Genome-wide expression profiling of aquaporin genes confer responses to abiotic and biotic stresses in Brassica rapa

Schematic representation of motif compositions in the BrAQP protein sequences. Different motifs, numbered 1–10, are displayed in different colored boxes. The names of all members are displayed on the left, while the length of the motif is shown in the scale at the bottom of the figure. (PPTX 269 kb

Overexpression of BrSAC1 encoding a phosphoinositide phosphatase isolated from Chinese cabbage (Brassica rapa L) improved tolerance to cold, dehydration, and salt stresses in transgenic tobacco

This study demonstrates the isolation and characterization of cDNA encoding a phosphoinositide phosphatase (PIP) from a stem cell cDNA library of Chinese cabbage (Brassica rapa) seedling. The full length gene (BrSAC1; GenBank accession no., GU434275) contained 1999 base pairs (bp), with an open reading frame of 1785 bp, encoding a polypeptide of 594 amino acids with a predicted molecular weight of 65 kDa, including a putative N-terminal signal peptide (the signal peptide counted within the 594 residues). Other regions found within the sequence include a conserved KXKXX COPI-binding motif and a consensus Cx5R (T/S) catalytic motif. BrSAC1 protein shares 92% identity with AtSac1B, and 86% identity with AtRHD4 at the amino acid level. Gene expression analyses revealed that BrsSAC1 was constitutively expressed at high levels in the pistil, stamen and flower bud, whereas it was expressed at low levels in the leaf and stem. In addition to injury, BrSAC1 expression was also induced in response to different types of stress condition, namely cold, desiccation, salt, submergence, abscisic acid and heavy metals. Overexpression of BrSAC1 in transgenic tobacco conferred tolerance to cold, dehydration, and salt stress at the seed germination/seedling stage as reflected by the percentage of germination/green seedlings, the fresh weight of seedlings and their development pattern. Our data suggest that BrSAC1 is an important stress response determinant in plants.Keywords: Abiotic stress, Brassica rapa, phosphoinositide phosphatase, transgenic plantAfrican Journal of Biotechnology Vol. 12(15), pp. 1782-179

Identification and characterization of longevity assurance gene related to stress resistance in Brassica

Brassica is a very important vegetable group worldwide and different stresses are a major concern for these crops. Enhancement of resistance against biotic and abiotic stresses by exploiting stress resistance related genes offers the most efficient approach to address this concern. In this study, a stress resistance related gene was identified from the full-length cDNA library of Brassica rapa cv. Osome, which was determined to be Brassica longevity assurance protein (BrLAP) after sequence analysis. A comparison study of this gene showed a high degree of homology with other stress resistance related longevity assurance genes and was shown to be expressed in all organs during all of the developmental growth stages. In addition, this gene significantly responded after cold, drought and ABA stress treatments in Chinese cabbage. All these data revealed that this gene may be involved in plant resistance against stresses.Keywords: Brassica rapa, longevity assurance gene, gene expression, biotic and abiotic stres

Overexpression of the pineapple fruit bromelain gene (BAA) in transgenic Chinese cabbage ( Brassica rapa ) results in enhanced resistance to bacterial soft rot

Bromelain is a crude protein extract obtained from pineapple stems, which comprises a variety of proteolytic enzymes. It exhibits potential therapeutic activities against trauma, inflammation, autoimmune diseases and malignant disorders. In this study, we cloned BAA1 (the gene encoding fruit bromelain) into a plant expression vector that was then used to transform Brassica rapa and overexpress BAA1 under the control of the cauliflower mosaic virus (CaMV) 35S promoter. We demonstrate that constitutive overexpression of BAA1 in B. rapa confers enhanced resistance to the soft rot pathogen Pectobacterium carotovorum ssp. carotovorum . These results suggest that it could be utilized for protecting plants from attack by bacterial pathogens

Molecular characterization and spatiotemporal expression of prohormone convertase 2 in the Pacific abalone, Haliotis discus hannai.

Prohormone convertases (PCs) are subtilisin-like proteases responsible for the intracellular processing of prohormones and proneuropeptides in vertebrates and invertebrates. The full-length PC2 cDNA sequence was cloned from pleuropedal ganglion of Haliotis discus hannai, consisted of 2254-bp with an open reading frame of 1989-bp and encoded a protein of 662 amino acid residues. The architecture of Hdh PC2 displayed key features of PCs, including a signal peptide, a pro-segment domain with sites for autocatalytic activation, a catalytic domain, and a pro-protein domain (P-domain). It shares the highest homology of its amino acid sequence with the PC2 from H. asinina and to lesser extent with that of Homo sapiens and Rana catesbeiana PC2. Sequence alignment analysis indicated that Hdh PC2 was highly conserved in the catalytic domain, including a catalytic triad of serine proteinases of the subtilisin family at positions Asp-195, His-236, and Ser-412. The cloned sequence contained a canonical integrin binding sequence, and four cysteine residues involved in the formation of an intramolecular disulfide link. Phylogenetic analysis revealed that the Hdh PC2 is robustly clustered with the Has PC2. Quantitative PCR assay demonstrated that the Hdh PC2 was predominantly expressed in the pleuropedal ganglion rather than in other examined tissues. Although PC2 mRNA was expressed throughout the gametogenetic cycle of male and female abalone, the expression level was significantly higher in the ripening stage of female abalone. Also, a significantly higher expression was observed in the pleuropedal ganglion and gonadal tissues at a higher effective accumulative temperature (1000°C). In situ hybridization revealed that the PC2 mRNA expressing neurosecretory cells were distributed in the cortex region of the pleuropedal ganglion. According to the results, it can be concluded that pleuropedal ganglion is the highest site of PC2 activity, and this enzyme might be involved in the abalone reproduction process

Characterization of Insulin-Like Growth Factor Binding Protein 7 (Igfbp7) and Its Potential Involvement in Shell Formation and Metamorphosis of Pacific Abalone, Haliotis discus hannai

Insulin-like growth factor binding proteins (IGFBPs) are secreted proteins that play an important role in IGF regulation of growth and development of vertebrate and invertebrates. In this study, the IGFBP7 gene was cloned and characterized from mantle tissues of H. discus hannai, and designated as Hdh IGFBP7. The full-length cDNA sequence transcribed from the Hdh IGFBP7 gene was 1519-bp long with an open reading frame of 720-bp corresponding to a putative polypeptide of 239 amino acids. The molecular mass of its mature protein was approximately 23.44 KDa with an estimated isoelectric point (pI) of 5.35, and it shared significant homology with IGFBP7 gene of H. madaka. Hdh IGFBP7 has a characteristic IGFBP N-terminal domain (22–89 aa), a kazal-type serine proteinase inhibitor domain (77–128), and an immunoglobulin-like C2 domain (144–223). Furthermore, twelve cysteine residues and a signature motif of IGFBPs (XCGCCXXC) were found in its N-terminal domain. Phylogenetic analysis revealed that Hdh IGFBP7 was aligned with IGFBP7 of H. madaka. Tissue distribution analysis showed that the mRNA of Hdh IGFBP7 was expressed in all examined tissues, with the highest expression level observed in the mantle and gill tissues. The expression level of Hdh IGFBP7 mRNA was relatively higher at the juvenile stage during its metamorphosis period. In situ hybridization showed that Hdh IGFBP7 transcript was expressed in epithelial cells of the dorsal mantle pallial and mucus cells of the branchial epithelium in gill. These results provide basic information for future studies on the role of IGFBP7 in IGF regulation of shell growth, development and metamorphosis of abalone

Expression of salicylic acid-related genes in Brassica oleracea var. capitata during Plasmodiophora brassicae infection

Brassica oleracea var. capitata (cabbage) is an important vegetable crop in Asian countries such as Korea, China and Japan. Cabbage production is severely affected by clubroot disease caused by the soil-borne plant pathogen Plasmodiophora brassicae. During clubroot development, methyl salicylate (MeSA) is biosynthesized from salicylic acid (SA) by methyltransferase. In addition, methyl salicylate esterase (MES) plays a major role in the conversion of MeSA back into free SA. The interrelationship between MES and methytransferases during clubroot development has not been fully explored. To begin to examine these relationships, we investigated the expression of MES genes in disease- susceptible and -resistant plants during clubroot development. We identified three MES-encoding genes potentially involved in the defense against pathogen attack. We found that SS1 was upregulated in both the leaves and roots of B. oleracea during P. brassicae infection. These results support the conclusion that SA biosynthesis is suppressed during pathogen infection in resistant plants. We also characterized the expression of a B. oleracea BSMT gene, which appears to be involved in glycosylation rather than MeSA biosynthesis. Our results provide insight into the functions and interactions of genes for MES and methyltransferase during infection. Taken together, our findings indicate that MES genes are important candidates for use to control clubroot diseases.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Characterization and abiotic stress-responsive expression analysis of SGT1 genes in Brassica oleracea

SGT1 genes are involved in enhancing plant responses to various biotic and abiotic stresses. Brassica oleracea is known to contain two types of SGT1 genes, namely Suppressor of G2 allele of SKP1 and Suppressor of GCR2. In this study, through systematic analysis, four putative SGT1 genes were identified and characterized in B. oleracea. In phylogenetic analysis, the genes were formed BolSGT1a, BolSGT1b (both Suppressor of G2 allele of SKP1 types) and BolSGT1 (Suppressor of GCR2) groups. Functional domain analysis and organ-specific expression patterns suggested possible roles for BolSGT1 genes during stress conditions. BolSGT1 genes showed significant changes in expression in response to heat, cold, drought, salt, or ABA treatment. Interaction network analysis supported the expression analysis, and showed that the BolSGT1a and BolSGT1b genes are strongly associated with co-regulators during stress conditions. However, the BolSGT1 gene not showed any strong association. Hence, BolSGT1 might be a stress resistance-related gene that functions without a co-regulator. Our results show that BolSGT1 genes are potential target genes to improve B. oleracea resistance to abiotic stresses such as heat, cold, and salt.The accepted manuscript in pdf format is listed with the files at the bottom of this page. The presentation of the authors' names and (or) special characters in the title of the manuscript may differ slightly between what is listed on this page and what is listed in the pdf file of the accepted manuscript; that in the pdf file of the accepted manuscript is what was submitted by the author

Molecular Modeling of Myrosinase from Brassica oleracea: A Structural Investigation of Sinigrin Interaction

Myrosinase, which is present in cruciferous plant species, plays an important role in the hydrolysis of glycosides such as glucosinolates and is involved in plant defense. Brassicaceae myrosinases are diverse although they share common ancestry, and structural knowledge about myrosinases from cabbage (Brassica oleracea) was needed. To address this, we constructed a three-dimensional model structure of myrosinase based on Sinapis alba structures using Iterative Threading ASSEmbly Refinement server (I-TASSER) webserver, and refined model coordinates were evaluated with ProQ and Verify3D. The resulting model was predicted with β/α fold, ten conserved N-glycosylation sites, and three disulfide bridges. In addition, this model shared features with the known Sinapis alba myrosinase structure. To obtain a better understanding of myrosinase–sinigrin interaction, the refined model was docked using Autodock Vina with crucial key amino acids. The key nucleophile residues GLN207 and GLU427 were found to interact with sinigrin to form a hydrogen bond. Further, 20-ns molecular dynamics simulation was performed to examine myrosinase–sinigrin complex stability, revealing that residue GLU207 maintained its hydrogen bond stability throughout the entire simulation and structural orientation was similar to that of the docked state. This conceptual model should be useful for understanding the structural features of myrosinase and their binding orientation with sinigrin