Haiyan Shi

Physiological and Molecular Responses to Salt Stress in Wild Emmer and Cultivated Wheat

Salinity severely influences growth and grain yield of wheat. Modern breeding efforts have contributed to severe loss of genetic diversity and reduced tolerance to salt stress in cultivated plants. Wild emmer wheat (Triticum dicoccoides), the progenitor of cultivated wheat, is well-adapted to a wide range of environments and exhibits tolerance to abiotic stress. However, there is lack of fundamental knowledge of the mechanism of salt stress tolerance in wild emmer wheat and how it differs from that of the cultivated wheat. By screening wild emmer genotypes, we identified a promising salt-tolerant line from Gitit in the eastern Samaria steppes. We investigated the physiological difference of wild emmer and cultivated wheats in response to salt stress. Our results revealed that salt stress resulted in an increase in lipid peroxidation (malondialdehyde) content and electrolyte leakage, to a greater extent in cultivated wheat genotype, Zheng 9023, than in salt-resistant wild emmer wheat genotype 18-35, but the latter had higher relative dry weight. Differential expression analysis showed that higher transcript induction folds of genes encoding transcription factor were detected in the resistant plants (wild emmer) than in sensitive plants (cultivated wheat) after salt treatment. In conclusion, wild emmer wheat demonstrated better tolerance to salt stress than cultivated wheat, and the higher tolerance of wild emmer wheat is because of high expression of stress-responsive genes encoding transcription factor, including NAC2F, NAC8, DREB3A, MYB3R, and MYB2A. Therefore, our results suggest that wild emmer wheat is an important germplasm for salt tolerance breeding in cultivated wheat.

Two Pear Glutathione S-Transferases Genes Are Regulated during Fruit Development and Involved in Response to Salicylic Acid, Auxin, and Glucose Signaling

Two genes encoding putative glutathione S-transferase proteins were isolated from pear (Pyrus pyrifolia) and designated PpGST1 and PpGST2. The deduced PpGST1 and PpGST2 proteins contain conserved Glutathione S-transferase N-terminal domain (GST_N) and Glutathione S-transferase, C-terminal domain (GST_C). Using PCR amplification technique, the genomic clones corresponding to PpGST1 and PpGST2 were isolated and shown to contain two introns and a singal intron respectively with typical GT/AG boundaries defining the splice junctions. Phylogenetic analysis clearly demonstrated that PpGST1 belonged to Phi class of GST superfamilies and had high homology with apple MdGST, while PpGST2 was classified into the Tau class of GST superfamilies. The expression of PpGST1 and PpGST2 genes was developmentally regulated in fruit. Further study demonstrated that PpGST1 and PpGST2 expression was remarkably induced by glucose, salicylic acid (SA) and indole-3-aceticacid (IAA) treatments in pear fruit, and in diseased fruit. These data suggested that PpGST1 and PpGST2 might be involved in response to sugar, SA, and IAA signaling during fruit development of pear.

Two pear auxin-repressed protein genes, PpARP1 and PpARP2, are predominantly expressed in fruit and involved in response to salicylic acid signaling

Two genes encoding putative auxin-repressed proteins, designated PpARP1 and PpARP2, were isolated from pear (Pyrus pyrifolia). The deduced PpARP1 and PpARP2 proteins contain the typical auxin_repressed Dormancy/auxin associated protein domains. Phylogenetic analyses clearly demonstrated that PpARP1 has high homology with pear ARPs except PpARP2 which belongs to another clade. Using PCR amplification technique, the genomic clones corresponding to PpARP1 and PpARP2 were isolated and shown to contain two introns with typical GT/AG boundaries defining the splice junctions. PpARP1 and PpARP2 genes were predominantly expressed in fruit. PpARP1 transcripts were also detected in shoots, petals, and anthers, but a relatively low expression signal was detected in young leaves. PpARP2 transcripts were also detected in other tissues. In particular, expression of PpARP1 and PpARP2 was developmentally regulated in fruit. Further study demonstrated that PpARP1 and PpARP2 expression in pear fruit was remarkably inhibited by salicylic acid and induced by IAA. These data suggested that PpARP1 and PpARP2 might be involved in the response to salicylic acid signaling during fruit development of pear. The study of the pear PpARP genes also demonstrated a link between salicylic acid and auxin signaling.

ABA Is Involved in Regulation of Cold Stress Response in Bermudagrass

As a representative warm-season grass, Bermudagrass [Cynodon dactylon (L). Pers.] is widely used in turf systems. However, low temperature remarkably limits its growth and distribution. ABA is a crucial phytohormone that has been reported to regulate much important physiological and biochemical processes in plants under abiotic stress. Therefore, the objective of this study was to figure out the effects of ABA on the cold-sensitive (S) and cold-resistant (R) Bermudagrass genotypes response to cold stress. In this study, the plants were treated with 100 μM ABA solution and exposed to 4°C temperature. After 7 days of cold treatment, the electrolyte leakage (EL), malonaldehyde (MDA) and H2O2 content were significantly increased in both genotypes compared with control condition, and these values were higher in R genotype than those of S genotype, respectively. By contrast, exogenous ABA application decreased the electrolyte leakage (EL), MDA and H2O2 content in both genotypes compared with those plants without ABA treatment under cold treatment condition. In addition, exogenous ABA application increased the levels of chlorophyll a fluorescence transient curve for both genotypes, and it was higher in R genotype than that of S genotype. Analysis of photosynthetic fluorescence parameters revealed that ABA treatment improved the performance of photosystem II under cold condition, particularly for the R genotype. Moreover, cold stress significantly increased δ13C values for both genotypes, while it was alleviated by exogenous ABA. Additionally, exogenous ABA application altered the expression of ABA- or cold related genes, including ABF1, CBF1, and LEA. In summary, exogenous ABA application enhanced cold resistance of both genotypes by maintaining cell membrane stability, improving the process of photosystem II, increasing carbon isotopic fractionation under cold stress, and more prominently in R genotype compared with S genotype.

TdCBL6, a calcineurin B-like gene from wild emmer wheat (Triticum dicoccoides), is involved in response to salt and low-K+ stresses

The calcineurin B-like proteins (CBLs), a unique family of calcium sensors in plants, have been shown to be involved in abiotic stresses, such as salt, drought and cold. Although extensive studies and remarkable progress have been made in Arabidopsis (Arabidopsis thaliana) CBLs, very little is known about the role of CBL genes in wheat. In this study, a CBL gene, designated TdCBL6, was cloned and characterized from wild emmer wheat (Triticum dicoccoides), the progenitor of cultivated wheat. Sequence alignment revealed that TdCBL6 shares high sequence homology with rice OsCBL6. Phylogenetic analysis also revealed that TdCBL6 protein has the closest evolutionary relationship with rice OsCBL6 protein. TdCBL6 transcription was induced by NaCl, polyethylene glycol and abscisic acid. Further differential expression analysis revealed that TdCBL6 expression was much higher in the salt-tolerant line than in the salt-sensitive line when they were subjected to salt treatment. Transgenic Arabidopsis ectopic expression of the TdCBL6 gene displayed higher levels of photosynthetic efficiency (Fv/Fm) and lower ion leakage (EL) than wild-type (WT) plants under NaCl stress conditions. Moreover, TdCBL6-overexpressing lines showed low-K+ (LK)-sensitive phenotypes compared with WT plants. Further experiments revealed that ectopic expression of TdCBL6 resulted in reduction of H2O2 content, and affected expression of K+-responsive/H2O2-regulated genes under LK stress. Taken together, we demonstrated that heterologous expression of TdCBL6 in Arabidopsis confers salt tolerance by reducing membrane injury and improving photosynthetic efficiency, and that the TdCBL6 gene may be involved in response to LK stress by regulating the reactive oxygen species-mediated LK signaling pathway.

Pear PIP1 gene is regulated during fruit development and is invovled in response to salicylic acid and ethylene

Shi, H., Wang, Y., Zhang, D., Chen, L. and Zhang, Y. 2015. Pear PIP1 gene is regulated during fruit development and is invovled in response to salicylic acid and ethylene. Can. J. Plant Sci. 95: 77-85. Plasma membrane intrinsic proteins (PIPs), a subfamily of aquaporins, are widely implicated in plant growth and development. A gene encoding a plasma membrane intrinsic protein and designated PpPIP1 was isolated from pear (Pyrus pyrifolia). Using PCR amplification techniques, the genomic clone corresponding to PpPIP1 was isolated and shown to contain three introns with typical GT/AG boundaries defining the splice junctions. The deduced PpPIP1 protein contains the conserved features of PIPs: six transmembrane a-helices, a major intrinsic protein domain, and a conserved asparagine-proline-alanine (NPA) signature sequence. Phylogenetic analyses clearly demonstrated that PpPIP1 has the highest homology with apple (Malus×domestica) MdPIP1a and Malus hupehensis MhPIP1-1. PpPIP1 transcripts were mainly detected in young leaves, shoots, petals and mesocarp of fruit, but a relatively low expression signal was detected in anthers. In particular, expression of PpPIP1 was developmentally regulated in fruit. Further study demonstrated that PpPIP1 expression in pear fruit was down-regulated by salicylic acid (SA) and up-regulated by ethylene. These data suggest that PpPIP1 may be involved in the response to SA and ethylene during fruit development, which would provide valuable information for water permeability studies in pear.

Pear IAA1 gene encoding an auxin-responsive Aux/IAA protein is involved in fruit development and response to salicylic acid

Shi, H., Wang, Y., Li, Z., Zhang, D., Zhang, Y., Xiang, D., Li, Y. and Zhang, Y. 2014. Pear IAA1 gene encoding an auxin-responsive Aux/IAA protein is involved in fruit development and response to salicylic acid. Can. J. Plant Sci. 94: 263-271. Auxin-responsive Aux/IAA proteins are rapidly auxin-induced, short-lived proteins that act as repressors for the auxin response factor (ARF)-activated gene expression. A gene encoding an Aux/IAA protein and designated PpIAA1 was isolated from pear (Pyrus pyrifolia). Using PCR amplification techniques, the genomic clone corresponding to PpIAA1 was isolated and shown to contain three introns with typical GT/AG boundaries defining the splice junctions. The deduced PpIAA1 protein contains the conserved features of indole-3-acetic acids (IAA): four Aux/IAA conserved domains, Aux/IAA family domain, Aux/IAA-ARF dimerization domain profile, and conserved nuclear localization signal (NLS) motifs. Phylogenetic analyses clearly demonstrated PpIAA1 has the highest homology with grape VvIAA. PpIAA1 was preferentially expressed in fruit, and moderate expression was found in anthers. Relatively low expression signal was detected in other tissues including shoots, leaves, and petals. Moreover, expression of PpIAA1 was developmentally regulated in fruit. Further study demonstrated that PpIAA1 expression in pear fruit was remarkably regulated by salicylic acid and IAA. The data suggest that PpIAA1 might be involved in the interplay between IAA and salicylic acid signaling pathway during the fruit development of pear.

Cloning, characterization and expression analysis of a 1-aminocyclopropane-1-carboxylate synthase gene from pear

Shi, H., Zhang, Y. and Chen, L. 2013. Cloning, characterization and expression analysis of a 1-aminocyclopropane-1-carboxylate synthase gene from pear. Can. J. Plant Sci. 93: 465-471. In this study, a cDNA clone encoding putative 1-aminocyclopropane-1-carboxylate (ACC) synthase (ACS) that catalyzes the conversion of S-adenosyl-L-methionine to ACC in ethylene biosynthetic pathway was isolated from a cDNA library produced using mRNA from pear (Pyrus pyrifolia). The cDNA clone, designated PpACS2, comprised an open reading frame of 1, 341 bp encoding a protein of 446 amino acids that shares high similarity with the known plant ACSs. Using PCR amplification technique, a genomic clone (GenBank accession number: KC146402) corresponding to PpACS2 was isolated and shown to contain two introns. The PpACS2 gene product shared 97% identity with an ACC synthase from pear (Pyrus communis). Phylogenetic analyses clearly placed the gene product in the ACC synthase cluster of plant ACS superfamily tree. Quantitative RT-PCR analysis indicated that the PpACS2 gene was preferentially expressed in young pear leaves and shoots. The transcript of PpACS2 gene was accumulated at relatively high levels in anthers, but no signal was detected in the petals and mesocarp of pear. These results suggest that the PpACS2 may participate in the regulation of ethylene production in pear leaves, shoots, and anthers.