Altered Leaf Shape Research Articles

Characterization of the KNOTTED1-like HOMEOBOX gene family in kiwifruit and functional analysis of AcKNOX11 related to plant growth, flowering, and melatonin-mediated germination inhibition

KNOX (KNOTTED1-like HOMEOBOX) plays essential roles in extensive plant developmental processes, although information on the KNOX gene has not been documented in kiwifruit (Actinidia chinensis). Here, we identified 19 KNOX gene (AcKNOX) members from the kiwifruit genome. The AcKNOX could be divided into three categories, and most members contain four conserved domains. AcKNOX promoters contain abundant CREs that respond to hormonal and stress cues. AcKNOX genes express in multiple tissues, and some of these genes respond to salt stress. It has been found in this study that AcKNOX11, selectively expressed in flower bud and shoot, activated during the lateral bud development and could be evoked by ABA and melatonin treatments. The AcKNOX11 protein located in the nucleus can be predicted to interact with other protein partners. Functional analysis revealed that AcKNOX11 delayed flowering, reduced plant height, altered leaf shape, and inhibited seed germination by ABA-melatonin crosstalk in Arabidopsis. This first thorough analysis of the AcKNOX genes will provide a more in-depth understanding of AcKNOX genes in kiwifruit and lay important theoretical basis for future functional dissection.

The strigolactone receptor SlDWARF14 plays a role in photosynthetic pigment accumulation and photosynthesis in tomato.

Tomato DWARF14 regulates the development of roots, shoot branches and leaves, and also plays a role in photosynthetic pigment accumulation and photosynthetic capacity. Strigolactones (SLs) are a novel class of plant hormones. DWARF14 (D14) is the only SL receptor identified to date, but it is not functionally analyzed in tomato (Solanum lycopersicum). In the present study, we identified the potential SL receptor in tomato by bioinformatic analysis, which was designated as SlD14. SlD14 was expressed in roots, stems, flowers and developing fruits, with the highest expression level in leaves. sld14 mutant plants produced by the CRISPR/Cas9 system displayed reduced plant height and root biomass, increased shoot branching and altered leaf shape comparing with WT plants. The cytokinin biosynthetic gene ISOPENTENYLTRANSFERASE 3 (SlIPT3), auxin biosynthetic genes FLOOZY (SlFZY) and TRYPTOPHAN AMINOTRANSFERASE RELATED 1 (SlTAR1) and several auxin transport genes SlPINs, which are involved in branch formation, showed higher expression levels in the sld14 plant stem. In addition, sld14 plants exhibited light-green leaves, reduced chlorophyll and carotenoid contents, abnormal chloroplast structure and reduced photosynthetic capacity. Transcriptomic analysis showed that the transcript levels of six chlorophyll biosynthetic genes, three carotenoid biosynthetic genes and numerous chlorophyll a/b-binding protein genes were decreased in sld14 plants. These results suggest that tomato SL receptor gene SlD14 not only regulates the development of roots, shoot branches and leaves, but also plays a role in regulating photosynthetic pigment accumulation and photosynthetic capacity.

Сорт мяты перечной Розовская Арома

The original plant of the peppermint variety Rozovskaya Aroma was obtained by vegetative propagation of cuttings of a mutant shoot with traits of increased length and altered leaf shape, discovered in 2010 on the variety Pamyat' Reznikovoy. As a result of vegetative propagation of cuttings, the breeding line L-9.24.455 was obtained, which received the commercial name "Rozovskaya Aroma". Based on the results of the variety trial of 2017–2019, the variety Rozovskaya Aroma exceeded the standard variety Vesna by 5.93 t/ha in terms of biomass yield. The shape of the bush of the variety Rozovskaya Aroma is semi-closed. Plant height is up to 95 cm. Leafiness degree 57.8%. The shape of the leaf blades is oval-rhombic, the color is green. The surface of the leaf is smooth, without pubescence. The size of the leaf blades is 6.0–7.0 × 2.5–3.5 cm. The inflorescence is loose, spike-shaped, with spaced whorls. The length of the inflorescences is 14–16 cm. Flowers are almost sterile. The color of the corolla of the flower is light pink. Stamens have undeveloped pollen sacs, and are longer than the corolla. The shape of the teeth of the calyx is triangular. The pubescence of the calyx is almost absent. The peppermint variety Rozovskaya Aroma is distinguished by a high content of essential oil in biomass – up to 4.70% and high yield of essential oil – up to 93.9 kg/ha. In 2020, the variety Ro-zovskaya Aroma was included in the State Register of Breeding Achievements and approved for cultivation in all regions of the Russian Federation.

PtrARF2.1 Is Involved in Regulation of Leaf Development and Lignin Biosynthesis in Poplar Trees.

Auxin response factors (ARFs) are important regulators modulating the expression of auxin-responsive genes in various biological processes in plants. In the Populus genome, a total of 39 ARF members have been identified, but their detailed functions are still unclear. In this study, six poplar auxin response factor 2 (PtrARF2) members were isolated from P. trichocarpa. Expression pattern analysis showed that PtrARF2.1 is highly expressed in leaf tissues compared with other PtrARF2 genes and significantly repressed by exogenous auxin treatment. PtrARF2.1 is a nuclear-localized protein without transcriptional activation activity. Knockdown of PtrARF2.1 by RNA interference (RNAi) in poplars led to the dwarf plant, altered leaf shape, and reduced size of the leaf blade, while overexpression of PtrARF2.1 resulted in a slight reduction in plant height and the similar leaf phenotype in contrast to the wildtype. Furthermore, histological staining analysis revealed an ectopic deposition of lignin in leaf veins and petioles of PtrARF2.1-RNAi lines. RNA-Seq analysis showed that 74 differential expression genes (DEGs) belonging to 12 transcription factor families, such as NAM, ATAF and CUC (NAC), v-myb avian myeloblastosis viral oncogene homolog (MYB), ethylene response factors (ERF) and basic helix–loop–helix (bHLH), were identified in PtrARF2.1-RNAi leaves and other 24 DEGs were associated with the lignin biosynthetic pathway. Altogether, the data indicate that PtrARF2.1 plays an important role in regulating leaf development and influences the lignin biosynthesis in poplars.

HEADLESS, a WUSCHEL homolog, uncovers novel aspects of shoot meristem regulation and leaf blade development in Medicago truncatula.

The formation and maintenance of the shoot apical meristem (SAM) are critical for plant development. However, the underlying molecular mechanism of regulating meristematic cell activity is poorly understood in the model legume Medicago truncatula. Using forward genetic approaches, we identified HEADLESS (HDL), a homolog of Arabidopsis WUSCHEL, required for SAM maintenance and leaf development in M. truncatula. Disruption of HDL led to disorganized specification and arrest of the SAM and axillary meristems, resulting in the hdl mutant being locked in the vegetative phase without apparent stem elongation. hdl mutant leaves are shorter in the proximal-distal axis due to reduced leaf length elongation, which resulted in a higher blade width/length ratio and altered leaf shape, uncovering novel phenotypes undescribed in the Arabidopsis wus mutant. HDL functions as a transcriptional repressor by recruiting MtTPL through its conserved WUS-box and EAR-like motif. Further genetic analysis revealed that HDL and STENOFOLIA (STF), a key regulator of M. truncatula lamina outgrowth, act independently in leaf development although HDL could recruit MtTPL in the same manner as STF does. Our results indicate that HDL has conserved and novel functions in regulating shoot meristems and leaf shape in M. truncatula, providing new avenues for understanding meristem biology and plant development.

Heterologous expression of rice 9-cis-epoxycarotenoid dioxygenase 4 (OsNCED4) in Arabidopsis confers sugar oversensitivity and drought tolerance

BackgroundThe 9-cis-epoxycarotenoid dioxygenases OsNCED4 was cloned from rice in conjunction with OsNCED 1-3 and 5, of which 3 has been shown to function in ABA biosynthesis and alteration of leaf morphology. In higher plants, NCEDs have been shown to be key enzymes controlling ABA biosynthesis and belong to a differentially expressed gene family. Aside from OsNCED3, it remains largely unknown if other OsNCED genes are involved in ABA biosynthesis in rice. Thus, transgenic Arabidopsis plants overexpressing OsNCED4 were generated in the 129B08/nced3 mutant background to explore OsNCED4 function in ABA biosynthesis.ResultsHeterologous expression of OsNCED4 in Arabidopsis increased ABA levels and altered plant size and leaf shape, delayed seed germination, caused sugar oversensitivity in post-germination growth, and enhanced tolerance to drought. The native OsNCED3 and OsNCED4 promoters were expressed in an overlapping pattern in rice seeds and young seedlings, suggesting possible functional redundancy between OsNCED3 and OsNCED4. At the one-leaf stage, similar regulation of OsNCED3 and OsNCED4 gene expression in roots or leaves in response to moderate salt stress (150 mM NaCl) was observed.ConclusionLike OsNCED3, OsNCED4 is functionally active in ABA biosynthesis in rice. OsNCED3 and OsNCED4 might play redundant roles in controlling ABA biosynthesis in rice, as suggested by GUS staining assay, but this should be further analyzed through complementation of rice NCED knockout mutants.

Differential Influences of Leaf Tip Trimming on Light Interception and Dry Matter Production in Tomato Dutch Cultivar Gourmet and Japanese Cultivar Momotaro York

To investigate the influence of morphological changes in individual leaves of tomato on light interception and dry matter (DM) production, we altered leaf shape by trimming leaflets of young or mature leaves of the Dutch cultivar Gourmet and the Japanese cultivar Momotaro York. Young leaves 5-cm long were trimmed of their first and second leaflets from the leaf apex. Mature leaves were similarly trimmed at ≥71 days after transplanting (DAT). The individual leaf area (LA) of intact ‘Momotaro York’ leaves was significantly larger than that of ‘Gourmet’. Light–photosynthesis curves of the cultivars were almost identical. Mature-trimmed plants of both cultivars had a smaller individual LA and a smaller leaf area index (LAI), and a greater light-extinction coefficient (LEC). Although there was no significant difference in light-use efficiency (LUE) (i.e., DM production per unit intercepted solar radiation) in ‘Gourmet’ between trimming stages, LUE of ‘Momotaro York’ was decreased significantly by young-leaf trimming. Trimming of young leaves significantly decreased the LEC in ‘Gourmet’ but increased it in ‘Momotaro York’. Although leaf trimming would be impractical for commercial cultivation, these results may provide with a clue for breeding for yield improvement.

The ANGULATA7 gene encodes a DnaJ-like zinc finger-domain protein involved in chloroplast function and leaf development in Arabidopsis.

The characterization of mutants with altered leaf shape and pigmentation has previously allowed the identification of nuclear genes that encode plastid-localized proteins that perform essential functions in leaf growth and development. A large-scale screen previously allowed us to isolate ethyl methanesulfonate-induced mutants with small rosettes and pale green leaves with prominent marginal teeth, which were assigned to a phenotypic class that we dubbed Angulata. The molecular characterization of the 12 genes assigned to this phenotypic class should help us to advance our understanding of the still poorly understood relationship between chloroplast biogenesis and leaf morphogenesis. In this article, we report the phenotypic and molecular characterization of the angulata7-1 (anu7-1) mutant of Arabidopsis thaliana, which we found to be a hypomorphic allele of the EMB2737 gene, which was previously known only for its embryonic-lethal mutations. ANU7 encodes a plant-specific protein that contains a domain similar to the central cysteine-rich domain of DnaJ proteins. The observed genetic interaction of anu7-1 with a loss-of-function allele of GENOMES UNCOUPLED1 suggests that the anu7-1 mutation triggers a retrograde signal that leads to changes in the expression of many genes that normally function in the chloroplasts. Many such genes are expressed at higher levels in anu7-1 rosettes, with a significant overrepresentation of those required for the expression of plastid genome genes. Like in other mutants with altered expression of plastid-encoded genes, we found that anu7-1 exhibits defects in the arrangement of thylakoidal membranes, which appear locally unappressed.

Inheritance of Leaf Tip Shape and Fringed Leaf Margin in Sunflower

AbstractAfter mutagenic treatment of sunflower seeds two traits of altered leaf shape were found – obtuse shape of leaf tip and strongly sinuate (fringed) leaf margin. To study the inheritance pattern of these traits, the mutant lines were crossed with the source lines and between themselves. It was shown that the traits of obtuse leaf and fringed leaf are recessive and inherited as monogenic traits when mating with the source line. In crossings of mutants, all F

Coupled enhancer and coding sequence evolution of a homeobox gene shaped leaf diversity.

Here we investigate mechanisms underlying the diversification of biological forms using crucifer leaf shape as an example. We show that evolution of an enhancer element in the homeobox gene REDUCED COMPLEXITY (RCO) altered leaf shape by changing gene expression from the distal leaf blade to its base. A single amino acid substitution evolved together with this regulatory change, which reduced RCO protein stability, preventing pleiotropic effects caused by its altered gene expression. We detected hallmarks of positive selection in these evolved regulatory and coding sequence variants and showed that modulating RCO activity can improve plant physiological performance. Therefore, interplay between enhancer and coding sequence evolution created a potentially adaptive path for morphological evolution.

Insights into Interspecific Hybridization Events in Allotetraploid Cotton Formation from Characterization of a Gene-Regulating Leaf Shape.

The morphology of cotton leaves varies considerably. Phenotypes, including okra, sea-island, super-okra, and broad leaf, are controlled by a multiple allele locus, L2 Okra leaf (L2°) is an incomplete mutation that alters leaf shape by increasing the length of lobes with deeper sinuses. Using a map-based cloning strategy, we cloned the L2 locus gene, which encodes a LATE MERISTEM IDENTITY 1 (LMI1)-like transcription factor (GhOKRA). Silencing GhOKRA leads to a change in phenotype from okra to broad leaf. Overexpression of GhOKRA in Arabidopsis thaliana greatly increases the degree of the leaf lobes and changes the leaf shape. Premature termination of translation in GhOKRA results in the production of broad leaves. The sequences of OKRA from diploid progenitor D-genome species, and wild races and domesticated allotetraploid cottons in Gossypium hirsutum show that a premature termination mutation occurred before and after the formation of tetraploid cotton, respectively. This study provides genomic insights into the two interspecific hybridization events: one produced the present broad leaf and another formed okra leaf phenotype with complete OKRA, that occurred during allotetraploid cotton formation.

Light-induced indeterminacy alters shade avoiding tomato leaf morphology

Plants sense the foliar shade of competitors and alter their developmental programs through the shade-avoidance response. Internode and petiole elongation, and changes in overall leaf area and leaf mass per area, are the stereotypical architectural responses to foliar shade in the shoot. However, changes in leaf shape and complexity in response to shade remain incompletely, and qualitatively, described. Using a meta-analysis of more than 18,000 previously published leaflet outlines, we demonstrate that shade avoidance alters leaf shape in domesticated tomato (Solanum lycopersicum) and wild relatives. The effects of shade avoidance on leaf shape are subtle with respect to individual traits but are combinatorially strong. We then seek to describe the developmental origins of shade-induced changes in leaf shape by swapping plants between light treatments. Leaf size is light responsive late into development, but patterning events, such as stomatal index, are irrevocably specified earlier. Observing that shade induces increases in shoot apical meristem size, we then describe gene expression changes in early leaf primordia and the meristem using laser microdissection. We find that in leaf primordia, shade avoidance is not mediated through canonical pathways described in mature organs but rather through the expression of KNOTTED1-LIKE HOMEOBOX and other indeterminacy genes, altering known developmental pathways responsible for patterning leaf shape. We also demonstrate that shade-induced changes in leaf primordium gene expression largely do not overlap with those found in successively initiated leaf primordia, providing evidence against classic hypotheses that shaded leaf morphology results from the prolonged production of juvenile leaf types.

Ectopic expression of Dendrobium EREB5 gene in Arabidopsis influences leaf morphology

The Dendrobium genus belongs to the Orchidaceae family and includes a broad range of terrestrial, epiphytic, and lithophytic plants. The genus includes many well-loved ornamental plants and valuable sources of traditional Chinese herbal medicines. Although popular and useful, their slow growth and vulnerability to environmental stress has strongly limited their more widespread adoption. The aim of our research is to expand the understanding of the regulation of Dendrobium growth and development. In this study, the gene encoding EREB5, a member of the APETALA2/ethylene-responsive element-binding protein (AP2/EREBP) family of transcription factors, was cloned from Dendrobium officinale. The full-length ORF of 558 bp, is predicted to code for a protein of 185 amino acids. The putative protein contains one copy of the well-conserved AP2 domain and a typical EAR motif, present in members of the ethylene response factor (ERF) subfamily. Arabidopsis thaliana plants ectopically expressing the DoEREB5 gene showed several phenotypic differences from wild-type plants, including altered leaf shape and petiole size. Expression of a DoEREB5 RNAi construct in Arabidopsis led to reduced levels of the putative Arabidopsis ortholog, AtERF9. Plants with reduced levels of AtERF9 showed an increase in chlorophyll content and photosynthetic rates compared to wild-type and heterologous ectopic DoEREB5 expression plants. Photosynthetic rates in the RNAi plants were higher than those in WT plants under both high and low light. Our results have provided clues as to the in vivo role of the Dendrobium gene, DoEREB5, suggesting that it is involved in the important processes of leaf development and function.

Control of organ shape and size by YUAN1 in Arabidopsis thaliana

Control of organ shape and size by cell proliferationandcell expansion is a fundamental process in plant development. However, little is known about the molecular mechanisms that set the shape and size of determinate organs in plants. We have previously demonstrated that the Arabidopsis gene DA1 controls the final size of organs by restricting cell proliferation. Through an activation tagging screen for modifiers of da1-1, we have identified a semi-dominant mutant (yuan1-1D) with altered leaf shape and size. The yuan1-1D mutation results in reduced plant height, short and round leaves and short petioles due to defects in cell elongation. YUAN1 encodes a PHD zinc finger domain-containing protein. The GFP-YUAN1 fusion protein is localized to the nucleus. Overexpression of YUAN1 leads to round leaves and short petioles. Genetic analyses show that YUAN1 acts independently of DA1, ROTUNDIFOLIA 3 (ROT3) and ROTUNDIFOLIA4 (ROT4) to influence leaf shape and size. Collectively, our findings show that Arabidopsis YUAN1, a PHD zinc finger domain-containing protein, controls organ shape and size by restricting cell elongation, and give insight into how plants control their organ shape and size.

Transcriptomic analysis of incised leaf-shape determination in birch

Plant researchers have focused much attention on leaf shape because of its importance in the identification. To evaluate the impact of intraspecies leaf-shape variation on the transcriptome, a series of Betula pendula ‘Dalecarlica’ and B. pendula saplings were generated through tissue culture. The leaf shapes and transcriptomes of B. pendula ‘Dalecarlica’ clones were compared with those of B. pendula clones. The leaf shape of B. pendula ‘Dalecarlica’ was incised and that of B. pendula was ovate. Transcriptome data revealed numerous changes in gene expression between B. pendula ‘Dalecarlica’ and B. pendula, including upregulation of 8767 unigenes and downregulation of 8379 unigenes in B. pendula ‘Dalecarlica’. A pathway analysis revealed that the transport and signal transduction of auxin were altered in ‘Dalecarlica’, which may have contributed to its altered leaf shape. These results shed light on variation in birch leaf shape and help identify important genes for the genetic engineering of birch trees.

The Arabidopsis IDD14, IDD15, and IDD16 Cooperatively Regulate Lateral Organ Morphogenesis and Gravitropism by Promoting Auxin Biosynthesis and Transport

The plant hormone auxin plays a critical role in regulating various aspects of plant growth and development, and the spatial accumulation of auxin within organs, which is primarily attributable to local auxin biosynthesis and polar transport, is largely responsible for lateral organ morphogenesis and the establishment of plant architecture. Here, we show that three Arabidopsis INDETERMINATE DOMAIN (IDD) transcription factors, IDD14, IDD15, and IDD16, cooperatively regulate auxin biosynthesis and transport and thus aerial organ morphogenesis and gravitropic responses. Gain-of-function of each IDD gene in Arabidopsis results in small and transversally down-curled leaves, whereas loss-of-function of these IDD genes causes pleiotropic phenotypes in aerial organs and defects in gravitropic responses, including altered leaf shape, flower development, fertility, and plant architecture. Further analyses indicate that these IDD genes regulate spatial auxin accumulation by directly targeting YUCCA5 (YUC5), TRYPTOPHAN AMINOTRANSFERASE of ARABIDOPSIS1 (TAA1), and PIN-FORMED1 (PIN1) to promote auxin biosynthesis and transport. Moreover, mutation or ectopic expression of YUC suppresses the organ morphogenic phenotype and partially restores the gravitropic responses in gain- or loss-of-function idd mutants, respectively. Taken together, our results reveal that a subfamily of IDD transcription factors plays a critical role in the regulation of spatial auxin accumulation, thereby controlling organ morphogenesis and gravitropic responses in plants.

Downregulation of multiple CDK inhibitor ICK/KRP genes upregulates the E2F pathway and increases cell proliferation, and organ and seed sizes in Arabidopsis

The ICK/KRP cyclin-dependent kinase (CDK) inhibitors are important plant cell cycle factors sharing only limited similarity with the metazoan CIP/KIP family of CDK inhibitors. Little is known about the specific functions of different ICK/KRP genes in planta. In this study, we created double and multiple mutants from five single Arabidopsis ICK/KRP T-DNA mutants, and used a set of 20 lines for the functional investigation of the important gene family. There were gradual increases in CDK activity from single to multiple mutants, indicating that ICK/KRPs act as CDK inhibitors under normal physiological conditions in plants. Whereas lower-order mutants showed no morphological phenotypes, the ick1ick2ick6ick7 and ick1ick2ick5ick6ick7 mutants had a slightly altered leaf shape. The quintuple mutant had larger cotyledons, leaves, petals and seeds than the wild-type control. At the cellular level, the ICK/KRP mutants had more but smaller cells in all the organs examined. These phenotypic effects became more apparent as more ICK/KRPs were downregulated, suggesting that to a large extent ICK/KRPs function in plants redundantly in a dosage-dependent manner. Analyses also revealed increased expression of E2F-dependent genes, and elevated RBR1 as well as an increased level of phospho-RBB1 protein in the quintuple mutant. Thus, downregulation of multiple ICK/KRP genes increases CDK activity, upregulates the E2F pathway and stimulates cell proliferation, resulting in increased cell numbers, and larger organs and seeds.

Regulation of Leaf Morphology by MicroRNA394 and its Target LEAF CURLING RESPONSIVENESS

The present study identified Arabidopsis miR394 and its target, an F-box (SKP1-Cullin/CDC53-F-box) gene At1g27340 (here referred to as LEAF CURLING RESPONSIVENESS, LCR), for regulation of leaf curling-related morphology. The loss-of-function lcr mutants exhibit pleiotropic defects with semi-dwarfism, altered leaf shape and a shorter stem. Overexpression of an miR394-resistant version of LCR under the 35S promoter (35S:m5LCR) and target mimicry MIM394 resulted in a curled-down leaf defect. Conversely, transgenic plants overexpressing 35S:MIR394a/b display a curled-up leaf phenotype. Detailed analyses show that there is a certain level of LCR that is optimal for leaf morphology, but lower or higher levels lead to abnormal leaf development, indicating that expression of miR394 in the leaf lamina is necessary for proper leaf morphology. Because the phytohormone auxin plays a crucial role in leaf morphogenesis and patterning, the DR5-GUS reporter gene was used to monitor the auxin response. We show that DR5 expression patterns in lcr and 35S::m5LCR plants were significantly different from those in the wild type. Also, overexpression of LCR in 35S::m5LCR plants drastically decreased the expression of the auxin-responsive genes IAA3, AXR3 and IAMT1, whereas increased expression of the genes was found in 35S::MIR394a plants. These results indicate that miR394 and its target LCR are involved in the regulation of leaf development.

Regulation of Leaf Morphology by MicroRNA394 and its Target LEAF CURLING RESPONSIVENESS

The present study identified Arabidopsis miR394 and its target, an F-box (SKP1-Cullin/CDC53-F-box) gene At1g27340 (here referred to as LEAF CURLING RESPONSIVENESS, LCR), for regulation of leaf curling-related morphology. The loss-of-function lcr mutants exhibit pleiotropic defects with semi-dwarfism, altered leaf shape and a shorter stem. Overexpression of an miR394-resistant version of LCR under the 35S promoter (35S:m5LCR) and target mimicry MIM394 resulted in a curled-down leaf defect. Conversely, transgenic plants overexpressing 35S:MIR394a/b display a curled-up leaf phenotype. Detailed analyses show that there is a certain level of LCR that is optimal for leaf morphology, but lower or higher levels lead to abnormal leaf development, indicating that expression of miR394 in the leaf lamina is necessary for proper leaf morphology. Because the phytohormone auxin plays a crucial role in leaf morphogenesis and patterning, the DR5-GUS reporter gene was used to monitor the auxin response. We show that DR5 expression patterns in lcr and 35S::m5LCR plants were significantly different from those in the wild type. Also, overexpression of LCR in 35S::m5LCR plants drastically decreased the expression of the auxin-responsive genes IAA3, AXR3 and IAMT1, whereas increased expression of the genes was found in 35S::MIR394a plants. These results indicate that miR394 and its target LCR are involved in the regulation of leaf development.

Identification of differentially expressed genes in a spontaneous altered leaf shape mutant of the navel orange [Citrus sinensis (L.) Osbeck

Most of the economically important citrus cultivars have originated from bud mutations. Leaf shape and structure are important factors that impact plant photosynthesis. We found a spontaneous bud mutant exhibiting a narrow leaf phenotype in navel orange [Citrus sinensis (L.) Osbeck]. To identify and characterize the genes involved in the formation of this trait, we performed suppression subtractive hybridization (SSH) and macroarray analysis. A total of 221 non-redundant differentially expressed transcripts were obtained. These transcripts included cell wall- and microtubule-related genes and two transcription factor-encoding genes, yabby and wox, which are crucial for leaf morphogenesis. Many highly redundant transcripts were associated with stress responses, while others, encoding caffeic acid 3-O-methyltransferase (EC 2.1.1.68) and a myb-like transcription factor, might be involved in the lignin pathway, which produces a component of secondary walls. Furthermore, real-time quantitative RT-PCR was performed for selected genes to validate the quality of the expressed sequence tags (ESTs) from the SSH libraries. This study represents an attempt to investigate the molecular mechanism associated with a leaf shape mutation, and its results provide new clues for understanding leaf shape mutations in citrus.