LATERAL ORGAN BOUNDARIES DOMAIN Proteins Research Articles

Genome-wide characterization of LBD transcription factors in switchgrass (Panicum virgatum L.) and the involvement of PvLBD12 in salt tolerance.

PvLBD12 enhanced the salt tolerance by increasing proline accumulation, improving K+ accumulation, and decreasing reactive oxygen species level in switchgrass. Abiotic stresses are the serious factors which limit plant development and productivity and restrict the agricultural economy. It is important, therefore, to understand the mechanism of abiotic tolerance in plants. Lateral organ boundaries domain (LBD) proteins as plant-specific transcription factors play important function in plant lateral organ development, plant regeneration, and abiotic stress. In our study, we identify 69 LBD members from switchgrass genome-wide sequences and classify them based on their homology with LBD proteins in Arabidopsis. RT-qPCR showed that PvLBD genes had different expression patterns under abiotic stress conditions, indicating that they play important roles in various stress. PvLBD12 was selected as a candidate gene for further functional analysis because it had the highest expression level under salt stress. Overexpression of PvLBD12 enhanced salt tolerance by altering a wide range of physiological responses (like increased proline accumulation, reduced malondialdehyde production, improved K+ accumulation, and reduced Na+ absorption) in switchgrass. Some stress response genes such as proline biosynthesis gene PvP5CS1, vacuolar Na+(K+)/H+ antiporter gene PvNHX1, two key ROS-scavenging enzyme genes PvCAT and PvSOD were all upregulated in PvLBD12 overexpression lines. Taken together, PvLBD12 plays a pivotal role in response to salt stress by increasing proline accumulation, improving K+ accumulation, reducing Na+ absorption, and decreasing reactive oxygen species level. It will be better to understand the potential biological functions of LBD genes in other plants.

Genome-wide analysis of the LBD family in rice: Gene functions, structure and evolution

Recent evidence suggests that LATERAL ORGAN BOUNDARIES DOMAIN (LBD) proteins are involved in different developmental processes of plants. Although the roles of LBD proteins in root development, leaf development and plant defense have been well summarized, their functional diversity and regulation mechanisms are still unclear. One of the reasons for the above problems is the lack of selection and classification of functional protein features of LBD genes. Combined with the existing research results, we found that LBD genes have similar features and mechanics and tend to be in the same phylogenetic branch. Research on the function of the LBD gene can expand our understanding of the diversity and function of LBD proteins. Therefore, to fully understand this large family, it is necessary to review functional studies through in-depth phylogenetic analysis of more genome-available species.

PheLBD29, an LBD transcription factor from Moso bamboo, causes leaf curvature and enhances tolerance to drought stress in transgenic Arabidopsis

The lateral organ boundaries domain (LBD), a unique family of transcription factors in higher plants, plays a key role in plant growth and development, and stress response. Here, we report on the novel lateral organ boundaries domain (LBD) gene PheLBD29, a nuclear localization protein that can bind the conserved GCCCCG sequence, as determined by electrophoretic mobility shift assay (EMSA). PheLBD29 was highly expressed in blade leaf and significantly induced by polyethylene glycol (PEG). Overexpression of PheLBD29 leads to small and abaxially rolled leaves in Arabidopsis, and anatomically, 35S:PheLBD29 Arabidopsis leaves showed transformation of adaxial cells into abaxial cells. Moreover, overexpression of PheLBD29 in Arabidopsis increased plant tolerance to drought stress, by accumulation of more soluble sugars, less malondialdehyde (MDA), and had lower REL levels under drought stress. Transient expression assay revealed PheLBD29 directly bound to the promoter region of RAB18. In addition, 35S:PheLBD29 Arabidopsis showed higher sensitivity to abscisic acid (ABA) than the wild type. Therefore, we conclude that PheLBD29 may participate in the ABA-dependent signaling pathway to improve drought tolerance. Our study provides new evidence for a Moso bamboo LBD protein regulatory module in leaf curvature and drought resistance.

ZmLBD2 a maize (Zea mays L.) lateral organ boundaries domain (LBD) transcription factor enhances drought tolerance in transgenicArabidopsis thaliana.

Maize (Zea mays L.) is an annual gramineous herb and is among the world’s most important crop species. Drought is the main factor contributing to maize yield reduction. The lateral organ boundaries domain (LBD) proteins belong to a class of higher-plant-specific transcription factors. LBD proteins usually include the highly conserved lateral organ boundaries (LOB) domains that play essential roles in plant growth and response to biotic stresses. However, few studies have addressed the biological functions of LBD genes associated with maize response to drought. Here we cloned the ZmLBD2 gene from maize and described its role in combating drought. Investigating ZmLBD2 subcellular localization, we show that it localizes to the cell nucleus and can specifically bind with inverted repeats of “GCGGCG”. Under drought stress, Arabidopsis thaliana overexpressing ZmLBD2 performed better than the wild-type plants in terms of seed germination rates, root length, relative water content, fresh weight, chlorophyll content, proline content, and antioxidant enzyme content. Arabidopsis overexpressing ZmLBD2 contained less MDA, H2O2, and than the wild-type plants. Our protein-protein interaction results indicate an interaction between the ZmLBD2 and ZmIAA5 genes. In conclusion, the ZmLBD2 gene positively regulates H2O2 homeostasis in plants, strengthening drought resistance.

An auxin-responsive transcription factor AbLBD1 promotes the development of lateral roots and reduces the biosynthesis of tropane alkaloids in Atropa belladonna

Atropa belladonna produces anticholinergic tropane alkaloids (TAs), such as hyoscyamine and scopolamine, which are synthesized in its lateral roots. Yet, the regulation of lateral root growth and TA biosynthesis remain poorly understood. In this study, an auxin-responsive transcription factor (AbLBD1), encoding a lateral organ boundaries domain protein (LBD), was functionally characterized from A. belladonna. Overexpression of AbLBD1 effectively promoted the initiation of lateral roots and adventitious roots of A. belladonna. The root biomass of AbLBD1-overexpressing lines was 1.48 to 2.45 fold higher that of the control. However, the contents of hyoscyamine and scopolamine were markedly reduced in AbLBD1-overexpressing lines. Gene expression analysis showed that the expression levels of TAs biosynthesis genes, namely AbPMT, AbPYKS, AbCYP82M3, and AbH6H, were greatly decreased in AbLBD1-overpressing lines. In conclusion, AbLBD1 plays a positive role in regulating lateral root development but a negative one in regulating TAs biosynthesis in A. belladonna. This study proved for the first time that AbLBD1 was a key auxin response regulator in root, which promotes initiation of lateral root and inhibits the biosynthesis of tropane alkaloids.

CHARACTERIZATION AND IN SILICO ANALYSES OF RTCS GENE FROM SUGARCANE ENCODING LOB PROTEIN FAMILY OF TRANSCRIPTION FACTORS: A KEY REGULATOR OF SHOOT-BORNE ROOT INITIATION

RTCS (Rootless concerning Crown and Seminal roots) is a member of the plant-specific LATERAL ORGAN BOUNDARIES (LOB) domain (LBD) protein family of transcription factors. The RTCS gene has been identified as a key regulator of shoot borne root initiation in maize. In this study an attempt has been made to characterize RTCS from a wild related genus of sugarcane Erianthus arundinaceus and compared to the RTCS gene isolated from commercial sugarcane hybrid genotypes. The candidate gene has been successfully isolated from two genotypes of E. arundinaceus (SES 288 and IND 04- 1335: EaRTCS) and two commercial sugarcane hybrid genotypes (Co 775 and Co 86032: ShRTCS). The isolated RTCS full length gene showed significant variation between EaRTCS and ShRTCS at the DNA and protein level. The RTCS amino acid sequence of all the four genotypes indicated the presence of the characteristic structural features of LBD proteins such as a C-motif, LOB domain sequence and leucine zipper-like dimerization domain. Prediction and analysis of secondary structure of RTCS protein in SES 288 and IND 04-1335 showed the presence of 65% alpha helix, 1% β strand and 38% disorder, while sugarcane commercial genotypes showed 58 % alpha helix, 1% β strand and 47% disorder. Both the sequences of EaRTCS and ShRTCS was found to be closely related to Zea mays RTCS.

Phylogeny and Functions of LOB Domain Proteins in Plants.

Lateral organ boundaries (LOB) domain (LBD) genes, a gene family encoding plant-specific transcription factors, play important roles in plant growth and development. At present, though there have been a number of genome-wide analyses on LBD gene families and functional studies on individual LBD proteins, the diverse functions of LBD family members still confuse researchers and an effective strategy is required to summarize their functional diversity. To further integrate and improve our understanding of the phylogenetic classification, functional characteristics and regulatory mechanisms of LBD proteins, we review and discuss the functional characteristics of LBD proteins according to their classifications under a phylogenetic framework. It is proved that this strategy is effective in the anatomy of diverse functions of LBD family members. Additionally, by phylogenetic analysis, one monocot-specific and one eudicot-specific subclade of LBD proteins were found and their biological significance in monocot and eudicot development were also discussed separately. The review will help us better understand the functional diversity of LBD proteins and facilitate further studies on this plant-specific transcription factor family.

Comprehensive characterization and gene expression patterns of LBD gene family in Gossypium

Main conclusionA comprehensive account of the LBD gene family of Gossypium was provided in this work. Expression analysis and functional characterization revealed that LBD genes might play different roles in G. hirsutum and G. barbadense.The Lateral Organ Boundaries Domain (LBD) proteins comprise a plant-specific transcription factor family, which plays crucial roles in physiological processes of plant growth, development, and stress tolerance. In the present work, a systematical analysis of LBD gene family from two allotetraploid cotton species, G. hirsutum and G. barbadense, together with their genomic donor species, G. arboreum and G. raimondii, was conducted. There were 131, 128, 62, and 68 LBDs identified in G. hirsutum, G. barbadense, G. arboreum and G. raimondii, respectively. The LBD proteins could be classified into two main classes, class I and class II, based on the structure of their lateral organ boundaries domain and traits of phylogenetic tree, and class I was further divided into five subgroups. The gene structure and motif composition analyses conducted in both G. hirsutum and G. barbadense revealed that LBD genes kept relatively conserved within the subfamilies. Synteny analysis suggested that segmental duplication acted as an important mechanism in expansion of the cotton LBD gene family. Cis-element analysis predicated the possible functions of LBD genes. Public RNA-seq data were investigated to analyze the expression patterns of cotton LBD genes in various tissues as well as gene expression under abiotic stress treatments. Furthermore, RT-qPCR results found that GhLBDs had various expression regulation under MeJA treatments. Expression analysis indicated the differential functions of cotton LBD genes in response to abiotic stress and hormones.

Structural and functional insights into the LBD family involved in abiotic stress and flavonoid synthases in Camellia sinensis

Lateral organ boundaries domain (LBD) proteins are plant-specific transcription factors that play a crucial role in growth and development, as well as metabolic processes. However, knowledge of the function of LBD proteins in Camellia sinensis is limited, and no systematic investigations of the LBD family have been reported. In this study, we identified 54 LBD genes in Camellia sinensis. The expression patterns of CsLBDs in different tissues and their transcription responses to exogenous hormones and abiotic stress were determined by RNA-seq, which showed that CsLBDs may have diverse functions. Analysis of the structural gene promoters revealed that the promoters of CsC4H, CsDFR and CsUGT84A, the structural genes involved in flavonoid biosynthesis, contained LBD recognition binding sites. The integrative analysis of CsLBD expression levels and metabolite accumulation also suggested that CsLBDs are involved in the regulation of flavonoid synthesis. Among them, CsLOB_3, CsLBD36_2 and CsLBD41_2, localized in the nucleus, were selected for functional characterization. Yeast two-hybrid assays revealed that CsLBD36_2 and CsLBD41_2 have self-activation activities, and CsLOB_3 and CsLBD36_2 can directly bind to the cis-element and significantly increase the activity of the CsC4H, CsDFR and CsUGT84A promoter. Our results present a comprehensive characterization of the 54 CsLBDs in Camellia sinensis and provide new insight into the important role that CsLBDs play in abiotic and flavonoid biosynthesis.

Structural analysis reveals a “molecular calipers” mechanism for a LATERAL ORGAN BOUNDARIES DOMAIN transcription factor protein from wheat

LATERAL ORGAN BOUNDARIES DOMAIN (LBD) proteins, a family of plant-specific transcription factors harboring a conserved Lateral Organ Boundaries (LOB) domain, are regulators of plant organ development. Recent studies have unraveled additional pivotal roles of the LBD protein family beyond defining lateral organ boundaries, such as pollen development and nitrogen metabolism. The structural basis for the molecular network of LBD-dependent processes remains to be deciphered. Here, we solved the first structure of the homodimeric LOB domain of Ramosa2 from wheat (TtRa2LD) to 1.9 Å resolution. Our crystal structure reveals structural features shared with other zinc-finger transcriptional factors, as well as some features unique to LBD proteins. Formation of the TtRa2LD homodimer relied on hydrophobic interactions of its coiled-coil motifs. Several specific motifs/domains of the LBD protein were also involved in maintaining its overall conformation. The intricate assembly within and between the monomers determined the precise spatial configuration of the two zinc fingers that recognize palindromic DNA sequences. Biochemical, molecular modeling, and small-angle X-ray scattering experiments indicated that dimerization is important for cooperative DNA binding and discrimination of palindromic DNA through a molecular calipers mechanism. Along with previously published data, this study enables us to establish an atomic-scale mechanistic model for LBD proteins as transcriptional regulators in plants.

Genome-wide identification, classification and expression pattern of LBD gene family in Camellia sinensis

The plant-specific LBD (LATERAL ORGAN BOUNDARIES DOMAIN) genes belong to a major family of transcription factors that encode a zinc finger-like domain. This family plays crucial roles in plant development, including lateral organ development, pathogen response, pollen development, plant regeneration and hormone response. However, there is no detailed information of LBD genes in the tea plant. Tea plant [Camellia sinensis (L.) O. Kuntze] is a perennial evergreen woody crop that is cultivated worldwide. Tea leaves are used as a non-alcoholic beverage, which is known for its beneficial ingredients and multiple flavours. In this study, a total of 31 CsLBD genes were identified and characterized based on the genome of C. sinensis. CsLBD factors and Arabidopsis counterparts were clustered into 10 subgroups after phylogenetic analysis. Conserved motif analysis revealed that CsLBD proteins with a close evolutionary relationship possessed uniform or similar motif compositions. The interaction networks of LBD proteins in C. sinensis were established according to the data in Arabidopsis. Seven CsLBD genes were examined through qRT-PCR (quantitative real time polymerase chain reaction) in tea plant cultivar, ‘Longjing43’. The expression profiles of these CsLBD genes were investigated in different tissues (i.e. root, stem, leaf and flower) and subjected to five hormonal stimuli (i.e. abscisic acid, gibberellin acid, ethephon, methyl jasmonate and salicylic acid). This study provides an extensive overview of the CsLBD family genes and a valuable foundation for future functional studies of CsLBD factors.

Coiled-coil motif in LBD16 and LBD18 transcription factors are critical for dimerization and biological function in arabidopsis

ABSTRACTThe LATERAL ORGAN BOUNDARIES (LOB) DOMAIN (LBD) gene family members encode a class of plant-specific transcription factors that play important roles in many different aspects of plant growth and development. The LBD proteins contain a conserved LOB domain harboring a Leu zipper-like coiled-coil motif, which has been predicted to mediate protein-protein interactions among the LBD family members. Dimerization of transcription factors is crucial for the modulation of their DNA-binding affinity, specificity, and diversity, contributing to the transcriptional regulation of distinct cellular and biological responses. Our various molecular and biochemical experiments with genetic approaches on LBD16 and LBD18, which are known to control lateral root development in Arabidopsis, demonstrated that the conserved Leu or Val residues in the coiled-coil motifs of these transcription factors are critical for their dimerization as well as the transcriptional regulation to display their biological functions during lateral root formation. We further showed that beside the coiled-coil motif, the carboxyl-terminal region in LBD18 acts as an additional dimerization domain. These findings provide a molecular framework for the homo- and hetero-dimerization of the LBD family proteins for displaying their distinct and diverse biological functions in plants.

Identification and expression analysis of <i>LATERAL ORGAN BOUNDARIES DOMAIN</i> (<i>LBD</i>) transcription factor genes in <i>Fragaria vesca </i>

The LATERAL ORGAN BOUNDARIES DOMAIN (LBD) gene family encodes plant-specific transcription factors that play crucial roles in the growth and development in many plant species. However, no systematic study of LBD genes has been conducted in strawberry. In this study, 35 LBD (FvLBD) genes were identified in the diploid woodland strawberry genome (Fragaria vesca L.). These LBD proteins could be classified into two groups based on the structure of their lateral organ boundaries domain. The promoters of FvLBD genes contain different regulatory elements associated with potential response to different environmental stimuli and plant developmental signals. Furthermore, we analysed the expression patterns of the LBD genes during the callus formation in strawberry and the results suggested that FvLBD16 might play a prominent role in the regulation of callus formation. In addition, we investigated the expression profiles of FvLBD genes during early fruit development based on transcriptome data. We found that some Fv...

OsLBD37 and OsLBD38, two class II type LBD proteins, are involved in the regulation of heading date by controlling the expression of Ehd1 in rice

LATERAL ORGAN BOUNDARIES DOMAIN (LBD) proteins are plant-specific transcription factors which are involved in many biological process of plant development, including lateral organ development, photomorphogenesis, pathogen response, pollen development and plant regeneration. Here, we report new functions of LBD proteins that participate in the regulation of heading date and yield in rice. OsLBD37 and OsLBD38 are two class II type LBD proteins, function as transcriptional activators. Overexpression of OsLBD37 and OsLBD38 separately causes delayed heading date and increased yield. Both OsLBD37 and OsLBD38 are expressed in rhythmic pattern, and their proteins are localized in the nucleus. Further analysis revealed that OsLBD37 and OsLBD38 delayed heading date by down-regulating the expression of the florigen genes Hd3a and RFT1 through key regulator of heading date Ehd1. These data indicates that OsLBD37 and OsLBD38, may function as novel regulators of heading date and crop yield in rice.

Characterization of the LBD gene family in Brachypodium: a phylogenetic and transcriptional study.

An unambiguous nomenclature is proposed for the twenty-eight-member LOB domain transcription factor family in Brachypodium . Expression analysis provides unique transcript patterns that are characteristic of a wide range of organs and plant parts. LOB (lateral organ boundaries)-domain proteins define a family of plant-specific transcription factors involved in developmental processes from embryogenesis to seed production. They play a crucial role in shaping the plant architecture through coordinating cell fate at meristem to organ boundaries. Despite their high potential importance, our knowledge of them is limited, especially in the case of monocots. In this study, we characterized LOB domain protein coding genes (LBDs) of Brachypodium distachyon, a model plant for grasses, and present their phylogenetic relationships and an overall spatial expression study. In the Brachypodium genome database, 28 LBDs were found and then classified based on the presence of highly conserved LOB domain motif. Their transcript amounts were measured via quantitative real-time RT-PCR in 37 different plant parts from root tip to generative organs. Comprehensive phylogenetic analysis suggests that there are neither Brachypodium- nor monocot-specific lineages among LBDs, but there are differences in terms of complexity of subclasses between monocots and dicots. Although LBDs in Brachypodium have wide variation of tissue-specific expression and relative transcript levels, overall expression patterns show similarity to their counterparts in other species. The varying transcript profiles we observed support the hypothesis that Brachypodium LBDs have diverse but conserved functions in plant organogenesis.

Expression and Protein Interaction Analyses Reveal Combinatorial Interactions of LBD Transcription Factors During Arabidopsis Pollen Development.

LATERAL ORGAN BOUNDARIES DOMAIN (LBD) transcription factor gene family members play key roles in diverse aspects of plant development. LBD10 and LBD27 have been shown to be essential for pollen development in Arabidopsis thaliana. From the previous RNA sequencing (RNA-Seq) data set of Arabidopsis pollen, we identified the mRNAs of LBD22, LBD25 and LBD36 in addition to LBD10 and LBD27 in Arabidopsis pollen. Here we conducted expression and cellular analysis using GFP:GUS (green fluorescent protein:β-glucuronidase) reporter gene and subcellular localization assays using LBD:GFP fusion proteins expressed under the control of their own promoters in Arabidopsis. We found that these LBD proteins display spatially and temporally distinct and overlapping expression patterns during pollen development. Bimolecular fluorescence complementation and GST (glutathione S-transferase) pull-down assays demonstrated that protein-protein interactions occur among the LBDs exhibiting overlapping expression during pollen development. We further showed that LBD10, LBD22, LBD25, LBD27 and LBD36 interact with each other to form heterodimers, which are localized to the nucleus in Arabidopsis protoplasts. Taken together, these results suggest that combinatorial interactions among LBD proteins may be important for their function in pollen development in Arabidopsis.

Identification and characterization of Lateral Organ Boundaries Domain genes in mulberry, Morus notabilis

Genes from the plant specific Lateral Organ Boundaries Domain (LBD) family encode transcriptional regulators that have a variety of functions in various physiological and developmental processes. In the present study, 31 LBD genes were identified in the mulberry genome. The genome features of all MnLBD genes and phylogenetic studies with Arabidopsis LBD protein sequences, accompanied by the expression analysis of each of the Morus LBD genes provide insights into the functional prediction of mulberry LBDs. The genome-wide surveys of the current mulberry genome have resulted in the identification of catalogs of MnLBD genes that may function in the development of leaf, root, and secondary metabolism in Morus sp.

LOB Domain Proteins: Beyond Lateral Organ Boundaries.

LATERAL ORGAN BOUNDARIES DOMAIN (LBD) proteins defined by a conserved LATERAL ORGAN BOUNDARIES (LOB) domain are key regulators of plant organ development. Recent studies have expanded their functional diversity beyond the definition of lateral organ boundaries to pollen development, plant regeneration, photomorphogenesis, pathogen response, and specific developmental functions in non-model plants, such as poplar and legumes. The identification of a range of upstream regulators, protein partners, and downstream targets of LBD family members has unraveled the molecular networks of LBD-dependent processes. Moreover, it has been demonstrated that LBD proteins have essential roles in integrating developmental changes in response to phytohormone signaling or environmental cues. As we discuss here, these novel discoveries of LBD functions and their molecular contexts promote a better understanding of this plant-specific transcription factor family.

Transcriptome Analysis of Tomato Flower Pedicel Tissues Reveals Abscission Zone-Specific Modulation of Key Meristem Activity Genes

Tomato flower abscises at the anatomically distinct abscission zone that separates the pedicel into basal and apical portions. During abscission, cell separation occurs only at the abscission zone indicating distinctive molecular regulation in its cells. We conducted a transcriptome analysis of tomato pedicel tissues during ethylene promoted abscission. We found that the abscission zone was the most active site with the largest set of differentially expressed genes when compared with basal and apical portions. Gene Ontology analyses revealed enriched transcription regulation and hydrolase activities in the abscission zone. We also demonstrate coordinated responses of hormone and cell wall related genes. Besides, a number of ESTs representing homologs of key Arabidopsis shoot apical meristem activity genes were found to be preferentially expressed in the abscission zone, including WUSCHEL (WUS), KNAT6, LATERAL ORGAN BOUNDARIES DOMAIN PROTEIN 1(LBD1), and BELL-like homeodomain protein 1 (BLH1), as well as tomato axillary meristem genes BLIND (Bl) and LATERAL SUPPRESSOR (Ls). More interestingly, the homologs of WUS and the potential functional partner OVATE FAMILIY PROTEIN (OFP) were subsequently down regulated during abscission while Bl and AGL12 were continuously and specifically induced in the abscission zone. The expression patterns of meristem activity genes corroborate the idea that cells of the abscission zone confer meristem-like nature and coincide with the course of abscission and post-abscission cell differentiation. Our data therefore propose a possible regulatory scheme in tomato involving meristem genes that may be required not only for the abscission zone development, but also for abscission.

Boundary issues in regulation and evolution of wood formation

Boundary issues in regulation and evolution of wood formation