Suvi K. Broholm

A TCP domain transcription factor controls flower type specification along the radial axis of the Gerbera (Asteraceae) inflorescence.

Several key processes in plant development are regulated by TCP transcription factors. CYCLOIDEA-like (CYC-like) TCP domain proteins have been shown to control flower symmetry in distantly related plant lineages. Gerbera hybrida, a member of one of the largest clades of angiosperms, the sunflower family (Asteraceae), is an interesting model for developmental studies because its elaborate inflorescence comprises different types of flowers that have specialized structures and functions. The morphological differentiation of flower types involves gradual changes in flower size and symmetry that follow the radial organization of the densely packed inflorescence. Differences in the degree of petal fusion further define the distinct shapes of the Gerbera flower types. To study the role of TCP transcription factors during specification of this complex inflorescence organization, we characterized the CYC-like homolog GhCYC2 from Gerbera. The expression of GhCYC2 follows a gradient along the radial axis of the inflorescence. GhCYC2 is expressed in the marginal, bilaterally symmetrical ray flowers but not in the centermost disk flowers, which are nearly radially symmetrical and have significantly less fused petals. Overexpression of GhCYC2 causes disk flowers to obtain morphologies similar to ray flowers. Both expression patterns and transgenic phenotypes suggest that GhCYC2 is involved in differentiation among Gerbera flower types, providing the first molecular evidence that CYC-like TCP factors take part in defining the complex inflorescence structure of the Asteraceae, a major determinant of the familys evolutionary success.

Identification of target genes for a MYB-type anthocyanin regulator in Gerbera hybrida

Genetic modification of the flavonoid pathway has been used to produce novel colours and colour patterns in ornamental plants as well as to modify the nutritional and pharmaceutical properties of food crops. It has been suggested that co-ordinate control of multiple steps of the pathway with the help of regulatory genes would lead to a more predictable control of metabolic flux. Regulation of anthocyanin biosynthesis has been studied in a common ornamental plant, Gerbera hybrida (Asteraceae). An R2R3-type MYB factor, GMYB10, shares high sequence similarity and is phylogenetically grouped together with previously characterized regulators of anthocyanin pigmentation. Ectopic expression of GMYB10 leads to strongly enhanced accumulation of anthocyanin pigments as well as to an altered pigmentation pattern in transgenic gerbera plants. Anthocyanin analysis indicates that GMYB10 specifically induces cyanidin biosynthesis in undifferentiated callus and in vegetative tissues. Furthermore, in floral tissues enhanced pelargonidin production is detected. Microarray analysis using the gerbera 9K cDNA array revealed a highly predicted set of putative target genes for GMYB10 including new gene family members of both early and late biosynthetic genes of the flavonoid pathway. However, completely new candidate targets, such as a serine carboxypeptidase-like gene as well, as two new MYB domain factors, GMYB11 and GMYB12, whose exact function in phenylpropanoid biosynthesis is not clear yet, were also identified.

Patterns of MADS-box gene expression mark flower-type development in Gerbera hybrida (Asteraceae)

BackgroundThe inflorescence of the cut-flower crop Gerbera hybrida (Asteraceae) consists of two principal flower types, ray and disc, which form a tightly packed head, or capitulum. Despite great interest in plant morphological evolution and the tractability of the gerbera system, very little is known regarding genetic mechanisms involved in flower type specification. Here, we provide comparative staging of ray and disc flower development and microarray screening for differentially expressed genes, accomplished via microdissection of hundreds of coordinately developing flower primordia.ResultsUsing a 9K gerbera cDNA microarray we identified a number of genes with putative specificity to individual flower types. Intrestingly, several of these encode homologs of MADS-box transcription factors otherwise known to regulate flower organ development. From these and previously obtained data, we hypothesize the functions and protein-protein interactions of several gerbera MADS-box factors.ConclusionOur RNA expression results suggest that flower-type specific MADS protein complexes may play a central role in differential development of ray and disc flowers across the gerbera capitulum, and that some commonality is shared with known protein functions in floral organ determination. These findings support the intriguing conjecture that the gerbera flowering head is more than a mere floral analog at the level of gene regulation.

Functional characterization of B class MADS-box transcription factors in Gerbera hybrida

According to the classical ABC model, B-function genes are involved in determining petal and stamen development. Most core eudicot species have B class genes belonging to three different lineages: the PI, euAP3, and TM6 lineages, although both Arabidopsis and Antirrhinum appear to have lost their TM6-like gene. Functional studies were performed for three gerbera (Gerbera hybrida) B class MADS-box genes—PI/GLO-like GGLO1, euAP3 class GDEF2, and TM6-like GDEF1—and data are shown for a second euAP3-like gene, GDEF3. In phylogenetic analysis, GDEF3 is a closely related paralogue of GDEF2, and apparently stems from a duplication common to all Asteraceae. Expression analysis and transgenic phenotypes confirm that GGLO1 and GDEF2 mediate the classical B-function since they determine petal and stamen identities. However, based on assays in yeast, three B class heterodimer combinations are possible in gerbera. In addition to the interaction of GGLO1 and GDEF2 proteins, GGLO1 also pairs with GDEF1 and GDEF3. This analysis of GDEF1 represents the first functional characterization of a TM6-like gene in a core eudicot species outside Solanaceae. Similarly to its relatives in petunia and tomato, the expression pattern and transgenic phenotypes indicate that GDEF1 is not involved in determination of petal identity, but has a redundant role in regulating stamen development.

Functional diversification of duplicated CYC2 clade genes in regulation of inflorescence development in Gerbera hybrida (Asteraceae)

The complex inflorescences (capitula) of Asteraceae consist of different types of flowers. In Gerbera hybrida (gerbera), the peripheral ray flowers are bilaterally symmetrical and lack functional stamens while the central disc flowers are more radially symmetrical and hermaphroditic. Proteins of the CYC2 subclade of the CYC/TB1-like TCP domain transcription factors have been recruited several times independently for parallel evolution of bilaterally symmetrical flowers in various angiosperm plant lineages, and have also been shown to regulate flower-type identity in Asteraceae. The CYC2 subclade genes in gerbera show largely overlapping gene expression patterns. At the level of single flowers, their expression domain in petals shows a spatial shift from the dorsal pattern known so far in species with bilaterally symmetrical flowers, suggesting that this change in expression may have evolved after the origin of Asteraceae. Functional analysis indicates that GhCYC2, GhCYC3 and GhCYC4 mediate positional information at the proximal-distal axis of the inflorescence, leading to differentiation of ray flowers, but that they also regulate ray flower petal growth by affecting cell proliferation until the final size and shape of the petals is reached. Moreover, our data show functional diversification for the GhCYC5 gene. Ectopic activation of GhCYC5 increases flower density in the inflorescence, suggesting that GhCYC5 may promote the flower initiation rate during expansion of the capitulum. Our data thus indicate that modification of the ancestral network of TCP factors has, through gene duplications, led to the establishment of new expression domains and to functional diversification.

Floral developmental genetics of Gerbera (Asteraceae)

Abstract Inflorescence development in the angiosperm family Asteraceae has distinct features not found in the traditional model systems (e.g., Arabidopsis, Petunia , and Zea ). In Gerbera hybrida , inflorescences are composed of morphologically different types of flowers tightly packed into a flower head (capitulum) that overtly resembles a single flower. Individual floral organs, such as pappus bristles (sepals), are developmentally specialized, petals and anthers form fused structures, stamens are aborted in marginal flowers, and ovaries are located inferior to other floral organs. These specific features have made Gerbera a rewarding target for comparative studies. We have shown that Gerbera MADS‐box genes that group phylogenetically with B‐ and C‐function genes from Arabidopsis and Antirrhinum function in organ determination in a very similar manner to their respective homologs from these model plants. However, MADS‐box genes encoding proteins that interact with those of the ABC‐genes behave differently. In Arabidopsis , three SEPALLATA ( SEP ) genes have redundant functions and are needed for development of petals, stamens, and carpels. Homologs of these SEP genes are found in Gerbera ( GRCD1, GRCD2 ), but they show functional specialization. GRCD1 is necessary for stamen development, but not for petal or carpel development. Similarly, GRCD2 has a homeotic function restricted to carpel development. Remarkably, downregulation of the latter also results in floral reversion (which occurs in ovaries) and alters inflorescence architecture by switching off terminal, determinate growth. This integrated SEP‐like control over reproductive meristem fate has not been detected in the well‐known model systems, which have a different carpel design and normally bear indeterminate inflorescences. Moreover, the organization of flowers on the Gerbera capitulum reveals the presence of a radial morphogenetic gradient that appears to regulate ABC and other MADS‐box genes differentially in a cell‐nonautonomous manner. As such, there is some commonality in gene regulatory features between Gerbera flowers and inflorescences, which suggests that Gerbera capitula are more than simple analogs of the flowers they bear.

Characterization of SQUAMOSA-like genes in Gerbera hybrida, including one involved in reproductive transition

BackgroundThe flowering process in plants proceeds through the induction of an inflorescence meristem triggered by several pathways. Many of the genes associated with both the flowering process and floral architecture encode transcription factors of the MADS domain family. Gerbera, a member of the sunflower family, Asteraceae, bears compressed inflorescence heads (capitula) with three different flower types characterized by differences in both sexuality and floral symmetry. To understand how such a complex inflorescence structure is achieved at the molecular level, we have characterized the array of Gerbera MADS box genes. The high number of SQUAMOSA-like genes in Gerbera compared to other model species raised the question as to whether they may relate to Gerberas complex inflorescence structure and whether or not a homeotic A function is present.ResultsIn this paper we describe six Gerbera genes related to the SQUAMOSA/APETALA1/FRUITFULL genes of snapdragon and Arabidopsis. Based on phylogenetic analysis of the entire gene lineage, our data indicates that GSQUA1 and GSQUA3 are members of the SQUA/AP1 clade, while GSQUA2, GSQUA4, GSQUA5 and GSQUA6 are co-orthologs of the Arabidopsis FUL gene. GSQUA1/GSQUA3 and GSQUA4/GSQUA5/GSQUA6, respectively, represent several gene duplication events unknown in the model systems that may be specific to either Gerbera or Asteraceae. GSQUA genes showed specific expression profiles. GSQUA1, GSQUA2, and GSQUA5 were inflorescence abundant, while GSQUA3, GSQUA4, and GSQUA6 expression was also detected in vegetative organs. Overexpression of GSQUA2 in Gerbera led to accelerated flowering, dwarfism and vegetative abnormalities, all new and specific phenomena observed in transgenic Gerbera plants with modified MADS box gene expression.ConclusionsBased on expression patterns, none of the Gerbera SQUA-like genes are likely to control flower organ identity in the sense of the floral A function. However, our data shows that the FUL-like gene GSQUA2 plays a vital role in meristem transition. The roles of other GSQUA-genes in Gerbera floral development are intriguing, but require still further study.

Alternate wiring of a KNOXI genetic network underlies differences in leaf development of A. thaliana and C. hirsuta

Two interrelated problems in biology are understanding the regulatory logic and predictability of morphological evolution. Here, we studied these problems by comparing Arabidopsis thaliana, which has simple leaves, and its relative, Cardamine hirsuta, which has dissected leaves comprising leaflets. By transferring genes between the two species, we provide evidence for an inverse relationship between the pleiotropy of SHOOTMERISTEMLESS (STM) and BREVIPEDICELLUS (BP) homeobox genes and their ability to modify leaf form. We further show that cis-regulatory divergence of BP results in two alternative configurations of the genetic networks controlling leaf development. In C. hirsuta, ChBP is repressed by the microRNA164A (MIR164A)/ChCUP-SHAPED COTYLEDON (ChCUC) module and ChASYMMETRIC LEAVES1 (ChAS1), thus creating cross-talk between MIR164A/CUC and AS1 that does not occur in A. thaliana. These different genetic architectures lead to divergent interactions of network components and growth regulation in each species. We suggest that certain regulatory genes with low pleiotropy are predisposed to readily integrate into or disengage from conserved genetic networks influencing organ geometry, thus rapidly altering their properties and contributing to morphological divergence.

Genome sequencing and population genomic analyses provide insights into the adaptive landscape of silver birch

Silver birch (Betula pendula) is a pioneer boreal tree that can be induced to flower within 1 year. Its rapid life cycle, small (440-Mb) genome, and advanced germplasm resources make birch an attractive model for forest biotechnology. We assembled and chromosomally anchored the nuclear genome of an inbred B. pendula individual. Gene duplicates from the paleohexaploid event were enriched for transcriptional regulation, whereas tandem duplicates were overrepresented by environmental responses. Population resequencing of 80 individuals showed effective population size crashes at major points of climatic upheaval. Selective sweeps were enriched among polyploid duplicates encoding key developmental and physiological triggering functions, suggesting that local adaptation has tuned the timing of and cross-talk between fundamental plant processes. Variation around the tightly-linked light response genes PHYC and FRS10 correlated with latitude and longitude and temperature, and with precipitation for PHYC. Similar associations characterized the growth-promoting cytokinin response regulator ARR1, and the wood development genes KAK and MED5A.

Evolutionary Co-option of Floral Meristem Identity Genes for Patterning of the Flower-like Asteraceae Inflorescence

Highly conserved genes that regulate the identity of single flowers in conventional plant models regulate the unique inflorescence architecture of the evolutionarily successful Asteraceae plant family. The evolutionary success of Asteraceae, the largest family of flowering plants, has been attributed to the unique inflorescence architecture of the family, which superficially resembles an individual flower. Here, we show that Asteraceae inflorescences (flower heads, or capitula) resemble solitary flowers not only morphologically but also at the molecular level. By conducting functional analyses for orthologs of the flower meristem identity genes LEAFY (LFY) and UNUSUAL FLORAL ORGANS (UFO) in Gerbera hybrida, we show that GhUFO is the master regulator of flower meristem identity, while GhLFY has evolved a novel, homeotic function during the evolution of head-like inflorescences. Resembling LFY expression in a single flower meristem, uniform expression of GhLFY in the inflorescence meristem defines the capitulum as a determinate structure that can assume floral fate upon ectopic GhUFO expression. We also show that GhLFY uniquely regulates the ontogeny of outer, expanded ray flowers but not inner, compact disc flowers, indicating that the distinction of different flower types in Asteraceae is connected with their independent evolutionary origins from separate branching systems.