Cloning and Expression Analysis of bZIP Transcription Factor from Finger Millet (Eleusine coracana L.) Under Abiotic Stress Conditions

Basic leucine zipper (bZIP) transcription factors comprise one of the largest gene families in plants. They play a key role in almost every aspect of plant growth and development and also in biotic and abiotic stress tolerance. In this study, we were attempted to study characterization of bZIP, a transcription factor from a climate smart cereal finger millet (Eleusine coracana L.). Seeds of Eleusine coracana (finger millet) was purchase from local market and were grown under field conditions drought and salt stress conditions. In this study, EcbZIP gene was isolated from finger millet, cloned into DH5α cells, screened by using colony PCR and expression analysis in response to two abiotic stresses was carried out by using qRT PCR. EcbZIP coding DNA sequence and protein sequence were retrieved from NCBI Nucleotide Database and Genpept of Accession number KP033192.1 and AJP67539.1 and validated by using SMART (simple modular architecture tool) Domain Tool. Cloning and expression studies were carried out using standardized molecular biology protocol. Results depicted that EcbZIP transcription factor showed significant upregulation under both salt and drought stress conditions, indicating that it plays an important role in tolerance towards these stresses. In conclusion, expression analysis of bZIP gene from finger millet seed cultivar ML-365 showed 5-fold upregulation to salt stress to drought stress and 8-fold upregulation to salt stress. Hence, it can serve as a candidate gene for improving abiotic stress tolerance and can be helpful in enhancing the crop productivity under stress conditions.

Heat shock transcription factor (Hsf) gene family in common bean (Phaseolus vulgaris): genome-wide identification, phylogeny, evolutionary expansion and expression analyses at the sprout stage under abiotic stress

Background Common bean (Phaseolus vulgaris) is an essential crop with high economic value. The growth of this plant is sensitive to environmental stress. Heat shock factor (Hsf) is a family of antiretroviral transcription factors that regulate plant defense system against biotic and abiotic stress. To date, few studies have identified and bio-analyzed Hsfs in common bean. Results In this study, 30 Hsf transcription factors (PvHsf1–30) were identified from the PFAM database. The PvHsf1–30 belonged to 14 subfamilies with similar motifs, gene structure and cis-acting elements. The Hsf members in Arabidopsis, rice (Oryza sativa), maize (Zea mays) and common bean were classified into 14 subfamilies. Collinearity analysis showed that PvHsfs played a role in the regulation of responses to abiotic stress. The expression of PvHsfs varied across different tissues. Moreover, quantitative real-time PCR (qRT-PCR) revealed that most PvHsfs were differentially expressed under cold, heat, salt and heavy metal stress, indicating that PvHsfs might play different functions depending on the type of abiotic stress. Conclusions In this study, we identified 30 Hsf transcription factors and determined their location, motifs, gene structure, cis-elements, collinearity and expression patterns. It was found that PvHsfs regulates responses to abiotic stress in common bean. Thus, this study provides a basis for further analysis of the function of PvHsfs in the regulation of abiotic stress in common bean.

Genome-wide identification and evolution of WNK kinases in Bambusoideae and transcriptional profiling during abiotic stress in Phyllostachys edulis

With-no-lysine (WNK) kinases play vital roles in abiotic stress response, circadian rhythms, and regulation of flowering time in rice, Arabidopsis, and Glycine max. However, there are no previous reports of WNKs in the Bambusoideae, although genome sequences are available for diploid, tetraploid, and hexaploid bamboo species. In the present study, we identified 41 WNK genes in five bamboo species and analysed gene evolution, phylogenetic relationship, physical and chemical properties, cis-elements, and conserved motifs. We predicted the structure of PeWNK proteins of moso bamboo and determined the exposed, buried, structural and functional amino acids. Real-time qPCR analysis revealed that PeWNK5, PeWNK7, PeWNK8, and PeWNK11 genes are involved in circadian rhythms. Analysis of gene expression of different organs at different developmental stages revealed that PeWNK genes are tissue-specific. Analysis of various abiotic stress transcriptome data (drought, salt, SA, and ABA) revealed significant gene expression levels in all PeWNKs except PeWNK11. In particular, PeWNK8 and PeWNK9 were significantly down- and up-regulated, respectively, after abiotic stress treatment. A co-expression network of PeWNK genes also showed that PeWNK2, PeWNK4, PeWNK7, and PeWNK8 were co-expressed with transcriptional regulators related to abiotic stress. In conclusion, our study identified the PeWNKs of moso bamboo involved in circadian rhythms and abiotic stress response. In addition, this study serves as a guide for future functional genomic studies of the WNK genes of the Bambusoideae.

Comprehensive Identification and Expression Analysis of CRY Gene Family in Gossypium

Background: The cryptochromes (CRY) comprise a specific blue light receptor for plants and animals, which play crucial roles in physiological processes of plant growth, development, and stress tolerance. Results: In the present work, a systematical analysis of CRY gene family from five allotetraploid cotton species, G. hirsutum, G. barbadense, G. tomentosum, G. mustelinum and G. darwinii together with seven diploid species. There were 18, 17, 17, 17, and 17 CRYs identified in G. hirsutum, G. barbadense, G. tomentosum, G. mustelinum and G. darwinii, respectively, whereas five to nine CRY genes were identified in the diploid species. Phylogenetic analysis of the protein-coding sequences revealed that CRY genes from the allotetraploids G. hirsutum and G. barbadense, three diploid cotton species (G. raimondii, G. herbaceum, and G. arboreum), and Arabidopsis thaliana could be classified into seven clades. Synteny analysis suggested that the homoeolog of G. hirsutum Gh_A02G0384 has undergone an evolutionary loss event in the other four allotetraploid cotton species. Cis-element analysis predicated the possible functions of CRY genes in G. hirsutum. Public RNA-seq data were investigated to analyze the expression patterns of G. hirsutum CRY genes in various tissues as well as gene expressions under abiotic stress treatments. Conclusion: These results indicated the possible functions of G. hirsutum CRY genes in differential tissues as well as in response to abiotic stress during the cotton plants life cycle.

Genome-wide identification of the Liriodendron chinense WRKY gene family and its diverse roles in response to multiple abiotic stress

Background Liriodendron chinense (Lchi) is a tree species within the Magnoliaceae family and is considered a basal angiosperm. The too low or high temperature or soil drought will restrict its growth as the adverse environmental conditions, thus improving L. chinense abiotic tolerance was the key issues to study. WRKYs are a major family of plant transcription factors known to often be involved in biotic and abiotic stress responses. So far, it is still largely unknown if and how the LchiWRKY gene family is tied to regulating L. chinense stress responses. Therefore, studying the involvement of the WRKY gene family in abiotic stress regulation in L. chinense could be very informative in showing how this tree deals with such stressful conditions. Results In this research, we performed a genome-wide analysis of the Liriodendron chinense (Lchi) WRKY gene family, studying their classification relationships, gene structure, chromosomal locations, gene duplication, cis-element, and response to abiotic stress. The 44 members of the LchiWRKY gene family contain a significant amount of sequence diversity, with their lengths ranging from 525 bp to 40,981 bp. Using classification analysis, we divided the 44 LchiWRKY genes into three phylogenetic groups (I, II, II), with group II then being further divided into five subgroups (IIa, IIb, IIc, IId, IIe). Comparative phylogenetic analysis including the WRKY families from 17 plant species suggested that LchiWRKYs are closely related to the Magnolia Cinnamomum kanehirae WRKY family, and has fewer family members than higher plants. We found the LchiWRKYs to be evenly distributed across 15 chromosomes, with their duplication events suggesting that tandem duplication may have played a major role in LchiWRKY gene expansion model. A Ka/Ks analysis indicated that they mainly underwent purifying selection and distributed in the group IId. Motif analysis showed that LchiWRKYs contained 20 motifs, and different phylogenetic groups contained conserved motif. Gene ontology (GO) analysis showed that LchiWRKYs were mainly enriched in two categories, i.e., biological process and molecular function. Two group IIc members (LchiWRKY10 and LchiWRKY37) contain unique WRKY element sequence variants (WRKYGKK and WRKYGKS). Gene structure analysis showed that most LchiWRKYs possess 3 exons and two different types of introns: the R- and V-type which are both contained within the WRKY domain (WD). Additional promoter cis-element analysis indicated that 12 cis-elements that play different functions in environmental adaptability occur across all LchiWRKY groups. Heat, cold, and drought stress mainly induced the expression of group II and I LchiWRKYs, some of which had undergone gene duplication during evolution, and more than half of which had three exons. LchiWRKY33 mainly responded to cold stress and LchiWRKY25 mainly responded to heat stress, and LchiWRKY18 mainly responded to drought stress, which was almost 4-fold highly expressed, while 5 LchiWRKYs (LchiWRKY5, LchiWRKY23, LchiWRKY14, LchiWRKY27, and LchiWRKY36) responded equally three stresses with more than 6-fold expression. Subcellular localization analysis showed that all LchiWRKYs were localized in the nucleus, and subcellular localization experiments of LchiWRKY18 and 36 also showed that these two transcription factors were expressed in the nucleus. Conclusions This study shows that in Liriodendron chinense, several WRKY genes like LchiWRKY33, LchiWRKY25, and LchiWRKY18, respond to cold or heat or drought stress, suggesting that they may indeed play a role in regulating the tree’s response to such conditions. This information will prove a pivotal role in directing further studies on the function of the LchiWRKY gene family in abiotic stress response and provides a theoretical basis for popularizing afforestation in different regions of China.

Differential Physio-Biochemical and Metabolic Responses of Peanut (Arachis hypogaea L.) under Multiple Abiotic Stress Conditions

The frequency and severity of extreme climatic conditions such as drought, salinity, cold, and heat are increasing due to climate change. Moreover, in the field, plants are affected by multiple abiotic stresses simultaneously or sequentially. Thus, it is imperative to compare the effects of stress combinations on crop plants relative to individual stresses. This study investigated the differential regulation of physio-biochemical and metabolomics parameters in peanut (Arachis hypogaea L.) under individual (salt, drought, cold, and heat) and combined stress treatments using multivariate correlation analysis. The results showed that combined heat, salt, and drought stress compounds the stress effect of individual stresses. Combined stresses that included heat had the highest electrolyte leakage and lowest relative water content. Lipid peroxidation and chlorophyll contents did not significantly change under combined stresses. Biochemical parameters, such as free amino acids, polyphenol, starch, and sugars, significantly changed under combined stresses compared to individual stresses. Free amino acids increased under combined stresses that included heat; starch, sugars, and polyphenols increased under combined stresses that included drought; proline concentration increased under combined stresses that included salt. Metabolomics data that were obtained under different individual and combined stresses can be used to identify molecular phenotypes that are involved in the acclimation response of plants under changing abiotic stress conditions. Peanut metabolomics identified 160 metabolites, including amino acids, sugars, sugar alcohols, organic acids, fatty acids, sugar acids, and other organic compounds. Pathway enrichment analysis revealed that abiotic stresses significantly affected amino acid, amino sugar, and sugar metabolism. The stress treatments affected the metabolites that were associated with the tricarboxylic acid (TCA) and urea cycles and associated amino acid biosynthesis pathway intermediates. Principal component analysis (PCA), partial least squares-discriminant analysis (PLS-DA), and heatmap analysis identified potential marker metabolites (pinitol, malic acid, and xylopyranose) that were associated with abiotic stress combinations, which could be used in breeding efforts to develop peanut cultivars that are resilient to climate change. The study will also facilitate researchers to explore different stress indicators to identify resistant cultivars for future crop improvement programs.