C-class Gene Research Articles

Reduced Expression of C-class Genes Is Associated with the Multiple-petal Phenotype in <i>Nelumbo nucifera</i>

Reduced Expression of C-class Genes Is Associated with the Multiple-petal Phenotype in <i>Nelumbo nucifera</i>

Effect of temperature on expression of B- and C-class MADS-box genes in flower buds of sweet cherry

The occurrence of double fruits in sweet cherry (Prunus avium) is due to abnormal differentiation of pistil primordia in the previous growing season. Double pistil formation is caused by high temperature above 30°C during flower differentiation. Furthermore, extremely high temperature induces double pistil formation as well as homeotic transformation of stamens into pistil-like structures. Therefore, the B and C classes of homeotic genes, which control stamen and pistil formations, may be involved in these phenomena. In this study, we investigated the effect of temperature on the expression levels of B- and C-class MADS-box genes in flower buds. On July 20, when the buds had formed sepal and petal primordia, 'Satohnishiki' sweet cherry trees were brought into sunlit growth chambers with controlled temperatures at 25 and 35°C. Shortly after the onset of the treatment (3 and 10 days), spur buds were collected, and the expression levels of the APETALA3 (AP3)-like and PISTILLATA (PI)-like B-class genes and the AGAMOUS (AG)-like and SHATTERPROOF (SHP)-like C-class genes in the buds were analysed using real-time PCR. The transcriptional levels of PaTM6 (TM6 lineage of AP3) and PaAG at 35°C were much higher than those at 25°C. On the other hand, the high temperature considerably reduced the transcriptional levels of PaPI and PaSHP. We expected that increases in the expression level of C-class genes and decreases in that of B-class genes may be involved in double pistil formation and pistil-like stamens due to high temperature. However, only the changes in the expression levels of PaAG and PaPI genes were consistent with our expectations. Next, we plan to compare the expression levels of these genes among cultivars with different frequency of double pistils.

Dosage imbalance of B- and C-class genes causes petaloid-stamen relating to F1 hybrid variation

BackgroundGreat advances have been achieved in our understanding of flower development and evolution since the establishment of the ABC model. However, it remains a challenge to define the exact context of organ identity in the component interactions of the ABC model.ResultsThrough hybridization, we detected a homeotic mutant in Petrocosmea (Gesneriaceae) uniquely displayed by the ‘petaloid-stamen’ in the third whorl with petal identity. Comparative Real-time PCR analyses demonstrate that both two B-class genes DEF2 and GLO are excessively expressed while the transcripts of the C-class gene PLE are reduced in the third floral whorl in the mutant compared to that in the wild-type F1 hybrids. Further allele-specific expression (ASE) analyses indicate that an allele-specific change in PgPLE might be responsible for up-regulation of both B-class genes and down-regulation of the C-class gene in the petaloid-stamen mutants.ConclusionsOur findings suggest that the petaloid-stamen is consequent upon an evident dosage imbalance between B- and C-class products that is probably triggered by a cis-regulatory change. In addition, the genetic pathway for the floral organ identity might be in parallel with that for the floral symmetry. The extreme variation in hybrids further suggests that interspecific hybridization may represent a major factor for evolutionary innovation and diversification in plants.

How to Evolve a Perianth: A Review of Cadastral Mechanisms for Perianth Identity.

The flower of angiosperms is considered to be a major evolutionary innovation that impacted the whole biome. In particular, two properties of the flower are classically linked to its ecological success: bisexuality and a differentiated perianth with sepals and petals. Although the molecular basis for floral organ identity is well understood in extant species and summarized in the famous ABC model, how perianth identity appeared during evolution is still unknown. Here we propose that cadastral mechanisms that maintain reproductive organ identities to the center of the flower could have supported perianth evolution. In particular, repressing B- and C-class genes expression toward the inner whorls of the flower, is a key process to isolate domains with sepal and petal identity in the outer whorls. We review from the literature in model species the diverse regulators that repress B- and C-class genes expression to the center of the flower. This review highlights the existence of both unique and conserved repressors between species, and possible candidates to investigate further in order to shed light on perianth evolution.

Differential Gene Expression Caused by the F and M Loci Provides Insight Into Ethylene-Mediated Female Flower Differentiation in Cucumber.

In cucumber (Cucumis sativus L.), the differentiation and development of female flowers are important processes that directly affect the fruit yield and quality. Sex differentiation is mainly controlled by three ethylene synthase genes, F (CsACS1G), M (CsACS2), and A (CsACS11). Thus, ethylene plays a key role in the sex differentiation in cucumber. The “one-hormone hypothesis” posits that F and M regulate the ethylene levels and initiate female flower development in cucumber. Nonetheless, the precise molecular mechanism of this process remains elusive. To investigate the mechanism by which F and M regulate the sex phenotype, three cucumber near-isogenic lines, namely H34 (FFmmAA, hermaphroditic), G12 (FFMMAA, gynoecious), and M12 (ffMMAA, monoecious), with different F and M loci were generated. The transcriptomic analysis of the apical shoots revealed that the expression of the B-class floral homeotic genes, CsPI (Csa4G358770) and CsAP3 (Csa3G865440), was immensely suppressed in G12 (100% female flowers) but highly expressed in M12 (∼90% male flowers). In contrast, CAG2 (Csa1G467100), which is an AG-like C-class floral homeotic gene, was specifically highly expressed in G12. Thus, the initiation of female flowers is likely to be caused by the downregulation of B-class and upregulation of C-class genes by ethylene production in the floral primordium. Additionally, CsERF31, which was highly expressed in G12, showed temporal and spatial expression patterns similar to those of M and responded to the ethylene-related chemical treatments. The biochemical experiments further demonstrated that CsERF31 could directly bind the promoter of M and promote its expression. Thus, CsERF31 responded to the ethylene signal derived from F and mediated the positive feedback regulation of ethylene by activating M expression, which offers an extended “one-hormone hypothesis” of sex differentiation in cucumber.

The Greenish Flower Phenotype of Habenaria radiata (Orchidaceae) Is Caused by a Mutation in the SEPALLATA-Like MADS-Box Gene HrSEP-1.

In Arabidopsis thaliana, the E-class SEPALLATA (SEP) genes are generally expressed across all floral whorls. These genes play fundamental roles in floral organ fate determination during development by interacting with other MADS-box gene products, such as those from A-, B-, and C-class genes. However, the function of SEP genes in orchid remains obscure. Here, we analyzed a mutant orchid cultivar with greenish flowers in Habenaria radiata and found that this phenotype is caused by the absence of SEP function. Wild type H. radiata flowers contain a column and two perianth whorls consisting of three greenish sepals, two white petals, and a lip (labellum). By contrast, the flowers of H. radiata cultivar ‘Ryokusei’ appear greenish, with three normal sepals in whorl 1, two greenish petals and a lip in whorl 2, and several sepaloid organs and a ventral column in whorls 3 and 4. We isolated two SEP-like genes (HrSEP-1 and HrSEP-2) and two AGAMOUS-like genes (HrAG-1 and HrAG-2) from wild type H. radiata and compared their expression in the wild type vs. the mutant cultivar. HrAG-1 and HrAG-2 were expressed in the column in the wild type, whereas these genes were expressed in the ventral column and in sepaloid organs that had been converted from a column in ‘Ryokusei.’ HrSEP-1 and HrSEP-2 were expressed in all floral organs in the wild type. However, in the mutant cultivar, HrSEP-2 was expressed in all floral organs, while HrSEP-1 expression was not detected. Thus, we analyzed the genomic structures of HrSEP-1 in the wild type and ‘Ryokusei’ and identified a retrotransposon-like element in its first exon in ‘Ryokusei.’ Yeast two-hybrid assays demonstrated that HrSEP-1 interacts with HrDEF, HrAG-1, and HrAG-2. These results indicate that the mutant phenotype of ‘Ryokusei’ flowers is caused by the loss of function of HrSEP-1. Therefore, this gene plays an important role in column, lip, and petal development in H. radiata flowers.

Functional Genomics and Genetic Control of Flower and Fruit Development in Medicago truncatula: An Overview.

A-, B-, and C-class genes code for MADS-box transcription factors required for floral organ identity in angiosperms. Other members of the family are also crucial to ensure proper carpel and fruit development. Development of genetic and genomic tools for Medicago truncatula has allowed its use as model system to study the genetic control of flower and fruit development in legumes. M. truncatula contains a single A-class gene, four B-function genes, and three C-class genes in its genome. This has made possible to do extensive functional characterization of these MADS-box transcription factors using gene expression analyses, protein-protein interactions, and forward and reverse genetic approaches. We have demonstrated the functions of these MADS-box transcription factors and the respective contributions of paralogous gene pairs to M. truncatula floral development. We have also defined the evolutionary outcomes of each duplicated pairs thus testing theoretical framework of several models about the evolution by gene duplication. Moreover, we have also studied the function of MADS-box fruit genes and how they may have contributed to the diversification of pod morphology within the Medicago genus. Our findings not only have contributed to increase knowledge in the field of the genetic control of flower and fruit development but also have provided a more complete understanding of the complexity of evolution by gene duplication and protein sequence diversification.

Expression Analyses of ABCDE Model Genes and Changes in Levels of Endogenous Hormones in Chinese Cabbage Exhibiting Petal-Loss

Abnormal formation of floral organs affects plant reproduction and can directly interfere with the progress of breeding programs. Using PCR amplification, ABCDE model genes BraAP2, BraAP3, BraPI, BraAG, BraSHP, and BraSEP were isolated from Chinese cabbage (Brassica rapa L. ssp. pekinensis). We examined six development stages of floral buds collected from Chinese cabbage and compared between a line demonstrating normal flowering (A-8) and two mutated lines that exhibited plants having petal-loss (A-16 and A-17). The expression of ABCDE model genes has been analyzed by qRT-PCR. Compared with flower buds of petal-loss plants and normal plants, the expression of A-class gene BraAP2 was significantly decreased during the first to fourth stages, C-class gene BraAG expression was significantly decreased during the first to fifth stages, and D-class gene BraSHP expression was significantly decreased during the first to third stages. Furthermore, B-class gene BraAP3 and BraPI and E-class gene BraSEP expressions were significantly decreased during all six stages of petal-loss plants compared with normal plants. Enzyme-linked immunosorbent assays detected nine endogenous phytohormones during all stages examined here. Except for the second-stage and third-stage buds, levels of the auxin IAA and cytokinin dhZR were always higher in the petal-loss plants than the normal plants at corresponding time points. Meanwhile, concentrations of GA1+3 at the first, fourth, and fifth stages were higher in the petal-loss plants than in the normal plants. Our results provide a theoretical basis for future exploration of the molecular mechanism that determines petal loss and the effects that hormones have on such development in Chinese cabbage plants.

Isolation and characterization of the C-class MADS-box gene from the distylous pseudo-cereal Fagopyrum esculentum

In the model species Arabidopsis thaliana, the floral homeotic C-class gene AGAMOUS (AG) specifies reproductive organ (stamen and carpels) identity and floral meristem determinacy. Gene function analyses in other core eudicots species reveal functional conservation, subfunctionalization and function switch of the C-lineage in this clade. To identify the possible roles of AG-like genes in regulating floral development in distylous species with dimorphic flowers (pin and thrum) and the C function evolution, we isolated and identified an AG ortholog from Fagopyrum esculentum (buckwheat, Family Polygonaceae), an early diverging species of core eudicots preceding the rosids-asterids split. Protein sequence alignment and phylogenetic analysis grouped FaesAG into the euAG lineage. Expression analysis suggested that FaesAG expressed exclusively in developing stamens and gynoecium of pin and thrum flowers. Moreover, FaesAG expression reached a high level in both pin and thrum flowers at the time when the stamens were undergoing rapidly increased in size and microspore mother cells were in meiosis. FaesAG was able to substitute for the endogenous AG gene in specifying stamen and carpel identity and in an Arabidopsis ag-1 mutant. Ectopic expression of FaesAG led to very early flowering, and produced a misshapen inflorescence and abnormal flowers in which sepals had converted into carpels and petals were converted to stamens. Our results confirmed establishment of the complete C-function of the AG orthologous gene preceding the rosids-asterids split, despite the distinct floral traits present in early- and late-diverging lineages of core eudicot angiosperms.

Isolation and characterization of a floral homeotic gene in Fraxinus nigra causing earlier flowering and homeotic alterations in transgenic Arabidopsis

Reproductive sterility, which can be obtained by manipulating floral organ identity genes, is an important tool for gene containment of genetically engineered trees. In Arabidopsis, AGAMOUS (AG) is the only C-class gene responsible for both floral meristem determinacy and floral organ identity, and its mutations produce sterility. As a first step in an effort to develop transgenic sterile black ash (Fraxinus nigra), an AG ortholog in black ash (FnAG) was isolated using reverse transcription polymerase chain reaction and rapid amplification of cDNA ends. Analysis of the deduced amino acid sequence showed a typical MIKC structure of type II plant MADS-box protein with a highly conserved MADS-domain. Phylogenetic analysis revealed that FnAG had a close relationship with AG orthologs from other woody species. FnAG transcript was mainly expressed in reproductive tissues, but rarely detected in the vegetative tissues, consistent with the ABC model for floral development. A functional analysis was performed by ectopic expression of FnAG driven by the CaMV 35S promoter in transgenic Arabidopsis. Transformed plants showed homeotic conversions of carpeloid sepals and stamenoid petals. Curled leaves, reduced plant size, and earlier flowering were also observed in transgenic Arabidopsis. These data indicated that the FnAG functions in the same way as AG in Arabidopsis. These results provide the framework for targeted genome editing of black ash, an ecologically and economically important wetland species.

Transcriptomic Analysis Implies That GA Regulates Sex Expression via Ethylene-Dependent and Ethylene-Independent Pathways in Cucumber (Cucumis sativus L.).

Sex differentiation of flower buds is an important developmental process that directly affects fruit yield of cucumber (Cucumis sativus L.). Plant hormones, such as gibberellins (GAs) and ethylene can promote development of male and female flowers, respectively, however, the regulatory mechanisms of GA-induced male flower formation and potential involvement of ethylene in this process still remain unknown. In this study, to unravel the genes and gene networks involved in GA-regulated cucumber sexual development, we performed high throughout RNA-Seq analyses that compared the transcriptomes of shoot tips between GA3 treated and untreated gynoecious cucumber plants. Results showed that GA3 application markedly induced male flowers but decreased ethylene production in shoot tips. Furthermore, the transcript levels of M (CsACS2) gene, ethylene receptor CsETR1 and some ethylene-responsive transcription factors were dramatically changed after GA3 treatment, suggesting a potential involvement of ethylene in GA-regulated sex expression of cucumber. Interestingly, GA3 down-regulated transcript of a C-class floral homeotic gene, CAG2, indicating that GA may also influence cucumber sex determination through an ethylene-independent process. These results suggest a novel model for hormone-mediated sex differentiation and provide a theoretical basis for further dissection of the regulatory mechanism of male flower formation in cucumber.Statement: We reveal that GA can regulate sex expression of cucumber via an ethylene-dependent manner, and the M (CsACS2), CsETR1, and ERFs are probably involved in this process. Moreover, CAG2, a C-class floral homeotic gene, may also participate in GA-modulated cucumber sex determination, but this pathway is ethylene-independent.

Interactions between FLORAL ORGAN NUMBER4 and floral homeotic genes in regulating rice flower development.

The floral meristem (FM) is self-maintaining at the early stages of flower development, but it is terminated when a fixed number of floral organs are produced. The FLORAL ORGAN NUMBER4 (FON4; also known as FON2) gene, an ortholog of Arabidopsis CLAVATA3 (CLV3), is required for regulating FM size and determinacy in rice. However, its interactions with floral homeotic genes remain unknown. Here, we report the genetic interactions between FON4 and floral homeotic genes OsMADS15 (an A-class gene), OsMADS16 (also called SUPERWOMAN1, SPW1, a B-class gene), OsMADS3 and OsMADS58 (C-class genes), OsMADS13 (a D-class gene), and OsMADS1 (an E-class gene) during flower development. We observed an additive phenotype in the fon4 double mutant with the OsMADS15 mutant allele dep (degenerative palea). The effect on the organ number of whorl 2 was enhanced in fon4 spw1. Double mutant combinations of fon4 with osmads3, osmads58, osmads13, and osmads1 displayed enhanced defects in FM determinacy and identity, respectively, indicating that FON4 and these genes synergistically control FM activity. In addition, the expression patterns of all the genes besides OsMADS13 had no obvious change in the fon4 mutant. This work reveals how the meristem maintenance gene FON4 genetically interacts with C, D, and E floral homeotic genes in specifying FM activity in monocot rice.

Deep into the Aristolochia Flower: Expression of C, D, and E-Class Genes in Aristolochia fimbriata (Aristolochiaceae).

Aristolochia fimbriata (Aristolochiaceae) is a member of an early diverging lineage of flowering plants and a promising candidate for evo-devo studies. Aristolochia flowers exhibit a unique floral synorganization that consists of a monosymmetric and petaloid calyx formed by three congenitally fused sepals, and a gynostemium formed by the congenital fusion between stamens and the stigmatic region of the carpels. This floral ground plan atypical in the magnoliids can be used to evaluate the role of floral organ identity MADS-box genes during early flower evolution. In this study, we present in situ hybridization experiments for the homologs of the canonical C-, D-, and E-class genes. Spatiotemporal expression of the C-class gene AfimAG is restricted to stamens, ovary, and ovules, suggesting a conserved stamen and carpel identity function, consistent with that reported in core-eudicots and monocots. The D-class gene AfimSTK is detected in the anthers, the stigmas, the ovary, the ovules, the fruit, and the seeds, suggesting conserved roles in ovule and seed identity and unique roles in stamens, ovary, and fruit development. In addition, AfimSTK expression patterns in areas of organ abscission and dehiscence zones suggest putative roles linked to senescence processes. We found that both E-class genes are expressed in the anthers and the ovary; however, AfimSEP2 exhibits higher expression compared to AfimSEP1. These findings provide a comprehensive picture of the ancestral expression patterns of the canonical MADS-box floral organ identity genes and the foundations for further comparative analyses in other magnoliids.

Molecular cloning, characterization and expression of WAG-2 alternative splicing transcripts in developing spikes of Aegilops tauschii.

WAG-2 is a C-class MADS-box gene, which is orthologous to AGAMOUS (AG) in Arabidopsis. The AG group C-class MADS-box genes are involved in stamen and pistil identity. In this study, two WAG-2 transcripts, namely, WAG-2f and WAG- 2g, were isolated and characterized from Aegilops tauschii. The open reading frames of WAG-2f and WAG-2g were 825 and 822 bp, respectively, encoding 275 and 274 amino acid residues. BLAST searches of partial WAG-2 genomic sequence against the draft sequence of Ae. tauschii genome database revealed the complex structure of WAG-2 gene, which consisted of seven exons and six introns. The WAG-2f and WAG-2g cDNAs were two alternative splicing transcripts. The alternative splicing events were produced by an alternative 5' splice site. The expression level of WAG-2f transcript, which was extremely weak in young spikes of floret primordium formation stage, increased as the spikes developed. The highest expression was observed in the spikes at the anther separation stage. Low expression levels of WAG-2f were also detected at the tetrad stage. The WAG- 2g transcript was expressed at all four stages of spike development but at a relatively low level. The expression pattern of the two transcripts was distinctly different during floral development, thereby suggesting a functional divergence.

The Rosa chinensis cv. Viridiflora Phyllody Phenotype Is Associated with Misexpression of Flower Organ Identity Genes.

Phyllody is a flower abnormality in which leaf-like structures replace flower organs in all whorls. Here, we investigated the origin and the molecular mechanism of phyllody phenotype in Rosa chinensis cv. Viridiflora, an ancient naturally occurring Chinese mutant cultivar. Reciprocal grafting experiments and microscopy analyses, demonstrated that the phyllody phenotype in Viridiflora is not associated with phytoplasmas infection. Transcriptome comparisons by the mean of RNA-Seq identified 672 up-regulated and 666 down-regulated genes in Viridiflora compared to its closely related genotype R. chinensis cv. Old Blush. A fraction of these genes are putative homologs of genes known to be involved in flower initiation and development. We show that in flower whorl 2 of Viridiflora, a down-regulation of the floral organ identity genes RcPISTILLATA (RcPI), RcAPETALA3 (RcAP3) and RcSEPALLATA3 (RcSEP3), together with an up-regulation of the putative homolog of the gene SUPPRESSOR of OVEREXPRESSION of CONSTANS1 (RcSOC1) are likely at the origin of the loss of petal identity and leaf-like structures formation. In whorl 3 of Viridiflora, ectopic expression of RcAPETALA2 (RcAP2) along with the down regulation of RcPI, RcAP3, and RcSEP3 is associated with loss of stamens identity and leaf-like structures formation. In whorl 4, the ectopic expression of RcAP2 associated with a down-regulation of RcSEP3 and of the C-class gene RcAGAMOUS correlate with loss of pistil identity. The latter also suggested the antagonist effect between the A and C class genes in the rose. Together, these data suggest that modified expression of the ABCE flower organ identity genes is associated with the phyllody phenotype in the rose Viridiflora and that these genes are important for normal flower organs development.

The origin of exon 3 skipping of paternal GLOBOSA pre-mRNA in some Nicotiana tabacum lines correlates with a point mutation of the very last nucleotide of the exon

In plants, genome duplication followed by genome diversification and selection is recognized as a major evolutionary process. Rapid epigenetic and genetic changes that affect the transcription of parental genes are frequently observed after polyploidization. The pattern of alternative splicing is also frequently altered, yet the related molecular processes remain largely unresolved. Here, we study the inheritance and expression of parental variants of three floral organ identity genes in allotetraploid tobacco. DEFICIENS and GLOBOSA are B-class genes, and AGAMOUS is a C-class gene. Parental variants of these genes were found to be maintained in the tobacco genome, and the respective mRNAs were present in flower buds in comparable amounts. However, among five tobacco cultivars, we identified two in which the majority of paternal GLOBOSA pre-mRNA transcripts undergo exon 3 skipping, producing an mRNA with a premature termination codon. At the DNA level, we identified a G-A transition at the very last position of exon 3 in both cultivars. Although alternative splicing resulted in a dramatic decrease in full-length paternal GLOBOSA mRNA, no phenotypic effect was observed. Our finding likely serves as an example of the initiation of homoeolog diversification in a relatively young polyploid genome.

Isolation and characterization of the C-class MADS-box gene involved in the formation of double flowers in Japanese gentian.

BackgroundGenerally, double-flowered varieties are more attractive than single-flowered varieties in ornamental plants. Japanese gentian is one of the most popular floricultural plants in Japan, and it is desirable to breed elite double-flowered cultivars. In this study, we attempted to characterize a doubled-flower mutant of Japanese gentian. To identify the gene that causes the double-flowered phenotype in Japanese gentian, we isolated and characterized MADS-box genes.ResultsFourteen MADS-box genes were isolated, and two of them were C-class MADS-box genes (GsAG1 and GsAG2). Both GsAG1 and GsAG2 were categorized into the PLE/SHP subgroup, rather than the AG/FAR subgroup. In expression analyses, GsAG1 transcripts were detected in the second to fourth floral whorls, while GsAG2 transcripts were detected in only the inner two whorls. Transgenic Arabidopsis expressing GsAG1 lacked petals and formed carpeloid organs instead of sepals. Compared with a single-flowered gentian cultivar, a double-flowered gentian mutant showed decreased expression of GsAG1 but unchanged expression of GsAG2. An analysis of the genomic structure of GsAG1 revealed that the gene had nine exons and eight introns, and that a 5,150-bp additional sequence was inserted into the sixth intron of GsAG1 in the double-flowered mutant. This insert had typical features of a Ty3/gypsy-type LTR-retrotransposon, and was designated as Tgs1. Virus-induced gene silencing of GsAG1 by the Apple latent spherical virus vector resulted in the conversion of the stamen to petaloid organs in early flowering transgenic gentian plants expressing an Arabidopsis FT gene.ConclusionsThese results revealed that GsAG1 plays a key role as a C-functional gene in stamen organ identity. The identification of the gene responsible for the double-flowered phenotype will be useful in further research on the floral morphogenesis of Japanese gentian.Electronic supplementary materialThe online version of this article (doi:10.1186/s12870-015-0569-3) contains supplementary material, which is available to authorized users.

Flower development schedule andAGAMOUS-like gene expression patterns in two morphs ofNigella damascena(Ranunculaceae) differing in floral architecture

Love-in-a-mist (Nigella damascena) is an annual species of Ranunculaceae native to the Mediterranean Basin, characterized by delicate flowers lying on long lacy bracts. Two floral morphs of N. damascena, designated [P] and [T], differ in the identity and number of perianth organs and in the position of the perianth–androecium boundary on the meristem. They both occur in the wild. Here we describe a precise comparative schedule of floral development in the two morphs. We divided the sequence of developmental events affecting the floral meristem into six stages and related them to the height of the elongating stem and to the time elapsed after the beginning of stem elongation. In addition, we characterized the expression pattern of C-class genes in floral organs of both morphs in an attempt to better characterize the differences between the two floral groundplans. In the [T] morph an expansion of the expression domain of AGAMOUS (AG) paralogues outside the fertile organs was observed, correlating with the change in identity of the inner perianth organs. Expression of AG-like genes in the sepal-like organs suggests these are not identical to true sepals at the molecular level. The morpho-temporal framework we have defined will allow us to compare various gene expression profiles at targeted developmental stages in both morphs, providing further insight into the molecular control of the floral dimorphism in N. damascena and into the processes underlying the transition from a differentiated (bipartite) to an undifferentiated (unipartite) perianth.

Interactions of OsMADS1 with Floral Homeotic Genes in Rice Flower Development

During reproductive development, rice plants develop unique flower organs which determine the final grain yield. OsMADS1, one of SEPALLATA-like MADS-box genes, has been unraveled to play critical roles in rice floral organ identity specification and floral meristem determinacy. However, the molecular mechanisms underlying interactions of OsMADS1 with other floral homeotic genes in regulating flower development remains largely elusive. In this work, we studied the genetic interactions of OsMADS1 with B-, C-, and D-class genes along with physical interactions among their proteins. We show that the physical and genetic interactions between OsMADS1 and OsMADS3 are essential for floral meristem activity maintenance and organ identity specification; while OsMADS1 physically and genetically interacts with OsMADS58 in regulating floral meristem determinacy and suppressing spikelet meristem reversion. We provided important genetic evidence to support the neofunctionalization of two rice C-class genes (OsMADS3 and OsMADS58) during flower development. Gene expression profiling and quantitative RT-PCR analyses further revealed that OsMADS1 affects the expression of many genes involved in floral identity and hormone signaling, and chromatin immunoprecipitation (ChIP)–PCR assay further demonstrated that OsMADS17 is a direct target gene of OsMADS1. Taken together, these results reveal that OsMADS1 has diversified regulatory functions in specifying rice floral organ and meristem identity, probably through its genetic and physical interactions with different floral homeotic regulators.

Positive selection on the K domain of the AGAMOUS protein in the Zingiberales suggests a mechanism for the evolution of androecial morphology.

BackgroundThe ABC model of flower development describes the molecular basis for specification of floral organ identity in model eudicots such as Arabidopsis and Antirrhinum. According to this model, expression of C-class genes is linked to stamen and gynoecium organ identity. The Zingiberales is an order of tropical monocots in which the evolution of floral morphology is characterized by a marked increase in petaloidy in the androecium. Petaloidy is a derived characteristic of the ginger families and seems to have arisen in the common ancestor of the ginger clade. We hypothesize that duplication of the C-class AGAMOUS (AG) gene followed by divergence of the duplicated AG copies during the diversification of the ginger clade lineages explains the evolution of petaloidy in the androecium. In order to address this hypothesis, we carried out phylogenetic analyses of the AG gene family across the Zingiberales and investigated patterns of gene expression within the androecium.ResultsPhylogenetic analysis supports a scenario in which Zingiberales-specific AG genes have undergone at least one round of duplication. Gene duplication was immediately followed by divergence of the retained copies. In particular, we detect positive selection in the third alpha-helix of the K domain of Zingiberales AGAMOUS copy 1 (ZinAG-1). A single fixed amino acid change is observed in ZinAG-1 within the ginger clade when compared to the banana grade. Expression analyses of AG and APETALA1/FRUITFULL (AP1/FUL) in Musa basjoo is similar to A- and C-class gene expressions in the Arabidopsis thaliana model, while Costus spicatus exhibits simultaneous expression of AG and AP1/FUL in most floral organs. We propose that this novel expression pattern could be correlated with the evolution of androecial petaloidy within the Zingiberales.ConclusionsOur results present an intricate story in which duplication of the AG lineage has lead to the retention of at least two diverged Zingiberales-specific copies, ZinAG-1 and Zingiberales AGAMOUS copy 2 (ZinAG-2). Positive selection on ZinAG-1 residues suggests a mechanism by which AG gene divergence may explain observed morphological changes in Zingiberales flowers. Expression data provides preliminary support for the proposed mechanism, although further studies are required to fully test this hypothesis.