Genetic Variability in Recovery Growth and Synthesis of Stress Proteins in Response to Polyethylene Glycol and Salt Stress in Finger Millet

Marked differences were found among 28 finger millet genotypes(Eleusine coracana Gaertn.) in acquired tolerance to osmoticstress as assessed by the recovery of root growth from severestress [-1·2 MPa polyethylene glycol, (PEG) or 400 mMNaCl]. However, these differences in tolerance were observedonly when the seedlings were subjected to a preceding mild inductionstress (-0·6 MPa PEG or 200 mM NaCl). In two contrastinggenotypes, synthesis of stress-induced proteins was studied.Proteins with apparent molecular weight of 70-72, 52, 37, 34and 23 kDa were synthesized in the highly responsive genotype(GE 415) and poorly responsive (VL 481) genotype following amild induction stress (200 mM NaCl). However, GE-415 synthesizeda 54 kDa protein that was not observed in VL-481. Addition ofabscisic acid (ABA) to the induction medium containing 200 mMNaCl enhanced the acquired tolerance of finger millet seedlingsover those without ABA in association with the appearance ofseveral ABA-responsive proteins. GE-415 required much less ABAthan VL-481 to obtain the same response. With 10 µM ABA+ 200 mM, A NaCl induction stress, GE-415 had significantlyhigher endogenous ABA. In association with higher levels ofABA, GE-415 had greater recovery root growth following severestress from 600 mM NaCl. Pretreatment with 10 µM ABA +200 mM NaCl induced several proteins with apparent molecularweights of 70-72, 54, 45, 36, 29 and 21 kDa in both genotypes.Qualitatively, GE-415 synthesized a unique 23-24 kDa proteinand quantitatively there was significantly more of the 21 kDaprotein in GE-415 compared to VL-481. The results indicate thatthe synthesis of stress proteins is correlated with the observedvariation in acquired tolerance of the two genotypes.Copyright1995, 1999 Academic Press Eleusine coracana Gaertn., salinity, polyethylene glycol, stress proteins, ABA, ABA-responsive proteins, finger millet seedlings     

Construction and application of EST library from Setaria italica in response to dehydration stress

Foxtail millet is a gramineous crop with low water requirement. Despite its high water use efficiency, less attention has been paid to the molecular genetics of foxtail millet. This article reports the construction of subtracted cDNA libraries from foxtail millet seedlings under dehydration stress and the expression profile analysis of 1947 UniESTs from the subtracted cDNA libraries by a cDNA microarray. The results showed that 95 and 57 ESTs were upregulated by dehydration stress, respectively, in roots and shoots of seedlings and that 10 and 27 ESTs were downregulated, respectively, in roots and shoots. The expression profile analysis showed that genes induced in foxtail millet roots were different from those in shoots during dehydration stress and that the early response to dehydration stress in foxtail millet roots was the activation of the glycolysis metabolism. Moreover, protein degradation pathway may also play a pivotal role in drought-tolerant responses of foxtail millet. Finally, Northern blot analysis validated well the cDNA microarray data.     

A Full-Length Dof1 Transcription Factor of Finger Millet and its Response to a Circadian Cycle

DOF1 (DNA binding with one finger) plays an important role in regulating C/N metabolism in cereals. In order to validate its role in the regulation of nitrogen use efficiency (NUE) and photosynthetic efficiency in finger millet, 5′–3′ RACE PCR was performed to obtain and characterize full-length Dof1 genes of high and low grain protein finger millet genotypes. The full-length DOF1 ORFs were both 1,284 nt long and were 98.8 % similar over 427 amino acids containing the characteristic Dof domain. Comparison of both the EcDof1 protein sequences with the Dof1 of other cereals revealed high sequence similarity to the Dof1 of rice. Southern hybridization carried out using the probe developed from the region encoding the highly variable C-terminal region of EcDof1 showed the presence of four copies of the DOF1 gene in finger millet, which might explain the high NUE and photosynthetic performance of finger millet. Since the genes involved in C/N metabolism are regulated diurnally and play crucial roles in determining grain protein content during grain filling, the diurnal expression of EcDOF1 was assessed in two finger millet genotypes (GE 3885 and GE 1437) with differing grain protein content (13.8 % and 6.15 % respectively). It was found that EcDOF1 exhibited diurnal regulation and peak differential pattern expression with early phasing in GE3885 and late phasing in GE1437. Differential expression of DOF1 might alter the regulation of genes involved in C/N metabolism affecting grain protein composition of finger millet genotypes.     

Biological seed priming mitigates the effects of water stress in sunflower seedlings

The sunflower (Helianthus annuus L. cv. PAC 36) seedlings were inoculated with plant growth promoting rhizobacteria (PGPR), viz. Azotobacter chroococcum (A+), Bacillus polymyxa (B+), separately and in combination of the two (AB+). Relative water content and seedling growth were maximum in AB+ seedlings under control. Water stress significantly decreased the RWC, growth and dry mass of non-inoculated seedlings. However, inoculated seedlings maintained higher growth even under water stress. Pigments and protein contents decreased under water stress, but higher amount of the same was observed in stressed AB+ seedlings. Enhanced activity of nitrate reductase was recorded in AB+ seedlings with maximum in control. Water stress significantly decreased the nitrate reductase activity. A significant increase in the activity of superoxide dismutase (SOD) in leaves was recorded under water stress except in B+ with maximum increase in non-inoculated seedlings. Catalase (CAT) activity decreased in stressed non-inoculated seedlings while increased in the leaves of A+ and AB+ seedlings. Almost similar trends were recorded for both leaves and cotyledons. PGPR improved the water status in stressed seedlings and thereby physiological and biochemical parameters and thus ameliorated the severe effects of water stress.     

Inhibition of ABA-mediated Responses by Dithiothreitol in Plants

In plants, abscisic acid (ABA)-mediated responses during abiotic stress, growth, and development have been well studied. Many chemicals which modulate ABA responses have been identified. In this study, we report that dithiothreitol (DTT), an inducer of endoplasmic reticulum (ER) stress, can overcome ABA-mediated responses in plants. In rice seedlings, combined treatment of ABA and DTT increased shoot growth compared to ABA alone. The phenotype correlated with the expression pattern of ABA and ER stress-responsive genes. In finger millet, increase in root growth was observed in combined treatment, compared to ABA treatment. Experiments using dimethyl sulfoxide indicated that the phenotype observed was specific to DTT. Priming of germinated rice seeds with DTT followed by salinity stress indicated that DTT can mask the ABA effect. In ABA bioassay using cotton petioles, an increase in intact petioles in combined treatment of ABA and DTT was observed compared to ABA treatment. The expression of OsWRKY48, an ABA-responsive gene, was down-regulated in combined treatment, indicating that the target of DTT-induced ER stress is upstream of OsWRKY48 in the ABA signaling pathway. The study demonstrated that DTT-induced ER stress can be a potential mechanism to regulate ABA-mediated responses in plants.

     

Abscisic Acid induces anaerobiosis tolerance in corn

Flooding is a frequently occurring environmental stress that can severely affect plant growth. This study shows that treatment of corn (Zea mays L.) seedlings with abscisic acid (ABA) increases their tolerance to anoxia 10-fold over untreated seedlings and twofold over seedlings treated with water. Corn seedlings stressed anoxically for 1 day showed only 8% survival when planted in vermiculite. Pretreatment of root tips with 100 micromolar ABA or water for 24 hours before the 1 day anoxic stress increased the anoxic survivability of seedlings to 87% and 47%, respectively. Cycloheximide (5 milligrams per liter), added together with ABA, reduced the seedling survival rate, indicating that the induction of anoxic tolerance in corn by ABA was partly a result of the synthesis of new proteins. ABA treatment induced a threefold increase in alcohol dehydrogenase enzyme activity in corn roots. However, after 24 h of anoxia, alcohol dehydrogenase enzyme activity between the ABA-pretreated and non-pretreated corn roots was not significantly different. The results indicated that ABA played an important role in inducing anoxic tolerance in corn and that the induced tolerance was probably mediated by an increase in alcohol dehydrogenase enzyme activity before the anoxic stress.     

Heat Shock Proteins of Sorghum [Sorghum bicolor (L.) Moench] and Pearl Millet [Pennisetum glaucum (L.) R.Br.] Cultivars with Differing Heat Tolerance at Seedling Establishment Stage

The production of heat shock proteins was compared in sorghumand pearl millet genotypes differing in seedling establishmentcharacteristics under heat stress. Two major heat shock proteins(hsps) of apparent mol. wt. 65 kD and 62 kD were seen in allthe genotypes of sorghum tested when the incubation temperatureof the 40 h seedlings was altered from 35 ?C to 45 ?C for 2h. Under identical conditions, pearl millet genotypes showedmore hsps and the apparent mol. wt. of these ranged from 30–70kD. The hsp bands were more prominent in whole seedlings androots as compared to plumules. Differences in the productionof hsps were seen in sorghum and pearl millet genotypes withcontrasting heat tolerance at seedling establishment stage butthe significance of these needs to be studied further. Key words: Heat shock proteins, sorghum, genotypic differences     

Simple sequence repeat (SSR) analysis in relation to calcium transport and signaling genes reveals transferability among grasses and a conserved behavior within finger millet genotypes

Being an excellent source of calcium, finger millet crop has nutraceutical importance. Mineral accumulation, being a polygenic trait, becomes essential to target potential candidate genes directly or indirectly involved in the regulation of calcium transport and signaling in cereals and might have influence on grain calcium accumulation. In view of this, genic microsatellite markers were developed from the coding and non-coding sequences of calcium signaling and transport genes viz. calcium transporters (channels; ATPases and antiporters), calcium-binding proteins and calcium-regulated protein kinases available in rice and sorghum. In total, 146 genic "simple sequence repeat" (SSR) primers were designed and evaluated for cross-transferability across a panel of nine grass species including finger millet. The average transferability of genic SSR markers from sorghum to other grasses was highest (73.2 %) followed by rice (63.4 %) with an overall average of 68.3 % which establishes the importance of these major crops as a useful resource of genomic information for minor crops. The transfer rate of SSR markers was also correlated with the phylogenetic relationship (or genetic relatedness) of the species. Primers with successful amplification in finger millet were further used to screen for polymorphism across a set of high and low calcium containing genotypes. The results reveal a conserved behavior across the finger millet genotypes indicating that the mineral transport and the storage machinery largely remain conserved in plants and even SSR variations in them remain suppressed during the course of evolution. Single nucleotide polymorphism and differential expression patterns of candidate genes, therefore, might be a plausible reason to explain variations in grain calcium contents among finger millet genotypes.     

Changes in physiology and biochemistry of mottle streak virus infected finger millet plants

A significant reduction in the growth parameters viz., plant height, number of tillers, number of productive tillers, leaves, leaf area, 1000 grain weight and grain yield were observed in the mottle streak virus infected finger millet plants compared to healthy finger millet plants. The germination and vigour of seedlings from the seeds of infected plants were reduced. Physiological changes in finger millet as a result of virus infection were investigated. The chlorophyll pigments ‘a’ and ‘b’ as well as total chlorophyll were reduced due to mottle streak infection. The virus infection led to increased total sugar, starch, soluble protein and phenol contents. The mineral metabolism of infected plants showed a reduction in nitrogen, phosphorus, potassium, magnesium, calcium and iron.     

Comparative analysis of solid-state bioprocessing and enzymatic treatment of finger millet for mobilization of bound phenolics

The present work investigates the probable bioprocessing technique to mobilize the bound phenolics naturally found in finger millet cell wall for enriching it with dietary antioxidants. Comparative study was performed between the exogenous enzymatic treatment and solid-state fermentation of grain (SSF) with a food grade organism Rhizopus oryzae. SSF results indicated that at the 6th day of incubation, total phenolic content (18.64 mg gallic acid equivalent/gds) and antioxidant property (DPPH radical scavenging activity of 39.03 %, metal chelating ability of 54 % and better reducing power) of finger millet were drastically enhanced when fermented with GRAS filamentous fungi. During the enzymatic bioprocessing, most of the phenolics released during the hydrolysis, leached out into the liquid portion rather than retaining them within the millet grain, resulting in overall loss of dietary antioxidant. The present study establishes the most effective strategy to enrich the finger millet with phenolic antioxidants.     

谷子幼苗对土壤铅、铬的生长响应及吸收积累的差异性

采用盆栽土培试验,研究了谷子幼苗对土壤中不同含量铅（Pb）、铬（Cr）的生长响应和吸收积累的差异性。结果表明,在所试浓度（50～800 mg·kg^-1）范围内,Pb、Cr在谷子幼苗地上部和地下部的积累量存在较大差异,幼苗重金属的吸收富集和转运系数均为Pb大于Cr、Pb、Cr胁迫对幼苗生物量的影响表现为低浓度的促进和高浓度的抑制作用,但Cr对生物量的影响比Pb更强。相同处理条件下,幼苗茎叶中可溶性蛋白质、DNA含量和增色效应对Pb、Cr的响应也有明显差别,Cr对幼苗的生理毒性和DNA损伤效应的作用强度大于Pb。     

Expression analysis of stress responsive pathway genes linked to drought hardiness in an adapted crop, finger millet (Eleusine coracana)

Drought is one of the major stresses limiting plant growth and productivity. Drought tolerance is regulated by multiple plant traits and examining the tolerance mechanisms from adapted species would assist in identification of novel pathways and superior genes. Since cellular tolerance is one of the major traits in drought acclimation we made in this study, an attempt to prospect candidate genes associated with the trait in drought hardy crop plant, finger millet (Eleusine coracana (L.) Gaertn). A novel gravimetric approach was employed to simulate field level drought stress for examining stress responsiveness of a few selected genes implicated in different stress response pathways. Gene expression was studied initially by e-northern analysis, and subsequently in leaf tissues experiencing different levels of drought stress by semi-quantitative and quantitative RT-PCR. A few stress responsive genes identified include metallothionein, farnesylated protein ATFP6, protein phosphatase 2A, RISBZ4 and farnesyl pyrophosphate synthase which probably have crucial roles in imparting hardiness to finger millet. Taken together the results suggest that multiple cellular tolerance pathways operate in a coordinated manner in drought tolerant crops.     

A novel rice C2H2-type zinc finger protein lacking DLN-box/EAR-motif plays a role in salt tolerance

A cDNA for the gene ZFP182, encoding a C2H2-type zinc finger protein, was cloned from rice by RT-PCR. ZFP182 codes an 18.2 kDa protein with two C2H2-type zinc finger motifs, one nuclear localization signal and one Leu-rich domain. The DLN-box/EAR-motif, which exists in most of plant C2H2-type zinc finger proteins, does not exist in ZFP182. The expression analysis showed that ZFP182 gene was constitutively expressed in leaves, culms, roots and spikes at the adult rice plants, and markedly induced in the seedlings by cold (4 °C), 150 mM NaCl and 0.1 mM ABA treatments. The approximate 1.4 kb promoter region of ZFP182 gene was fused into GUS reporter gene and transformed into tobacco. The histochemical analysis revealed that GUS expression could not be detected in transformed tobacco seedlings under normal conditions, but strongly observed in tobacco leaf discs and the vascular tissue of roots treated with NaCl or KCl. Expression of ZFP182 in transgenic tobacco and overexpression in rice increased plant tolerance to salt stress. These results demonstrated that ZFP182 might be involved in plant responses to salt stress.