Lateral Organ Boundaries Domain Family Research Articles

Overexpression Analysis of PtrLBD41 Suggests Its Involvement in Salt Tolerance and Flavonoid Pathway in Populus trichocarpa.

Soil salinization is a significant environmental stress factor, threatening global agricultural yield and ecological security. Plants must effectively cope with the adverse effects of salt stress on survival and successful reproduction. Lateral Organ Boundaries (LOB) Domain (LBD) genes, a gene family encoding plant-specific transcription factors (TFs), play important roles in plant growth and development. Here, we identified and functionally characterized the LBD family TF PtrLBD41 from Populus trichocarpa, which can be induced by various abiotic stresses, including salt, dehydration, low temperature, and Abscisic Acid (ABA). Meanwhile, transgenic plants overexpressing PtrLBD41 showed a better phenotype and higher tolerance than the wild-type (WT) plants under salt stress treatment. Transcriptome analysis found that the differentially expressed genes (DEGs) between the WT and overexpression (OE) line were enriched in the flavonoid biosynthetic process, in which chalcone synthases (CHS), naringenin 3-dioxygenase (F3H), and chalcone isomerase (CHI) were significantly up-regulated under salt stress conditions through qRT-PCR analysis. Therefore, we demonstrate that PtrLBD41 plays an important role in the tolerance to salt stress in P. trichocarpa.

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.

Genome-Wide Analysis of Soybean Lateral Organ Boundaries Domain Gene Family Reveals the Role in Phytophthora Root and Stem Rot

The plant-specific lateral organ boundaries (LOB) domain (LBD) proteins, a family of transcription factors, play important roles in plant growth and development, as well as in responses to various stresses. However, little is known about the functions of LBD genes in soybean (Glycine max). In this study, we investigated the evolution and classification of the LBD family in soybean by a phylogenetic tree of the LBD gene family from 16 species. Phylogenetic analysis categorized these proteins into two classes (Class I and Class II) with seven subgroups. Moreover, we found that all the 18 LBD ancestors in angiosperm were kept in soybean, common bean genomes, and genome-wide duplication, suggesting the main force for the expansion of LBD from common bean to soybean. Analysis of gene expression profiling data indicated that 16 GmLBD genes were significantly induced at different time points after inoculation of soybean plants (cv. Huachun 6) with Phytophthora sojae (P. sojae). We further assessed the role of four highly upregulated genes, GmLBD9, GmLBD16, GmLBD23, and GmLBD88, in plant defense in soybean hairy roots using the transient overexpression and knockdown assays. The results showed that GmLBD9 and GmLBD23 negatively regulate plant immunity against P. sojae, whereas GmLBD16 and GmLBD88 positively manipulate plant immunity against P. sojae. Collectively, our findings expand our knowledge of the origin and evolution of the GmLBD gene family in soybean and promote the potential application of these genes in soybean genetic improvement.

Genome-Wide Identification, Evolution, and Expression Analysis of LBD Transcription Factor Family in Bread Wheat (Triticum aestivum L.).

The lateral organ boundaries domain (LBD) genes, as the plant-specific transcription factor family, play a crucial role in controlling plant architecture and stress tolerance. Although it has been thoroughly characterized in many species, the LBD family was not well studied in wheat. Here, the wheat LBD family was systematically investigated through an in silico genome-wide search method. A total of 90 wheat LBD genes (TaLBDs) were identified, which were classified into class I containing seven subfamilies, and class II containing two subfamilies. Exon–intron structure, conserved protein motif, and cis-regulatory elements analysis showed that the members in the same subfamily shared similar gene structure organizations, supporting the classification. Furthermore, the expression patterns of these TaLBDs in different types of tissues and under diverse stresses were identified through public RNA-seq data analysis, and the regulation networks of TaLBDs involved were predicted. Finally, the expression levels of 12 TaLBDs were validated by quantitative PCR (qPCR) analysis and the homoeologous genes showed differential expression. Additionally, the genetic diversity of TaLBDs in the landrace population showed slightly higher than that of the genetically improved germplasm population while obvious asymmetry at the subgenome level. This study not only provided the potential targets for further functional analysis but also contributed to better understand the roles of LBD genes in regulating development and stress tolerance in wheat and beyond.

A Novel Lateral Organ Boundary-domain Factor CmLBD2 Positively Regulates Pollen Development by Activating CmACOS5 in Chrysanthemum morifolium.

Male sterility, as a common reproductive characteristic in plants, plays an important role in breeding, in which pollen abortion is a key factor leading to male sterility. Here, based on a low expression level gene CmACOS5 in transcriptome of pollen abortive chrysanthemum, a new transcription factor CmLBD2 of the Lateral Organ Boundaries Domain family, which could bind the promoter of CmACOS5 by yeast one-hybrid library was screened. This study revealed the origin and expression pattern of CmLBD2 in chrysanthemum and verified the functions of two genes in pollen development by transgenic means. Inhibiting the expression of CmACOS5 or CmLBD2 can lead to a large reduction in pollen and even abortion in chrysanthemum. Using yeast one-/two-hybrid, electrophoretic mobility shift assays, and luciferase reporter assays, it was verified that CmLBD2 directly binds to the promoter of CmACOS5. These results suggest that LBD2 is a novel, key transcription factor regulating pollen development. This result will provide a new research background for enriching the function of LBD family proteins and also lay a new foundation for the breeding of male sterile lines and the mechanism of pollen development.

Overexpression of a Pak Choi Gene, BcAS2, Causes Leaf Curvature in Arabidopsis thaliana.

The LBD (Lateral Organ Boundaries Domain) family are a new group of plant-specific genes, which encode a class of transcription factors containing conserved Lateral Organization Boundary (LOB) domains, and play an important role in regulating the adaxial–abaxial polarity of plant leaves. In Arabidopsis thaliana, ASYMMETRIC LEAVES 2 (AS2) has a typical LOB domain and is involved in determining the adaxial cell fate. In this study, we isolated the BcAS2 gene from the pak choi cultivar “NHCC001”, and analyzed its expression pattern. The results showed that the BcAS2 encoded a protein made up of 202 amino acid residues which were located in the nucleus and cytomembrane. The Yeast two-hybrid system (Y2H) assay indicated that BcAS2 interacts with BcAS1-1 and BcAS1-2 (the homologous genes of AS1 gene in pak choi). In the transgenic Arabidopsis thaliana that overexpressed BcAS2 gene, it presented an abnormal phenotype with a curly shape. Taken together, our findings not only validate the function of BcAS2 in leaf development in Arabidopsis thaliana, but also contribute in unravelling the molecular regulatory mechanism of BcAS2, which fulfills a special role by forming complexes with BcAS1-1/2 in the establishment of the adaxial–abaxial polarity of the lateral organs in pak choi.

CRISPR/Cas9 targeted mutagenesis of SlLBD40, a lateral organ boundaries domain transcription factor, enhances drought tolerance in tomato

The LATERAL ORGAN BOUNDARIES DOMAIN (LBD)-containing genes are plant-specific genes that play important roles in lateral organ development. In this study, we identified LBD40 (Solyc02g085910), which belongs to subfamily II of the LBD family of genes in tomato. LBD40 was highly expressed in roots and fruit. LBD40 expression was significantly induced by PEG and salt. Moreover, SlLBD40 expression was induced by methyl jasmonate treatment, while SlLBD40 expression could not be induced in the jasmonic acid-insensitive1 (jai1) mutant or MYC2-silenced plants, in which jasmonic acid (JA) signaling was disrupted. These findings demonstrate that SlLBD40 expression was dependent on JA signaling and that it might be downstream of SlMYC2, which is the master transcription factor in the JA signal transduction pathway. Overexpressing and CRISPR/Cas9 mediated knockout transgenic tomato plants were generated to explore SlLBD40 function. The drought tolerance test showed that two SlLBD40 knockout lines wilted slightly, while SlLBD40 overexpressing plants suffered severe wilting. The statistical water loss rate and midday leaf water potential also confirmed that knockout of SlLBD40 improved the water-holding ability of tomato under drought conditions. Taken together, our study demonstrates that SlLBD40, involved in JA signaling, was a negative regulator of drought tolerance and that knockout of SlLBD40 enhanced drought tolerance in tomato. This study also provides a novel function of SlLBD40, which belongs to subfamily II of LBD genes.

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.

Genome-wide characterisation and expression profiling of the LBD family in Salvia miltiorrhiza reveals the function of LBD50 in jasmonate signaling and phenolic biosynthesis

Jasmonates (JAs) regulate growth, development, and secondary metabolism in many economically significant crops. The lateral organ boundaries domain (LBD) protein family is a class of plant-specific transcription factors involved in the regulation of lateral organ development and jasmonate signaling. However, the functions of LBD genes in Salvia miltiorrhiza Bunge, an emerging model medicinal plant, are unclear. In this study, a total of 51 LBD transcription factor genes were identified in the genome of S. miltiorrhiza, and were classified by phylogenetic analysis into seven subfamilies. The expression profile revealed that 10 of the LBD genes are tissue-specific and JA-inducible. Of the 51 LBD transcription factor genes, 11 % (six out of 51) were potentially involved in the biosynthesis of tanshinones or phenolic acids. Correlation analysis showed that the expression of SmLBD9, SmLBD13, SmLBD21, and SmLBD50 were observably regulated in JA zim domain (JAZ) proteins transgenic S. miltiorrhiza, indicating their critical roles in the JA signaling pathway. Further functional analysis indicated that the overexpression of SmLBD50 inhibited the synthesis of total phenolics, total flavonoids, and anthocyanins in S. miltiorrhiza. Consistent with these phytochemical changes, the expression of phenolic biosynthetic genes was also stimulated. Yeast two-hybrid (Y2H) assays indicated that the SmLBD50 protein interacts with SmJAZ1, SmMYB36/97, SmbHLH37 and SmMYC2a/2b, four important transcription factors involved in JA signaling. In conclusion, our study off ;ers the first genome-wide overview and annotation of the LBD transcription factor family in S. miltiorrhiza and provides a foundation for understanding the molecular basis and regulatory mechanisms of SmLBD50 in relation to the JA response and phenolic metabolism in S. miltiorrhiza.

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.

Genome-Wide Analysis of the Lateral Organ Boundaries Domain (LBD) Gene Family in Solanum tuberosum.

Lateral organ boundaries domain (LBD) proteins belong to a particular class of transcription factors of lateral organ boundary (LOB) specific domains that play essential roles in plant growth and development. However, a potato phylogenetic analysis of the LBD family has not been fully studied by scholars and researchers. In this research, bioinformatics methods and the growth of potatoes were used to identify 43 StLBD proteins. We separated them into seven subfamilies: Ia, Ib, Ic, Id, Ie, IIa and IIb. The number of amino acids encoded by the potato LBD family ranged from 94 to 327. The theoretical isoelectric point distribution ranged from 4.16 to 9.12 Kda, and they were distributed among 10 chromosomes. The results of qRT-PCR showed that the expression levels of StLBD2-6 and StLBD3-5 were up-regulated under drought stress in the stem. The expression levels of StLBD1-5 and StLBD2-6 were down-regulated in leaves. We hypothesized that StLBD1-5 was down-regulated under drought stress, and that StLBD2-6 and StLBD3-5 up-regulation might help to maintain the normal metabolism of potato and enhance the potatoes’ resistance to drought.

Lateral Organ Boundaries Domain 19 (LBD19) negative regulate callus formation in Arabidopsis

Callus is a remarkable regeneration tissue. The genes correlated with root development can be involved in regulating callus development in higher plants. Several Lateral Organ Boundaries Domain (LBD) transcription factors are known as the key regulators in callus induction, and they also participate in the development of lateral root. Here we took a systemic analysis approach to investigate the expression pattern of many LBD family members during callus induction on CIM medium. We found that most of the LBD members were upregulated on CIM medium but LBD19 was different from other family members showing increased expression at early stage but decreased expression later on. We were unable to obtain the loss of function mutant of lbd19 plant, so we generated the overexpression transgenic lines of LBD19. The overexpression of LBD19 caused obvious phenotypic changes in vegetative development and suppressed callus formation. The quantitative RT-PCR analysis results revealed that the expression of most genes that promote callus formation were down-regulated in callus of oeLBD19. These findings suggest that LBD19 may plays an important function in coordinating callus formation in Arabidopsis along with other LBD members.

Genome-wide Analysis of LBD (LATERAL ORGAN BOUNDARIES Domain) Gene Family in Brassica rapa

ABSTRACT LOB (lateral organ boundaries)-domain proteins define a family of plant-specific transcription factors involved in developmental process from embryogenesis to seed production. They play a crucial role in shaping the plant architecture through coordinating cell fate at meristem to organ boundaries. Identification of LBD genes from Brassica rapa genome, and analysis of phylogeny，gene structure, chromosome location, phylogenetic and tissue expression pattern analysis of LBD family genes in Chinese cabbage will be useful to the functions identification of plant LBD genes. Based on Brassica rapa genome database and bioinformatic method, Chinese cabbage LBD family genes were identified and the genes were sequenced. A phylogenetic tree was created using the MEGA5 program. Gene structure and chromosomes location were done by MapDraw, GSDS and Clustal X. Expression pattern of LBD genes at different development stages was analyzed based on RNA-seq. A total of 62 LBD genes were identified and could be classified into two classes and four subclasses according to the gene structure and conserved domain phylogeny relationship. Distribution mapping showed that the predicted LBDs were unevenly localized on all the 10 chromosomes, suggesting that they have an extensive distribution on the Brassica rapa chromosomes. Most of the LBD genes had differential expression pattern and showed highly diverse tissue-specific expression and functional diversity. To our knowledge, this is the first report of a genome wide analysis of the Brassica rapa LBD gene family, which would provide valuable information for understanding the classification and putative functions of the gene family.

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.

Cloning and elucidation of the functional role of apple MdLBD13 in anthocyanin biosynthesis and nitrate assimilation

The plant specific lateral organ boundaries domain (LBD) family of transcription factors (TFs) is involved in many aspects of plant growth and development. In this study, MdLBD13, a nitrate-induced LBD family gene from the apple (Malus × domestica), was isolated and characterized. Overexpression of MdLBD13 repressed anthocyanin biosynthesis by reducing expression of the structural genes associated with the flavonoid pathway. Overexpression ofMdLBD13 also repressed expression of the N–responsive genes that are required for nitrate uptake, transport and assimilation, resulting in reduced nitrate content and nitrate reductase activity in apple calli, as well as in Arabidopsis. Ectopic expression of MdLBD13 also promoted lateral root development in transgenic Arabidopsis. These results demonstrate that MdLBD13acts as a negative regulator in anthocyanin biosynthesis and nitrate utilization.

Identification and expression of Aux/IAA, ARF, and LBD family transcription factors in Boehmeria nivea

Auxin controls numerous processes in plant development and auxin/indoleacetic acid (Aux/IAA), an auxin response factor (ARF), and a lateral organ boundaries domain (LBD) were considered as early auxin response transcription factors (TFs). Till now, no Aux/IAA, ARF, and LBD TFs were identified in ramie (Boehmeria nivea L. Gaud). In this study, we used Arabidopsis and mulberry sequences as query to search against the ramie transcriptome database and the searched sequences were analyzed for a full-length coding sequence. In total, we obtained 16 BnAux/IAA, 14 BnARF, and 16 BnLBD TFs on which evolutionary analysis and expression profiling were conducted. Analysis of sequence conservation revealed close evolution relationships between ramie and mulberry. Expression analysis shows these genes were actively expressed in major ramie tissues, and several were auxin responsive. The expressions of these genes were also investigated under drought and a high temperature, main abiotic stresses during ramie life cycle. We found that most genes of the three families were stress-responsive and showed distinct expression patterns under the drought and high temperature stresses.

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.

Boundary issues in regulation and evolution of wood formation

Boundary issues in regulation and evolution of wood formation