Patricia S. Springer

The Lateral Organ Boundaries Gene Defines a Novel, Plant-Specific Gene Family

The LATERAL ORGAN BOUNDARIES(LOB) gene in Arabidopsis defines a new conserved protein domain. LOB is expressed in a band of cells at the adaxial base of all lateral organs formed from the shoot apical meristem and at the base of lateral roots. LOB encodes a predicted protein that does not have recognizable functional motifs, but that contains a conserved domain (the LOB domain) that is present in 42 other Arabidopsis proteins and in proteins from a variety of other plant species. Proteins showing similarity to the LOB domain were not found outside of plant databases, indicating that this unique protein may play a role in plant-specific processes. Genes encoding LOB domain proteins are expressed in a variety of temporal- and tissue-specific patterns, suggesting that they may function in diverse processes. Loss-of-function LOB mutants have no detectable phenotype under standard growth conditions, suggesting thatLOB is functionally redundant or required during growth under specific environmental conditions. Ectopic expression ofLOB leads to alterations in the size and shape of leaves and floral organs and causes male and female sterility. The expression of LOB at the base of lateral organs suggests a potential role for LOB in lateral organ development.

The Arabidopsis LATERAL ORGAN BOUNDARIES–Domain Gene ASYMMETRIC LEAVES2 Functions in the Repression of KNOX Gene Expression and in Adaxial-Abaxial Patterning

The normal development of lateral organs of the shoot requires the simultaneous repression of meristem-specific genes and the activation of organ-specific genes. ASYMMETRIC LEAVES2 (AS2) is required for the development of normal leaf shape and for the repression of KNOX genes in the leaf. AS2 is a member of the recently identified, plant-specific LATERAL ORGAN BOUNDARIES (LOB)–domain gene family. Expression of AS2 at high levels resulted in repression of the KNOX homeobox genes BREVIPEDICELLUS, KNAT2, and KNAT6 but not of the related SHOOT MERISTEMLESS gene. Overexpression of AS2 also led to a perturbation of normal adaxial-abaxial asymmetry in lateral organs, resulting in the replacement of abaxial cell types with adaxial cell types. These results indicate that AS2 is sufficient to induce adaxial cell fate and repress KNOX gene expression.

Gene traps: tools for plant development and genomics.

Classical genetic approaches to gene identification rely on disruption of a gene leading to a recognizable phenotype. This approach continues to be an extremely successful one, yielding mutations that result in overt phenotypes reflecting the function of the corresponding gene. Not all genes,

Architecture of floral branch systems in maize and related grasses

The external appearance of flowering plants is determined to a large extent by the forms of flower-bearing branch systems, known as inflorescences, and their position in the overall structure of the plant. Branches and branching patterns are produced by tissues called shoot apical meristems. Thus, inflorescence architecture reflects meristem number, arrangement and activity, and the duration of meristem activity correlates with branch length. The inflorescences of maize, unlike those of related grasses such as rice and sorghum, predominantly lack long branches, giving rise to the tassel and familiar corncob. Here we report the isolation of the maize ramosa1 gene and show that it controls inflorescence architecture. Through its expression in a boundary domain near the nascent meristem base, ramosa1 imposes short branch identity as branch meristems are initiated. A second gene, ramosa2, acts through ramosa1 by regulating ramosa1 gene expression levels. ramosa1 encodes a transcription factor that appears to be absent in rice, is heterochronically expressed in sorghum, and may have played an important role in maize domestication and grass evolution.

LATERAL ORGAN BOUNDARIES defines a new family of DNA-binding transcription factors and can interact with specific bHLH proteins

Conserved in a variety of evolutionarily divergent plant species, LOB DOMAIN (LBD) genes define a large, plant-specific family of largely unknown function. LBD genes have been implicated in a variety of developmental processes in plants, although to date, relatively few members have been assigned functions. LBD proteins have previously been predicted to be transcription factors, however supporting evidence has only been circumstantial. To address the biochemical function of LBD proteins, we identified a 6-bp consensus motif recognized by a wide cross-section of LBD proteins, and showed that LATERAL ORGAN BOUNDARIES (LOB), the founding member of the family, is a transcriptional activator in yeast. Thus, the LBD genes encode a novel class of DNA-binding transcription factors. Post-translational regulation of transcription factors is often crucial for control of gene expression. In our study, we demonstrate that members of the basic helix–loop–helix (bHLH) family of transcription factors are capable of interacting with LOB. The expression patterns of bHLH048 and LOB overlap at lateral organ boundaries. Interestingly, the interaction of bHLH048 with LOB results in reduced affinity of LOB for the consensus DNA motif. Thus, our studies suggest that bHLH048 post-translationally regulates the function of LOB at lateral organ boundaries.

Brassinosteroids regulate organ boundary formation in the shoot apical meristem of Arabidopsis

Spatiotemporal control of the formation of organ primordia and organ boundaries from the stem cell niche in the shoot apical meristem (SAM) determines the patterning and architecture of plants, but the underlying signaling mechanisms remain poorly understood. Here we show that brassinosteroids (BRs) play a key role in organ boundary formation by repressing organ boundary identity genes. BR-hypersensitive mutants display organ-fusion phenotypes, whereas BR-insensitive mutants show enhanced organ boundaries. The BR-activated transcription factor BZR1 directly represses the CUP-SHAPED COTYLEDON (CUC) family of organ boundary identity genes. In WT plants, BZR1 accumulates at high levels in the nuclei of central meristem and organ primordia but at a low level in organ boundary cells to allow CUC gene expression. Activation of BR signaling represses CUC gene expression and causes organ fusion phenotypes. This study uncovers a role for BR in the spatiotemporal control of organ boundary formation and morphogenesis in the SAM.

KANADI1 regulates adaxial–abaxial polarity in Arabidopsis by directly repressing the transcription of ASYMMETRIC LEAVES2

Lateral organ polarity in Arabidopsis is regulated by antagonistic interactions between genes that promote either adaxial or abaxial identity, but the molecular basis of this interaction is largely unknown. We show that the adaxial regulator ASYMMETRIC LEAVES2 (AS2) is a direct target of the abaxial regulator KANADI1 (KAN1), and that KAN1 represses the transcription of AS2 in abaxial cells. Mutation of a single nucleotide in a KAN1 binding site in the AS2 promoter causes AS2 to be ectopically expressed in abaxial cells, resulting in a dominant, adaxialized phenotype. We also show that the abaxial expression of KAN1 is mediated directly or indirectly by AS2. These results demonstrate that KAN1 acts as a transcriptional repressor and that mutually repressive interactions between KAN1 and AS2 contribute to the establishment of adaxial–abaxial polarity in plants.

Arabidopsis LATERAL ORGAN BOUNDARIES negatively regulates brassinosteroid accumulation to limit growth in organ boundaries

Leaves and flowers begin life as outgrowths from the edges of shoot apical meristems. Stem cell divisions in the meristem center replenish cells that are incorporated into organ primordia at the meristem periphery and leave the meristem. Organ boundaries, regions of limited growth that separate forming organs from the meristem, serve to isolate these two domains and are critical for coordination of organogenesis and meristem maintenance. Boundary formation and maintenance are poorly understood processes, despite the identification of a number of boundary-specific transcription factors. Here we provide genetic and biochemical evidence that the Arabidopsis thaliana transcription factor LATERAL ORGAN BOUNDARIES (LOB) negatively regulates accumulation of the plant steroid hormone brassinosteroid (BR) in organ boundaries. We found that ectopic expression of LOB results in reduced BR responses. We identified BAS1, which encodes a BR-inactivating enzyme, as a direct target of LOB transcriptional activation. Loss-of-function lob mutants exhibit organ fusions, and this phenotype is suppressed by expression of BAS1 under the LOB promoter, indicating that BR hyperaccumulation contributes to the lob mutant phenotype. In addition, LOB expression is BR regulated; therefore, LOB and BR form a feedback loop to modulate local BR accumulation in organ boundaries to limit growth in the boundary domain.

LATERAL ORGAN FUSION1 and LATERAL ORGAN FUSION2 function in lateral organ separation and axillary meristem formation in Arabidopsis

Plant organs are generated from meristems throughout development. Patterning and elaboration of organ primordia occur as a result of organized cell division and expansion, processes that are likely to be controlled, in part, by meristem-derived signals. Communication between the meristem and lateral organs is crucial for meristem maintenance and organ patterning, and organ boundaries are thought to be important for mediating this communication. Arabidopsis thaliana LATERAL ORGAN FUSION1 (LOF1) encodes a MYB-domain transcription factor that is expressed in organ boundaries. lof1 mutants display defects in organ separation as a result of abnormal cell division and expansion during early boundary formation. lof1 mutants also fail to form accessory shoot meristems. Mutations in the closely related LATERAL ORGAN FUSION2 (LOF2) gene enhance the lof1 phenotype, such that lof1 lof2 double mutants display additional fusion defects. Genetic interactions with the CUP-SHAPED COTYLEDON genes CUC2 and CUC3 revealed a role for LOF1 in both organ separation and axillary meristem formation. Expression of the meristem determinant STM was reduced in lof1 mutant paraclade junctions and lof1 enhanced the weak stm-10 mutant, such that double mutants had severe defects in meristem maintenance and organ separation. Our data implicate LOF1 and LOF2 in boundary specification, meristem initiation and maintenance, and organ patterning.

The Arabidopsis gene PROLIFERA is required for proper cytokinesis during seed development.

Abstract. The Arabidopsis thaliana (L.) Heynh. gene PROLIFERA (PRL) is a member of the MCM family of genes that are required for DNA replication during the S phase of the cell cycle. PRL is expressed in dividing cells throughout plant development. During reproductive development, PRL is expressed in both the developing megaspore mother cells and microspore mother cells, but is not expressed in the developing microgametophyte, suggesting that it does not function in the final haploid divisions leading to the production of a mature pollen grain. Disruption of PRL leads to megagametophyte and embryo lethality. prl mutant embryos arrest at a variety of stages, and often show defects in cytokinesis. Multinucleate cells and non-stereotypical cell division planes are commonly observed in developing prl mutant embryos, although mcm mutations in other organisms have not been reported to affect cytokinesis. These observations suggest that PRL may play a role in cytokinesis that is distinct from its role in regulating DNA replication. Additionally, a novel cytokinesis checkpoint that monitors cell cycle progression may exist in Arabidopsis.