Promoter of a salinity and cold stress-induced MCM6 DNA helicase from pea.
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ABSTRACT:
SUBMITTER: Dang HQ
PROVIDER: S-EPMC3257780 | biostudies-literature |
REPOSITORIES: biostudies-literature
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Project description:Helicases are motor proteins that can transiently catalyze the unwinding of energetically stable duplex DNA or RNA molecules by using ATP hydrolysis as the source of energy. Many helicases share a core region of highly conserved sequence motifs, and belong to the rapidly growing DEAD-box protein family. Pea DNA helicase 45 (PDH45), that exhibits striking homology with eukaryotic translation initiation factor 4A (eIF4A), contains ATP-dependent DNA and RNA helicase, DNA-dependent ATPase, and ATP-binding activities. The transcript of the PDH45 gene was reported to be upregulated in pea plant in response to high salinity, cold stress, abscisic acid (ABA), dehydration and early wounding. The first direct evidence that overexpression of PDH45 confers salinity stress tolerance without yield loss has also been reported. A promoter analysis of PDH45 gene has not been studied. The cis-regulatory elements present on promoter region of the gene act as binding sites for RNA polymerase and transcription factors and control the regulation of gene expression. Here we report the promoter of the PDH45 gene that contains stress-responsive cis-regulatory elements which may be responsible for regulating the expression of PDH45 under abiotic stress conditions.
| S-EPMC3256367 | biostudies-literature
Project description:Salinity stress is one of the major factors negatively affecting growth and productivity in living organisms including plants and bacteria resulting in significant losses worldwide. Therefore, it would be fruitful to develop salinity stress tolerant useful species and also to understand the mechanism of stress tolerance. The pea DNA helicase 45 (PDH45) is a DNA and RNA helicase, homologous to eukaryotic translation initiation factor 4A (eIF-4A) and is involved in various processes including protein synthesis, maintaining the basic activities of the cell, upregulation of topoisomerase I activity and salinity stress tolerance in plant, but its role in salinity stress tolerance in bacteria has not heretofore been studied. This study provides an evidence for a novel function of the PDH45 gene in high salinity (NaCl) stress tolerance in bacteria (Eschericia coli, BL21 cells) also. Furthermore, it has been shown that the functionally active PDH45 gene is required to show the stress tolerance in bacteria because the single mutants (E183G or R363Q) and the double mutant (E183G + R363Q) of the gene could not confer the same function. The response was specific to Na+ ions as the bacteria could not grow in presence of LiCl. This study suggests that the cellular response to high salinity stress across prokaryotes and plant kingdom is conserved and also helps in our better understanding of mechanism of stress tolerance in bacteria and plants. It could also be very useful in developing high salinity stress tolerant useful bacteria of agronomic importance. Overall, this study provides an evidence for a novel function of the PDH45 gene in high salinity stress tolerance in bacteria.
| S-EPMC3258049 | biostudies-literature
Project description:Pea mini-chromosome maintenance 6 (MCM6) single subunit (93 kDa) forms homohexamer (560 kDa) and contains an ATP-dependent and replication fork stimulated 3' to 5' DNA unwinding activity along with intrinsic DNA-dependent ATPase and ATP-binding activities [Plant Mol. Biol. 2010; DOI: 10.1007/s11103-010-9675-7]. Here, we have determined the effect of various DNA-binding agents, such as actinomycin, nogalamycin, daunorubicin, doxorubicin, distamycin, camptothecin, cyclophosphamide, ellipticine, VP-16, novobiocin, netropsin, cisplatin, mitoxantrone and genistein on the DNA unwinding and ATPase activities of the pea MCM6 DNA helicase. The results show that actinomycin and nogalamycin inhibited the DNA helicase (apparent Ki values of 10 and 1 μM, respectively) and ATPase (apparent Ki values of 100 and 17 μM, respectively) activities. Although, daunorubicin and doxorubicin also inhibited the DNA helicase activity of pea MCM6, but with less efficiency; however, these could not inhibit the ATPase activity. These results suggest that the intercalation of the inhibitors into duplex DNA generates a complex that impedes translocation of MCM6, resulting in the inhibitions of the activities. This study could be useful in our better understanding of the mechanism of plant nuclear DNA helicase unwinding.
| S-EPMC3142409 | biostudies-literature
Project description:Salt tolerance is an important trait that is required to overcome salinity-induced reduction in plant productivity. We have reported previously the isolation of a pea DNA helicase 45 (PDH45) that exhibits striking homology with the eukaryotic translation initiation factor eIF-4A. Here, we report that PDH45 mRNA is induced in pea seedlings in response to high salt, and its overexpression driven by a constitutive cauliflower mosaic virus-(35)S promoter in tobacco plants confers salinity tolerance, thus suggesting a previously undescribed pathway for manipulating stress tolerance in crop plants. The T(0) transgenic plants showed high levels of PDH45 protein in normal and stress conditions, as compared with WT plants. The T(0) transgenics also showed tolerance to high salinity as tested by a leaf disk senescence assay. The T(1) transgenics were able to grow to maturity and set normal viable seeds under continuous salinity stress without any reduction in plant yield in terms of seed weight. Measurement of Na(+) ions in different parts of the plant showed higher accumulation in the old leaves and negligible accumulation in seeds of T(1) transgenic lines as compared with the WT plants. The possible mechanism of salinity tolerance is discussed. Overexpression of PDH45 provides a possible example of the exploitation of DNA/RNA unwinding pathways for engineering salinity tolerance without affecting yield in crop plants.
| S-EPMC544286 | biostudies-literature
Project description:Mcm proteins play an essential role in eukaryotic DNA replication, but their biochemical functions are poorly understood. Recently, we reported that a DNA helicase activity is associated with an Mcm4-Mcm6-Mcm7 (Mcm4,6,7) complex, suggesting that this complex is involved in the initiation of DNA replication as a DNA-unwinding enzyme. In this study, we have expressed and isolated the mouse Mcm2, 4,6,7 proteins from insect cells and characterized various mutant Mcm4,6,7 complexes in which the conserved ATPase motifs of the Mcm4 and Mcm6 proteins were mutated. The activities associated with such preparations demonstrated that the DNA helicase activity is intrinsically associated with the Mcm4,6,7 complex. Biochemical analyses of these mutant Mcm4,6,7 complexes indicated that the ATP binding activity of the Mcm6 protein in the complex is critical for DNA helicase activity and that the Mcm4 protein may play a role in the single-stranded DNA binding activity of the complex. The results also indicated that the two activities of DNA helicase and single-stranded DNA binding can be separated.
| S-EPMC84885 | biostudies-literature
Project description:BackgroundThe DEAD-box helicases are required mostly in all aspects of RNA and DNA metabolism and they play a significant role in various abiotic stresses, including salinity. The p68 is an important member of the DEAD-box proteins family and, in animal system, it is involved in RNA metabolism including pre-RNA processing and splicing. In plant system, it has not been well characterized. Here we report the cloning and characterization of p68 from pea (Pisum sativum) and its novel function in salinity stress tolerance in plant.ResultsThe pea p68 protein self-interacts and is localized in the cytosol as well as the surrounding of cell nucleus. The transcript of pea p68 is upregulated in response to high salinity stress in pea. Overexpression of p68 driven by constitutive cauliflower mosaic virus-35S promoter in tobacco transgenic plants confers enhanced tolerances to salinity stress by improving the growth, photosynthesis and antioxidant machinery. Under stress treatment, pea p68 overexpressing tobacco accumulated higher K+ and lower Na+ level than the wild-type plants. Reactive oxygen species (ROS) accumulation was remarkably regulated by the overexpression of pea p68 under salinity stress conditions, as shown from TBARS content, electrolyte leakage, hydrogen peroxide accumulation and 8-OHdG content and antioxidant enzyme activities.ConclusionsTo the best of our knowledge this is the first direct report, which provides the novel function of pea p68 helicase in salinity stress tolerance. The results suggest that p68 can also be exploited for engineering abiotic stress tolerance in crop plants of economic importance.
| S-EPMC4039504 | biostudies-literature
Project description:The ability to modify RNA secondary structure is crucial for numerous cellular processes. We have characterized two RNA helicase genes, crhB and crhC, which are differentially expressed in the cyanobacterium Anabaena sp. strain PCC 7120. crhC transcription is limited specifically to cold shock conditions while crhB is expressed under a variety of conditions, including enhanced expression in the cold. This implies that both RNA helicases are involved in the cold acclimation process in cyanobacteria; however, they presumably perform different roles in this adaptation. Although both CrhB and CrhC belong to the DEAD box subfamily of RNA helicases, CrhC encodes a novel RNA helicase, as the highly conserved SAT motif is modified to FAT. This alteration may affect CrhC function and its association with specific RNA targets and/or accessory proteins, interactions required for cold acclimation. Primer extension and analysis of the 5' untranslated region of crhC revealed the transcriptional start site, as well as a number of putative cold shock-responsive elements. The potential role(s) performed by RNA helicases in the acclimation of cyanobacteria to cold shock is discussed.
| S-EPMC93569 | biostudies-literature
Project description:We have shown previously that CrhC is a unique member of the DEAD-box family of RNA helicases whose expression occurs specifically under conditions of cold stress. Here we show that recombinant His-tagged CrhC, purified from Escherichia coli, is an ATP-independent RNA binding protein possessing RNA-dependent ATPase activity which is stimulated most efficiently by rRNA and polysome preparations. RNA strand displacement assays indicate that CrhC possesses RNA unwinding activity that is adenosine nucleotide specific. Unwinding of partially duplexed RNA proceeds in the 5'-->3' but not the 3'-->5' direction using standard assay conditions. Immunoprecipitation and far-western analysis indicate that CrhC is a component of a multisubunit complex, interacting specifically with a 37 kDa polypeptide. We propose that CrhC unwinds cold-stabilized secondary structure in the 5'-UTR of RNA during cold stress.
| S-EPMC110790 | biostudies-literature
Project description:To understand physiological mechanisms of cold acclimation in pea, we performed a transcriptomique analysis in order to compare the response to LT treatment in two varieties, one being cold tolerant (Champagne) and the other cold sensitive (Terese).
2010-04-01 | GSE19209 | GEO
Project description:Imparting tolerance to abiotic stresses is of global importance as they inflict significant yield losses in field as well as in vegetable crops. Transcriptional activators, including helicases are identified to play a pivotal role in stress mitigation. Helicases, also known as molecular motors, are involved in myriad cellular processes that impart intrinsic tolerance to abiotic stresses in plants. Our study demonstrates the potential of a Pea DNA Helicase 45 (PDH45), in combating multiple abiotic stresses in chili. We harnessed Agrobacterium-mediated in planta transformation strategy for the generation of stable, single copy transgenic events. Precise molecular detection of the transgenes by sqRT-PCR coupled with genomic Southern analysis revealed variation in the integration of PDH45 at distinct loci in independent transgenic events. Characterization of five promising transgenic events showed both improved response to an array of simulated abiotic stresses and enhanced expression of several stress-responsive genes. While survival and recovery of transgenic events were significantly higher under gradual moisture stress conditions, under imposition of moderate stress, the transgenic events exhibited invigorated growth and productivity with concomitant improvement in water use efficiency (WUE). Thus, our study, unequivocally demonstrated the cardinal role of PDH45 in alleviating multiple abiotic stresses in chili.
| S-EPMC5459802 | biostudies-literature