are involved in the activation of these processes .
- According to the evidence found, during fertilization the pollen tube interacts with various types of sporophytic cells that support its growth and guide it towards the ovular surface. In the same way, the female gametophyte cells orient the tube and control the delivery of the male gametes. These events lead to cell-cell signaling and communication cascades, which are activated to maximize reproductive success.
- In the case of pollen germination in orchids, there were no reports that alluded to a specific type of molecule implicit in the signaling of said process, it would be believed that the signaling pathways that are activated in the process of fertilization described in the dicotyledons could be similar for those subfamilies where the pollen dispersion is in monada, however when the dispersion is done using pollen aggregates, it is necessary to deepen biochemical studies and in vivo and in vitro level, achieving a full identification of the cellular signaling networks.
- The study of sexual reproduction in orchids has received little attention , despite the enormous ecological and ornamental interest of this family, since it is still not very clear what are the cellular and molecular mechanisms that regulate the interactions that are established between pollen and the pistil during the progammic phase of these plants.
- Due to the diversity of pollen aggregates that the orchids exhibit, it is necessary to identify the type of communication between the pollen grain and the glandular stigmatic cells, at the moment of pollen reception. Identifying the type of stigma implicit in the reception, will be useful to understand the cellular signaling that induces pollen germination, since in this case the process will be mediated by liquid secretions or, conversely, by proteins and secreted waxes.