KNOTTED1-like Homeobox Proteins Research Articles

Genome-wide identification of the CclKNOX gene family and functional characterization of CclKNOX3 and CclKNOX5 in citrus shoot development

KNOTTED1-LIKE HOMEOBOX (KNOX) genes are a relatively conserved gene family in plants, and have important regulatory functions in the morphological development of plants. In this study, a total of 9 KNOX proteins were found from citrus and divided into two classes, Class I and Class II. Subsequently, their gene structures, protein characteristics and sequence features were analyzed. Spatial expression analysis showed that most of the genes exhibited a constitutive expression pattern in different tissues of Citrus clementina. Interestingly, the expression pattern of CclKNOX3 was different from other Class I members, as it was mainly expressed in stems, lateral buds and shoot tips. Overexpression of CclKNOX3 led to strong differentiation of shoot apical meristem and curled leaves in transgenic tobacco and lemon. In addition, CclKNOX3 was found to interact with CclKNOX5, localized in the nucleus and have transcriptional inhibitory activity. Further transgenic analysis found that 35S:CclKNOX5 transgenic tobacco showed similar phenotype with 35S:CclKNOX3. Furthermore, yeast one hybrid and luciferase assay confirmed that CclKNOX3 and CclKNOX5 were regulated by CclKNOX4. This study expands our knowledge about the citrus KNOX family and provides valuable informations for future studies on shoot developmental regulation.

Identification and expression analysis of the KNOX genes during organogenesis and stress responseness in Camellia sinensis (L.) O. Kuntze.

Tea plant (Camellia sinensis L.), whose leaves are the major reproductive organs, has been cultivated and consumed widely for its economic and health benefits. The Knotted1-like Homeobox (KNOX) proteins play significant roles in leaf morphology formation and development. However, the functions of KNOX proteins in tea plants are still unknown. Here, 11 CsKNOX genes from the tea plants were cloned and divided into Class I, II, and KNATM clades based on their protein sequences. These 11 CsKNOX genes were mapped on 8 out of 15 tea plant chromosomes, all localized in the nucleus. Specific spatiotemporal expression patterns of CsKNOX genes were found in various tissues and different development periods of buds, flowers, and roots of tea plants. Meanwhile, transcript levels of CsKNOX in tea leaves were strongly correlated with the accumulation of flavan-3-ols and proanthocyanidins. It was found that most of the CsKNOX genes could respond to drought, salt, cold, and exogenous MeJA and GA3 by analysis of transcriptomics data and promoter elements. The protein interaction analysis showed that CsKNOX could cooperate with CsAS1 and other critical functional proteins. In conclusion, this research provided the basic information for the functions of the CsKNOX family during organogenesis and stress response in tea plants, which was necessary for further functional characterization verification.

Prunus Knotted-like Genes: Genome-Wide Analysis, Transcriptional Response to Cytokinin in Micropropagation, and Rootstock Transformation.

Knotted1-like homeobox (KNOX) transcription factors are involved in plant development, playing complex roles in aerial organs. As Prunus species include important fruit tree crops of Italy, an exhaustive investigation of KNOX genes was performed using genomic and RNA-seq meta-analyses. Micropropagation is an essential technology for rootstock multiplication; hence, we investigated KNOX transcriptional behavior upon increasing 6-benzylaminopurine (BA) doses and the effects on GF677 propagules. Moreover, gene function in Prunus spp. was assessed by Gisela 6 rootstock transformation using fluorescence and peach KNOX transgenes. Based on ten Prunus spp., KNOX proteins fit into I-II-M classes named after Arabidopsis. Gene number, class member distribution, and chromosome positions were maintained, and exceptions supported the diversification of Prunus from Cerasus subgenera, and that of Armeniaca from the other sections within Prunus. Cytokinin (CK) cis-elements occurred in peach and almond KNOX promoters, suggesting a BA regulatory role in GF677 shoot multiplication as confirmed by KNOX expression variation dependent on dose, time, and interaction. The tripled BA concentration exacerbated stress, altered CK perception genes, and modified KNOX transcriptions, which are proposed to concur in in vitro anomalies. Finally, Gisela 6 transformation efficiency varied (2.6-0.6%) with the genetic construct, with 35S:GFP being more stable than 35S:KNOPE1 lines, which showed leaf modification typical of KNOX overexpression.

Genome-Wide Identification and Expression Pattern Analysis of KNOX Gene Family in Orchidaceae.

The establishment of lateral organs and subsequent plant architecture involves factors intrinsic to the stem apical meristem (SAM) from which they are derived. KNOTTED1-LIKE HOMEOBOX (KNOX) genes are a family of plant-specific homeobox transcription factors that especially act in determining stem cell fate in SAM. Although KNOXs have been studied in many land plants for decades, there is a dearth of knowledge on KNOX’s role in Orchidaceae, the largest and most diverse lineage of flowering plants. In this study, a total of 32 putative KNOX genes were identified in the genomes of five orchid species and further designated into two classes (Class I and Class II) based on phylogenetic relationships. Sequence analysis showed that most orchid KNOX proteins retain four conserved domains (KNOX1, KNOX2, ELK, and Homeobox_KN). Comparative analysis of gene structure showed that the exon–intron structure is conserved in the same clade but most orchids exhibited longer intron, which may be a unique feature of Orchidaceae. Cis-elements identified in the promoter region of orchid KNOXs were found mostly enriched in a function of light responsiveness, followed by MeJA and ABA responsiveness, indicative of their roles in modulating light and phytohormones. Collinear analysis unraveled a one-to-one correspondence among KNOXs in orchids, and all KNOX genes experienced strong purifying selection, indicating the conservation of this gene family has been reinforced across the Orchidaceae lineage. Expression profiles based on transcriptomic data and real-time reverse transcription–quantitative PCR (RT-qPCR) revealed a stem-specific expression of KNOX Class I genes and a broader expression pattern of Class II genes. Taken together, our results provided a comprehensive analysis to uncover the underlying function of KNOX genes in Orchidaceae.

Evolutionary Relationships and Divergence of KNOTTED1-Like Family Genes Involved in Salt Tolerance and Development in Cotton (Gossypium hirsutum L.).

The KNOX (KNOTTED1-like homeobox) transcription factors play an important role in leaf, shoot apical meristem and seed development and respond to biotic and abiotic stresses. In this study, we analyzed the diversity and evolutionary history of the KNOX gene family in the genome of tetraploid cotton (Gossypium hirsutum). Forty-four putative KNOX genes were identified. All KNOX genes from seven higher plant species were classified into KNOXI, KNOXII, and KNATM clades based on a phylogenetic analysis. Chromosomal localization and collinearity analysis suggested that whole-genome duplication and a polyploidization event contributed to the expansion of the cotton KNOX gene family. Analyses of expression profiles revealed that the GhKNOX genes likely responded to diverse stresses and were involved in cotton growth developmental processes. Silencing of GhKNOX2 enhanced the salt tolerance of cotton seedlings, whereas silencing of GhKNOX10 and GhKNOX14 reduced seedling tolerance to salt stress. Silencing of GhSTM3 influenced the cotton flowering time and plant development. These findings clarify the evolution of the cotton KNOX gene family and provide a foundation for future functional studies of KNOX proteins in cotton growth and development and response to abiotic stresses.

Genome wide identification of StKNOX gene family and characterization of their expression in Solanum tuberosum

The KNOTTED1-LIKE HOMEOBOX (KNOX) transcription factors (TF) play an important role in growth and developmental stages in potato. However, only few reports are available regarding the KNOX gene family in potato. In this study, 17 KNOX genes were detected in the potato genome. The gene structure, protein characteristics, promoter analysis, synthetic regions, gene duplication, gene expression in developmental stages, and chromosomal location were investigated. Based on their phylogenetic relationships, the StKNOX family members were categorized into three classes. Analysis of gene expression showed that the tissue expression patterns of StKNOX genes in the same group were similar but varied among groups. Analysis of transcription factor binding site (TFBS) and microRNA in the promoter region and coding sequence of StKNOX genes revealed the presence of putative TFBS and 49 StmiRNAs. Moreover, variable frequencies of TFSB and miRNA in StKNOX genes could indicate that these genes control different developmental stages and are involved in complex regulatory mechanisms. Our results showed that some members of Class II (StKNOX14) have the highest number of TFBS and are highly expressed in 19 potato tissues. Our findings showed that these genes could be involved in the regulation of plant growth and tuber development in S.tuberosum . The results of this study will be useful in further investigations on the functional role of potato StKNOX genes. • KNOX proteins are homeodomain transcription factors that have a critical role in morphogenesis. • Some members of KNAT2/6 (KNOX6/7/8) have the highest number of TFBS and are highly expressed in plant roots. • KNOX genes could be involved in regulation of plant growth and tuber development in S.tuberosum.

Integrated analysis of high-throughput sequencing data shows abscisic acid-responsive genes and miRNAs in strawberry receptacle fruit ripening

The perception and signal transduction of the plant hormone abscisic acid (ABA) are crucial for strawberry fruit ripening, but the underlying mechanism of how ABA regulates ripening-related genes has not been well understood. By employing high-throughput sequencing technology, we comprehensively analyzed transcriptomic and miRNA expression profiles simultaneously in ABA- and nordihydroguaiaretic acid (NDGA, an ABA biosynthesis blocker)-treated strawberry fruits with temporal resolution. The results revealed that ABA regulated many genes in different pathways, including hormone signal transduction and the biosynthesis of secondary metabolites. Transcription factor genes belonging to WRKY and heat shock factor (HSF) families might play key roles in regulating the expression of ABA inducible genes, whereas the KNOTTED1-like homeobox protein and Squamosa Promoter-Binding-like protein 18 might be responsible for ABA-downregulated genes. Additionally, 20 known and six novel differentially expressed miRNAs might be important regulators that assist ABA in regulating target genes that are involved in versatile physiological processes, such as hormone balance regulation, pigments formation and cell wall degradation. Furthermore, degradome analysis showed that one novel miRNA, Fa_novel6, could degrade its target gene HERCULES1, which likely contributed to fruit size determination during strawberry ripening. These results expanded our understanding of how ABA drives the strawberry fruit ripening process as well as the role of miRNAs in this process.

DIVARICATA AND RADIALIS INTERACTING FACTOR (DRIF) also interacts with WOX and KNOX proteins associated with wood formation in Populus trichocarpa.

DIVARICATA AND RADIALIS INTERACTING FACTOR (DRIF) from snapdragon (Antirrhinum majus) is a MYB/SANT protein that interacts with related MYB/SANT proteins, RADIALIS and DIVARICATA, through its N-terminal MYB/SANT domain. In addition to the MYB/SANT domain, DRIF contains a C-terminal domain of unknown function (DUF3755). Here we describe novel protein-protein interactions involving a poplar (Populus trichocarpa) homolog of DRIF, PtrDRIF1. In addition to interacting with poplar homologs of RADIALIS (PtrRAD1) and DIVARICATA (PtrDIV4), PtrDRIF1 interacted with members of other families within the homeodomain-like superfamily, including PtrWOX13c, a WUSCHEL-RELATED HOMEOBOX protein, and PtrKNAT7, a KNOTTED1-LIKE HOMEOBOX protein. PtrRAD1 and PtrDIV4 interacted with the MYB/SANT-containing N-terminal portion of PtrDRIF1, whereas DUF3755 was both necessary and sufficient for interactions with PtrWOX13c and PtrKNAT7. Of the two MYB/SANT domains present in PtrDIV4, only the N-terminal MYB/SANT domain interacted with PtrDRIF1. GFP-PtrDRIF1 expressed alone or with PtrRAD1 localized to the cytoplasm, whereas co-expression of GFP-PtrDRIF1 with PtrDIV4, PtrWOX13c or PtrKNAT7 resulted in nuclear localization of GFP-PtrDRIF1. Modified yeast two-hybrid (Y2H) and bimolecular fluorescence complementation (BiFC) experiments using PtrDRIF1 as a bridge protein revealed that PtrDRIF1 simultaneously interacted with PtrRAD1 and PtrWOX13c, but could not form a heterotrimeric complex when PtrDIV4 was substituted for PtrRAD1. Moreover, a Y2H competition assay indicated that PtrKNAT7 inhibits the interaction between PtrRAD1 and PtrDRIF1. The discovery of an additional protein-protein interaction domain in DRIF proteins, DUF3755, and its ability to form heterodimers and heterotrimers involving MYB/SANT and wood-associated homeodomain proteins, implicates DRIF proteins as mediators of a broader array of processes than previously reported.

Evolution, diversification, and expression of KNOX proteins in plants.

The KNOX (KNOTTED1-like homeobox) transcription factors play a pivotal role in leaf and meristem development. The majority of these proteins are characterized by the KNOX1, KNOX2, ELK, and homeobox domains whereas the proteins of the KNATM family contain only the KNOX domains. We carried out an extensive inventory of these proteins and here report on a total of 394 KNOX proteins from 48 species. The land plant proteins fall into two classes (I and II) as previously shown where the class I family seems to be most closely related to the green algae homologs. The KNATM proteins are restricted to Eudicots and some species have multiple paralogs of this protein. Certain plants are characterized by a significant increase in the number of KNOX paralogs; one example is Glycine max. Through the analysis of public gene expression data we show that the class II proteins of this plant have a relatively broad expression specificity as compared to class I proteins, consistent with previous studies of other plants. In G. max, class I protein are mainly distributed in axis tissues and KNATM paralogs are overall poorly expressed; highest expression is in the early plumular axis. Overall, analysis of gene expression in G. max demonstrates clearly that the expansion in gene number is associated with functional diversification.

Identification of warm day and cool night conditions induced flowering-related genes in a Phalaenopsis orchid hybrid by suppression subtractive hybridization

The influence of warm day and cool night conditions on induction of spikes in Phalaenopsis orchids has been studied with respect to photosynthetic efficiency, metabolic cycles and physiology. However, molecular events involved in spike emergence induced by warm day and cool night conditions are not clearly understood. We examined gene expression induced by warm day and cool night conditions in the Phalaenopsis hybrid Fortune Saltzman through suppression subtractive hybridization, which allowed identification of flowering-related genes in warm day and cool night conditions in spikes and leaves at vegetative phase grown under warm daily temperatures. In total, 450 presumably regulated expressed sequence tags (ESTs) were identified and classified into functional categories, including metabolism, development, transcription factor, signal transduction, transportation, cell defense, and stress. Furthermore, database comparisons revealed a notable number of Phalaenopsis hybrid Fortune Saltzman ESTs that matched genes with unknown function. The expression profiles of 24 genes (from different functional categories) have been confirmed by quantitative real-time PCR in induced spikes and juvenile apical leaves. The results of the real-time PCR showed that, compared to the vegetative apical leaves, the transcripts of genes encoding flowering locus T, AP1, AP2, KNOX1, knotted1-like homeobox protein, R2R3-like MYB, adenosine kinase 2, S-adenosylmethionine synthetase, dihydroflavonol 4-reductase, and naringenin 3-dioxygenase accumulated significantly higher levels, and genes encoding FCA, retrotransposon protein Ty3 and C3HC4-type RING finger protein accumulated remarkably lower levels in spikes of early developmental stages. These results suggested that the genes of two expression changing trends may play positive and negative roles in the early floral transition of Phalaenopsis orchids. In conclusion, spikes induced by warm day and cool night conditions were complex in Phalaenopsis orchids; nevertheless, several molecular flowering pathway-related genes were found. The acquired data form the basis for a molecular understanding of spike induction by warm day and cool night conditions in Phalaenopsis orchids.

Compound leaf development in the palmChamaedorea elegansis KNOX‐independent

How a leaf acquires its shape is a major and largely unresolved question in plant biology. This problem is particularly complex in the case of compound leaves, where the leaf blade is subdivided into leaflets. In many eudicots with compound leaves, class I KNOTTED1-LIKE HOMEOBOX (KNOX) genes are upregulated in the leaf primordium and promote leaflet initiation, while KNOX genes are restricted to the shoot apical meristem in simple-leaved plants. In monocots, however, little is known about the extent of KNOX contribution to compound leaf development, and we aimed to address this issue in the palm Chamaedorea elegans. We investigated the accumulation pattern of KNOX proteins in shoot apical meristems and leaf primordia of the palm C. elegans using immunolocalization experiments. KNOX proteins accumulated in vegetative and inflorescence apical meristems and in the subtending stem tissue, but not in the plicated regions of the leaf primordia. These plicated areas form during primary morphogenesis and are the only meristematic tissue in the developing primordium. In addition, KNOX proteins did not accumulate in any region of the developing leaf during secondary morphogenesis, when leaflets separate to create the final pinnately compound leaf. The compound leaf character in palms, C. elegans in particular and likely other pinnately compound palms, does not depend on the activities of KNOX proteins.

KNOX genes: versatile regulators of plant development and diversity

Knotted1-like homeobox (KNOX) proteins are homeodomain transcription factors that maintain an important pluripotent cell population called the shoot apical meristem, which generates the entire above-ground body of vascular plants. KNOX proteins regulate target genes that control hormone homeostasis in the meristem and interact with another subclass of homeodomain proteins called the BELL family. Studies in novel genetic systems, both at the base of the land plant phylogeny and in flowering plants, have uncovered novel roles for KNOX proteins in sculpting plant form and its diversity. Here, we discuss how KNOX proteins influence plant growth and development in a versatile context-dependent manner.

A developmental framework for dissected leaf formation in the Arabidopsis relative Cardamine hirsuta

The developmental basis for the generation of divergent leaf forms is largely unknown. Here we investigate this problem by studying processes that distinguish development of two related species: Arabidopsis thaliana, which has simple leaves, and Cardamine hirsuta, which has dissected leaves with individual leaflets. Using genetics, expression studies and cell lineage tracing, we show that lateral leaflet formation in C. hirsuta requires the establishment of growth foci that form after leaf initiation. These growth foci are recruited at the leaf margin in response to activity maxima of auxin, a hormone that polarizes growth in diverse developmental contexts. Class I KNOTTED1-like homeobox (KNOX) proteins also promote leaflet initiation in C. hirsuta, and here we provide evidence that this action of KNOX proteins is contingent on the ability to organize auxin maxima via the PINFORMED1 (PIN1) auxin efflux transporter. Thus, differential deployment of a fundamental mechanism polarizing cellular growth contributed to the diversification of leaf form during evolution.

KNOXLost theOX: TheArabidopsis KNATMGene Defines a Novel Class of KNOX Transcriptional Regulators Missing the Homeodomain

Three amino acid loop extension (TALE) homeodomain transcriptional regulators play a central role in plant and animal developmental programs. Plant KNOTTED1-like homeobox (KNOX) and animal Myeloid ecotropic viral integration site (MEIS) proteins share a TALE homeodomain and a MEINOX (MEIS-KNOX) domain, suggesting that an ancestral MEINOX-TALE protein predates the divergence of plants from fungi and animals. In this study, we identify and characterize the Arabidopsis thaliana KNATM gene, which encodes a MEINOX domain but not a homeodomain. Phylogenetic analysis of the KNOX family places KNATM in a new class and shows conservation in dicotyledons. We demonstrate that KNATM selectively interacts with Arabidopsis BELL TALE proteins through the MEINOX domain. The homeodomain is known to be necessary for KNOX-KNOX interaction. On the contrary, KNATM specifically dimerizes with the KNOX protein BREVIPEDICELLUS through an acidic coiled-coil domain. KNATM is expressed in proximal-lateral domains of organ primordia and at the boundary of mature organs; in accordance, genetic analyses identify a function for KNATM in leaf proximal-distal patterning. In vivo domain analyses highlighted KNATM functional regions and revealed a role as transcriptional regulator. Taken together, our data reveal a homeodomain-independent mechanism of KNOX dimerization and transcriptional regulation.

ASYMMETRIC LEAVES1 and auxin activities converge to repressBREVIPEDICELLUSexpression and promote leaf development inArabidopsis

Leaf development in higher plants requires the specification of leaf initials at the flanks of a pluripotent structure termed the shoot apical meristem. In Arabidopsis, this process is facilitated by negative interactions between class I KNOTTED1-like homeobox (KNOX) and ASYMMETRIC LEAVES1 (AS1) transcription factors, such that KNOX proteins are confined to the meristem and AS1 to leaf initials. Sites of leaf inception are also defined by local accumulation of the hormone auxin; however, it is unknown how auxin and AS1 activities are integrated to control leaf development. Here, we show that auxin and AS1 pathways converge to repress expression of the KNOX gene BREVIPEDICELLUS (BP) and thus promote leaf fate. We also demonstrate that regulated auxin gradients control leaf shape in a KNOX-independent fashion and that inappropriate KNOX activity in leaves perturbs these gradients, hence altering leaf shape. We propose that regulatory interactions between auxin, AS1 and KNOX activities may both direct leaf initiation and sculpt leaf form.

Ectopic Expression of KNOTTED1-Like Homeobox Protein Induces Expression of Cytokinin Biosynthesis Genes in Rice

Some phytohormones such as gibberellins (GAs) and cytokinins (CKs) are potential targets of the KNOTTED1-like homeobox (KNOX) protein. To enhance our understanding of KNOX protein function in plant development, we identified rice (Oryza sativa) genes for adenosine phosphate isopentenyltransferase (IPT), which catalyzes the rate-limiting step of CK biosynthesis. Molecular and biochemical studies revealed that there are eight IPT genes, OsIPT1 to OsIPT8, in the rice genome, including a pseudogene, OsIPT6. Overexpression of OsIPTs in transgenic rice inhibited root development and promoted axillary bud growth, indicating that OsIPTs are functional in vivo. Phenotypes of OsIPT overexpressers resembled those of KNOX-overproducing transgenic rice, although OsIPT overexpressers did not form roots or ectopic meristems, both of which are observed in KNOX overproducers. Expression of two OsIPT genes, OsIPT2 and OsIPT3, was up-regulated in response to the induction of KNOX protein function with similar kinetics to those of down-regulation of GA 20-oxidase genes, target genes of KNOX proteins in dicots. However, expression of these two OsIPT genes was not regulated in a feedback manner. These results suggest that OsIPT2 and OsIPT3 have unique roles in the developmental process, which is controlled by KNOX proteins, rather than in the maintenance of bioactive CK levels in rice. On the basis of these findings, we concluded that KNOX protein simultaneously decreases GA biosynthesis and increases de novo CK biosynthesis through the induction of OsIPT2 and OsIPT3 expression, and the resulting high-CK and low-GA condition is required for formation and maintenance of the meristem.

KNOX Action in Arabidopsis Is Mediated by Coordinate Regulation of Cytokinin and Gibberellin Activities

The shoot apical meristem (SAM) is a pluripotent group of cells that gives rise to the aerial parts of higher plants. Class-I KNOTTED1-like homeobox (KNOX) transcription factors promote meristem function partly through repression of biosynthesis of the growth regulator gibberellin (GA). However, regulation of GA activity cannot fully account for KNOX action. Here, we show that KNOX function is also mediated by cytokinin (CK), a growth regulator that promotes cell division and meristem function. We demonstrate that KNOX activity is sufficient to rapidly activate both CK biosynthetic gene expression and a SAM-localized CK-response regulator. We also show that CK signaling is necessary for SAM function in a weak hypomorphic allele of the KNOX gene SHOOTMERISTEMLESS (STM). Additionally, we provide evidence that a combination of constitutive GA signaling and reduced CK levels is detrimental to SAM function. Our results indicate that CK activity is both necessary and sufficient for stimulating GA catabolic gene expression, thus reinforcing the low-GA regime established by KNOX proteins in the SAM. We propose that KNOX proteins may act as general orchestrators of growth-regulator homeostasis at the shoot apex of Arabidopsis by simultaneously activating CK and repressing GA biosynthesis, thus promoting meristem activity.

CORKSCREW1 Defines a Novel Mechanism of Domain Specification in the Maize Shoot

In higher plants, determinate leaf primordia arise in regular patterns on the flanks of the indeterminate shoot apical meristem (SAM). The acquisition of leaf form is then a gradual process, involving the specification and growth of distinct domains within the three leaf axes. The recessive corkscrew1 (cks1) mutation of maize (Zea mays) disrupts both leaf initiation patterns in the SAM and domain specification within the mediolateral and proximodistal leaf axes. Specifically, cks1 mutant leaves exhibit multiple midribs and leaf sheath tissue differentiates in the blade domain. Such perturbations are a common feature of maize mutants that ectopically accumulate KNOTTED1-like homeobox (KNOX) proteins in leaf tissue. Consistent with this observation, at least two knox genes are ectopically expressed in cks1 mutant leaves. However, ectopic KNOX proteins cannot be detected. We therefore propose that CKS1 primarily functions within the SAM to establish boundaries between meristematic and leaf zones. Loss of gene function disrupts boundary formation, impacts phyllotactic patterns, and leads to aspects of indeterminate growth within leaf primordia. Because these perturbations arise independently of ectopic KNOX activity, the cks1 mutation defines a novel component of the developmental machinery that facilitates leaf-versus-shoot development in maize.

VAAMANA--a BEL1-like homeodomain protein, interacts with KNOX proteins BP and STM and regulates inflorescence stem growth in Arabidopsis.

Plant shoot growth depends on the activity of the shoot apical meristem (SAM), where organ primordia are initiated. In turn, the function of the SAM depends on the activities of homeodomain (HD) proteins of the knotted1-like homeobox (KNOX) class [Long et al., Nature 379 (1996) 66; Vollbrecht et al., Development 127 (2000) 3161]. In plants, KNOX proteins have been shown to interact specifically with the BEL1-like (BELL) class of homeodomain proteins [Bellaoui et al., Plant Cell 13 (2001) 2455; Muller et al., Plant 27 (2001) 13; Smith et al., Proc. Natl. Acad. Sci. USA 99 (2002) 9579], through a domain conserved between plants and animals. We have isolated a mutation in a BELL homeobox gene VAAMANA (VAN) that causes a dwarf phenotype. In addition, van inflorescence stems have clusters of cauline leaves; typically three are produced at each node. VAN interacts specifically with the class I KNOX proteins SHOOTMERISTEMLESS (STM), BREVIPEDICELLUS (BP), and KNAT6 (K6), and nuclear localisation of a VAN-GFP fusion depends on co-expression of STM or BP in tobacco leaves. This suggests that localisation of VAN, like that of the animal PBC homeodomain protein [Rieckhof et al., Cell 91 (1997) 171; Berthelsen et al., Genes Dev. 13 (1999) 946], is also regulated by interaction with a partner homeodomain protein.

The interaction of two homeobox genes, BREVIPEDICELLUS and PENNYWISE, regulates internode patterning in the Arabidopsis inflorescence.

Plant architecture results from the activity of the shoot apical meristem, which initiates leaves, internodes, and axillary meristems. KNOTTED1-like homeobox (KNOX) genes are expressed in specific patterns in the shoot apical meristem and play important roles in plant architecture. KNOX proteins interact with BEL1-like (BELL) homeodomain proteins and together bind a target sequence with high affinity. We have obtained a mutation in one of the Arabidopsis BELL genes, PENNYWISE (PNY), that appears phenotypically similar to the KNOX mutant brevipedicellus (bp). Both bp and pny have randomly shorter internodes and display a slight increase in the number of axillary branches. The double mutant shows a synergistic phenotype of extremely short internodes interspersed with long internodes and increased branching. PNY is expressed in inflorescence and floral meristems and overlaps with BP in a discrete domain of the inflorescence meristem where we propose the internode is patterned. The physical association of the PNY and BP proteins suggests that they participate in a complex that regulates early patterning events in the inflorescence meristem.