Ectopic expression of SUPERMAN suppresses development of petals and stamens

The floral regulatory gene SUPERMAN (SUP) encodes a C2H2 type zinc finger protein that is required for maintaining boundaries between floral organs in Arabidopsis. It has been proposed that the main function of SUP is to balance cell proliferation in the third and fourth whorl of developing flowers, thereby maintaining the boundaries between the two whorls. To gain further insight into the function of SUP, we have ectopically expressed SUP using the promoter of APETALA1 (AP1), a gene that is initially expressed throughout floral meristems and later becomes restricted to the first and second whorls. Flowers of AP1::SUP plants have fewer floral organs, consistent with an effect of SUP on cell proliferation. In addition, the AP1::SUP transgene caused the conversion of petals to sepals and suppressed the development of stamens. The expression of the B function homeotic gene APETALA3 (AP3) and its regulator UNUSUAL FLORAL ORGANS (UFO) were delayed and reduced in AP1::SUP flowers. However, SUP does not act merely through UFO, as constitutive expression of UFO did not rescue the defects in petal and stamen development in AP1::SUP flowers. Together, these results suggest that SUP has both indirect and direct effects on the expression of B function homeotic genes.     

Negative regulation of the Arabidopsis homeotic gene AGAMOUS by the APETALA2 product.

We characterized the distribution of AGAMOUS (AG) RNA during early flower development in Arabidopsis. Mutations in this homeotic gene cause the transformation of stamens to petals in floral whorl 3 and of carpels to another ag flower in floral whorl 4. We found that AG RNA is present in the stamen and carpel primordia but is undetectable in sepal and petal primordia throughout early wild-type flower development, consistent with the mutant phenotype. We also analyzed the distribution of AG RNA in apetela2 (ap2) mutant flowers. AP2 is a floral homeotic gene that is necessary for the normal development of sepals and petals in floral whorls 1 and 2. In ap2 mutant flowers, AG RNA is present in the organ primordia of all floral whorls. These observations show that the expression patterns of the Arabidopsis floral homeotic genes are in part established by regulatory interactions between these genes.     

Identification of class B and class C floral organ identity genes from rice plants

The functions of two rice MADS-box genes were studied by the loss-of-function approach. The first gene, OsMADS4, shows a significant homology to members in the PISTILLATA (PI) family, which is required to specify petal and stamen identity. The second gene, OsMADS3, is highly homologous to the members in the AGAMOUS (AG) family that is essential for the normal development of the internal two whorls, the stamen and carpel, of the flower. These two rice MADS box cDNA clones were connected to the maize ubiquitin promoter in an antisense orientation and the fusion molecules were introduced to rice plants by the Agrobacterium-mediated transformation method. Transgenic plants expressing antisense OsMADS4 displayed alterations of the second and third whorls. The second-whorl lodicules, which are equivalent to the petals of dicot plants in grasses, were altered into palea/lemma-like organs, and the third whorl stamens were changed to carpel-like organs. Loss-of-function analysis of OsMADS3 showed alterations in the third and fourth whorls. In the third whorl, the filaments of the transgenic plants were changed into thick and fleshy bodies, similar to lodicules. Rather than making a carpel, the fourth whorl produced several abnormal flowers. These phenotypes are similar to those of the agamous and plena mutants in Arabidopsis and Antirrhinum, respectively. These results suggest that OsMADS4 belongs to the class B gene family and OsMADS3 belongs to the class C gene family of floral organ identity determination.     

Comparative transcript profiling of a male sterile cybrid pummelo and its fertile type revealed altered gene expression related to flower development

Male sterile and seedless characters are highly desired for citrus cultivar improvement. In our breeding program, a male sterile cybrid pummelo, which could be considered as a variant of male fertile pummelo, was produced by protoplast fusion. Herein, ecotopic stamen primordia initiation and development were detected in this male sterile cybrid pummelo. Histological studies revealed that the cybrid showed reduced petal development in size and width, and retarded stamen primordia development. Additionally, disorganized cell proliferation was also detected in stamen-like structures (fused to petals and/or carpel). To gain new insight into the underlying mechanism, we compared, by RNA-Seq analysis, the nuclear gene expression profiles of floral buds of the cybrid with that of fertile pummelo. Gene expression profiles which identified a large number of differentially expressed genes (DEGs) between the two lines were captured at both petal primordia and stamen primordia distinguishable stages. For example, nuclear genes involved in nucleic acid binding and response to hormone synthesis and metabolism, genes required for floral bud identification and expressed in particular floral whorls. Furthermore, in accordance with flower morphology of the cybrid, expression of PISTILLATA (PI) was reduced in stamen-like structures, even though it was restricted to correct floral whorls. Down-regulated expression of APETALA3 (AP3) coincided with that of PI. These finding indicated that, due to their whorl specific effects in flower development, citrus class-B MADS-box genes likely constituted 'perfect targets' for CMS retrograde signaling, and that dysfunctional mitochondria seemed to cause male sterile phenotype in the cybrid pummelo.     

FLORAL DEVELOPMENT OF HYDROCHARIS MORSUS-RANAE (HYDROCHARITACEAE)

In both male and female flowers of H. morsus-ranae the primordia of the floral appendages appear in an acropetal succession consisting of alternating trimerous whorls. In the male flower a whorl of sepals is followed by a whorl of petals, three whorls of stamens, and a whorl of filamentous staminodes. The mature androecial arrangement therefore consists of two antisepalous stamen whorls, an antipetalous whorl of stamens, and antipetalous staminodes. Shortly before anthesis, basal meristematic upgrowth between filaments of adjacent whorls produces paired stamens, joining Whorls 1 and 3, and Whorl 2 with the staminodial whorl. A central domelike structure develops between the closely appressed filaments of the inner stamen and staminodial whorl, giving the structure a lobed appearance. After petal inception in the female flower a whorl of antisepalous staminodes develop, each of which may bifurcate to form a pair of staminodes. During staminode development a girdling primordium arises by upgrowth at the periphery of the floral apex. The girdling primordium rapidly forms six gynoecial primordia, which then go on to produce six free styles with bifid stigmas. Intercalary meristem activity, below the point of floral appendage attachment, leads to the production of a syncarpous inferior ovary with six parietal placentae. The styles and carpels remain open along their ventral sutures. During the final stages of female floral development, several hundred ovules develop along the carpel walls, and three nectaries develop dorsally and basally on the three antipetalous styles.     

宽叶泽苔草的花器官发生

通过扫描电镜观察了宽叶泽苔草Caldesia grandisSamuel.的花器官发生。宽叶泽苔草 的萼片3枚,逆时针螺旋向心发生 ;花瓣3枚,呈一轮近同时发生,未观察到花瓣_雄蕊复合原基;雄蕊、心皮原基皆轮状向心 发生,最先近同时发生的6枚原基全部发育成雄蕊,随后发生的6枚原基早期并无差别,在发 育过程中逐渐出现形态差异,直至其中1－4枚发育成心皮,其余的发育成雄蕊;而后的几轮 心皮原基,6枚一轮,陆续向心相间发生。本文揭示了3枚萼片螺旋状的发生方式,并推测这种螺旋方式是泽泻科植物进化过程中保留下来     

The ASK1 gene regulates development and interacts with the UFO gene to control floral organ identity in Arabidopsis.

Normal flower development likely requires both specific and general regulators. We have isolated an Arabidopsis mutant ask1-1 (for -Arabidopsis skp1-like1-1), which exhibits defects in both vegetative and reproductive development. In the ask1-1mutant, rosette leaf growth is reduced, resulting in smaller than normal rosette leaves, and internodes in the floral stem are shorter than normal. Examination of cell sizes in these organs indicates that cell expansion is normal in the mutant, but cell number is reduced. In the mutant, the numbers of petals and stamens are reduced, and many flowers have one or more petals with a reduced size. In addition, all mutant flowers have short stamen filaments. Furthermore, petal/stamen chimeric organs are found in many flowers. These results indicate that the ASK1 gene affects the size of vegetative and floral organs. The ask1 floral phenotype resembles somewhat that of the Arabidopsis ufo mutants in that both genes affect whorls 2 and 3. We therefore tested for possible interactions between ASK1 and UFO by analyzing the phenotypes of ufo-2 ask1-1 double mutant plants. In these plants, vegetative development is similar to that of the ask1-1 single mutant, whereas the floral defects are more severe than those in either single mutant. Interior to the first whorl, the double mutant flowers have more sepals or sepal-like organs than are found in ufo-2, and less petals than ask1-1. Our results suggest that ASK1 interacts with UFO to control floral organ identity in whorls 2 and 3. This is very intriguing because ASK1 is very similar in sequence to the yeast SKP1 protein and UFO contains an F-box, a motif known to interact with SKP1 in yeast. Although the precise mechanism of ASK1 and UFO action is unknown, our results support the hypothesis that these two proteins physically interact in vivo.     

Regulation of SUP expression identifies multiple regulators involved in arabidopsis floral meristem development

During the course of flower development, floral homeotic genes are expressed in defined concentric regions of floral meristems called whorls. The SUPERMAN (SUP, also called FLO10) gene, which encodes a C2H2-type zinc finger protein, is involved in maintenance of the stamen/carpel whorl boundary (the boundary between whorl 3 and whorl 4) in Arabidopsis. Here, we show that the regulation of SUP expression in floral meristems is complex, consisting of two distinct phases, initiation and maintenance. The floral meristem identity gene LEAFY (LFY) plays a role in the initiation phase through at least two pathways, which differ from each other in the involvement of two homeotic genes, APETALA3 (AP3) and PISTILLATA (PI). AP3, PI, and another homeotic gene, AGAMOUS (AG), are further required for SUP expression in the later maintenance phase. Aside from these genes, there are other as yet unidentified genes that control both the temporal and spatial patterns of SUP expression in whorl 3 floral meristems. SUP appears to act transiently, probably functioning to trigger a genetic circuit that creates the correct position of the whorl 3/whorl 4 boundary.     

Expression levels of meristem identity and homeotic genes are modified by nuclear-mitochondrial interactions in alloplasmic male-sterile lines of Brassica napus

Homeotic conversions of anthers were found in cytoplasmic male sterile (CMS) plants of Brassica napus derived from somatic hybrids of B. napus and Arabidopsis thaliana. CMS line flowers displayed petals reduced in size and width and stamens replaced by carpelloid structures. In order to investigate when these developmental aberrations appeared, flower development was analysed histologically, ultrastructurally and molecularly. Disorganized cell divisions were detected in the floral meristems of the CMS lines at stage 4. As CMS is associated with mitochondrial aberrations, ultrastructural analysis of the mitochondria in the floral meristems was performed. Two mitochondrial populations were found in the CMS lines. One type had disrupted cristae, while the other resembled mitochondria typical of B. napus. Furthermore, expression patterns of genes expressed in particular floral whorls were determined. In spite of the aberrant development of the third whorl organs, BnAP3 was expressed as in B. napus during the first six stages of development. However, the levels of BnPI were reduced. At later developmental stages, the expression of both BnAP3 and BnPI was strongly reduced. Interestingly the expression levels of genes responsible for AP3 and PI activation such as LFY, UFO and ASK1 were higher in the CMS lines, which indicates that activation of B-genes in the CMS lines does not occur as in B. napus. Disrupted and dysfunctional mitochondria seem to be one of the first aberrations manifested in CMS which result in a retrograde influence of the expression levels of genes responsible for the second and third whorl organ differentiation.     

重瓣紫蓝大岩桐组培苗的花同源异型现象（简报）

Six types of floral homeotic variants of in vitro seedlings were observed in doubleflower sinningia. Type I, red and green mosaic petals exist in the outermost whorl of petal-whorls, 2.38%. Type II, the outermost whorl of petal-whorls exhibit green petals with thin yellow edge, 25.0%. Type III, green petals exist in the innermost side of normal red petal whorls, 1.78%. Type IV, multiple whorls of green petals exist in the inner side of normal sepals, no stamen and carpel, 1.67%. Type V, it exhibits duplicated whorls of sepals in the outermost, 7.14%. Type VI, it exists multiple whorls of green sepals, no petal, stamen and carpel, 0.12%. The total percentage of all types of floral homeotic variants is up to 38.1%. The distribution of nodal site of homeotic flowers were analyzed, and the results showed that the homeotic flower occurred mainly at the fourth and fifth nodes.     

红花玉兰MwAG基因在花发育不同时期的表达

MwAG基因是调控红花玉兰(Magnolia wufengensis)雌雄蕊发育的关键转录因子。采用半定量RT-PCR、Northern blot杂交和实时荧光定量PCR技术检测了MwAG基因在红花玉兰花芽形态分化几个关键时期表达的组织特异性和表达水平的变化。研究结果表明, MwAG基因仅在红花玉兰雌雄蕊中表达, 而在幼叶、外轮花被和内轮花被中不表达。在花器官形态分化过程中, MwAG基因在雌雄蕊原基分化期和雌雄蕊成熟期均维持在一个较高的水平, 且在雄蕊中的表达波峰早于雌蕊, 这与雌雄蕊形态分化的时间基本吻合; 在花芽分化早期, 相同大小的花芽, 瓣数越多, MwAG基因在雌雄蕊中的表达量越低, 其结果与不同瓣数雌雄蕊分化的时间一致, 即瓣数越多, 雌雄蕊分化越晚。     

Interactions between gene activity and cell layers during floral development

The DEFICIENS (DEF) gene is required for establishing petal and stamen identity in Antirrhinum and is expressed in all three layers of the floral meristem in whorls 2 and 3. Expression of DEF in a subset of meristem layers gives rise to organs with characteristic shapes and cell types, reflecting altered patterns and levels of DEF gene activity. To determine how the contributions of layers and gene activity interact, we exploited a DEF allele which carries a transposon insertion in the MADS box region to generate periclinal chimeras expressing alleles with different activities. By comparing the phenotype, development and expression patterns of these chimeras we show that expression of DEF in L1 makes a major contribution to morphology in whorl 2, irrespective of the allele. By contrast L1 expression is largely unable to rescue whorl 3, possibly because of a non-autonomous inhibitor of DEF activity in this whorl.     

Development of the cymose inflorescence, cupulum and flower of Psoralea pinnata (Leguminosae: Papilionoideae: Psoraleeae)

The inflorescence of Psoralea pinnata sensu lato is composed of dichasial cymes of three to seven flowers in each bract axil. However, the main inflorescence terminal is indeterminate. Cymes are very unusual among legumes, occurring in perhaps six or seven genera. The 'cupulum' is a unique structure found only in Psoralea among legumes. While highly variable in form, it serves as a protective structure around the calyx during development. Developmentally the cupulum results from fusion by intercalary growth of three to four successive bracts on the pedicel. Paired bracteoles are also produced above the cupulum and below the calyx, but they are very reduced and easily overlooked. Their presence is evidence that the cupulum is not part of the flower proper. Organogeny in P. pinnata is unidirectional in each floral whorl, starting on the abaxia! side, except for a minor exception in precocity of the vexillary (adaxial) stamen arising with the two lateral stamens of the same inner whorl. There is no overlap between whorls. The carpel is initiated early, during or just after petal initiation and before stamen initiation.     

Floral development of the model legume <Emphasis Type="Italic">Medicago truncatula</Emphasis>: ontogeny studies as a tool to better characterize homeotic mutations

This work provides new evidence of the complex genetic regulation necessary to accomplish flower development in legumes. Using scanning electron microscopy (SEM) analysis, we have characterized the early developmental events of the wild type Medicago truncatula flower and selected morphological characters as markers to break it down into eight different developmental stages. The order of floral organ initiation in M. truncatula and pea (Pisum sativum L.), in contrast to Arabidopsis and Antirrhinum, is unidirectional in all whorls starting from the abaxial position of the flower with a high degree of overlap. Another main difference is the existence of four common primordia from which petals and stamens differentiate. The formation of common primordia, as opposed to discrete petal and stamen primordia, has been described in many legume and non-legume plants. The main differences between pea and M. truncatula floral ontogeny are in carpel and fruit development. We also used these morphological markers as tools to characterize early alterations in the flower development of a male-sterile M. truncatula floral homeotic mutant named mtapetala. This mutant displays a phenotype resembling those of weak class B mutants with homeotic conversions of floral organ whorls 2 and 3 into sepaloid and carpelloid structures, respectively. Ontogeny studies of the mtapetala mutant flowers showed similarities with the effects of previously described loss-of-B-function mutations. Differences between ontogeny of wild type and mtapetala flowers could not be detected during the first stages (1-5) of flower development. In late stage 5, abnormal-shaped petals with acute lobes and trichomes as well as abnormal-shaped stamens were visible in whorls 2 and 3. At stage 6, the morphology of petals began to change, developing enlarged sepaloid structures bearing trichomes and first the antesepalous stamens and then the antepetalous stamens began to differentiate carpelloid anthers from filaments. Third whorl organs presented different degrees of carpelloidy. The present study should provide tools for the characterization and comparative analyses of new Medicago floral homeotic mutants and could be useful in elucidating how floral organ identity functions work in legumes.     

Male and female flowers of the dioecious plant sorrel show different patterns of MADS box gene expression.

Male and female flowers of the dioecious plant sorrel (Rumex acetosa) each produce three whorls of developed floral organs: two similar whorls of three perianth segments and either six stamens (in the male) or a gynoecium consisting of a fertile carpel and two sterile carpels (in the female). In the developing male flower, there is no significant proliferation of cells in the center of the flower, in the position normally occupied by the carpels of a hermaphrodite plant. In the female flower, small stamen primordia are formed. To determine whether the organ differences are associated with differences in the expression of organ identity genes, cDNA clones representing the putative homologs of B and C function MADS box genes were isolated and used in an in situ hybridization analysis. The expression of RAD1 and RAD2 (two different DEFICIENS homologs) in males and females was confined to the stamen whorl; the lack of expression in the second, inner perianth whorl correlated with the sepaloid nature of the inner whorl of perianth segments. Expression of RAP1 (a PLENA homolog) occurred in the carpel and stamen whorls in very young flower primordia from both males and females. However, as soon as the inappropriate set of organs ceased to develop, RAP1 expression became undetectable in those organs. The absence of expression of RAP1 may be the cause of the arrest in organ development or may be a consequence.     

A novel role of BELL1-like homeobox genes, PENNYWISE and POUND-FOOLISH, in floral patterning

Flowers are determinate shoots comprised of perianth and reproductive organs displayed in a whorled phyllotactic pattern. Floral organ identity genes display region-specific expression patterns in the developing flower. In Arabidopsis, floral organ identity genes are activated by LEAFY (LFY), which functions with region-specific co-regulators, UNUSUAL FLORAL ORGANS (UFO) and WUSCHEL (WUS), to up-regulate homeotic genes in specific whorls of the flower. PENNYWISE (PNY) and POUND-FOOLISH (PNF) are redundant functioning BELL1-like homeodomain proteins that are expressed in shoot and floral meristems. During flower development, PNY functions with a co-repressor complex to down-regulate the homeotic gene, AGAMOUS (AG), in the outer whorls of the flower. However, the function of PNY as well as PNF in regulating floral organ identity in the central whorls of the flower is not known. In this report, we show that combining mutations in PNY and PNF enhance the floral patterning phenotypes of weak and strong alleles of lfy, indicating that these BELL1-like homeodomain proteins play a role in the specification of petals, stamens and carpels during flower development. Expression studies show that PNY and PNF positively regulate the homeotic genes, APETALA3 and AG, in the inner whorls of the flower. Moreover, PNY and PNF function in parallel with LFY, UFO and WUS to regulate homeotic gene expression. Since PNY and PNF interact with the KNOTTED1-like homeodomain proteins, SHOOTMERISTEMLESS (STM) and KNOTTED-LIKE from ARABIDOPSIS THALIANA2 (KNAT2) that regulate floral development, we propose that PNY/PNF-STM and PNY/PNF-KNAT2 complexes function in the inner whorls to regulate flower patterning events.     

OVERLAPPING ORGAN INITIATION AND COMMON PRIMORDIA IN FLOWERS OF PISUM SATIVUM (LEGUMINOSAE: PAPILIONOIDEAE)

The floral ontogeny of Pisum sativum shows a vertical order of succession of sepals, petals plus carpel, antesepalous stamens, and antepetalous stamens. Within each whorl, unidirectional order is followed among the organs, beginning on the abaxial side of the flower, as in most papilionoids. Unusual features include the four common primordia which precede initiation of discrete petal and antesepalous stamen primordia, and the marked overlap of organ initiations between whorls which are usually separately initiated. The stamens arise in free condition, then become diadelphous by intercalary growth at the base of nine stamens, and finally become pseudomonadelphous by surface fusion between the vexillary stamen filament and the adjacent edges of the filament tube. The early initiation of the carpel is not unique among papilionoids, but is somewhat unusual.     

AINTEGUMENTA promotes petal identity and acts as a negative regulator of AGAMOUS

The Arabidopsis AINTEGUMENTA (ANT) gene has been shown previously to be involved in ovule development and in the initiation and growth of floral organs. Here, we show that ANT acts in additional processes during flower development, including repression of AGAMOUS (AG) in second whorl cells, promotion of petal epidermal cell identity, and gynoecium development. Analyses of ap2-1 ant-6 double mutants reveal that ANT acts redundantly with AP2 to repress AG in second whorl cells. The abaxial surface of ant petals contains features such as stomata and elongated, interdigitated cells that are not present on wild-type petals. The loss of petal identity in these second whorl cells does not result from ectopic AG expression, suggesting that ANT acts in a pathway promoting petal cell identity that is independent of its role in repression of AG. These data suggest that ANT may function as a class A gene.