Plants expressing a miR164-resistant CUC2 gene reveal the importance of post-meristematic maintenance of phyllotaxy in Arabidopsis

In plants, the arrangement of organs along the stem (phyllotaxy) follows a predictable pattern. Recent studies have shown that primordium position at the meristem is governed by local auxin gradients, but little is known about the subsequent events leading to the phyllotaxy along the mature stem. We show here that plants expressing a miR164-resistant CUP-SHAPED COTYLEDON2 (CUC2) gene have an abnormal phyllotactic pattern in the fully grown stem, despite the pattern of organ initiation by the meristem being normal. This implies that abnormal phyllotaxy is generated during stem growth. These plants ectopically express CUC2 in the stem, suggesting that the proper timing of CUC2 expression is required to maintain the pattern initiated in the meristem. Furthermore, by carefully comparing the phyllotaxy in the meristem and along the mature inflorescence in wild types, we show that such deviation also occurs during wild-type development, although to a smaller extent. We therefore suggest that the phyllotactic pattern in a fully grown stem results not only from the organogenetic activity of the meristem, but also from the subsequent growth pattern during stem development.     

The Arabidopsis FILAMENTOUS FLOWER gene is required for flower formation.

A screen for mutations affecting flower formation was carried out and several filamentous flower (fil) alleles were identified. In fil mutants, floral primordia occasionally give rise to pedicels lacking flowers at their ends. This defect is dramatically enhanced in fil rev double mutants, in which every floral primordium produces a flowerless pedicel. These data suggest that the FIL and REV genes are required for an early step of flower formation, possibly for the establishment of a flower-forming domain within the floral primordium. The FIL gene is also required for establishment of floral meristem identity and for flower development. During flower development, the FIL gene is required for floral organ formation in terms of the correct numbers and positions; correct spatial activity of the AGAMOUS, APETALA3, PISTILLATA and SUPERMAN genes; and floral organ development.     

CHANGE IN EPILOBIUM PHYLLOTAXY DURING REPRODUCTIVE TRANSITION

The ontogeny of Epilobium hirsutum grown under natural summer photoperiod in a glasshouse was divided into vegetative, early transitional, transitional, and floral stages. Bijugate phyllotaxy, common to both the vegetative and early transitional stages, is transformed into spiral phyllotaxy during the transitional stage by an initial change in the divergence angle of a single primordium inserted at a unique level on the shoot. Leaf primordia subsequently are inserted in a spiral arrangement in the indeterminate floral shoot apex. The early transitional shoot apical meristem is about 1.5 times the volume of the vegetative meristem but expands at about two-thirds the relative plastochron rate of volume increment of the vegetative meristem. There are progressive decreases in the plastochron and relative plastochron rates of radial and vertical shoot growth through ontogeny. Relative chronological rates of shoot growth, however, are not altered during ontogeny. Spiral transformation results from changes in the relative points of insertion of leaf primordia on the shoot meristem. These changes are accompanied by an increased rate of primordia initiation on a more circular shoot meristem. The change in phyllotaxy during ontogeny is similar to that which was artificially induced by chemical modification of auxin concentration gradients in the shoot apex, with the additional feature that there is an initial increase in the volume of the shoot meristem prior to the natural spiral transformation. Size of the shoot apical meristem, however, appears to have little influence on Epilobium phyllotaxy; but the geometric shape of the meristem is well correlated with bijugate to spiral transformations. This suggests that geometric parameters of the shoot meristem should be considered in theoretical models of phyllotaxy.     

Studies of aberrant phyllotaxy1 Mutants of Maize Indicate Complex Interactions between Auxin and Cytokinin Signaling in the Shoot Apical Meristem

One of the most fascinating aspects of plant morphology is the regular geometric arrangement of leaves and flowers, called phyllotaxy. The shoot apical meristem (SAM) determines these patterns, which vary depending on species and developmental stage. Auxin acts as an instructive signal in leaf initiation, and its transport has been implicated in phyllotaxy regulation in Arabidopsis (Arabidopsis thaliana). Altered phyllotactic patterns are observed in a maize (Zea mays) mutant, aberrant phyllotaxy1 (abph1, also known as abphyl1), and ABPH1 encodes a cytokinin-inducible type A response regulator, suggesting that cytokinin signals are also involved in the mechanism by which phyllotactic patterns are established. Therefore, we investigated the interaction between auxin and cytokinin signaling in phyllotaxy. Treatment of maize shoots with a polar auxin transport inhibitor, 1-naphthylphthalamic acid, strongly reduced ABPH1 expression, suggesting that auxin or its polar transport is required for ABPH1 expression. Immunolocalization of the PINFORMED1 (PIN1) polar auxin transporter revealed that PIN1 expression marks leaf primordia in maize, similarly to Arabidopsis. Interestingly, maize PIN1 expression at the incipient leaf primordium was greatly reduced in abph1 mutants. Consistently, auxin levels were reduced in abph1, and the maize PIN1 homolog was induced not only by auxin but also by cytokinin treatments. Our results indicate distinct roles for ABPH1 as a negative regulator of SAM size and a positive regulator of PIN1 expression. These studies highlight a complex interaction between auxin and cytokinin signaling in the specification of phyllotactic patterns and suggest an alternative model for the generation of altered phyllotactic patterns in abph1 mutants. We propose that reduced auxin levels and PIN1 expression in abph1 mutant SAMs delay leaf initiation, contributing to the enlarged SAM and altered phyllotaxy of these mutants.     

榛属（桦木科）花序及花的形态发生

在扫描电镜下观察了桦木科榛属榛、毛榛和滇榛的花序和花的形态发生过程。榛属雌花序由多个小聚伞花序螺旋状排列组成;每个小花序原基分化出1枚初级苞片和一团小花序原基分生组织,由小花序原基分生组织分化形成2个花原基;每个花原基分化出2个心皮原基,形成二心皮雌蕊;雌蕊基部有2层花被原基,内层花被原基环状,外层花被发生于花原基近轴面和远轴面,近轴面和远轴面的花被不均等分化,外层花被发生早于内层花被。雄花序为柔荑状,由多个小聚伞花序螺旋状排列组成。每个小花序原基分化出1枚初级苞片和一团小花序原基分生组织,由小花序原基分生组织分化出2枚次级苞片和4。6个雄蕊原基,形成4—6枚雄蕊,每个雄蕊具4个药囊,在雄蕊原基分化形成4药囊雄蕊过程中．出现雄蕊原基纵裂。并且花丝纵裂至基部。为进一步全面探讨桦木科属间系统演化关系提供了证据。     

榛属 (桦木科) 花序及花的形态发生

在扫描电镜下观察了桦木科榛属榛、毛榛和滇榛的花序和花的形态发生过程。榛属雌花序由多个小聚伞花序螺旋状排列组成;每个小花序原基分化出1枚初级苞片和一团小花序原基分生组织,由小花序原基分生组织分化形成2个花原基;每个花原基分化出2个心皮原基,形成二心皮雌蕊;雌蕊基部有2层花被原基,内层花被原基环状,外层花被发生于花原基近轴面和远轴面,近轴面和远轴面的花被不均等分化,外层花被发生早于内层花被。雄花序为柔荑状,由多个小聚伞花序螺旋状排列组成。每个小花序原基分化出1枚初级苞片和一团小花序原基分生组织,由小花序原基分生组织分化出2枚次级苞片和4~6个雄蕊原基,形成4~6枚雄蕊,每个雄蕊具4个药囊,在雄蕊原基分化形成4药囊雄蕊过程中,出现雄蕊原基纵裂,并且花丝纵裂至基部。为进一步全面探讨桦木科属间系统演化关系提供了证据。     

Patterns of auxin transport and gene expression during primordium development revealed by live imaging of the Arabidopsis inflorescence meristem

BACKGROUND: Plants produce leaf and flower primordia from a specialized tissue called the shoot apical meristem (SAM). Genetic studies have identified a large number of genes that affect various aspects of primordium development including positioning, growth, and differentiation. So far, however, a detailed understanding of the spatio-temporal sequence of events leading to primordium development has not been established. RESULTS: We use confocal imaging of green fluorescent protein (GFP) reporter genes in living plants to monitor the expression patterns of multiple proteins and genes involved in flower primordial developmental processes. By monitoring the expression and polarity of PINFORMED1 (PIN1), the auxin efflux facilitator, and the expression of the auxin-responsive reporter DR5, we reveal stereotypical PIN1 polarity changes which, together with auxin induction experiments, suggest that cycles of auxin build-up and depletion accompany, and may direct, different stages of primordium development. Imaging of multiple GFP-protein fusions shows that these dynamics also correlate with the specification of primordial boundary domains, organ polarity axes, and the sites of floral meristem initiation. CONCLUSIONS: These results provide new insight into auxin transport dynamics during primordial positioning and suggest a role for auxin transport in influencing primordial cell type.     

Shoot organization genes regulate shoot apical meristem organization and the pattern of leaf primordium initiation in rice

The mechanism regulating the pattern of leaf initiation was analyzed by using shoot organization (sho) mutants derived from three loci (SHO1, SHO2, and SHO3). In the early vegetative phase, sho mutants show an increased rate of leaf production with random phyllotaxy. The resulting leaves are malformed, threadlike, or short and narrow. Their shoot apical meristems are relatively low and wide, that is, flat shaped, although their shape and size are highly variable among plants of the same genotype. Statistical analysis reveals that the shape of the shoot meristem rather than its size is closely correlated with the variations of plastochron and phyllotaxy. Rapid and random leaf production in sho mutants is correlated with the frequent and disorganized cell divisions in the shoot meristem and with a reduction of expression domain of a rice homeobox gene, OSH1. These changes in the organization and behavior of the shoot apical meristems suggest that sho mutants have fewer indeterminate cells and more determinate cells than wild type, with many cells acting as leaf founder cells. Thus, the SHO genes have an important role in maintaining the proper organization of the shoot apical meristem, which is essential for the normal initiation pattern of leaf primordia.     

Reversion of flowering in Glycine Max (Fabaceae)

Photoperiodic changes, if occurring before a commitment to flowering is established, can alter the morphological pattern of plant development. In this study, Glycine max (L.) Merrill cv. Ransom plants were initially grown under an inductive short-day (SD) photoperiod to promote flower evocation and then transferred to a long-day (LD) photoperiod to delay flower development by reestablishing vegetative growth (SD-LD plants). Some plants were transferred back to SD after 4-LD exposures to repromote flowering (SD-LD-SD plants). Alterations in organ initiation patterns, from floral to vegetative and back to floral, are characteristic of a reversion phenomenon. Morphological features that occurred at the shoot apical meristem in SD, LD, SD-LD, and SD-LD-SD plants were observed using scanning electron microscopy (SEM). Reverted plants initiated floral bracts and resumed initiation of trifoliolate leaves in the two-fifths floral phyllotaxy prior to terminal inflorescence development. When these plants matured, leaf-bract intermediates were positioned on the main stem instead of trifoliolate leaves. Plants transferred back to a SD photoperiod flowered earlier than those left in LD conditions. Results indicated that in plants transferred between SDs and LDs, photoperiod can influence organ initiation in florally evoked, but not committed, G. max plants.     

CHANGES IN EPILOBIUM PHYLLOTAXY INDUCED BY N-1-NAPHTHYLPHTHALAMIC ACID AND α-4-CHLOROPHENOXYISOBUTYRIC ACID

Preliminary studies establishing relationships between leaf plastochron index and Epilobium hirsutum L. shoot growth provide a method for rigorous selection of plants utilized in experiments designed to test the working hypothesis that endogenous auxin gradient interactions are factors of phyllotactic control in this species. Application of N-1-naphthylphthalamic acid (NPA), an auxin transport inhibitor, to one of the youngest bijugate primordia on the shoot meristem results in increased growth of the treated primordium. Fasciation between the treated primordium and one of the next primordia to be initiated alters relative vertical spacing of primordia. Angular shifts between subsequent primordia result in spiral transformation of Epilobium bijugate phyllotaxy. Application of α-4-chlorophenoxyisobutyric acid (CPIB), an auxin antagonist, to one of the youngest bijugate primordia on the shoot meristem results in decreased growth of the treated primordium that alters both radial and vertical spacing of primordia. This is followed by angular shifts between subsequent primordia resulting in spiral transformation of the bijugate phyllotaxy. Changes in the growth parameters of NPA- and CPIB-treated shoots are similar. Relative plastochron rates of radial and vertical shoot growth of induced spiral shoots are about half those of lanolin paste control shoots, as are the plastochrons and relative plastochron rates of leaf elongation. Treated shoot meristems have eccentricities of 0.5 as compared to bijugate control meristem eccentricities of 0.7. No significant difference is apparent between basal transverse areas of treated and control shoot meristems. The relative chronological rates of growth of treated shoots are not significantly different from those rates of control shoots. Spiral transformation results from changes in relative positions of leaf primordia insertion on the shoot meristem, not from changes in growth of treated shoots. These changes are accompanied by an increased rate of leaf initiation on a more circular shoot meristem. Existing theoretical models of phyllotaxy are discussed in relation to these chemically induced changes of Epilobium leaf arrangement.     

Initiation of axillary and floral meristems in Arabidopsis

Shoot development is reiterative: shoot apical meristems (SAMs) give rise to branches made of repeating leaf and stem units with new SAMs in turn formed in the axils of the leaves. Thus, new axes of growth are established on preexisting axes. Here we describe the formation of axillary meristems and floral meristems in Arabidopsis by monitoring the expression of the SHOOT MERISTEMLESS and AINTEGUMENTA genes. Expression of these genes is associated with SAMs and organ primordia, respectively. Four stages of axillary meristem development and previously undefined substages of floral meristem development are described. We find parallels between the development of axillary meristems and the development of floral meristems. Although Arabidopsis flowers develop in the apparent absence of a subtending leaf, the expression patterns of AINTEGUMENTA and SHOOT MERISTEMLESS RNAs during flower development suggest the presence of a highly reduced, "cryptic" leaf subtending the flower in Arabidopsis. We hypothesize that the STM-negative region that develops on the flanks of the inflorescence meristem is a bract primordium and that the floral meristem proper develops in the "axil" of this bract primordium. The bract primordium, although initially specified, becomes repressed in its growth.     

The indeterminate floral apex1 gene regulates meristem determinacy and identity in the maize inflorescence

Meristems may be determinate or indeterminate. In maize, the indeterminate inflorescence meristem produces three types of determinate meristems: spikelet pair, spikelet and floral meristems. These meristems are defined by their position and their products. We have discovered a gene in maize, indeterminate floral apex1 (ifa1) that regulates meristem determinacy. The defect found in ifa1 mutants is specific to meristems and does not affect lateral organs. In ifa1 mutants, the determinate meristems become less determinate. The spikelet pair meristem initiates more than a pair of spikelets and the spikelet meristem initiates more than the normal two flowers. The floral meristem initiates all organs correctly, but the ovule primordium, the terminal product of the floral meristem, enlarges and proliferates, expressing both meristem and ovule marker genes. A role for ifa1 in meristem identity in addition to meristem determinacy was revealed by double mutant analysis. In zea agamous1 (zag1) ifa1 double mutants, the female floral meristem converts to a branch meristem whereas the male floral meristem converts to a spikelet meristem. In indeterminate spikelet1 (ids1) ifa1 double mutants, female spikelet meristems convert to branch meristems and male spikelet meristems convert to spikelet pair meristems. The double mutant phenotypes suggest that the specification of meristems in the maize inflorescence involves distinct steps in an integrated process.     

Activation of Basal Cells of the Apical Meristem during Sepal Formation in Tomato

Although great advances have been made in research on the regulation of primordium fate in the floral meristem, our understanding of the molecular events occurring during the floral transition remains incomplete. Via a careful analysis of the expression patterns of five genes encoding housekeeping functions during the floral transition in tomato (using both in situ hybridization and enzyme histochemistry), we identified a particular phase of floral development (sepal initiation) at which cells located toward the base of the meristem show a high level of cellular metabolism, whereas cells at the tip of the meristem dome show little activity. At other stages of floral development, the probes used showed genespecific patterns of expression generally consistent with our previous investigation of the vegetative apical meristem. Our data, in conjunction with other reports in the literature, enabled us to postulate that relative activation of basal cells of the meristem may be of general occurrence during the transition to flowering. Such a hypothesis could account for recent observations using periclinal chimeras on the effect of L3 genotypes on flower development and have a bearing on the expected mechanism by which the number of primordia generated by a floral meristem is regulated.     

The Evergreen gene is essential for flower initiation in carnation

One of the leading cut-flower crops in the world, the greenhouse carnation (Dianthus caryophyllus), has been subjected to intense breeding efforts for the past few hundred years. As an ornamental crop, flowering and flower architecture are major breeding targets that are constantly in demand. In an ongoing breeding program aimed at improving these characteristics, two mutants heterozygous for a mutation in a gene termed evergreen (e) were identified. In these mutants, spike-like clusters of bracteoles subtend each flower. Genetic analysis of the mutants confirmed the semidominant nature of this nuclear mutation and that the two original mutants were allelic at the evergreen locus. In homozygous mutant plants, a more severe phenotype was observed. Flower formation was completely blocked and spikelike clusters of bracteoles did not subtend any flowers. Morphological characterization of mutant plants revealed that vegetative growth and inflorescence structure are not affected by the mutant allele. In plants heterozygous for the evergreen mutation, fertility, petal and pistil length, calyx diameter, and stamen number were not affected. However, flowers from these heterozygous plants had a reduced number of petals, suggesting an intriguing link between evergreen and the double flower (d) gene that determines petal number in carnation. The control by evergreen of bracteole formation, floral meristem initiation, and petal number in carnation is discussed in comparison to the recessive leafy (lfy) and floricaula (flo) mutants of Arabidopsis and Antirrhinum, respectively.     

SUPERMAN, a regulator of floral homeotic genes in Arabidopsis.

We describe a locus, SUPERMAN, mutations in which result in extra stamens developing at the expense of the central carpels in the Arabidopsis thaliana flower. The development of superman flowers, from initial primordium to mature flower, is described by scanning electron microscopy. The development of doubly and triply mutant strains, constructed with superman alleles and previously identified homeotic mutations that cause alterations in floral organ identity, is also described. Essentially additive phenotypes are observed in superman agamous and superman apetala2 double mutants. The epistatic relationships observed between either apetala3 or pistillata and superman alleles suggest that the SUPERMAN gene product could be a regulator of these floral homeotic genes. To test this, the expression patterns of AGAMOUS and APETALA3 were examined in superman flowers. In wild-type flowers, APETALA3 expression is restricted to the second and third whorls where it is required for the specification of petals and stamens. In contrast, in superman flowers, APETALA3 expression expands to include most of the cells that would normally constitute the fourth whorl. This ectopic APETALA3 expression is proposed to be one of the causes of the development of the extra stamens in superman flowers. The spatial pattern of AGAMOUS expression remains unaltered in superman flowers as compared to wild-type flowers. Taken together these data indicate that one of the functions of the wild-type SUPERMAN gene product is to negatively regulate APETALA3 in the fourth whorl of the flower. In addition, superman mutants exhibit a loss of determinacy of the floral meristem, an effect that appears to be mediated by the APETALA3 and PISTILLATA gene products.     

A nucleostemin-like GTPase required for normal apical and floral meristem development in Arabidopsis

Mammalian nucleostemin (NS) is preferentially expressed in stem cells and acts to promote cell cycle progression. In plants, stem cell activities have to be terminated during flower development, and this process requires the activation of AGAMOUS (AG) gene expression. Here, a nucleostemin-like 1 gene, NSN1, is shown to be required for flower development in Arabidopsis. The NSN1 mRNA was found in the inflorescence meristem and floral primordia, and its protein was localized to the nucleoli. Both heterozygous and homozygous plants developed defective flowers on inflorescences that were eventually terminated by the formation of carpelloid flowers. Overexpression of NSN1 resulted in loss of apical dominance and formation of defective flowers. Expression of the AG gene was found to be up-regulated in nsn1. The carpelloid flower defect of nsn1 was suppressed by the ag mutation in the nsn1 ag double mutant, whereas double mutants of nsn1 apetala2 (ap2) displayed enhanced defective floral phenotypes. These results suggest that in the delicately balanced regulatory network, NSN1 acts to repress AG and plays an additive role with AP2 in floral organ specification. As a midsize nucleolar GTPase, NSN1 represents a new class of regulatory proteins required for flower development in Arabidopsis.     

Genomic approach to study floral development genes in Rosa sp

Cultivated for centuries, the varieties of rose have been selected based on a number of flower traits. Understanding the genetic and molecular basis that contributes to these traits will impact on future improvements for this economically important ornamental plant. In this study, we used scanning electron microscopy and sections of meristems and flowers to establish a precise morphological calendar from early rose flower development stages to senescing flowers. Global gene expression was investigated from floral meristem initiation up to flower senescence in three rose genotypes exhibiting contrasted floral traits including continuous versus once flowering and simple versus double flower architecture, using a newly developed Affymetrix microarray (Rosa1_Affyarray) tool containing sequences representing 4765 unigenes expressed during flower development. Data analyses permitted the identification of genes associated with floral transition, floral organs initiation up to flower senescence. Quantitative real time PCR analyses validated the mRNA accumulation changes observed in microarray hybridizations for a selection of 24 genes expressed at either high or low levels. Our data describe the early flower development stages in Rosa sp, the production of a rose microarray and demonstrate its usefulness and reliability to study gene expression during extensive development phases, from the vegetative meristem to the senescent flower.     

铁木属(桦木科)花序及花的形态发生

在扫描电镜下观察了桦木科(Betulaceae)铁木属花序和花的形态发生过程。结果显示, 铁木雌花序由多个小聚伞花序螺旋状排列组成。每个小花序原基分化出1枚初级苞片和一团小花序原基分生组织, 由小花序原基分生组织分化形成1对次级苞片和2个花原基, 每个花原基分化出2个或3个心皮原基, 形成二心皮或三心皮雌蕊, 雌蕊基部有1层环状花被原基。雄花序为柔荑状, 由多个小聚伞花序螺旋状排列组成。每个小花序原基分化出1枚初级苞片和一团小花序原基分生组织, 由小花序原基分生组织分化出3个花原基分区, 位于中央的花原基分区, 分化形成5-6枚雄蕊原基, 两侧的花原基分区, 分别分化形成3-4枚雄蕊原基, 雄蕊原基分化形成四药囊雄蕊。雄蕊原基纵裂, 但花丝纵裂没有达到基部。