The CUP-SHAPED COTYLEDON1 gene of Arabidopsis regulates shoot apical meristem formation

In higher plants, molecular mechanisms regulating shoot apical meristem (SAM) formation and organ separation are largely unknown. The CUC1 (CUP-SHAPED COTYLEDON1) and CUC2 are functionally redundant genes that are involved in these processes. We cloned the CUC1 gene by a map-based approach, and found that it encodes a NAC-domain protein highly homologous to CUC2. CUC1 mRNA was detected in the presumptive SAM during embryogenesis, and at the boundaries between floral organ primordia. Surprisingly, overexpression of CUC1 was sufficient to induce adventitious shoots on the adaxial surface of cotyledons. Expression analyses in the overexpressor and in loss-of-function mutants suggest that CUC1 acts upstream of the SHOOT MERISTEMLESS gene.     

Arabidopsis CUP-SHAPED COTYLEDON3 regulates postembryonic shoot meristem and organ boundary formation

Overall shoot architecture in higher plants is highly dependent on the activity of embryonic and axillary shoot meristems, which are produced from the basal adaxial boundaries of cotyledons and leaves, respectively. In Arabidopsis thaliana, redundant functions of the CUP-SHAPED COTYLEDON genes CUC1, CUC2, and CUC3 regulate embryonic shoot meristem formation and cotyledon boundary specification. Their functional importance and relationship in postembryonic development, however, is poorly understood. Here, we performed extensive analyses of the embryonic and postembryonic functions of the three CUC genes using multiple combinations of newly isolated mutant alleles. We found significant roles of CUC2 and CUC3, but not CUC1, in axillary meristem formation and boundary specification of various postembryonic shoot organs, such as leaves, stems, and pedicels. In embryogenesis, all three genes make significant contributions, although CUC3 appears to possess, at least partially, a distinct function from that of CUC1 and CUC2. The function of CUC3 and CUC2 overlaps that of LATERAL SUPPRESSOR, which was previously shown to be required for axillary meristem formation. Our results reveal that redundant but partially distinct functions of CUC1, CUC2, and CUC3 are responsible for shoot organ boundary and meristem formation throughout the life cycle in Arabidopsis.     

Towards a functional understanding of cell growth dynamics in shoot meristem stem-cell niche

Shoot apical meristems (SAMs) harbor a set of stem-cells which supply cells for the development of all above-ground structures. A precise spatio-temporal control of growth patterns in stem-cells and the differentiating progeny is critical to maintain a stable set of stem-cells. In recent years, an array of approaches including molecular genetics, transient perturbations, live-imaging, image processing and mathematical modeling have been employed to study the cellular dynamics. In this article, we highlight recent studies that link cell–cell communication mechanisms to cell mechanics and overall growth control that govern stem-cell homeostasis and morphogenesis in SAMs.     

Gene NANA regulates cell proliferation in Arabidopsis thaliana shoot apical meristem without interaction with CLV1, CLV2, CLV3

A constancy of stem cell pool in shoot apical meristem of Arabidopsis thaliana is provided by a genetic regulation system with negative feedback loop based on the interaction of the gene WUS, which maintains indeterminate state of cells, with CLV genes, which restrict the level of WUS expression and stem cell pool size. clv mutations lead to an increase in the pool of stem cells in the apical and floral meristems and wus mutation leads to the opposite effect. Mutation na (nana), like wus mutation, causes premature termination of shoot apical meristem function, although it does not affect the activity of the flower meristem. To elucidate the role of NA in the control of shoot apical meristem functioning, the interaction of NA with CLV genes were investigated. Additive phenotype of double mutants na clv1, na clv2-1, and na clv3-2 indicates that the NA gene makes an independent contribution to the functioning of the shoot apical meristem. It is assumed that the NA gene controls apical meristem cell proliferation during the transition to the reproductive phase of plant development, acting much later and independently of the genes WUS-CLV.     

PIN-FORMED 1 regulates cell fate at the periphery of the shoot apical meristem

The process of organ positioning has been addressed, using the pin-formed 1 (pin1) mutant as a tool. PIN1 is a transmembrane protein involved in auxin transport in Arabidopsis. Loss of function severely affects organ initiation, and pin1 mutants are characterised by an inflorescence meristem that does not initiate any flowers, resulting in the formation of a naked inflorescence stem. This phenotype, combined with the proposed role of PIN1 in hormone transport, makes the mutant an ideal tool to study organ formation and phyllotaxis, and here we present a detailed analysis of the molecular modifications at the shoot apex caused by the mutation. We show that meristem structure and function are not severely affected in the mutant. Major alterations, however, are observed at the periphery of the pin1 meristem, where organ initiation should occur. Although two very early markers of organ initiation, LEAFY and AINTEGUMENTA, are expressed at the periphery of the mutant meristem, the cells are not recruited into distinct primordia. Instead a ring-like domain expressing those primordium specific genes is observed around the meristem. This ring-like domain also expresses a boundary marker, CUP-SHAPED COTYLEDON 2, involved in organ separation, showing that the zone at the meristem periphery has a hybrid identity. This implies that PIN1 is not only involved in organ outgrowth, but that it is also necessary for organ separation and positioning. A model is presented in which PIN1 and the local distribution of auxin control phyllotaxis.     

Cellular parameters of the shoot apical meristem in Arabidopsis.

The shoot apical meristem (SAM) is a small group of dividing cells that generate all of the aerial parts of the plant. With the goal of providing a framework for the analysis of Arabidopsis meristems at the cellular level, we performed a detailed morphometric study of actively growing inflorescence apices of the Landsberg erecta and Wassilewskija ecotypes. For this purpose, cell size, spatial distribution of mitotic cells, and the mitotic index were determined in a series of optical sections made with a confocal laser scanning microscope. The results allowed us to identify zones within the inflorescence SAM with different cell proliferation rates. In particular, we were able to define a central area that was four to six cells wide and had a low mitotic index. We used this technique to compare the meristem of the wild type with the enlarged meristems of two mutants, clavata3-1 (clv3-1) and mgoun2 (mgo2). One of the proposed functions of the CLV genes is to limit cell division rates in the center of the meristem. Our data allowed us to reject this hypothesis, because the mitotic index was reduced in the inflorescence meristem of the clv3-1 mutant. We also observed a large zone of slowly dividing cells in meristems of clv3-1 seedlings. This zone was not detectable in the wild type. These results suggest that the central area is increased in size in the mutant meristem, which is in line with the hypothesis that the CLV3 gene is necessary for the transition of cells from the central to the peripheral zone. Genetic and microscopic analyses suggest that mgo2 is impaired in the production of primordia, and we previously proposed that the increased size of the mgo2 meristem could be due to an accumulation of cells at the periphery. Our morphometric analysis showed that mgo2 meristems, in contrast to those of clv3-1, have an enlarged periphery with high cell proliferation rates. This confirms that clv3-1 and mgo2 lead to meristem overgrowth by affecting different aspects of meristem function.     

Photoperiod regulates flower meristem development in Arabidopsis thaliana

Photoperiod has been known to regulate flowering time in many plant species. In Arabidopsis, genes in the long day (LD) pathway detect photoperiod and promote flowering under LD. It was previously reported that clavata2 (clv2) mutants grown under short day (SD) conditions showed suppression of the flower meristem defects, namely the accumulation of stem cells and the resulting production of extra floral organs. Detailed analysis of this phenomenon presented here demonstrates that the suppression is a true photoperiodic response mediated by the inactivation of the LD pathway under SD. Inactivation of the LD pathway was sufficient to suppress the clv2 defects under LD, and activation of the LD pathway under SD conditions restored clv2 phenotypes. These results reveal a novel role of photoperiod in flower meristem development in Arabidopsis. Flower meristem defects of clv1 and clv3 mutants are also suppressed under SD, and 35S:CO enhanced the defects of clv3, indicating that the LD pathway works independently from the CLV genes. A model is proposed to explain the interactions between photoperiod and the CLV genes.     

Dual CLAVATA3 peptides in Arabidopsis shoot stem cell signaling

Plant shoot stem cell pool is constantly maintained by a negative feedback loop through peptide-receptor mediated signaling pathway. CLAVATA3 (CLV3) encode a 96 aminoacid protein which is processed to 12-amino-acid or arabinosylated 13-amino-acid peptides, acting as a ligand signal to regulate stem cell homeostasis in the shoot apical meristem (SAM). Although arabinosylated 13-amino-acid CLV3 peptide (CLV3p) shows more significant binding affinity to its receptors and biological activities in the SAM, the physiological function of two mature forms of CLV3p remained an unresolved puzzle in the past decade due to the technical difficulties of arabinosylation modification in the peptide synthesis. Here, we analyzed the role of two mature CLV3 peptides with newly synthesized arabinosylated peptide. Beside shoot meristem phenotypes, arabinosylated CLV3p showed the conventional trait of CLV2-dependent root growth inhibition. Moreover, both 12-amino-acid and arabinosylated 13-amino-acid CLV3 peptides have analogous activities in shoot stem cell signaling. Notably, we demonstrated that non-arabinosylated 12-amino acid CLV3p can affect shoot stem cell signaling at the physiological level unlike previously suggested (Ohyama et al. 2009; Shinohara and Matsubayashi 2013; Shinohara and Matsubayashi 2015). Therefore, these results support the physiological role of the 12-amino-acid CLV3p in shoot stem cell signaling in the deficient condition of arabinosylated 13-amino-acid CLV3p in Arabidopsis thaliana.