Sagadevan G. Mundree

An aldose reductase homolog from the resurrection plant Xerophyta viscosa Baker

Abstract. An aldose reductase homologue (ALDRXV4) was cloned from the resurrection plant Xerophyta viscosa Baker using complementation by functional sufficiency in Escherichia coli. A cDNA library constructed from X. viscosa leaves dehydrated to 85%, 37% and 5% relative water contents (RWC) was converted into an infective phagemid library. Escherichia coli (srl::Tn10) cells transformed with ds-pBluescript phagemids were selected on minimal medium plates supplemented with 1 mM isopropyl β-d-thiogalactopyranoside and 1.25 M sorbitol. Nine cDNA clones that conferred tolerance to the osmotically stressed E. coli cells were selected. The phagemid from one clone contained the ALDRXV4 insert. The E. coli cells expressing ALDRXV4 were capable of tolerating the osmotic stress, whereas control cultures were not. The ALDRXV4 insert contained an open reading frame that can code for 319 amino acids, and the predicted protein had a calculated Mr of 35,667. Amino acid sequence comparisons revealed significant similarity to several aldose reductases, with the highest similarity to aldose reductase proteins from Hordeum vulgare, Bromus inermis and Avena fatua, in the order of 66%, 65% and 65% respectively. Northern blot analysis revealed that ALDRXV4 was expressed only under dehydration conditions in X. viscosa leaves. Western blot analysis detected a protein of 36 kDa under dehydration conditions only. Aldose reductase activity levels in X. viscosa leaves increased as the leaf RWC decreased, whereas there was no significant change in aldose reductase activity in Sporobolus stafianus as the leaf RWC decreased.

An ultrastructural study using anhydrous fixation of Eragrostis nindensis, a resurrection grass with both desiccation-tolerant and -sensitive tissues

The ability of tissues to survive desiccation is common in seeds but rare in vegetative tissues. In this study the ultrastructure of hydrated and dehydrated tissues were examined at different stages of the life cycle of the resurrection grass, Eragrostis nindensis Ficalho & Hiern. Conventional fixation techniques are unsuitable for dry tissues as rehydration occurs during fixation in aqueous fixatives. Thus a cryofixation and freeze-substitution method was developed. As a result of the improved fixation methods, it was possible to identify the stage and nature of the damage in the desiccation-sensitive tissues. E. nindensis has desiccation-tolerant orthodox seeds, but the young seedlings are not tolerant to extreme water loss. However, like the seeds, most of the leaves of the adult plant are tolerant to desiccation (only the oldest outermost leaf on a tiller are not). Desiccation-induced damage in these outer leaves was observed in the later stage of dehydration, dominated by the appearance of abundant cell wall fractures (1 wall fracture per 50 μm2). Unlike the outer leaves, the leaves of seedlings appeared similar to those of the hydrated ones upon desiccation. Irreparable damage occurred on rehydration of these tissues possibly as a result of the absence of protection mechanisms observed during desiccation of the inner desiccation-tolerant leaves of the mature plants. The mesophyll tissues of these leaves become compact with extensive cell wall folding on drying. The bundle sheath cells maintained their shape with desiccation but became packed with small vacuoles.

Trehalose Accumulation Triggers Autophagy during Plant Desiccation

Global climate change, increasingly erratic weather and a burgeoning global population are significant threats to the sustainability of future crop production. There is an urgent need for the development of robust measures that enable crops to withstand the uncertainty of climate change whilst still producing maximum yields. Resurrection plants possess the unique ability to withstand desiccation for prolonged periods, can be restored upon watering and represent great potential for the development of stress tolerant crops. Here, we describe the remarkable stress characteristics of Tripogon loliiformis, an uncharacterised resurrection grass and close relative of the economically important cereals, rice, sorghum, and maize. We show that T. loliiformis survives extreme environmental stress by implementing autophagy to prevent Programmed Cell Death. Notably, we identified a novel role for trehalose in the regulation of autophagy in T.loliiformis. Transcriptome, Gas Chromatography Mass Spectrometry, immunoblotting and confocal microscopy analyses directly linked the accumulation of trehalose with the onset of autophagy in dehydrating and desiccated T. loliiformis shoots. These results were supported in vitro with the observation of autophagosomes in trehalose treated T. loliiformis leaves; autophagosomes were not detected in untreated samples. Presumably, once induced, autophagy promotes desiccation tolerance in T.loliiformis, by removal of cellular toxins to suppress programmed cell death and the recycling of nutrients to delay the onset of senescence. These findings illustrate how resurrection plants manipulate sugar metabolism to promote desiccation tolerance and may provide candidate genes that are potentially useful for the development of stress tolerant crops.

Some physiological comparisons between the resurrection grass, Eragrostis nindensis, and the related desiccation-sensitive species, E. curvula

Both the poikilochlorophyllous resurrection grass, Eragrostisnindensis, and the desiccation sensitive species, E.curvula, dehydrate to a relative water content (RWC) of less than5% in two weeks. On rewatering, most E. nindensisleaves (except the older, outer ones) rehydrate and resume normal metabolicactivity within a few days, whereas E. curvula does notrecover. There is a controlled loss of photosynthetic pigments, paralleled witha gradual shutdown in gas exchange during dehydration of E.nindensis. On rehydration respiration resumes almost immediately butphotosynthesis only restarts at 70% RWC by which time chlorophyll hasbeen resynthesised and anthocyanin content reduced. In contrast, photosyntheticactivity in E. curvula is maintained down to 40%RWC, after which further drying results in a sudden breakdown of thephotosynthetic system and its pigments. At this point, electrolyte leakage andincreases FV/FM decreases such that belowca. 40% RWC, metabolism is irreparably damaged.Interestingly, the older outer leaf in most tillers of E.nindensis does not rehydrate. These leaves show signs of membranedamage and curl in an irregular manner similar to those of E.curvula during dehydration.

Some Physiological and Molecular Insights into the Mechanisms of Desiccation Tolerance in the Resurrection Plant Xerophyta viscosa Baker

Few organisms are able to survive desiccation to the air-dried state. Nevertheless, there are a small number of species from every major class of plants (with the exception of gymnosperms) that exhibit this characteristic (Bewley and Oliver, 1992; Oliver and Bewley, 1997). Many of these “resurrection” plants (Gaff, 1971) are being studied in an attempt to understand the mechanisms which enable vegetative tissues to withstand desiccation, with the ultimate aim of identifying characteristics which may be used to produce crops with improved tolerance to osmotic stress.

Molecular characterization of XVT8, a stress-responsive gene from the resurrection plant Xerophyta viscosa Baker.

Xerophyta viscosa (Baker) is a monocotyledonousresurrection plant that is capable of tolerating extremes of desiccation. Uponrewatering, it rehydrates completely, assuming its full physiologicalactivities. Studies on changes in gene expression associated with dehydrationstress tolerance were conducted. A cDNA library was constructed from mRNAisolated from dehydrated X. viscosa leaves [85%,37% and 5% relative water content (RWC)].XVT8 represents one of 30 randomly selected clones thatwere differentially expressed when X. viscosa wasdehydrated. Sequence analysis of XVT8 revealed thatXVT8 exhibited 45% and 43% identity todehydrin proteins from Arabidopsis thaliana andPisum sativum respectively, at the amino acid level.XVT8 encodes a glycine -rich protein (27 kDa)which is largely hydrophilic and contains a hydrophobic segment at theC-terminus. Southern blot analysis confirmed the presence ofXVT8 in the X. viscosa genome.XVT8 transcripts accumulated in X.viscosa plants that were exposed to heat, low temperature anddehydration stresses, and to exogenous abscisic acid and ethylene. Theseresultsprovide direct evidence for the heat, low temperature, dehydration, abscisicacid and ethylene -dependent regulation of the XVT8 geneinX. viscosa.

A footprint of desiccation tolerance in the genome of Xerophyta viscosa

Desiccation tolerance is common in seeds and various other organisms, but only a few angiosperm species possess vegetative desiccation tolerance. These ‘resurrection species’ may serve as ideal models for the ultimate design of crops with enhanced drought tolerance. To understand the molecular and genetic mechanisms enabling vegetative desiccation tolerance, we produced a high-quality whole-genome sequence for the resurrection plant Xerophyta viscosa and assessed transcriptome changes during its dehydration. Data revealed induction of transcripts typically associated with desiccation tolerance in seeds and involvement of orthologues of ABI3 and ABI5, both key regulators of seed maturation. Dehydration resulted in both increased, but predominantly reduced, transcript abundance of genomic ‘clusters of desiccation-associated genes’ (CoDAGs), reflecting the cessation of growth that allows for the expression of desiccation tolerance. Vegetative desiccation tolerance in X. viscosa was found to be uncoupled from drought-induced senescence. We provide strong support for the hypothesis that vegetative desiccation tolerance arose by redirection of genetic information from desiccation-tolerant seeds.

Biofuels from food processing wastes

Food processing industry generates substantial high organic wastes along with high energy uses. The recovery of food processing wastes as renewable energy sources represents a sustainable option for the substitution of fossil energy, contributing to the transition of food sector towards a low-carbon economy. This article reviews the latest research progress on biofuel production using food processing wastes. While extensive work on laboratory and pilot-scale biosystems for energy production has been reported, this work presents a review of advances in metabolic pathways, key technical issues and bioengineering outcomes in biofuel production from food processing wastes. Research challenges and further prospects associated with the knowledge advances and technology development of biofuel production are discussed.

Photochemical and Antioxidant Responses in the Leaves of Xerophyta viscosa Baker and Digitaria sanguinalis L. under Water Deficit

In this study, photochemical and antioxidant responses of the monocotyledonous resurrection plant Xerophyta viscosa Baker and the crab grass Digitaria sanguinalis L. under water deficit were investigated as a function of time. Water deficit was imposed by withholding irrigation for 21 d. Gas exchange and chlorophyll a fluorescence analyses indicated that the dehydration treatment caused photoinhibition in both species. The reduction in the photosynthesis rate in both species during water deficit probably contributed to the decline in the photochemical efficiency of PSII and electron transport rate. However, the stomatal conductance of both species did not change during treatment whereas the intercellular CO2 pressure increased after 10 d of water deficit treatment. These observations could be related to nonstomatal limitations. The increasing net transpiration rate of both species may have contributed to leaf cooling because of water limitations. Prolonged water deficit resulted in photosynthetic pigment chlorophyll (a + b) and carotenoids content loss in only D. sanguinalis. Both species especially D. sanguinalis had increased the level of anthocyanin after 15 d of treatment, possibly to prevent the damaging effect of photooxidation. The total SOD activity of D. sanguinalis was significantly different from X. viscosa during the treatment. The total peroxidase activity in D. sanguinalis was significantly higher than in X. viscosa. X. viscosa acclimated to water deficit with no ultimate apparent oxidative damage due to endogenous protective mechanisms of resurrection. In case of D. sanguinalis, water deficit induced considerable stress and possibly caused some oxidative damage, despite the upregulation of protection mechanisms.

Physiological basis of salt stress tolerance in rice expressing the antiapoptotic gene SfIAP

Programmed cell death-associated genes, especially antiapoptosis-related genes have been reported to confer tolerance to a wide range of biotic and abiotic stresses in dicotyledonous plants such as tobacco (Nicotiana tabacum L.) and tomato (Solanum lycopersicum L.). This is the first time the antiapoptotic gene SfIAP was transformed into a monocotyledonous representative: rice (Oryza sativa L.). Transgenic rice strains expressing SfIAP were generated by the Agrobacterium-mediated transformation method and rice embryogenic calli, and assessed for their ability to confer tolerance to salt stress at both the seedling and reproductive stages using a combination of molecular, agronomical, physiological and biochemical techniques. The results show that plants expressing SfIAP have higher salt tolerance levels in comparison to the wild-type and vector controls. By preventing cell death at the onset of salt stress and maintaining the cell membranes integrity, SfIAP transgenic rice plants can retain plant water status, ion homeostasis, photosynthetic efficiency and growth to combat salinity successfully.