RESUMO
Major constituents of the plant cell walls are structural proteins that belong to the hydroxyproline-rich glycoprotein (HRGP) family. Leucine-rich repeat extensin (LRX) proteins contain a leucine-rich domain and a C-terminal domain with repetitive Ser-Pro3-5 motifs that are potentially to be O-glycosylated. It has been demonstrated that pollen-specific LRX8-LRX11 from Arabidopsis thaliana are necessary to maintain the integrity of the pollen tube cell wall during polarized growth. In HRGPs, including classical extensins (EXTs), and probably in LRXs, proline residues are converted to hydroxyproline by prolyl-4-hydroxylases (P4Hs), thus defining novel O-glycosylation sites. In this context, we aimed to determine whether hydroxylation and subsequent O-glycosylation of Arabidopsis pollen LRXs are necessary for their proper function and cell wall localization in pollen tubes. We hypothesized that pollen-expressed P4H4 and P4H6 catalyze the hydroxylation of the proline units present in Ser-Pro3-5 motifs of LRX8-LRX11. Here, we show that the p4h4-1 p4h6-1 double mutant exhibits a reduction in pollen germination rates and a slight reduction in pollen tube length. Pollen germination is also inhibited by P4H inhibitors, suggesting that prolyl hydroxylation is required for pollen tube development. Plants expressing pLRX11::LRX11-GFP in the p4h4-1 p4h6-1 background show partial re-localization of LRX11-green fluorescent protein (GFP) from the pollen tube tip apoplast to the cytoplasm. Finally, immunoprecipitation-tandem mass spectrometry analysis revealed a decrease in oxidized prolines (hydroxyprolines) in LRX11-GFP in the p4h4-1 p4h6-1 background compared with lrx11 plants expressing pLRX11::LRX11-GFP. Taken together, these results suggest that P4H4 and P4H6 are required for pollen germination and for proper hydroxylation of LRX11 necessary for its localization in the cell wall of pollen tubes.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Tubo Polínico , Prolil Hidroxilases , Arabidopsis/metabolismo , Arabidopsis/genética , Hidroxilação , Tubo Polínico/crescimento & desenvolvimento , Tubo Polínico/metabolismo , Tubo Polínico/genética , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Prolil Hidroxilases/metabolismo , Prolil Hidroxilases/genética , Parede Celular/metabolismoRESUMO
Most Aristolochiaceae species studied so far are from temperate regions, bearing self-compatible protogynous trap flowers. Although self-incompatibility has been suggested for tropical species, the causes of self-sterility in this family remain unknown. To fill this gap, we studied the pollination of the tropical Aristolochia esperanzae, including the physical and physiological anti-selfing mechanisms. Floral visitors trapped inside flowers were collected to determine the pollinators. Protogyny was characterized by observing the temporal expression of sexual phases and stigmatic receptivity tests. The breeding system was investigated using hand-pollination treatments. Pollen tube growth was observed using epifluorescence to identify the self-incompatibility mechanism. Flies were the most frequent visitors found inside A. esperanzae trap flowers, with individuals from the family Ulidiidae being potential pollinators since they carried pollen. The characteristic flower odour and presence of larvae indicate that A. esperanzae deceives flies through oviposition-site mimicry. Although this species showed incomplete protogyny, stigmatic receptivity decreased during the male phase, avoiding self-pollination. Fruits developed only after cross- and open pollination, indicating that the population is non-autonomous, non-apomictic, and self-sterile. This occurred through a delay in the growth of geitonogamous pollen tubes to the ovary and lower ovule penetration, indicating a late-acting self-incompatibility mechanism. Our findings expand the number of families in which late-acting self-incompatibility has been reported, demonstrating that it is more widespread than previously thought, especially when considering less-studied tropical species among the basal angiosperms.
Assuntos
Aristolochia , Flores , Polinização , Polinização/fisiologia , Flores/fisiologia , Aristolochia/fisiologia , Animais , Autoincompatibilidade em Angiospermas/fisiologia , Tubo Polínico/fisiologia , Tubo Polínico/crescimento & desenvolvimento , Frutas/fisiologia , Frutas/crescimento & desenvolvimento , Pólen/fisiologia , Dípteros/fisiologiaRESUMO
KEY MESSAGE: In Araucaria angustifolia, the seed scale is part of the ovule, the female gametophyte presents a monosporic origin and arises from a coenocytic tetrad, and the pollen tube presents a single axis. The seed cone of conifers has many informative features, and its ontogenetic data may help interpret relationships among function, development patterns, and homology among seed plants. We reported the seed cone development, from pollination to pre-fertilization, including seed scale, ovule ontogeny, and pollen tube growth in Araucaria angustifolia. The study was performed using light microscopy, scanning electron microscopy, and X-ray microcomputed tomography (µCT). During the pollination period, the ovule arises right after the seed scale has emerged. From that event to the pre-fertilization period takes about 14 months. Megasporogenesis occurs three weeks after ovule formation, producing a coenocytic tetrad. At the same time as the female gametophyte's first nuclear division begins, the pollen tube grows through the seed scale adaxial face. Until maturity, the megagametophyte goes through the free nuclei stage, cellularization stage, and cellular growth stage. Along its development, many pollen tubes develop in the nucellar tissue extending straight toward the female gametophyte. Our observations show that the seed scale came out of the same primordia of the ovule, agreeing with past studies that this structure is part of the ovule itself. The formation of a female gametophyte with a monosporic origin that arises from a coenocytic tetrad was described for the first time in conifers, and the three-dimensional reconstruction of the ovule revealed the presence of pollen tubes with only one axis and no branches, highlighting a new pattern of pollen tube growth in Araucariaceae.
Assuntos
Araucaria , Araucariaceae , Polinização , Tubo Polínico , Cone de Plantas , Brasil , Microtomografia por Raio-X , Sementes , Óvulo Vegetal , BiologiaRESUMO
MAIN CONCLUSIONS: Transmitting tissue cells of Luehea divaricata present a thick middle lamella and prominent convex lens-shaped thickenings of the cell wall that act as reservoir of energy for pollen tube growth. Luehea Milld. is a Neotropical genus with 18 species. This paper reports the study of the structure and the ultrastructure of the stigma and the style of Luehea divaricata Milld. using bright-field microscope and transmission electron microscope. Multiseriate papillae are observed in the stigma. The papillae cells are large with a content that stains intensively and the sub-stigmatic tissue resembles the style's transmitting tissue. L. divaricata has a closed style with an epidermis that presents raised stomata and multicellular trichomes. Numerous organelles and a large nucleus are present in the dense cytoplasm of the transmitting tissue cells. A relevant feature is that the cells of this tissue in the species studied present a very thick middle lamella with two zones of different electron density in the angle of contact between cells, and convex lens-shaped thickenings of the cell wall are prominent mainly also in these angles of contact. The growth of the pollen tube is initiated on the stigmatic papillae surface and continues growing in the middle lamella of the sub-stigmatic tissue and the transmitting tissue, mainly at the contact angle between cells. The present work is the first contribution to the knowledge of structure and ultrastructure of the stigma and style in the genus Luehea, as well as to the subfamily Grewioideae (Malvaceae).
Assuntos
Malvaceae , Núcleo Celular , Citoplasma , Tubo Polínico , Coloração e RotulagemRESUMO
Cassava breeding is hampered by high flower abortion rates that prevent efficient recombination among promising clones. To better understand the factors causing flower abortion and propose strategies to overcome them, we 1) analyzed the reproductive barriers to intraspecific crossing, 2) evaluated pollen-pistil interactions to maximize hand pollination efficiency, and 3) identified the population structure of elite parental clones. From 2016 to 2018, the abortion and fertilization rates of 5,748 hand crossings involving 91 parents and 157 progenies were estimated. We used 16,300 single nucleotide polymorphism markers to study the parents' population structure via discriminant analysis of principal components, and three clusters were identified. To test for male and female effects, we used a mixed model in which the environment (month and year) was fixed, while female and male (nested to female) were random effects. Regardless of the population structure, significant parental effects were identified for abortion and fertilization rates, suggesting the existence of reproductive barriers among certain cassava clones. Matching ability between cassava parents was significant for pollen grains that adhered to the stigma surface, germinated pollen grains, and the number of fertilized ovules. Non-additive genetic effects were important to the inheritance of these traits. Pollen viability and pollen-pistil interactions in cross- and self-pollination were also investigated to characterize pollen-stigma compatibility. Various events related to pollen tube growth dynamics indicated fertilization abnormalities. These abnormalities included the reticulated deposition of callose in the pollen tube, pollen tube growth cessation in a specific region of the stylet, and low pollen grain germination rate. Generally, pollen viability and stigma receptivity varied depending on the clone and flowering stage and were lost during flowering. This study provides novel insights into cassava reproduction that can assist in practical crossing and maximize the recombination of contrasting clones.
Assuntos
Manihot/genética , Óvulo Vegetal , Melhoramento Vegetal , Tubo Polínico , Polinização , Polimorfismo de Nucleotídeo ÚnicoRESUMO
All kingdoms of life have evolved tip-growing cells able to mine their environment or deliver cargo to remote targets. The basic cellular processes supporting these functions are understood in increasing detail, but the multiple interactions between them lead to complex responses that require quantitative models to be disentangled. Here, I review the equations that capture the fundamental interactions between wall mechanics and cell hydraulics starting with a detailed presentation of James Lockhart's seminal model. The homeostatic feedbacks needed to maintain a steady tip velocity are then shown to offer a credible explanation for the pulsatile growth observed in some tip-growing cells. Turgor pressure emerges as a central variable whose role in the morphogenetic process has been a source of controversy for more than 50 yr. I argue that recasting Lockhart's work as a process of chemical stress relaxation can clarify how cells control tip growth and help us internalise the important but passive role played by turgor pressure in the morphogenetic process.
Assuntos
Parede Celular , Tubo Polínico , Ciclo Celular , Diferenciação Celular , PolinizaçãoRESUMO
Proline-rich extensin-like receptor kinases (PERKs) belong to the hydroxyproline-rich glycoprotein (HRGP) superfamily known to be involved in many plant developmental processes. Here, we characterized two pollen-expressed PERKs from Arabidopsis thaliana, PERK5 and PERK12. Pollen tube growth was impaired in single and double perk5-1 perk12-1 loss of function mutants, with an impact on seed production. When the segregation was analysed, a male gametophytic defect was found, indicating that perk5-1 and perk12-1 mutants carry deficient pollen transmission. Furthermore, perk5-1 perk12-1 displayed an excessive accumulation of pectins and cellulose at the cell wall of the pollen tubes. Our results indicate that PERK5 and PERK12 are necessary for proper pollen tube growth, highlighting their role in cell wall assembly and reactive oxygen species homeostasis.
Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Tubo Polínico/crescimento & desenvolvimento , Prolina/metabolismo , Proteínas Quinases/metabolismo , Espécies Reativas de Oxigênio/metabolismoRESUMO
The pollen and pistil RALF peptides, along with multiple receptor-like kinases and leucine-rich repeat extensins, regulate pollen tube growth and the final burst within the ovule, where sperm cells are released for fertilisation to occur. This review introduces some new questions that arose about the regulation of this complex process.
Assuntos
Proteínas de Arabidopsis , Arabidopsis , Peptídeos , Pólen , Tubo PolínicoRESUMO
To achieve fertilization, pollen tubes have to protect and properly deliver sperm cells through the pistil to the ovules. Pollen tube growth is a representative example of polarized growth where new components of the cell wall and plasma membrane are continuously deposited at the tip of the growing cell. The integrity of the cell wall is of fundamental importance to maintain apical growth. For this reason, pollen tube growth has become an excellent model to study the role of polysaccharides and structural cell wall proteins involved in polar cell expansion. However, quantification of structural polysaccharides at the pollen tube cell wall has been challenging due to technical complexity and the difficulty of finding specific dyes. Here, we propose simple methods for imaging and quantification of callose, pectin , and cellulose using specific dyes such as Aniline Blue, Propidium Iodide, and Pontamine Fast Scarlet 4B.
Assuntos
Parede Celular/metabolismo , Celulose/análise , Glucanos/análise , Pectinas/análise , Tubo Polínico/metabolismo , Coloração e Rotulagem/métodos , Arabidopsis , Parede Celular/química , Microscopia de Fluorescência/métodos , Tubo Polínico/citologiaRESUMO
Pollen tubes require functional mitochondria in order to achieve fast and sustained growth. In addition, cell wall expansion requires a calcium gradient in the tube apex formed by a dedicated array of calcium pumps and channels. Most studies have traditionally focused on the molecular aspects of calcium interactions and transport across the pollen tube plasmalemma. However, calcium transients across mitochondrial membranes from pollen tubes are beginning to be studied. Here, we report the presence of a ruthenium red-sensitive mitochondrial calcium uniporter-like activity in tobacco pollen tubes with functional oxidative phosphorylation. The present study provides a framework to measure in situ specifics of mitochondrial transport and respiration in pollen tubes from different plants. The relevance of a mitochondrial calcium uniporter for pollen tube growth is discussed.