Homeodomain-leucine Zipper Class Research Articles

A Catalog of GNI-A1 Genes That Regulate Floret Fertility in a Diverse Bread Wheat Collection

Modifying inflorescence architecture improves grain number and grain weight in bread wheat (Triticum aestivum). Allelic variation in Grain Number Increase 1 (GNI-A1) genes, encoding a homeodomain leucine zipper class I transcription factor, influences grain number and yield. However, allelic information about GNI-A1 in diverse germplasms remains limited. Here, we investigated GNI-A1 alleles in a panel of 252 diverse bread wheat accessions (NBRP core collection and HRO breeder’s panel) by target resequencing. Cultivars carrying the reduced-function allele (105Y) were predominant in the NBRP panel, whereas the 105N functional allele was the major type in the HRO panel. Cultivars with the 105Y allele were distributed in Asian landraces but not in European genotypes. Association analysis demonstrated that floret fertility, together with grain size, were improved in cultivars in the NBRP core collection carrying the 105Y allele. These results imply that different alleles of GNI-A1 have been locally selected, with the 105Y allele selected in East Asia and the 105N allele selected in Europe.

Pepper homeobox abscisic acid signalling-related transcription factor 1, CaHAT1, plays a positive role in drought response.

Abscisic acid (ABA) signalling triggers drought resistance mediated by SNF1-related kinase 2s (SnRK2s), which transmits stress signals through the phosphorylation of several downstream factors. However, these kinases and their downstream targets remain elusive in pepper plants. This study aimed to isolate interacting partners of CaSnRK2.6, a homologue of Arabidopsis SnRK2.6/OST1. Among the candidate proteins, we identified a homeodomain-leucine zipper (HD-Zip) class II protein and named it CaHAT1 (Capsicum annuum homeobox ABA signalling related- transcription factor 1). CaHAT1-silenced pepper and -overexpression (OE) transgenic Arabidopsis plants were generated to investigate the in vivo function of CaHAT1 in drought response. Following the application of drought stress, CaHAT1-silenced pepper plants exhibited drought-sensitive phenotypes with reduced ABA-mediated stomatal closure and lower expression of stress-responsive genes compared with control plants. In contrast, CaHAT1-OE transgenic Arabidopsis plants showed the opposite phenotypes, including increased drought resistance and ABA sensitivity. CaHAT1, particularly its N-terminal consensus sequences, was directly phosphorylated by CaSnRK2.6. Furthermore, CaSnRK2.6 kinase activity and CaSnRK2.6-mediated CaHAT1 phosphorylation levels were enhanced by treatment with ABA and drought stress. Taken together, our results indicated that CaHAT1, which is the target protein of CaSnRK2.6, is a positive regulator of drought stress response. This study advances our understanding of CaHAT1-CaSnRK2.6 mediated defence mechanisms in pepper plants against drought stress.

Differential gene expression analysis of the resprouting process in Pinus canariensis provides new insights into a rare trait in conifers

Resprouting is crucial in population dynamics in response to wildfires or herbivory. This trait, common in angiosperms, is considered rare in conifers, being Pinus canariensis one of the few species able to resprout. We analyzed gene expression during wound-induced resprouting in 5 years-old Canarian pines. RNA was extracted at three different stages, including control samples from remote branches, representing immediate response to wounding (R0), resprouting initiation (R1), and elongation (R2), and then hybridized to a microarray designed with 15,000 cDNAs from Pinus canariensis transcriptome of meristematic activity. We found 1164 Differentially Expressed Genes (DEGs), of which 903 were significant in R0, 278 in R1, and 261 in R2. Genes related to defense- and stress-response were mainly found to be overexpressed at R0, while DEGs coding for transcription factors such as GROWTH-REGULATING FACTOR (GRF), FLOWERING-PROMOTING FACTOR (FPF), and the HOMEODOMAIN LEUCINE ZIPPER Class IV (HD-ZIP IV), mainly related to outgrowth processes and lateral organogenesis in plants, showed overexpression at R1 and R2, when new shoots were emerging. This data was compared to differential gene expression during apical growth in P. canariensis, suggesting similarities between vegetative apical growth and regulation of lateral shoot development in response to mechanical wounding, but also yielding 32 DEGs exclusively overexpressed at some point during resprouting stages (R1 and/or R2). The implication of such transcription factor families, with important roles in leaf and inflorescence development also in flowering plants, suggests underlying homologies between common lateral organogenesis processes in angiosperms and resprouting in gymnosperms.

The Arabidopsis NLP7-HB52/54-VAR2 pathway modulates energy utilization in diverse light and nitrogen conditions

In plants, nitrate is the dominant nitrogen (N)source and a critical nutrient signal regulating various physiological and developmental processes.1,2,3,4 Nitrate-responsive gene regulatory networks are widely believed to control growth, development, and life cycle in addition to N acquisition and utilization,1,2,3,4 and NIN-LIKE PROTEIN (NLP) transcriptional activators have been identified as the master regulators governing the networks.5,6,7 However, it remains to be elucidated how nitrate signaling regulates respective physiological and developmental processes. Here, we have identified a new nitrate-activated transcriptional cascade involved in chloroplast development and the maintenance of chloroplast function in Arabidopsis. This cascade consisting of NLP7 and two homeodomain-leucine zipper (HD-Zip) class I transcription factors, HOMEOBOX PROTEIN52 (HB52) and HB54,8,9 was responsible for nitrate- and light-dependent expression of VAR2 encoding the FtsH2 subunit of the chloroplast FtsH protease involved in the quality control of photodamaged thylakoid membrane proteins.10,11 Consistently, the nitrate-activated NLP7-HB52/54-VAR2 pathway underpinned photosynthetic light energy utilization, especially in high light environments. Furthermore, genetically enhancing the NLP7-HB52/54-VAR2 pathway resulted in improved light energy utilization under high light and low N conditions, a superior agronomic trait. These findings shed light on a new role of nitrate signaling and a novel mechanism for integrating information on N nutrient and light environments, providing a hint for enhancing the light energy utilization of plants in low N environments.

Genome‐wide characterization of homeodomain‐leucine zipper genes reveals RsHDZ17 enhances the heat tolerance in radish (Raphanus sativus L.)

Homeodomain-leucine zipper (HD-Zip) transcription factors are involved in various biological processes of plant growth, development, and abiotic stress response. However, how they regulate heat stress (HS) response remains largely unclear in plants. In this study, a total of 83 RsHD-Zip genes were firstly identified from the genome of Raphanus sativus. RNA-Seq, RT-qPCR and promoter activity assays revealed that RsHDZ17 from HD-Zip Class I was highly expressed under heat, salt, and Cd stresses. RsHDZ17 is a nuclear protein with transcriptional activity at the C-terminus. Ectopic overexpression (OE) of RsHDZ17 in Arabidopsis thaliana enhanced the HS tolerance by improving the survival rate, photosynthesis capacity, and scavenging for reactive oxygen species (ROS). In addition, transient OE of RsHDZ17 in radish cotyledons impeded cell injury and augmented ROS scavenging under HS. Moreover, yeast one-hybrid, dual-luciferase assay, and electrophoretic mobility shift assay revealed that RsHDZ17 could bind to the promoter of HSFA1e. Collectively, these pieces of evidence demonstrate that RsHDZ17 could play a positive role in thermotolerance, partially through up-regulation of the expression of HSFA1e in plants. These results provide novel insights into the role of HD-Zips in radish and facilitate genetical engineering and development of heat-tolerant radish in breeding programs.

The Arabidopsis HDZIP class II transcription factor ABA INSENSITIVE TO GROWTH 1 functions in leaf development.

Leaf laminar growth and adaxial-abaxial boundary formation are fundamental outcomes of plant development. Boundary and laminar growth coordinate the further patterning and growth of the leaf, directing the differentiation of cell types within the top and bottom domains and promoting initiation of lateral organs along their adaxial or abaxial axis. Leaf adaxial-abaxial polarity specification and laminar outgrowth are regulated by two transcription factors, REVOLUTA (REV) and KANADI (KAN). ABA INSENSITIVE TO GROWTH 1 (ABIG1) encodes a HOMEODOMAIN-LEUCINE ZIPPER (HD-ZIP) class II transcription factor and is a direct target of the adaxial-abaxial regulators REV and KAN. To investigate the role of ABIG1 in leaf development and in the establishment of polarity, we examined the phenotypes of both gain-of-function and loss-of-function mutants. Through genetic interaction analysis with REV and KAN mutants, we determined that ABIG1 plays a role in leaf laminar growth as well as in adaxial-abaxial polarity establishment. Genetic and physical interaction assays showed that ABIG1 interacts with the transcriptional TOPLESS corepressor. This study provides new evidence that ABIG1, another HD-ZIP II, facilitates growth through the corepressor TOPLESS.

The lipid-binding START domain regulates the dimerization of ATML1 via modulating the ZIP motif activity in Arabidopsis thaliana.

In Arabidopsis thaliana, the epidermis is the outermost cell layer composed of many specialized types of epidermal cells, such as pavement cells, trichomes, and guard cells. The homeodomain-leucine zipper (HD-ZIP) class Ⅳ transcription factors (TFs), which are unique to the plant kingdom, have been recognized as key regulators of epidermis development. Unlike animal HD proteins, which can bind to DNA as monomers, plant HD-ZIP class Ⅳ TFs bind to DNA as dimers, although little is known about the regulation of their dimerization process. Here, we show that the homodimerization of ARABIDOPSIS THALIANA MERISTEM LAYER 1 (ATML1) - HD-ZIP class Ⅳ TF that is required for protoderm development - is regulated by the lipid-binding steroidogenic acute regulatory protein-related lipid transfer (START) domain. We found that ATML1 forms homodimer through interaction via its ZIP motif in yeast and plant cells, although the interaction is abolished by generating a mutation into the lipid-binding START domain to disrupt the lipid-binding ability. These results suggest that lipidic ligands function as key regulators of protoderm development via modulating the dimerization of ATML1.

LF1 regulates the lateral organs polarity development in rice.

The patterning of adaxial-abaxial tissues plays a vital role in the morphology of lateral organs, which is maintained by antagonism between the genes that specify adaxial and abaxial tissue identity. The homeo-domain leucine zipper class III (HD-ZIP III) family genes regulate adaxial identity; however, little information is known about the physical interactions or transcriptionally regulated downstream genes of HD-ZIP III. In this study, we identified a dominant rice mutant, lateral floret 1 (lf1), which has defects in lateral organ polarity. LF1 encodes the HD-ZIP III transcription factor, which expressed in the adaxial area of lateral organs. LF1 can activate directly the expression of LITTLE ZIPPER family gene OsZPR4 and HD-ZIP II family gene OsHOX1, and OsZPR4 and OsHOX1 respectively interact with LF1 to form a heterodimer to repress the transcriptional activity of LF1. LF1 influences indole-3-acetic acid (IAA) content by directly regulating the expression of OsYUCCA6. Therefore, LF1 forms negative feedback loops between OsZPR4 and OsHOX1 to affect IAA content, leading to the regulation of lateral organs polarity development. These results reveal the cross-talk among HD-ZIP III, LITTLE ZIPPER, and HD-ZIP II proteins and provide new insights into the molecular mechanisms underlying the polarity development of lateral organs.

Water use efficiency and nutrient uptake of rice under soil water stress condition

One obstacle that can limit the growth and production of rice is low water availability. Therefore, a comprehensive study of the response of rice plants to drought is very important. The study was conducted in the greenhouse of the Faculty of Animal Husbandry and Agriculture, Diponegoro University, from April to August 2016. The study used a 3 x 3 factorial design with 3 replications. The first factor is three types of rice (Sidenuk, Way Apo Buru, Pepe) and the second factor is water stress treatment (KA <field capacity = enough water, field capacity (KL), water saturation (JA). The observed parameters are protein content, leaf levels rolling (number of leaves rolled), relative water content (RWC), water use efficiency (WUE), N, P, K uptake, rice protein content decreases with increasing water supply. The relative water content of rice plants decreases with increasing stress water that occurs in the limited water supply for the three types of rice Sidenuk rice has the lowest N seed content (2.1%) compared to Way Apo (3.0%) and Pepe (3.1%). The mineral content of P in both biomass and seeds is not really influenced by water stress, as well as K content of seeds. REFERENCES Badan Pusat Statistik. 2018. https://www.bps.go.id/dynamictable/2019/04/15/1608/luas-panen-produksi-dan-produktivitas-padi-menurut-provinsi-2018.html Darwesh, R, S,S. 2013. Improving growth of date palm plantlets grown undersalt stress with yeast and amino acids applications Annals of Agricultural Science (2013) 58(2), 247–256 Etienne P.I.D, S. Diquelou , M. Prudent, C. Salon., A. Maillard, and A.Ourry. 2018. Macro and Micronutrient Storage in Plants and Their Remobilization When Facing Scarcity: The Case of Drought. Agriculture 2018, 8, 14; 2-17. doi:10.3390/agriculture8010014 Farooq, M., S.M.A.Basra, A. 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Genome-wide characterization of NtHD-ZIP IV: different roles in abiotic stress response and glandular Trichome induction

BackgroundThe plant-specific homeodomain-leucine zipper class IV (HD-ZIP IV) gene family has been involved in the regulation of epidermal development.ResultsFifteen genes coding for HD-ZIP IV proteins were identified (NtHD-ZIP-IV-1 to NtHD-ZIP-IV-15) based on the genome of N. tabacum. Four major domains (HD, ZIP, SAD and START) were present in these proteins. Tissue expression pattern analysis indicated that NtHD-ZIP-IV-1, − 2, − 3, − 10, and − 12 may be associated with trichome development; NtHD-ZIP-IV-8 was expressed only in cotyledons; NtHD-ZIP-IV-9 only in the leaf and stem epidermis; NtHD-ZIP-IV-11 only in leaves; and NtHD-ZIP-IV-15 only in the root and stem epidermis. We found that jasmonates may induce the generation of glandular trichomes, and that NtHD-ZIP-IV-1, − 2, − 5, and − 7 were response to MeJA treatment. Dynamic expression under abiotic stress and after application of phytohormones indicated that most NtHD-ZIP IV genes were induced by heat, cold, salt and drought. Furthermore, most of these genes were induced by gibberellic acid, 6-benzylaminopurine, and salicylic acid, but were inhibited by abscisic acid. NtHD-ZIP IV genes were sensitive to heat, but insensitive to osmotic stress.ConclusionNtHD-ZIP IV genes are implicated in a complex regulatory gene network controlling epidermal development and abiotic stress responses. The present study provides evidence to elucidate the gene functions of NtHD-ZIP IVs during epidermal development and stress response.

Overexpressing the HD-Zip class II transcription factor EcHB1 from Eucalyptus camaldulensis increased the leaf photosynthesis and drought tolerance of Eucalyptus

Alteration in the leaf mesophyll anatomy by genetic modification is potentially a promising tool for improving the physiological functions of trees by improving leaf photosynthesis. Homeodomain leucine zipper (HD-Zip) transcription factors are candidates for anatomical alterations of leaves through modification of cell multiplication, differentiation, and expansion. Full-length cDNA encoding a Eucalyptus camaldulensis HD-Zip class II transcription factor (EcHB1) was over-expressed in vivo in the hybrid Eucalyptus GUT5 generated from Eucalyptus grandis and Eucalyptus urophylla. Overexpression of EcHB1 induced significant modification in the mesophyll anatomy of Eucalyptus with enhancements in the number of cells and chloroplasts on a leaf-area basis. The leaf-area-based photosynthesis of Eucalyptus was improved in the EcHB1-overexpression lines, which was due to both enhanced CO2 diffusion into chloroplasts and increased photosynthetic biochemical functions through increased number of chloroplasts per unit leaf area. Additionally, overexpression of EcHB1 suppressed defoliation and thus improved the growth of Eucalyptus trees under drought stress, which was a result of reduced water loss from trees due to the reduction in leaf area with no changes in stomatal morphology. These results gave us new insights into the role of the HD-Zip II gene.

Unleashing floret fertility in wheat through the mutation of a homeobox gene

Floret fertility is a key determinant of the number of grains per inflorescence in cereals. During the evolution of wheat (Triticum sp.), floret fertility has increased, such that current bread wheat (Triticum aestivum) cultivars set three to five grains per spikelet. However, little is known regarding the genetic basis of floret fertility. The locus Grain Number Increase 1 (GNI1) is shown here to be an important contributor to floret fertility. GNI1 evolved in the Triticeae through gene duplication. The gene, which encodes a homeodomain leucine zipper class I (HD-Zip I) transcription factor, was expressed most abundantly in the most apical floret primordia and in parts of the rachilla, suggesting that it acts to inhibit rachilla growth and development. The level of GNI1 expression has decreased over the course of wheat evolution under domestication, leading to the production of spikes bearing more fertile florets and setting more grains per spikelet. Genetic analysis has revealed that the reduced-function allele GNI-A1 contributes to the increased number of fertile florets per spikelet. The RNAi-based knockdown of GNI1 led to an increase in the number of both fertile florets and grains in hexaploid wheat. Mutants carrying an impaired GNI-A1 allele out-yielded WT allele carriers under field conditions. The data show that gene duplication generated evolutionary novelty affecting floret fertility while mutations favoring increased grain production have been under selection during wheat evolution under domestication.

Stochasticity in transcriptional expression of a negative regulator of Arabidopsis ABA network.

Stably heritable spatiotemporal co/over-expression of distinct transcriptional regulators across generations is a desired target as they signal traffic in the cell. Here, the stability and expression pattern of AtHB7 (Arabidopsis homeodomain-leucine zipper class I) cDNA was characterized in 220 random population of transformed tomato clones where no AtHB7 orthologous has been identified in to date. Integration of p35S::AtHB7 casette was tested by the amplification of the stretches (700/425bp) in the target by NPT II/AtHB7 oligos. Transcriptional expression pattern for the amplicons of the specific transcripts in the leaf tissues of transformants were determined by qRT-PCR. Transgene copy number was negatively correlated with transgene expression level, yet a majority of transformants (78%) carried single-copy of transgene. About 1:3 of the lines containing two-copy inserts showed less transcript expression. Heterologous CaMV 35S promoter drove AtHB7, illuminated no penalty on transgene expression levels, stability or plant phenotype under drought stress. Integration and expression analysis of transcription factors is of great significance for reliable prediction of gene dosing/functions in plant genomes so as to sustain breeding under abiotic stress to guarantee food security.

GhHB12, a HD-ZIP I Transcription Factor, Negatively Regulates the Cotton Resistance to Verticillium dahliae.

The homeodomain-leucine zipper (HD-ZIP) is a plant-specific transcription factor family that plays important roles in plant developmental processes in response to multiple stressors. We previously isolated a cotton HD-ZIP class I transcription factor gene, GhHB12, which is regulated by the circadian clock and photoperiodism. Furthermore, it regulates cotton architecture, phase transition, and photoperiod sensitivity. Here we report that GhHB12 was induced by methyl jasmonate (MeJA) and Verticillium dahliae infection. Additionally, stress-responsive elements were found in the GhHB12 promoter. Promoter fusion analysis showed that GhHB12 was predominantly expressed in primary roots and that it was induced by mechanical damage. Overexpression of GhHB12 increased susceptibility of the cotton plant to the fungal pathogens Botrytis cinerea and V. dahliae, which was coupled with suppression of the jasmonic acid (JA)-response genes GhJAZ2 and GhPR3. Our results suggest that GhHB12, a cotton stress-responsive HD-ZIP I transcription factor, negatively regulates cotton resistance to V. dahliae by suppressing JA-response genes.

Evidence for the Extensive Conservation of Mechanisms of Ovule Integument Development Since the Most Recent Common Ancestor of Living Angiosperms.

The ovules and seeds of most angiosperm groups are enclosed by two integuments, whose evolutionary origins are considerably separated in time, as the inner integument arose over 300 million years ago (MYA) in an ancestor of all living seed plants, while the outer integument arose, perhaps as recently as 164 MYA, in an ancestor of all living angiosperms. Studies of the model angiosperm Arabidopsis thaliana indicate that the mechanisms of development of the inner and outer integuments depend on largely different sets of molecular players. However, it was not known, in most cases, whether these differences were already present in early flowering plants, or arose later in the Arabidopsis lineage. Here, we analyze the expression patterns of integument regulators in Amborella trichopoda, the likely sister to all other living angiosperms. The data obtained indicate that regulators of the YABBY, KANADI, and homeodomain-leucine zipper class III transcription factor families have largely conserved their integument-specific expression profiles in the Amborella and Arabidopsis lineages since the most recent common ancestor (MRCA) of living angiosperms. We identified only one case, involving the paralogous genes ETTIN and AUXIN RESPONSE FACTOR4, in which integument-specific expression patterns had clearly diverged between Amborella and Arabidopsis. We use the data obtained to partially reconstruct molecular mechanisms of integument development in the MRCA of living angiosperms and discuss our findings in the context of alternative hypotheses for the origin of the angiosperm outer integument.

Overexpression of the class I homeodomain transcription factor TaHDZipI-5 increases drought and frost tolerance in transgenic wheat.

SummaryCharacterization of the function of stress‐related genes helps to understand the mechanisms of plant responses to environmental conditions. The findings of this work defined the role of the wheat TaHDZipI‐5 gene, encoding a stress‐responsive homeodomain–leucine zipper class I (HD‐Zip I) transcription factor, during the development of plant tolerance to frost and drought. Strong induction of TaHDZipI‐5 expression by low temperatures, and the elevated TaHDZipI‐5 levels of expression in flowers and early developing grains in the absence of stress, suggests that TaHDZipI‐5 is involved in the regulation of frost tolerance at flowering. The TaHDZipI‐5 protein behaved as an activator in a yeast transactivation assay, and the TaHDZipI‐5 activation domain was localized to its C‐terminus. The TaHDZipI‐5 protein homo‐ and hetero‐dimerizes with related TaHDZipI‐3, and differences between DNA interactions in both dimers were specified at 3D molecular levels. The constitutive overexpression of TaHDZipI‐5 in bread wheat significantly enhanced frost and drought tolerance of transgenic wheat lines with the appearance of undesired phenotypic features, which included a reduced plant size and biomass, delayed flowering and a grain yield decrease. An attempt to improve the phenotype of transgenic wheat by the application of stress‐inducible promoters with contrasting properties did not lead to the elimination of undesired phenotype, apparently due to strict spatial requirements for TaHDZipI‐5 overexpression.

Extreme Suppression of Lateral Floret Development by a Single Amino Acid Change in the VRS1 Transcription Factor.

Increasing grain yield is an endless challenge for cereal crop breeding. In barley (Hordeum vulgare), grain number is controlled mainly by Six-rowed spike 1 (Vrs1), which encodes a homeodomain leucine zipper class I transcription factor. However, little is known about the genetic basis of grain size. Here, we show that extreme suppression of lateral florets contributes to enlarged grains in deficiens barley. Through a combination of fine-mapping and resequencing of deficiens mutants, we have identified that a single amino acid substitution at a putative phosphorylation site in VRS1 is responsible for the deficiens phenotype. deficiens mutant alleles confer an increase in grain size, a reduction in plant height, and a significant increase in thousand grain weight in contemporary cultivated germplasm. Haplotype analysis revealed that barley carrying the deficiens allele (Vrs1.t1) originated from two-rowed types carrying the Vrs1.b2 allele, predominantly found in germplasm from northern Africa. In situ hybridization of histone H4, a marker for cell cycle or proliferation, showed weaker expression in the lateral spikelets compared with central spikelets in deficiens Transcriptome analysis revealed that a number of histone superfamily genes were up-regulated in the deficiens mutant, suggesting that enhanced cell proliferation in the central spikelet may contribute to larger grains. Our data suggest that grain yield can be improved by suppressing the development of specific organs that are not positively involved in sink/source relationships.

CaHDZ27, a Homeodomain-Leucine Zipper I Protein, Positively Regulates the Resistance to Ralstonia solanacearum Infection in Pepper.

Homeodomain-leucine zipper class I (HD-Zip I) transcription factors have been functionally characterized in plant responses to abiotic stresses, but their roles in plant immunity are poorly understood. Here, a HD-Zip I gene, CaHZ27, was isolated from pepper (Capsicum annum) and characterized for its role in pepper immunity. Quantitative real-time polymerase chain reaction showed that CaHDZ27 was transcriptionally induced by Ralstonia solanacearum inoculation and exogenous application of methyl jasmonate, salicylic acid, or ethephon. The CaHDZ27-green fluorescent protein fused protein was targeted exclusively to the nucleus. Chromatin immunoprecipitation demonstrated that CaHDZ27 bound to the 9-bp pseudopalindromic element (CAATAATTG) and triggered β-glucuronidase expression in a CAATAATTG-dependent manner. Virus-induced gene silencing of CaHDZ27 significantly attenuated the resistance of pepper plants against R. solanacearum and downregulated defense-related marker genes, including CaHIR1, CaACO1, CaPR1, CaPR4, CaPO2, and CaBPR1. By contrast, transient overexpression of CaHDZ27 triggered strong cell death mediated by the hypersensitive response and upregulated the tested immunity-associated marker genes. Ectopic CaHDZ27 expression in tobacco enhances its resistance against R. solanacearum. These results collectively suggest that CaHDZ27 functions as a positive regulator in pepper resistance against R. solanacearum. Bimolecular fluorescence complementation and coimmunoprecipitation assays indicate that CaHDZ27 monomers bind with each other, and this binding is enhanced significantly by R. solanacearum inoculation. We speculate that homodimerization of CaHZ27 might play a role in pepper response to R. solanacearum, further direct evidence is required to confirm it.

The homeodomain transcription factor TaHDZipI-2 from wheat regulates frost tolerance, flowering time and spike development in transgenic barley.

Homeodomain leucine zipper class I (HD-Zip I) transcription factors (TFs) play key roles in the regulation of plant growth and development under stresses. Functions of the TaHDZipI-2 gene isolated from the endosperm of developing wheat grain were revealed. Molecular characterization of TaHDZipI-2 protein included studies of its dimerisation, protein-DNA interactions and gene activation properties using pull-down assays, in-yeast methods and transient expression assays in wheat cells. The analysis of TaHDZipI-2 gene functions was performed using transgenic barley plants. It included comparison of developmental phenotypes, yield components, grain quality, frost tolerance and the levels of expression of potential target genes in transgenic and control plants. Transgenic TaHDZipI-2 lines showed characteristic phenotypic features that included reduced growth rates, reduced biomass, early flowering, light-coloured leaves and narrowly elongated spikes. Transgenic lines produced 25-40% more seeds per spike than control plants, but with 50-60% smaller grain size. In vivo lipid imaging exposed changes in the distribution of lipids between the embryo and endosperm in transgenic seeds. Transgenic lines were significantly more tolerant to frost than control plants. Our data suggest the role of TaHDZipI-2 in controlling several key processes underlying frost tolerance, transition to flowering and spike development.

Molecular interactions of the γ-clade homeodomain-leucine zipper class I transcription factors during the wheat response to water deficit

The γ-clade of class I homeodomain-leucine zipper (HD-Zip I) transcription factors (TFs) constitute members which play a role in adapting plant growth to conditions of water deficit. Given the importance of wheat (Triticum aestivum L.) as a global food crop and the impact of water deficit upon grain yield, we focused on functional aspects of wheat drought responsive HD-Zip I TFs. While the wheat γ-clade HD-Zip I TFs share significant sequence similarities with homologous genes from other plants, the clade-specific features in transcriptional response to abiotic stress were detected. We demonstrate that wheat TaHDZipI-3, TaHDZipI-4, and TaHDZipI-5 genes respond differentially to a variety of abiotic stresses, and that proteins encoded by these genes exhibit pronounced differences in oligomerisation, strength of DNA binding, and trans-activation of an artificial promoter. Three-dimensional molecular modelling of the protein-DNA interface was conducted to address the ambiguity at the central nucleotide in the pseudo-palindromic cis-element CAATNATTG that is recognised by all three HD-Zip I proteins. The co-expression of these genes in the same plant tissues together with the ability of HD-Zip I TFs of the γ-clade to hetero-dimerise suggests a role in the regulatory mechanisms of HD-Zip I dependent transcription. Our findings highlight the complexity of TF networks involved in plant responses to water deficit. A better understanding of the molecular complexity at the protein level during crop responses to drought will enable adoption of efficient strategies for production of cereal plants with enhanced drought tolerance.