| Plants are sessile organisms and hence cannot escape unfavorable environmental conditions within their life cycle, such as high salinity, drought, waterlog, temperature stress, various ion stresses and so on. To deal with the abiotic stresses, plants have developed a series of coordinated responses involving a complex variety of tolerance mechanisms that are activated by the expression of relevant genes.RNA helicases refer to enzymes that use energy derived from the hydrolysis of a nucleotide triphosphate to unwind double-stranded RNAs and modify RNA structure. As RNA molecules are prone to forming stable nonfunctional secondary structures, their proper function requires RNA chaperones. RNA helicases are prominent candidates for RNA chaperones because they can actively disrupt misfolded RNA structures to ensure correct folding. Helicases belong to a class of molecular motor proteins in yeast, animals and plants. RNA helicases are proved to be involved in every step of RNA metabolism, including nuclear transcription, pre-mRNA splicing, translation, RNA decay, ribosome biogenesis and assembly, nucleocytoplasmic transport, embryogenesis, cell division and differentiation. Because of their multiple functions, RNA helicases have been the research hotpot in life sciences.Several studies have shown the majority of RNA helicases is involved in plant growth and development and diverse physiological processes. In 2001, Seki et al. reported firstly that an Arabidopsis helicase gene AB050574 was induced by cold treatment, suggesting that helicases might play a role in plant stress response. Recently, an Arabidopsis DEAD box RNA helicase LOS4 localized in the cytoplasm and enriched at the nuclear rim was shown to be essential for mRNA export and important for development and stress responses in Arabidopsis. Another two DEAD box RNA helicases, STRS1 and STRS2, were shown to improve Arabidopsis responses to multiple abiotic stresses as negative regulators, such as salt, osmotic stress, heat stress and ABA. In response to abiotic stress, RNA helicase have been proved to regulate downstream genes expression on transcription, post-transcription and translation level. In Arabidopsis, RNA helicase is the biggest family, with many members and various functions. Nevertheless, the relationships between DExD/H-box RNA helicase and environmental stress and development are still largely unknown in Arabidopsis.Two DExD/H-box RNA helicases have been isolated and characterized from the Arabidopsis genome, which were designated as AtHELPS (AT3G46960) and AtRDEM (AT5G47010). Their expression pattern and function are analyzed in detail.(1) Arabidopsis RNA helicase AtHELPS possesses eight conserved motifs, I, Ia, Ib, II, III, IV, V and VI in N terminal; these characterizations indicate that AtHELPS protein has hallmarks of DExD/H-box RNA helicase.(2) Transient analysis with onion epidermal cells and stable analysis with AtHELPS-GFP transgenic lines by Confocal Microscopy revealed that AtHELPS is unexclusively localized in the whole cell, but higher expression level is detected in plasma membrane and nucleus than in cytoplasm.(3) By quantitative real-time reverse transcription-PCR (qRT-PCR) and promoter:β-glucouronidase (GUS) fusions analysis, AtHELPS is shown to mainly express in vascular tissues of leaves and roots in young seedlings. The expression of AtHELPS could be regulated by several stresses like salt, low K~+ and low temperature stress, especially by low K~+ stress. Moreover, the expression level of AtHELPS could be induced by zeatin but not by other hormones.(4) GO analysis based on the microarray data indicated that around half amount of genes with different expression patterns encode the protein kinase and transcription factors such as bHLH, WRKY, MYB etc. Among those differentially expressed genes, a certain amount of which encodes ion binding proteins, ion channel proteins and transporters. These data suggest that AtHELPS might participate in the abiotic stress responses and ion transportation.(5) AtHELPS regulated Arabidopsis responses and tolerances to salt stress. Our results indicate that helps mutant, OE6 and WT plants have different sensitivity to salt stress. In our experiments, we discovered both seedling and adult of helps mutant and OE6 plants showed no morphological or developmental differences compared to WT when grown under normal conditions. But the results suggest that the helps mutant plants are more tolerant to salt stress, whereas the OE6 plants are more sensitive to salt stress. qRT-PCR analysis showed that the expression levels of WRKY25 and WRKY33 were higher in the OE6 than those in wild-type and helps mutant plants under salt stress condition, but the expression of SOS family genes were not affected. We thus suggest that the DEVH box RNA helicase AtHELPS might be involved in the regulation of expression of WRKY25 and WRKY33 under salt stress condition. We also applied noninvasive ion-selective microelectrode ion flux measurements to clarify genotype differences of Na+ flux profiles from root meristem zones of Arabidopsis. The net Na+ flux in helps mutant, wild type and OE6 seedlings showed no difference under salt stress condition compared to normal condition, suggesting that AtHELPS might not be involved in regulating Na+ uptake and accumulation in Arabidopsis roots, but possibly involved in other ion uptake, assimilation and transportation.(6) AtHELPS regulated Arabidopsis responses and tolerances to low K~+ stress. Our results indicate that helps mutant, OE6 and WT plants have different sensitivity to low K~+ stress. The seed germination percentage and seedling fresh weight of the helps mutants were higher than those of wide-type and OE6 plants in the low K~+ condition, whereas no differences were observed among the three genotypes under normal conditions. Interestingly, qRT-PCR analysis showed that the expression of AKT1, CBL1/9, and CIPK23 in the helps mutants were consistently higher than those in WT and OE6 plants after low K~+ treatment. We thus suggest that the DEVH box RNA helicase AtHELPS might be involved in the regulation of the AKT1-mediated and CBL/CIPK-regulated K~+ uptake pathway under the low K~+ stress condition. We also applied noninvasive ion-selective microelectrode ion flux measurements to clarify genotype differences of K~+ flux profiles from root meristem zones of Arabidopsis. The net K~+-induced influx in helps mutants was higher than that of wild type and OE6 seedlings when they were exposed to potassium deprivation, suggesting that AtHELPS might be involved in regulating K~+ uptake in Arabidopsis roots via high-affinity transporters like AKT1.(7) AtRDEM plays an important role in the growth and development of Arabidopsis. OEAtRDEM exhibited obvious phenotypic alterations, such as lacked an apical dominance, shorter stem, flower abnormality, silique cluster and more shoot branching. These data reveal that AtRDEM plays an important role in plant development in Arabidopsis. By microarray analysis, many biological processes are affected by overexpressing AtRDEM, such as developmental progress, hormone metabolic pathway, response to abiotic or biotic stimulus, signal transduction etc. Among these changed genes, the expressions of genes involved in development, hormone responsive and metabolic are changed significantly in AtRDEM overexpression plants compared to WT plants, suggesting AtRDEM mediated Arabidopsis growth and development, most probably through hormone. In addition, we found that the expression level of Carotenoid Cleavage Dioxygenase 7 (CCD7) and Carotenoid Cleavage Dioxygenase 8 (CCD8) were enhanced in AtRDEM overexpression lines, but the cytochrome P450 monooxygenase located downstream of the strigolactone biosynthesis pathway was reduced. These results suggested that the novel hormone strigolactone might not accumulate and move acropetally to inhibit axillary bud outgrowth. Together, we speculate that the AtHELPS may be involved in regulating the strigolactone biosynthesis and signal transduction pathway.
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