Novel Drought-Inducible Genes in the Highly Drought-Tolerant Cowpea: Cloning of cDNAs and Analysis of the Expression of the Corresponding Genes

Ten cDNAs of genes that were induced by dehydration stress werecloned by differential screening from the highly drought-tolerantlegume, cowpea (Vigna unguiculata), a major crop in West Africa.The clones were collectively named CPRD (cowpea clones responsiveto dehydration). Northern blot analysis revealed that nine ofthe CPRD genes were induced by dehydration stress, but the timingof induction of mRNA synthesis varied among the CPRD genes.We analyzed the effects of other environmental stresses on theexpression of the CPRD8, CPRD14 and CPRD22 genes, and we foundthat these genes were strongly induced by high-salinity stressbut not by cold or heat stress. Drought-stressed cowpea plantsaccumulated abscisic acid (ABA) to a level that was 160 timeshigher than that in unstressed plants. The CPRD8 and CPRD22genes were induced to a significant extent by the applicationof exogenous ABA but the CPRD14 gene was not. These resultsindicate the existence of at least two signal-transduction pathwaysbetween the detection of water stress and the expression ofCPRD genes in cowpea. Sequence analysis of CPRD8 and CPRD22cDNAs revealed that they encoded putative proteins that wererelated to old yellow enzyme and group 2 LEA proteins, respectively.The protein encoded by CPRD14 exhibited sequence homology todihydroflavonol-4-reductase (DFR) and vestitone reductase (VR).Old yellow enzyme, DFR and VR have not been identified as drought-inducibleproteins in other plants, whereas LEA genes have been well characterizedas drought-inducible genes. The various gene products mightfunction to protect cells from environmental stress. (Received April 17, 1996; Accepted August 28, 1996)     

Correlation between the induction of a gene for Δ1-pyrroline-5-carboxylate synthetase and the accumulation of proline in Arabidopsis thaliana under osmotic stress

The isolation and characterization is reported of a cDNA for Δ¹-pyrroline-5-carboxylate (P5C) synthetase (cAtP5CS), an enzyme involved in the biosynthesis of proline, from a cDNA library prepared from a dehydrated rosette plant of Arabidopsis thaliana . Southern blot analysis suggested that only one copy of the corresponding gene ( AtP5CS ) is present in A. thaliana . The deduced amino acid sequence of the P5CS protein (AtP5CS) from A. thaliana exhibited 74% homology to that of the P5CS from Vigna aconitifolia . Northern blot analysis revealed that the gene for P5CS was induced by dehydration, high salt and treatment with ABA, while it was not induced by heat or cold treatment. Moreover, the simultaneous accumulation of proline was observed as a result of the former treatments in A. thaliana . A cDNA for P5C reductase (cAtP5CR) was also isolated from A. thaliana and Northern blot analysis was performed. The AtP5CR gene was not induced to a significant extent by dehydration or high-salt stress. These observations suggest that the AtP5CS gene plays a principal role in the biosynthesis of proline in A. thaliana under osmotic stress.     

Identification of a cis-regulatory region of a gene in Arabidopsis thaliana whose induction by dehydration is mediated by abscisic acid and requires protein synthesis

The effect of the ATP-dependent exonuclease AddAB complex on the structural stability of plasmid pGP1 inBacillus subtilis was studied. Using deletion mutagenesis and gene amplification techniques,B. subtilis strains were constructed either lacking or overproducing the AddAB complex, a key enzyme in homologous recombination. The deletion mutant possessed no residual ATP-dependent nuclease activity; in contrast, the nuclease activity was up to 30 times higher in lysates of strains carrying multiple copies of theaddAB genes in the chromosome. Southern blot analyses of these strains indicated that a linear relationship exists between the number of chromosomal gene copies and the level of AddAB activity. The structural stability of pGP1 was analyzed in the AddAB-deficient and over-producing backgrounds. Frequencies of deletion formation in the plasmid, as monitored by the expression of the pGP1-encodedpenP-lacZ fusion on media containing X-gal, were shown to be increased at least 25-fold in theaddAB knock-out mutant, whereas the stability of pGP1 was improved up to 15-fold in strains verproducing the AddAB enzyme. A possible explanation for these findings is that interactions between AddAB and plasmid molecules prevent the formation of secondary structures that constitute potential deletion target sites, and thereby enhance the structural stability of plasmids.     

Regulation of Levels of Proline as an Osmolyte in Plants under Water Stress

Compatible osmolytes are potent osmoprotectants that play arole in counteracting the effects of osmotic stress. Proline(Pro) is one of the most common compatible osmolytes in water-stressedplants. The accumulation of Pro in dehydrated plants is causedboth by activation of the biosynthesis of Pro and by inactivationof the degradation of Pro. In plants, L-Pro is synthesized fromL-glutamic acid (l-G1u) via     

Characterization of the gene for Δ1-pyrroline-5-carboxylate synthetase and correlation between the expression of the gene and salt tolerance in Oryza sativa L.

A cDNA for ¹-pyrroline-5-carboxylate (P5C) synthetase (cOsP5CS), an enzyme involved in the biosynthesis of proline, was isolated and characterized from a cDNA library prepared from 14-day-old seedlings of Oryza sativa cv. Akibare. The deduced amino acid sequence of the P5CS protein (OsP5CS) from O. sativa exhibited 74.2% and 75.5% homology to that of the P5CS from Arabidopsis thaliana and Vigna aconitifolia, respectively. Northern blot analysis revealed that the gene for P5CS (OsP5CS) was induced by high salt, dehydration, treatment of ABA and cold treatment, while it was not induced by heat treatment. Simultaneously, accumulation of proline was observed as a result of high salt treatment in O. sativa. Moreover, the levels of expression of OsP5CS mRNA and content of proline under salt stress condition were compared between a salt-tolerant cultivar, Dee-gee-woo-gen (DGWG) and a salt-sensitive breeding line, IR28. It was observed that the expression of the P5CS gene and the accumulation of proline in DGWG steadily increased, whereas those in IR28 increased slightly.