Recruitment of an ancient branching program to suppress carpel development in maize flowers

Carpels in maize undergo programmed cell death in half of the flowers initiated in ears and in all flowers in tassels. The HD-ZIP I transcription factor gene GRASSY TILLERS1 (GT1) is one of only a few genes known to regulate this process. To identify additional regulators of carpel suppression, we performed a gt1 enhancer screen and found a genetic interaction between gt1 and ramosa3 (ra3). RA3 is a classic inflorescence meristem determinacy gene that encodes a trehalose-6-phosphate (T6P) phosphatase (TPP). Dissection of floral development revealed that ra3 single mutants have partially derepressed carpels, whereas gt1;ra3 double mutants have completely derepressed carpels. Surprisingly, gt1 suppresses ra3 inflorescence branching, revealing a role for gt1 in meristem determinacy. Supporting these genetic interactions, GT1 and RA3 proteins colocalize to carpel nuclei in developing flowers. Global expression profiling revealed common genes misregulated in single and double mutant flowers, as well as in derepressed gt1 axillary meristems. Indeed, we found that ra3 enhances gt1 vegetative branching, similar to the roles for the trehalose pathway and GT1 homologs in the eudicots. This functional conservation over ∼160 million years of evolution reveals ancient roles for GT1-like genes and the trehalose pathway in regulating axillary meristem suppression, later recruited to mediate carpel suppression. Our findings expose hidden pleiotropy of classic maize genes and show how an ancient developmental program was redeployed to sculpt floral form.

Recruitment of an ancient branching program to suppress carpel development in maize flowers

Floral morphology is immensely diverse. One developmental process acting to shape this diversity is growth suppression. For example, grass flowers exhibit extreme diversity in floral sexuality, arising through differential suppression of stamens or carpels. In maize, carpels undergo programmed cell death in half of the flowers initiated in ears and in all flowers in tassels. The HD-ZIP I transcription factor gene GRASSY TILLERS1 (GT1) is one of only a few genes known to regulate this process. To identify additional regulators of carpel suppression, we performed a gt1 enhancer screen, and found a genetic interaction between gt1 and ramosa3 (ra3). RA3 is a classic inflorescence meristem determinacy gene that encodes a trehalose-6-phosphate (T6P) phosphatase (TPP). Dissection of floral development revealed that ra3 single mutants have partially derepressed carpels, whereas gt1; ra3 double mutants have completely derepressed carpels. Surprisingly, gt1 suppresses ra3 inflorescence branching, revealing a role for gt1 in meristem determinacy. Supporting these genetic interactions, GT1 and RA3 proteins colocalize to carpel nuclei in developing flowers. Global expression profiling revealed common genes misregulated in single and double mutant flowers, as well as in derepressed gt1 axillary meristems. Indeed, we found that ra3 enhances gt1 vegetative branching, similar to the roles for the trehalose pathway and GT1 homologs in the eudicots. This functional conservation over ~160 million years of evolution reveals ancient roles for GT1-like genes and the trehalose pathway in regulating axillary meristem suppression, later recruited to mediate carpel suppression. Our findings expose hidden pleiotropy of classic maize genes, and show how an ancient developmental program was redeployed to sculpt floral form.

Robust control of floral meristem determinacy by position-specific multifunctions of KNUCKLES

Floral organs are properly developed on the basis of timed floral meristem (FM) termination in Arabidopsis. In this process, two known regulatory pathways are involved. The WUSCHEL (WUS)-CLAVATA3 (CLV3) feedback loop is vital for the spatial establishment and maintenance of the FM, while AGAMOUS (AG)-WUS transcriptional cascades temporally repress FM. At stage 6 of flower development, a C2H2-type zinc finger repressor that is a target of AG, KNUCKLES (KNU), directly represses the stem cell identity gene WUS in the organizing center for FM termination. However, how the robust FM activity is fully quenched within a limited time frame to secure carpel development is not fully understood. Here, we demonstrate that KNU directly binds to the CLV1 locus and the cis-regulatory element on CLV3 promoter and represses their expression during FM determinacy control. Furthermore, KNU physically interacts with WUS, and this interaction inhibits WUS from sustaining CLV3 in the central zone. The KNU–WUS interaction also interrupts the formation of WUS homodimers and WUS–HAIRYMERISTEM 1 heterodimers, both of which are required for FM maintenance. Overall, our findings describe a regulatory framework in which KNU plays a position-specific multifunctional role for the tightly controlled FM determinacy.

Tomato CRABS CLAW paralogues interact with chromatin remodelling factors to mediate carpel development and floral determinacy

CRABS CLAW (CRC) orthologues play a crucial role in floral meristem (FM) determinacy and gynoecium formation across angiosperms, key developmental processes for ensuring successful plant reproduction and crop production. Here, we revealed that the incomplete penetrance and variable expressivity of the carpel-inside-carpel phenotype observed in flowers of the tomato fruit iterative growth (fig) mutant is due to a lack of function of a homologue of the CRC gene, Solanum lycopersicum CRCa (SlCRCa). Likewise, a comprehensive functional analysis of SlCRCa and SlCRCb paralogues, including Arabidopsis complementation experiments, allowed us to propose that they operate as positive regulators of FM determinacy by acting in a compensatory and partially redundant manner to safeguard the proper formation of flowers and fruits. Furthermore, we provide the first evidence for the role of putative CRC orthologues as members of the chromatin remodelling complex that terminates floral stem cell activity by repressing WUSCHEL expression.

A CYC-RAD-DIV-DRIF interaction likely pre-dates the origin of floral monosymmetry in Lamiales

BackgroundAn outstanding question in evolutionary biology is how genetic interactions defining novel traits evolve. They may evolve either by de novo assembly of previously non-interacting genes or by en bloc co-option of interactions from other functions. We tested these hypotheses in the context of a novel phenotype—Lamiales flower monosymmetry—defined by a developmental program that relies on regulatory interaction among CYCLOIDEA , RADIALIS , DIVARICATA , and DRIF gene products. In Antirrhinum majus (snapdragon), representing Lamiales, we tested whether components of this program likely function beyond their previously known role in petal and stamen development. In Solanum lycopersicum (tomato), representing Solanales which diverged from Lamiales before the origin of Lamiales floral monosymmetry, we additionally tested for regulatory interactions in this program. ResultsWe found that RADIALIS , DIVARICATA , and DRIF are expressed in snapdragon ovaries and developing fruit, similar to their homologs during tomato fruit development. Additionally, we found that a tomato CYCLOIDEA ortholog positively regulates a tomato RADIALIS ortholog. ConclusionOur results provide preliminary support to the hypothesis that the developmental program defining floral monosymmetry in Lamiales was co-opted en bloc from a function in carpel development. This expands our understanding of novel trait evolution facilitated by co-option of existing regulatory interactions.

The Relationship between AGAMOUS and Cytokinin Signaling in the Establishment of Carpeloid Features

Gynoecium development is dependent on gene regulation and hormonal pathway interactions. The phytohormones auxin and cytokinin are involved in many developmental programs, where cytokinin is normally important for cell division and meristem activity, while auxin induces cell differentiation and organ initiation in the shoot. The MADS-box transcription factor AGAMOUS (AG) is important for the development of the reproductive structures of the flower. Here, we focus on the relationship between AG and cytokinin in Arabidopsis thaliana, and use the weak ag-12 and the strong ag-1 allele. We found that cytokinin induces carpeloid features in an AG-dependent manner and the expression of the transcription factors CRC, SHP2, and SPT that are involved in carpel development. AG is important for gynoecium development, and contributes to regulating, or else directly regulates CRC, SHP2, and SPT. All four genes respond to either reduced or induced cytokinin signaling and have the potential to be regulated by cytokinin via the type-B ARR proteins. We generated a model of a gene regulatory network, where cytokinin signaling is mainly upstream and in parallel with AG activity.

Molecular Control of Carpel Development in the Grass Family

Carpel is the ovule-bearing female reproductive organ of flowering plants and is required to ensure its protection, an efficient fertilization, and the development of diversified types of fruits, thereby it is a vital element of most food crops. The origin and morphological changes of the carpel are key to the evolution and adaption of angiosperms. Progresses have been made in elucidating the developmental mechanisms of carpel establishment in the model eudicot plant Arabidopsis thaliana, while little and fragmentary information is known in grasses, a family that includes many important crops such as rice (Oryza sativa), maize (Zea mays), barley (Hordeum vulgare), and wheat (Triticum aestivum). Here, we highlight recent advances in understanding the mechanisms underlying potential pathways of carpel development in grasses, including carpel identity determination, morphogenesis, and floral meristem determinacy. The known role of transcription factors, hormones, and miRNAs during grass carpel formation is summarized and compared with the extensively studied eudicot model plant Arabidopsis. The genetic and molecular aspects of carpel development that are conserved or diverged between grasses and eudicots are therefore discussed.