Unpollinated Ovaries Used to Produce Doubled Haploid Lines in Durum Wheat.

The use of doubled haploid lines improves the efficiency of cultivar development and homozygous genotypes can be obtained in one generation, as opposed to conventional line production, which requires several cycles of self-pollination. However, in durum wheat (Triticum turgidum subsp. durum Desf.), the low efficiency of green plant regeneration and the very high frequency of albino plants hinder the application of this technique.We observed the success of using gynogenesis for durum wheat and the significant influence of growing conditions on ovary and callus development, and on plant regeneration. Our results suggested that the cold pretreatment for 2 weeks is efficient for durum wheat. Furthermore, the addition of 2,4-D, vitamins and glutamine, and the use of maltose as sugar source in media improved the ovary regeneration. We describe in this work an efficient method to regenerate green plants from in vitro durum wheat gynogenesis .

Mitochondrial heat-shock cognate protein 70 contributes to auxin-mediated embryo development.

In Arabidopsis thaliana, mitochondrial-localized heat-shock cognate protein 70-1 (mtHSC70-1) plays an important role in vegetativegrowth. However, whether mtHSC70-1 affects reproductive growth remains unknown. Here, we found that the mtHSC70-1 gene was expressed in the provascular cells of the embryo proper from the early heart stage onward during embryogenesis. Phenotypic analyses of mthsc70-1 mutants revealed that mtHSC70 deficiency leads to defective embryo development and that this effect is mediated by auxin. In addition to a dwarf phenotype, the mthsc70-1 mutant displayed defects in flower morphology, anther development, and embryogenesis. At early developmental stages, the mthsc70-1 embryos exhibited abnormal cell divisions in both embryo proper and suspensor cells. From heart stage onward, they displayed an abnormal shape such as with no or very small cotyledon protrusions, had aberrant number of cotyledons, or were twisted. These embryo defects were associated with reduced or ectopic expression of auxin responsive reporter DR5rev:GFP. Consistently, the expression of auxin biosynthesis and polar auxin transport genes were markedly altered in mthsc70-1. On the other hand, mitochondrial retrograde regulation (MRR) was enhanced in mthsc70-1. Treatment of wild-type plants with an inhibitor that activates mitochondrial retrograde signaling reduced the expression level of auxin biosynthesis and polar auxin transport genes and induced phenotypes similar to those of mthsc70-1. Taken together, our data reveal that loss of function of mtHSC70-1 induces MRR, which inhibits auxin biosynthesis and polar auxin transport, leading to abnormal auxin gradients and defective embryo development.

EMBRYO SAC DEVELOPMENT 1 affects seed setting rate in rice by controlling embryo sac development.

Seed setting rate is one of the critical factors that determine rice yield. Grain formation is a complex biological process, whose molecular mechanism is yet to be improved. Here we investigated the function of an OVATE family protein, Embryo Sac Development 1 (ESD1), in the regulation of seed setting rate in rice (Oryza sativa) by examining its loss-of-function mutants generated via clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated9 (Cas9) technology. ESD1 was predominantly expressed at Stage 6 of panicle development, especially in the ovules. esd1 mutants displayed reduced seed setting rates with normal stamen development and pollen tube growth but abnormal pistil group. Investigation of embryo sacs revealed that during the mitosis of functional megaspores, some egg cells degraded during differentiation in esd1 mutants, thereby hindering subsequent fertilization process and reducing seed setting rate. In addition, the transcriptional level of O. sativa anaphase-promoting complex 6, a reported embryo sac developing gene, was significantly reduced in esd1 mutants. These results support that ESD1 is an important modulator of ESD and seed setting rate in rice. Together, this finding demonstrates that ESD1 positively regulates the seed setting rate by controlling ESD in rice and has implications for the improvement of rice yield.

Gene expression trajectories during male and female reproductive development in balsam poplar (Populus balsamifera L.).

Plant reproductive development from the first appearance of reproductively committed axes through to floral maturation requires massive and rapid remarshalling of gene expression. In dioecious species such as poplar this is further complicated by divergent male and female developmental programs. We used seven time points in male and female balsam poplar (Populus balsamifera L.) buds and catkins representing the full annual flowering cycle, to elucidate the effects of time and sex on gene expression during reproductive development. Time (developmental stage) is dominant in patterning gene expression with the effect of sex nested within this. Here, we find (1) evidence for five successive waves of alterations to the chromatin landscape which may be important in setting the overall reproductive trajectory, regardless of sex. (2) Each individual developmental stage is further characterized by marked sex-differential gene expression. (3) Consistent sexually differentiated gene expression regardless of developmental stage reveal candidates for high-level regulators of sex and include the female-specific poplar ARR17 homologue. There is also consistent male-biased expression of the MADS-box genes PISTILLATA and APETALA3. Our work provides insights into expression trajectories shaping reproductive development, its potential underlying mechanisms, and sex-specific translation of the genome information into reproductive structures in balsam poplar.

Gene regulatory network controlling carpel number variation in cucumber.

The WUSCHEL-CLAVATA3 pathway genes play an essential role in shoot apical meristem maintenance and floral organ development, and under intense selection during crop domestication. The carpel number is an important fruit trait that affects fruit shape, size and internal quality in cucumber, but the molecular mechanism remains elusive. Here, we found that CsCLV3 expression was negatively correlated with carpel number in cucumber cultivars. CsCLV3-RNAi led to increased number of petals and carpels, whereas overexpression of CsWUS resulted in more sepals, petals and carpels, suggesting that CsCLV3 and CsWUS function as a negative and a positive regulator for carpel number variation, respectively. Biochemical analyses indicated that CsWUS directly bound to the promoter of CsCLV3 and activated its expression. Overexpression of CsFUL1A , a FRUITFULL-like MADS-box gene, resulted in more petals and carpels. CsFUL1A can directly bind to the CsWUS promoter to stimulate its expression. Furthermore, we found that auxin participated in carpel number variation in cucumber through interaction of CsARF14 with CsWUS. Therefore, we have identified a gene regulatory pathway involving CsCLV3, CsWUS, CsFUL1A and CsARF14 in determining carpel number variation in an important vegetable crop - cucumber.

WUSCHEL: a master regulator in plant growth signaling.

KEY MESSAGE: This review summarizes recent knowledge on functions of WUS and WUS-related homeobox (WOX) transcription factors in diverse signaling pathways governing shoot meristem biology and several other aspects of plant dynamics. Transcription factors (TFs) are master regulators involved in controlling different cellular and biological functions as well as diverse signaling pathways in plant growth and development. WUSCHEL (WUS) is a homeodomain transcription factor necessary for the maintenance of the stem cell niche in the shoot apical meristem, the differentiation of lateral primordia, plant cell totipotency and other diverse cellular processes. Recent research about WUS has uncovered several unique features including the complex signaling pathways that further improve the understanding of vital network for meristem biology and crop productivity. In addition, several reports bridge the gap between WUS expression and plant signaling pathway by identifying different WUS and WUS-related homeobox (WOX) genes during the formation of shoot (apical and axillary) meristems, vegetative-to-embryo transition, genetic transformation, and other aspects of plant growth and development. In this respect, the WOX family of TFs comprises multiple members involved in diverse signaling pathways, but how these pathways are regulated remains to be elucidated. Here, we review the current status and recent discoveries on the functions of WUS and newly identified WOX family members in the regulatory network of various aspects of plant dynamics.

Evaluation of in vitro genotoxic effects induced by in vitro anther culture conditions in sunflower.

Sunflower is a globally important oilseed, food, and ornamental crop. This study seeks to investigate the genotoxic effects of tissue culture parameters in sunflower calli tissues belongs to two genotypes obtained via anther culture. Anthers were pretreated with cold for 24 hours at 4°C and heat for 2 days at 35°C in the dark and plated onto media supplemented with different concentrations and combinations of 6-benzylaminopurine, 2,4-dichlorophenoxyacetic acid, α-naphthalene acetic acid and indole-3-acetic acid. Obtaining calli tissues were used to detect the DNA damage levels by Comet assay, evaluating changes on superoxide dismutase and guaiacol peroxidase activities derived from in vitro culture factors. 0.5 mg/L 2,4-dichlorophenoxyacetic acid and 2 mg/L α-naphthalene acetic acid from plant growth regulators showed acute genotoxic effect while 0.5 mg/L indole-3-acetic acid and 0.5 mg/L α-naphthalene acetic acid showed no genotoxic effect. Total protein content analysis of antioxidant enzymes revealed that although superoxide dismutase activity did not increase, Guaiacol peroxidase (GPOX) activity decreased in comparison to control. The obtained results have indicated that in vitro culture factors apparently lead to genotoxicity and oxidative stress.

RcAP1, a Homolog of APETALA1, is Associated with Flower Bud Differentiation and Floral Organ Morphogenesis in Rosa chinensis.

Rosa chinensis is one of the most popular flower plants worldwide. The recurrent flowering trait greatly enhances the ornamental value of roses, and is the result of the constant formation of new flower buds. Flower bud differentiation has always been a major topic of interest among researchers. The APETALA1 (AP1) MADS-box (Mcm1, Agamous, Deficiens and SRF) transcription factor-encoding gene is important for the formation of the floral meristem and floral organs. However, research on the rose AP1 gene has been limited. Thus, we isolated AP1 from Rosa chinensis 'Old Blush'. An expression analysis revealed that RcAP1 was not expressed before the floral primordia formation stage in flower buds. The overexpression of RcAP1 in Arabidopsis thaliana resulted in an early-flowering phenotype. Additionally, the virus-induced down-regulation of RcAP1 expression delayed flowering in 'Old Blush'. Moreover, RcAP1 was specifically expressed in the sepals of floral organs, while its expression was down-regulated in abnormal sepals and leaf-like organs. These observations suggest that RcAP1 may contribute to rose bud differentiation as well as floral organ morphogenesis, especially the sepals. These results may help for further characterization of the regulatory mechanisms of the recurrent flowering trait in rose.

Arabinogalactan proteins mediate intercellular crosstalk in the ovule of apple flowers.

KEY MESSAGE: AGP-rich glycoproteins mediate pollen-ovule interactions and cell patterning in the embryo sac of apple before and after fertilization. Glycoproteins are significant players in the dialog that takes place between growing pollen tubes and the stigma and style in the angiosperms. Yet, information is scarce on their possible involvement in the ovule, a sporophytic organ that hosts the female gametophyte. Apple flowers have a prolonged lapse of time between pollination and fertilization, offering a great system to study the developmental basis of glycoprotein secretion and their putative role during the last stages of the progamic phase and early seed initiation. For this purpose, the sequential pollen tube elongation within the ovary was examined in relation to changes in arabinogalactan proteins (AGPs) in the tissues of the ovule before and after fertilization. To evaluate what of these changes are developmentally regulated, unpollinated and pollinated flowers were compared. AGPs paved the pollen tube pathway in the ovules along the micropylar canal, and the nucellus entrance toward the synergids, which also developmentally accumulated AGPs at the filiform apparatus. Glycoproteins vanished from all these tissues following pollen tube passage, strongly suggesting a role in pollen-ovule interaction. In addition, AGPs marked the primary cell walls of the haploid cells of the female gametophyte, and they further built up in the cell walls of the embryo sac and developing embryo, layering the interactive walls of the three generations hosted in the ovule, the maternal sporophytic tissues, the female gametophyte, and the developing embryo.

Effects of High Temperature on Embryological Development and Hormone Profile in Flowers and Leaves of Common Buckwheat (Fagopyrum esculentum Moench).

Common buckwheat is a valuable crop, mainly due to the beneficial chemical composition of its seeds. However, buckwheat cultivation is limited because of unstable seed yield. The most important reasons for the low yield include embryo and flower abortion. The aim of this work is to verify whether high temperature affects embryological development in this plant species. The experiment was conducted on plants of a Polish cultivar 'Panda' and strain PA15, in which the percentage of degenerating embryo sacs was previously determined and amounted to 32% and 10%, respectively. The plants were cultivated in phytotronic conditions at 20 °C (control), and 30 °C (thermal stress). The embryological processes and hormonal profiles in flowers at various developmental stages (buds, open flowers, and wilted flowers) and in donor leaves were analyzed in two-month-old plants. Significant effects of thermal stress on the defective development of female gametophytes and hormone content in flowers and leaves were observed. Ovules were much more sensitive to high temperature than pollen grains in both genotypes. Pollen viability remained unaffected at 30 °C in both genotypes. The effect of temperature on female gametophyte development was visible in cv. Panda but not in PA15 buds. A drastic reduction in the number of properly developed embryo sacs was clear in open flowers at 30 °C in both genotypes. A considerable increase in abscisic acid in open flowers ready for fertilization may serve as a signal inducing flower senescence observed in the next few days. Based on embryological analyses and hormone profiles in flowers, we conclude that cv. 'Panda' is more sensitive to thermal stress than strain PA15, mainly due to a much earlier response to thermal stress involving impairment of embryological processes already in the flower buds.

Somatic Embryogenesis in Theobroma cacao L.

Theobroma cacao L. is a tropical tree originating in the Amazon, where it grows naturally in the shade of tropical rainforests. Cacao sub-products, such as butter and powder, are produced as principal components of chocolate and contain important nutritional compounds such as polyphenols and flavonoids. However, bean production is decreasing because plantations are antiquated and unproductive. Cacao propagation has been traditionally performed through classical propagation methods, such as grafting or rooted cuttings, but those methods are not sufficient to obtain large quantities of planting material with the desired genetic quality and optimal plant health. In the search for solutions to this problem, somatic embryogenesis (SE) is a vegetative method used for cacao propagation that has the potential to be explored. SE is a type of clonal propagation by which totipotent cells in the somatic tissue can develop into embryos and subsequently convert into plants.This method offers significant technological advantages because it is possible to obtain a large quantity of disease-free planting material with good agronomic characteristics and genetic stability. In T. cacao, tow techniques of in vitro micropropagation have been reported as direct and indirect SE. Indirect SE requires the additional step of cell dedifferentiation, unlike direct SE, which does not require this step. Here, we report a protocol using direct and indirect SE techniques using two types of culture methodologies-solid and liquid culture media.

Spatial dynamics of floral organ formation.

Understanding the emergence of biological structures and their changes is a complex problem. On a biochemical level, it is based on gene regulatory networks (GRN) consisting on interactions between the genes responsible for cell differentiation and coupled in a greater scale with external factors. In this work we provide a systematic methodological framework to construct Waddington's epigenetic landscape of the GRN involved in cellular determination during the early stages of development of angiosperms. As a specific example we consider the flower of the plant Arabidopsis thaliana. Our model, which is based on experimental data, recovers accurately the spatial configuration of the flower during cell fate determination, not only for the wild type, but for its homeotic mutants as well. The method developed in this project is general enough to be used in the study of the relationship between genotype-phenotype in other living organisms.

The calmodulin-like protein, CML39, is involved in regulating seed development, germination, and fruit development in Arabidopsis.

KEY MESSAGE: We show that the calcium sensor, CML39, is important in various developmental processes from seeds to mature plants. This study bridges previous work on CML39 as a stress-induced gene and highlights the importance of calcium signalling in plant development. In addition to the evolutionarily-conserved Ca2+ sensor, calmodulin (CaM), plants possess a large family of CaM-related proteins (CMLs). Using a cml39 loss-of-function mutant, we investigated the roles of CML39 in Arabidopsis and discovered a range of phenotypes across developmental stages and in different tissues. In mature plants, loss of CML39 results in shorter siliques, reduced seed number per silique, and reduced number of ovules per pistil. We also observed changes in seed development, germination, and seed coat properties in cml39 mutants in comparison to wild-type plants. Using radicle emergence as a measure of germination, cml39 mutants showed more rapid germination than wild-type plants. In marked contrast to wild-type seeds, the germination of developing, immature cml39 seeds was not sensitive to cold-stratification. In addition, germination of cml39 seeds was less sensitive than wild-type to inhibition by ABA or by treatments that impaired gibberellic acid biosynthesis. Tetrazolium red staining indicated that the seed-coat permeability of cml39 seeds is greater than that of wild-type seeds. RNA sequencing analysis of cml39 seedlings suggests that changes in chromatin modification may underlie some of the phenotypes associated with cml39 mutants, consistent with previous reports that orthologs of CML39 participate in gene silencing. Aberrant ectopic expression of transcripts for seed storage proteins in 7-day old cml39 seedlings was observed, suggesting mis-regulation of early developmental programs. Collectively, our data support a model where CML39 serves as an important Ca2+ sensor during ovule and seed development, as well as during germination and seedling establishment.

Auxin Response Dynamics During Wild-Type and entire Flower Development in Tomato.

The plant hormone auxin is a major regulator of plant development and response to environmental cues. Auxin plays a particularly central role in flower development, but the knowledge of its role of flower development in crop plants with fleshy fruits, such as tomato, is still scarce. Mutations in the Aux/IAA gene ENTIRE/Indole Acetic Acid 9 (E/IAA9) lead to the precocious development of young gynoecia into parthenocarpic fruits. Here, we compared the distribution of the auxin response sensor DR5::VENUS and the auxin efflux transporter PIN1 between the wild type and entire during successive stages of flower and fruit development. Up-regulation of the DR5::VENUS signal in the shoot apical meristem (SAM) was observed upon the transition to flowering, implicating a possible role for auxin in the transition from a vegetative SAM into an inflorescence meristem. DR5::VENUS was expressed in all initiating floral organs. Additionally, DR5::VENUS was highly expressed during gametogenesis, in both male and female organs, and in the developing seeds during embryogenesis. DR5::VENUS is expressed in functional cell layers such as the anther stomium and tapetum, suggesting that auxin plays a role in flower organ development and function. The entire mutation affected DR5::VENUS expression patterns during inflorescence formation and flower organ development, which correlated with phenotypic alterations. We also show dynamic distribution and localization of the auxin transporter PIN1 during flower and fruit organ development. These results emphasize the dynamic auxin response in inflorescence and flower development and suggest multiple roles of auxin in these processes.

Cytokinin-Auxin Crosstalk in the Gynoecial Primordium Ensures Correct Domain Patterning.

The Arabidopsis (Arabidopsis thaliana) gynoecium consists of two congenitally fused carpels made up of two lateral valve domains and two medial domains, which retain meristematic properties and later fuse to produce the female reproductive structures vital for fertilization. Polar auxin transport (PAT) is important for setting up distinct apical auxin signaling domains in the early floral meristem remnants allowing for lateral domain identity and outgrowth. Crosstalk between auxin and cytokinin plays an important role in the development of other meristematic tissues, but hormone interaction studies to date have focused on more accessible later-stage gynoecia and the spatiotemporal interactions pivotal for patterning of early gynoecium primordia remain unknown. Focusing on the earliest stages, we propose a cytokinin-auxin feedback model during early gynoecium patterning and hormone homeostasis. Our results suggest that cytokinin positively regulates auxin signaling in the incipient gynoecial primordium and strengthen the concept that cytokinin regulates auxin homeostasis during gynoecium development. Specifically, medial cytokinin promotes auxin biosynthesis components [YUCCA1/4 (YUC1/4)] in, and PINFORMED7 (PIN7)-mediated auxin efflux from, the medial domain. The resulting laterally focused auxin signaling triggers ARABIDOPSIS HISTIDINE PHOSPHOTRANSFER PROTEIN6 (AHP6), which then represses cytokinin signaling in a PAT-dependent feedback. Cytokinin also down-regulates PIN3, promoting auxin accumulation in the apex. The yuc1, yuc4, and ahp6 mutants are hypersensitive to exogenous cytokinin and 1-napthylphthalamic acid (NPA), highlighting their role in mediolateral gynoecium patterning. In summary, these mechanisms self-regulate cytokinin and auxin signaling domains, ensuring correct domain specification and gynoecium development.

Identification of pre-fertilization reproductive barriers and the underlying cytological mechanism in crosses among three petal-types of Jasminum sambac and their relevance to phylogenetic relationships.

Crosses among single-, double- and multi-petal jasmine cultivars (Jasminum sambac Aiton) are unable to easily generate hybrids. To identify the reproductive barriers restricting hybrid set, dynamic changes in jasmine pollen viability and pistil receptivity were compared at different flowering stages. Pollen-pistil interactions in six reciprocal crosses were also investigated to characterize pollen-stigma compatibility. Additionally, paraffin sections of pollinated embryo sacs were prepared for subsequent analyses of developmental status. Furthermore, pistil cell ultrastructural characteristics were observed to reveal cytological mechanism regulating pistil receptivity and the pollen-pistil interactions. We observed that pollen viability and stigma receptivity varied depending on petal phenotype and flowering stage and were easily lost during flowering. Different reciprocal crosses exhibited varied pollen-stigma compatibilities according to the pollen germination rates. Although some pollen grains germinated normally on maternal stigmas, the pollen tubes were arrested in the pistils and were unable to reach the ovaries. Additionally, the embryo sacs remained unfertilized until degenerating. Therefore, jasmine crosses are affected by pre-fertilization reproductive barriers. Low pollen fertility and poor stigma receptivity are detrimental to pollen germination and pollen-pistil compatibility, indicating they are two factors affecting hybrid set. Ultrastructural observation of the pistil cells revealed that cell death occurred during flowering. Thus, the early and rapid senescence of pistils is likely responsible for the decreased pistil receptivity and inhibited pollen tube growth. These findings may be relevant for future jasmine hybridizations. They provide new insights for the development of methods to overcome reproductive barriers and may also be useful for clarifying the phylogenetic relationships among jasmine cultivars with differing petal phenotypes.

Embryology of Pera (Peraceae, Malpighiales): systematics and evolutionary implications.

Pera is a neotropical genus that currently belongs to the family Peraceae. This circumscription resulted from an inclusion of the Rafflesiaceae between the old tribe Pereae and all other Euphorbiaceae, and wherein Pereae was elevated to family rank making Euphorbiaceae monophyletic again. These changes are necessary although Rafflesiaceae are holoparasitic with extremely reduced vegetative bodies and large flowers while Peraceae and Euphorbiaceae have well developed vegetative parts and reduced flowers. As the embryology of Peraceae was poorly known, and embryological processes are conservative, we studied the embryology of Pera glabrata, searching for similarities between Peraceae, Rafflesiaceae, and Euphorbiaceae that could support this grouping. Usual methods of light microscopy and scanning electron microscopy were utilised. The results show endothecium with reversed-T-shaped cells, prismatic crystals in the tapetum, and disintegrated aerenchymatous septum in the mature fruit as unique features for Peraceae and possibly apomorphies for the family. In addition to the unisexual flowers, porogamous fertilization is present and one ovule per carpel which may support the Peraceae-Rafflesiaceae-Euphorbiaceae clade. The comparative approach also suggests possible (syn-)apomorphies for linoids and phyllanthoids, only linoids, Rafflesiaceae, Euphorbiaceae, and Ixonanthaceae. The presence of a placental obturator found previously unknown in Peraceae emerged as a possible synapomorphy for the euphorbioids (including Ixonanthaceae, Linaceae, Phyllanthaceae, Picrodendraceae, Peraceae, Rafflesiaceae, and Euphorbiaceae), which appeared in a common ancestor of the group and has been lost in Rafflesiaceae.

MS23, a master basic helix-loop-helix factor, regulates the specification and development of the tapetum in maize.

Successful male gametogenesis involves orchestration of sequential gene regulation for somatic differentiation in pre-meiotic anthers. We report here the cloning of Male Sterile23 (Ms23), encoding an anther-specific predicted basic helix-loop-helix (bHLH) transcription factor required for tapetal differentiation; transcripts localize initially to the precursor secondary parietal cells then predominantly to daughter tapetal cells. In knockout ms23-ref mutant anthers, five instead of the normal four wall layers are observed. Microarray transcript profiling demonstrates a more severe developmental disruption in ms23-ref than in ms32 anthers, which possess a different bHLH defect. RNA-seq and proteomics data together with yeast two-hybrid assays suggest that MS23 along with MS32, bHLH122 and bHLH51 act sequentially as either homo- or heterodimers to choreograph tapetal development. Among them, MS23 is the earliest-acting factor, upstream of bHLH51 and bHLH122, controlling tapetal specification and maturation. By contrast, MS32 is constitutive and independently regulated and is required later than MS23 in tapetal differentiation.

Starch Turnover and Metabolism during Flower and Early Embryo Development.

The accumulation of starch within photosynthetic tissues and within dedicated storage organs has been characterized extensively in many species, and a function in buffering carbon availability or in fueling later growth phases, respectively, has been proposed. However, developmentally regulated starch turnover within heterotrophic tissues other than dedicated storage organs is poorly characterized, and its function is not well understood. Here, we report on the characterization of starch turnover during flower, early embryo, and silique development in Arabidopsis (Arabidopsis thaliana) using a combined clearing-staining technique on whole-mount tissue. Besides the two previously documented waves of transient starch accumulation in the stamen envelope, occurring during meiosis and pollen mitosis I, we identified a novel, third wave of starch amylogenesis/amylolysis during the last stages of stamen development. To gain insights into the underlying molecular mechanisms, we analyzed publicly available microarray data, which revealed a developmentally coordinated expression of carbohydrate transport and metabolism genes during these waves of transient starch accumulation. Based on this analysis, we characterized starch dynamics in mutants affecting hexose phosphate metabolism and translocation, and identified the Glc-6-phosphate/phosphate antiporter GPT1 as the putative translocator of Glc-6-phosphate for starch biosynthesis in reproductive tissues. Based on these results, we propose a model of starch synthesis within the pollen grain and discuss the nutrient transport route feeding the embryo within the developing seed.

Cytokinin response factors integrate auxin and cytokinin pathways for female reproductive organ development.

The developmental programme of the pistil is under the control of both auxin and cytokinin. Crosstalk between these factors converges on regulation of the auxin carrier PIN-FORMED 1 (PIN1). Here, we show that in the triple transcription factor mutant cytokinin response factor 2 (crf2) crf3 crf6 both pistil length and ovule number were reduced. PIN1 expression was also lower in the triple mutant and the phenotypes could not be rescued by exogenous cytokinin application. pin1 complementation studies using genomic PIN1 constructs showed that the pistil phenotypes were only rescued when the PCRE1 domain, to which CRFs bind, was present. Without this domain, pin mutants resemble the crf2 crf3 crf6 triple mutant, indicating the pivotal role of CRFs in auxin-cytokinin crosstalk.