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.

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.L eaves and flowers are produced from the periphery of the shoot apical meristem, a self-perpetuating structure containing a population of self-renewing stem cells. Stem cell divisions in the meristem center replenish the cells that are incorporated into organ primordia at the meristem periphery and exit the meristem (1). The balance between organogenesis and meristem maintenance is essential for continued organ formation, and the boundary between the meristem and organ primordia plays a key role in maintaining the integrity of the meristem and differentiating organs. Boundary cells are small and divide infrequently relative to cells in the adjacent regions; thus, the boundary is a discrete domain that is distinct from the meristem and organ primordia (2-4). During organ formation, inhibition of growth in the boundary allows formation of a cleft, which results in separation of the forming organ from the meristem. A number of boundary-specific transcription factors in several families act redundantly to specify organ boundary cell fate and meristem maintenance (5-11). Few targets of boundary-specific transcription factors have been identified, and little is known about the physiological and biochemical processes they regulate.Arabidopsis LATERAL ORGAN BOUNDARIES (LOB) encodes a member of the plant-specific LOB-domain transcription factor family and is expressed specifically in organ boundaries (12). To investigate the developmental function of LOB, we examined the consequence of increased and decreased LOB activity and used expression profiling to identify targets of LOB transcriptional regulation. We show that LOB negatively regulate...

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.A daxial-abaxial polarity in plants is specified by interactions between genes that individually specify either adaxial or abaxial identity (1, 2). In Arabidopsis, adaxial identity is specified by class III homeodomain leucine zipper (HD-ZIPIII) transcription factors (3, 4), the transcription factor ASYMMETRIC LEAVES2 (AS2) (5, 6), and the transacting siRNA tasiARF (7), whereas abaxial identity is specified by KANADI (KAN) (8, 9), YABBY (YAB) (10, 11), AUXIN RESPONSE FACTOR (ARF) (12), and LITTLE ZIPPER (ZPR) (13) transcription factors and by the miRNAs miR165/miR166 (14-16). Genetic analysis indicates that many of these genes interact antagonistically: loss-of-function mutations in adaxial genes typically produce an abaxialized phenotype that is accompanied by the expanded expression of abaxial genes, whereas loss-of-function mutations in abaxial genes produce an adaxialized phenotype that is associated with the expanded expression of adaxial genes; mutations or transgenes that produce ubiquitous expression of these genes have a phenotype opposite to that of the loss-offunction mutations. miR165/166 and tasiARF repress the expression of their targets, respectively, HD-ZIPIII genes and ARF3/ETTIN, by directing the cleavage of the transcripts of these genes (14, 17). The molecular basis for the antagonistic interactions between other adaxial and abaxial regulators is unknown.KAN1, KAN2, and KAN3 are members of the GARP family of transcription factors and act redundantly to promote abaxial identity in both lateral organs and the shoot axis. These genes are expressed in abaxial cells of lateral organs and peripheral cells of the hypocotyl and stem (9, 16). Loss-of-function mutations in individual genes have relatively weak effects on organ polarity (9, 18), but kan1 kan2 (8) and kan1 kan2 kan3 (16) mutants are strongly adaxialized and resemble plants ectopically expressing the HD-ZIPIII genes PHB, PHV, and REV (3,15) or the LOB gene AS2 (5, 6). Consistent with this observation, PHB is abaxially expressed in kan1 kan2 kan3 triple mutants (16). The effect of kan on the expression of AS2 has not been examined.Here, we show that KAN1 promotes abaxial identity by di...

The LATERAL ORGAN BOUNDARIES DOMAIN (LBD) gene family encodes plant-specific transcription factors. In this report, the LBD gene DOWN IN DARK AND AUXIN1 (DDA1), which is closely related to LATERAL ORGAN BOUNDARIES (LOB) and ASYMMETRIC LEAVES2 (AS2), was characterized. DDA1 is expressed primarily in vascular tissues and its transcript levels were reduced by exposure to exogenous indole-3-acetic acid (IAA or auxin) and in response to dark exposure. Analysis of a T-DNA insertion line, dda1-1, in which the insertion resulted in misregulation of DDA1 transcripts in the presence of IAA and in the dark revealed possible functions in auxin response and photomorphogenesis. dda1-1 plants exhibited reduced sensitivity to auxin, produced fewer lateral roots, and displayed aberrant hypocotyl elongation in the dark. Phenotypes resulting from fusion of a transcriptional repression domain to DDA1 suggest that DDA1 may act as both a transcriptional activator and a transcriptional repressor depending on the context. These results indicate that DDA1 may function in both the auxin signalling and photomorphogenesis pathways.

The Arabidopsis LOB-domain (LBD) gene family is composed by 43 members divided in two classes based on amino acid conservation within the LOB-domain. The LOB domain is known to be responsible for DNA binding and protein-protein interactions. There is very little functional information available for most genes in the LBD family and many lbd single mutants do not exhibit conspicuous phenotypes. One plausible explanation for the limited loss-of-function phenotypes observed in this family is that LBD genes exhibit significant functional redundancy. Here we discuss an example of one phylogenetic subgroup of the LBD family, in which genes that are closely related based on phylogeny exhibit distinctly different expression patterns and do not have overlapping functions. We discuss the challenges of using phylogenetic analyses to predict redundancy in gene families.