Browsing by Author "Lindsey, George G"
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- ItemOpen AccessThe anti-fungal and anti-oxidant properties of polyphenols extracted from the resurrection plant, Myrothamnus flabellifolia(2008) Shibambo, Segopotjo Linah; Lindsey, George G; Brandt, Wolf FIn this study the effects of M. flabellifolia polyphenols on growth of S. cerevisiae yeast strains was investigated. This study showed that M flabellifolia polyphenols inhibited growth of both the wild type and the Δhsp 12 yeast strains largely by binding protein in the growth medium. A decreased specific growth rate, reduced maximum biomass, and prolonged lag phase were observed for both strains.
- ItemOpen AccessASP 53, a 53 kDa cupin-containing protein from Acacia erioloba seeds that protects proteins against thermal denaturation(2004) Mtwisha, Linda; Lindsey, George G; Brandt, Wolf F; Farrant, Jill MIncludes bibliographical references (leaves 103-111).
- ItemRestrictedASP53, a thermostable protein from Acacia erioloba seeds that protects target proteins against thermal denaturation.(CSIRO Publishing, 2007) Mtwisha, Linda; Farrant, Jill M; Brandt, Wolf; Hlongwane, Caswell; Lindsey, George GASP53, a 53 kDa heat soluble protein, was identified as the most abundant protein in the mature seeds of Acacia erioloba E.Mey. Immunocytochemistry showed that ASP53 was present in the vacuoles and cell walls of the axes and cotyledons of mature seeds and disappeared coincident with loss of desiccation tolerance. The sequence of the ASP53 transcript was determined and found to be homologous to the double cupin domain-containing vicilin class of seed storage proteins. Mature seeds survived heating to 60◦C and this may be facilitated by the presence of ASP53. Circular dichroism spectroscopy demonstrated that the protein displayed defined secondary structure, which was maintained even at high temperature. ASP53 was found to inhibit all three stages of protein thermal denaturation. ASP53 decreased the rate of loss of alcohol dehydrogenase activity at 55◦C, decreased the rate of temperature-dependent loss of secondary structure of haemoglobin and completely inhibited the temperature-dependent aggregation of egg white protein.
- ItemOpen AccessBiolistic-mediated transformation of Eragrostis Curvula with the HSP 12 gene(2003) Ncanana, Sandile Welcome; Farrant, Jill M; Lindsey, George G; Brandt, Wolf FEragrostis curvula is a desiccation sensitive monocotyledonous plant and an economically important forage grass in southern Africa. This species has a potential to be improved for drought and salt resistance among other important agronomic traits. In this study, E. curvula was used as a model system to explore the feasibility of producing pasture grasses with increased tolerance to water deficit. To date, no reports have been published on transformation of this species. This study reports the transformation of E. curvula with Saccharomyces cerevisiae Hsp 12 gene using biolistic-mediated transformation. Firstly, a tissue culture protocol was established for E. curvula that was suitable for transformation studies. Although this species has been previously regenerated in vitro using inflorescence tissues, this study established new protocols that utilize leaf and seed as source of material. The aim of which was to find the best regenerable tissue that could be used for transformation studies. Plant regeneration was achieved from shoot explants cultured on MS medium supplemented with either 0.5 mg/I thidiazuron (TDZ) or 0.5 mg/I 6-benzylaminopurine (BAP) through the process of direct organogenesis. It was found that TDZ was the most effective cytokinin. Plant regeneration was also achieved from callus induced from immature leaves on MS medium supplemented with 2 mg/I 2,4- 0,0.01 mg/I BAP and 6 % (w/v) sucrose. Histological experiments performed gave clear evidence that plant regeneration from callus was through a process of indirect organogenesis. The regeneration protocol was combined with an optimized biolistic mediated transformation protocol using the PDS-I00/He apparatus of which both shoot explants and callus were used as target tissues. In the molecular aspect of the study, the Hsp 12 gene was ligated in the Sac I restriction site of pCAMBIAUbeeQ vector. The successful cloning of the Hsp 12 gene was confirmed by PCR and restriction endonuclease digestion. The resultant vector pCAMBIAUbeeQ Hsp 12 was purified and subsequently used for biolistic transformation of E. curvula. The regenerable shoot explants and callus tissue were bombarded with DNA (pCAMBIAUBeeQHsp12) coated on gold particles. As a comparative study, Agrobacterium tumefaciens was transformed with pCAMBIAUBeeQHsp12 vector for subsequent transformation of Nicotiana tabacum. Transient expression of GUS gene in transformed E. curvula shoot explants was visualized 72 h after bombardment. Optimized conditions for expression of GUS gene were gold micro projectiles, 7 cm travel distance and helium pressure of 9100 kPa. Transformed tissues were cultured on the regeneration medium without antibiotic selection. Putative transformants were generated and the presence of Hsp 12 gene was verified by PCR and its expression at RNA level was confirmed by RT-PCR. The presence of Hsp 12 protein in the transformed plants was analyzed using SDS-PAGE and MALDI-TOF peptide mass spectrometry.
- ItemOpen AccessCharacterization of polyphenols in leaves of four desiccation tolerant plant families(2005) Dzobo, Kevin; Lindsey, George G; Farrant, Jill M; De Wet, JacquesPolyphenols in plants are known to act as antioxidants, antimicrobials, antifungal, photoreceptors, visual attractors and as light screens. In this study polyphenols in angiosperms found in southern Africa and called resurrection (desiccation tolerant) plants were studied. These plants are Myrothamnus flabellifolius, Xerophyta viscosa, Xerophyta humilis, Xerophyta schlecterii, Xerophyta villosa. Craterostigma wilmsii, Craterostigma plantagineum, Craterostigma pumilum and Eragrostis nindensis. These plants are able to tolerate water stress without undergoing permanent damage. During drying these plants are subjected to different stresses and one such stress is oxidative stress. It has been suggested that polyphenols function as stress protectants in plant cells by scavenging reactive oxygen species (ROS) produced during a period of oxidative stress. In this study the total phenolic content and the related antioxidant capacity of the plants leaf extracts were analysed.
- ItemRestrictedDesiccation-induced ultrastructural and biochemical changes in the leaves of the resurrection plant Myrothamnus flabellifolia(CSIRO Publishing, 2007) Moore, John P; Hearshaw, Meredith; Ravenscroft, Neil; Lindsey, George G; Farrant, Jill M; Brandt, Wolf FLight microscopy and low-temperature scanning electron microscopy were used to systematically compare the surface and internal ultrastructures of hydrated and desiccated leaves of the resurrection plant Myrothamnus flabellifolia (Welw.). This revealed that leaf tissue underwent considerable shrinkage and collapse on desiccation but was supported by a framework of vascular and sclerenchymous tissue, which is responsible for the fan-like shape of the leaves. In addition, the leaf ribs were covered with wax and an internal wax cuticle was observed. Biochemical analysis showed that the cyanidin 3-glucoside content increased on desiccation as did the trehalose and sucrose contents. Salt deposits were observed at the apices of desiccated leaves in the proximity of hydathode-like structures. We propose that this might regulate the leaf salt content since decreased intracellular cation concentration was observed in desiccated leaves. We believe that these unique adaptations contribute to the remarkable desiccation-tolerance properties of this plant.
- ItemOpen AccessEffect of heat stress of the cell wall of the yeast, Saccharomyces cerevisiae : phosphorylation of ribosomal protein S10-B brought about by enzymes of the glycolytic pathways(2005) Zondi, Nondumiso Busisiwe Ntombikhona; Lindsey, George GThe cell wall is a dynamic and elastic structure that provides osmotic and physical protection to the yeast cell (l). As such, it forms the immediate site of contact between the yeast cell and its environment (2), and is essential for maintaining cell integrity and shape. Cell growth and development demand that the cell wall is not rigid and unchangeable as the cell needs to adjust the wall composition and structure during growth and development stages such as morphogenesis, flocculation, cell-cell recognition and pathogenicity, and in response to changes in environmental conditions (l, 2, 3, 4).
- ItemOpen AccessThe effect of osmotic stress and HSP12 on passive and active transport in the Yeast Saceharomyces cervisiae(2010) Kleinsmidt, Leela S; Lindsey, George G; Brandt, Wolf F
- ItemOpen AccessThe functional significance of Hsp12p and trehalose in desiccation and oxidative stress in the budding yeast Saccharomyces cerevisiae(2007) Shamrock, Vanessa J; Lindsey, George GThe preservation of yeast viability and vitality during storage in the desiccated state is fundamental as several industrial processes utilise this technology. The significance of the stress response protein and putative hydrophilin, Hsp 12p, was therefore examined in vivo under desiccation conditions.
- ItemOpen AccessThe inhibition of M-MLV and HIV-1 reverse transcriptase by polyphenols extracted from the resurrection plant Myrothamnus flabellifolia (Welw.)(2008) Kamng'ona, Arox Wadson; Brandt, Wolf F; Lindsey, George GPolyphenols have been shown to exhibit anti-viral activity in vitro, making them a promising starting point for the development of HIV treatment drugs. The main objective of this thesis was to assess the inhibitory effect of polyphenols extracted from Myrothamnus flabellifolia (Welw.) on M-MLV and HIV-I reverse transcriptases. The first part of the thesis was an attempt to isolate 3,4,5 tri-O-galloylquinic acid, the major polyphenol found in Namibian Myrothamnus flabellifolia plants. This polyphenol was successfully purified by column chromatography (Sephadex LH-20) and its purity was confirmed by HPLC and MALDI-TOF mass spectrometry. The second part of this thesis involved the development of a polymerase enzyme activity assay based on ethidium bromide fluorescence. A calibration curve for quantification of DNA was therefore prepared from the ethidium bromide fluorescence of Calf Thymus DNA. Results demonstrated that Calf Thymus DNA was a good standard for estimating the amount of cDNA synthesised during reverse transcription, thus enabling the monitoring of both M-ML V and HIV -1 reverse transcriptase activity. The reverse transcriptase activity assay was optimised using a poly (rA) template, an oligo (dTb primer and dTTP as a substrate. It was observed that the rate of catalysis for M-ML V and HIV -1 RTs decreased with increase in the concentration of dTTP, which suggested substrate inhibition. A decrease in M-MLV RT activity with increased substrate concentration was found to be due to depletion of Mg2+ ions by dTTP. True substrate inhibition was however observed for HIV-I RT, and analysis of the observed kinetics suggested the formation of an ineffective enzyme substrate complex with two substrate molecules binding to HIV -1 reverse transcriptase. A Hill coefficient of one was obtained at low dTTP concentration and less than one at high dTTP concentration, suggesting zero and negative cooperativity respectively. The final part of this thesis tested the inhibitory effect of pure and crude polyphenol fractions on the activity of M-MLV and HIV-1 RTs. Results showed that all polyphenol fractions inhibited M-ML V and HIV -I reverse transcriptase activity, with the highest inhibitory activity demonstrated by the fraction that contained pure 3,4,5 tri-O-galloylquinic acid. The 50 % inhibitory concentrations of 3,4,5 tri-O-galloylquinic acid was 0.5 μM for M-MLV RT and 34 μM for HIV-I RT. Lineweaver-Burk plots showed that 3,4,5 tri-O-galloylquinic acid inhibited both enzymes non-competitively. Pure non-competitive inhibition was observed for M-MLV RT and mixed non-competitive inhibition for HIV-I RT. Results showed that the binding of 3,4,5 tri-O-galloylquinic acid to M-MLV RT was irreversible, suggesting strong binding under the conditions tested. 3,4,5 Tri-O-galloylquinic acid, however, bound to HIV-I RT reversibly. A comparison of catalytic efficiencies showed that M-MLV RT was more efficient than HIV -1 RT under saturating substrate concentrations with Kcat (min-¹) values of II ± 3 and 1.31 ± 0.02 respectively. M-MLV RT and HIV-¹ RT were, however, equally efficient under limiting substrate concentrations with Kcat/Km (min-¹M-¹) values of 1.1 ± 0.3 x 10⁴ and 1.2 ± 0.2 x 10⁴ respectively.
- ItemOpen AccessInvestigating the mechanisms of desiccation tolerance in the resurrection plant, myrothamnus flabellifolius (WELW)(1999) Koonjul, Priyum K; Lindsey, George G; Brandt, Wolf F; Farrant, Jill MResurrection plants, including Myrothamnus flabellifolius, grow in shallow soil upon rocky outcrops where they experience regular periods of water stress. Associated with this is light stress. The presence of light under water limiting conditions can result in photo-oxidation which causes damage to plant tissues. M flabellifolius is a homoichlorophyllous plant and thus retains chlorophyll during desiccation. The mechanisms whereby this plant prevents photo-oxidation damage are not known and thus one of the objectives of this study was to characterise the chloroplasts and the changes they undergo during dehydration. It was shown that chloroplasts from M flabellifolius could only be isolated using trehalose gradients (instead of sucrose gradients) and were found to have a higher buoyant density than chloroplasts isolated from another resurrection plant, Craterostigma wilmsii. The latter had the same buoyant density as those isolated from the desiccation sensitive plant Pisum sativum. The increased buoyant density in M flabellifolius was ascribed to the unusual ultrastructure of the thylakoid membranes. The latter have a staggered conformation (staircase arrangement) rather than the discrete granal and intergranal conformation found in most plants.
- ItemOpen AccessInvestigating the role of a yeast membrane protein, HSP30 in tolerance to ethanol stress(2001) Tsekoa, Tsepo; Lindsey, George G; Brandt, Wolf FOne of the contributors of the widespread interest the yeast Saccharomyces cerevisiae has received is its ability to yield and tolerate high levels of ethanol. S. cerevisiae is able to grow and remain viable in growth media containing ethanol concentrations as high as 19.8% (w/v), a level that is toxic to many other microorganisms. Since production of ethanol is a normal event in the growth cycyle of S. cerevisiae, this organisms has evolved a number of mechanisms to cope with deleterious effects of ethanol. These include induction of heat shock proteins (HSPs). Among these, HSP30 is particularly interesting in that it is the only stress-induced protein known to be instrinsically bound to the yeast plasma membrane. Another ethanol induced HSP; HSP12 has previously been shown to have a peripheral plasma membrane localisation. It has further been shown that HSP12 protects liposomes against damage by ethanol. This study was initially aimed at investigating whether there is co-operation between HSP30 and HSP12 in this membrane protection role.
- ItemOpen AccessInvestigations into the role of histone H2A ubiquitination in chromatin(1999) Jason, Laure Jeanine Monique; Lindsey, George G; Brandt, Wolf F; Patterton, HughAn in vitro system was used to determine the effect of histone H2A ubiquitination on linker histone binding to mononucleosomes. Hybrid octamers containing either H2A or ubiquitinated H2A (uH2A) were reconstituted onto random sequence 167 bp DNA. The affinity of the resultant nucleosome cores for linker histone H1 was determined from nucleoprotein gel shifts, protein analyses and thermal denaturation. Ubiquitinated H2A did not inhibit linker histone binding to nucleosome cores. The effect of uH2A on nucleosome and chromatosoine positioning on a 208 bp Lytechinus variegatus 5S rDNA fragment was investigated using a combination of micrococcal nuclease digestion and subsequent restriction enzyme digestion of the core particle or chromatosome DNA. Nucleosomes and chromatosomes containing uH2A were found to occupy the same positions on the template DNA as those containing H2A. Chromatin folding of nucleosomal arrays containing either H2A or uH2A was analysed using a quantitative agarose gel electrophoresis system developed by Hansen and co-workers. The extent of folding of nucleosomal arrays containing uH2A was comparable to that of control nucleosomal arrays. A differential centrifugation assay was used to monitor the extent of divalent cation induced oligomerisation of reconstituted nucleosomal arrays. Nucleosomal arrays containing uH2A were found to oligomerise at a lower magnesium concentration than control arrays. As a first step towards studying the effects of H2A ubiquitination in linker histone-bound nucleosomal arrays, a novel method for linker histone reconstitution onto long chromatin stripped of linker histones was developed. The fidelity of linker histone reconstitution was assayed by micrococcal nuclease digestion, thermal denaturation and determination of the orientation of neighbouring linker histone molecules in extended chromatin. In a separate study, the relationship between the observed repeat length of chromatin and the rate of micrococcal nuclease digestion was investigated. The repeat length of the same starting chromatin preparation at equivalent extents of digestion was found to vary according to the rate of digestion.
- ItemOpen AccessIsolation and characterisation of a LEA-like protein from yeast (Saccharomyces cerevisiae)(1998) Mtwisha, Linda; Brandt, W B; Lindsey, George GLEA proteins are plant proteins that are characteristically hydrophilic and soluble at elevated temperature. The consistent correlation between desiccation tolerance in orthodox seed tissue and an accumulation of LEA proteins suggests that these proteins play an important role in protecting cells from desiccation induced damage. Yeast (Saccharomyces cerevisiae) has been known to desiccate as part of its normal growth cycle and to remain viable after long periods in the desiccated state. As a result of these properties this project was designed to investigate the presence of LEA-like proteins in yeast. A protein was isolated from baker’s yeast that fulfils the requirements for being a LEA protein. This protein, with a molecular mass of 11 kDa, was found to be the most prevalent heat soluble protein in the yeast extract. Antibodies raised against LEA group I proteins recognised this 11 kDa yeast protein in the total extract but failed to recognise the protein after heat treatment at 80°C for 10 min. Amino acid analysis showed that the ll kDa protein was highly hydrophilic - a characteristic of LEA proteins. The protein was partially sequenced (10 cycles) after CNBr digestion and the sequence obtained was compared with the sequence of known proteins in the Stanford databank. Only one protein, HSP 12, was identified to be 100 % homologous to the obtained sequence without the introduction of gaps. Despite a previous report that HSP 12 is a heat shock protein, HSP 12 was present in a reduced concentration in yeast grown at 37 °C compared with yeast grown at 30 °C. HSP 12 was found to increase in concentration after entry into stationary phase - a time when nutrients are limiting and the yeast is preparing to reduce its water content and sporulate. This might be considered equivalent to plant seed maturation - the stage when LEA proteins are synthesised. Moreover, growth conditions that have been reported to stimulate LEA protein biosynthesis in plants also stimulated HSP 12 synthesis in yeast. Purified HSP 12 was shown to inhibit thermal denaturation of yeast alcohol dehydrogenase (ADH) at elevated temperatures. This is a functional property of the pea seed p11 LEA group I protein. From the above results, it was therefore concluded that HSP 12 should be identified as a LEA-like protein rather than as a heat shock protein.
- ItemOpen AccessThe isolation and characterization of heat shock protein Hsp12 in Lipomyces starkeyi(2002) Mukwevho, Emmanuel; Brandt, Wolf F; Lindsey, George GThe stress response protein Hsp 12 is induced in S. cerevisiae cells upon exposure to salt stress, heat shock, ethanol, and upon entry to stationary phase (Mtwisha et aI., 1998). In this study, the occurrence of proteins related to Hsp12 was investigated in a number of yeasts (namely, Saccharomyces cerevisiae S288C, Schizosaccharomyces pombe, Debaromyces hansenii, Lipomyces starkeyi Y-2024, Saccharomyces cerevisiae IFO 23X7 (Kaokai), Zygosaccharomyces rouxii and Pichia sorbitophila. This was performed by selective protein extraction followed by SDS-P AGE and western blotting using a S. cerevisiae anti-Hsp 12 antibody. The results showed that almost all the yeasts investigated possessed a protein that had an identical migration to that of Hsp 12 with the exception of S. pombe, which contained a 9 kDa protein. Western blotting using the antiHsp 12 antibody cross-reacted only with the two S. cerevisiae species in addition to the 12 kDa protein from Lipomyces starkeyi of all the species investigated. MALDI-TOF peptide mass analysis after tryptic digestion of the L. starkeyi 12 kDa protein showed that a close sequence similarity existed to that of S. cerevisiae Hsp 12 and none to rest of the 12 kDa proteins isolated from all the other species investigated. In order to determine the sequence of the Hsp 12 protein, the L. starkeyi Hsp 12 gene was amplified using S. cerevisiae Hsp 12 primers. Gene sequencing of both S. cerevisiae and L. starkeyi Hsp 12 genes revealed three nucleotide differences existed between them. L. starkeyi Hsp 12 was found to be present in relatively small amounts during early growth stages but increased during log phase with a slight further increase during stationary phase. Increasing the salt concentration in the growth medium was found to induce Hsp 12. Increased levels of Hsp 12 appeared to confer a degree of protection during desiccation and subsequent rehydration of both L. starkeyi and S. cerevisiae.
- ItemOpen AccessThe localization, function and applications of the stress response protein Hsp12p in the yeast Saccharomyces cerevisiae(2006) Karreman, Robert Jan; Lindsey, George G; Brandt, Wolf FSince 1990, the yeast Saccharomyces cerevisiae small heat shock protein Hsp12p, has continuously appeared in data associated with stress responses in this organism. Hsp12p is expressed abundantly in response to a large variety of different stresses, but for many years has eluded researchers as to its function, primarily because the viability of yeast strains lacking HSP12 are unaffected by osmotic stress and heat shock. Subsequent studies indicated that Hsp12p played a role in the adaptation of the cell wall of Saccharomyces cerevisiae to conditions of stress. However, the exact in vivo localization, specific function and mediation of function of Hsp12p had yet to be elucidated. The localization of Hsp12p was determined by fusion to the green fluorescent reporter protein, Gfp2p and a combination of epifluorescent microscopy and confocal imagery. Chemical extraction revealed that Hsp12p was present in the cell wall while fluorescent imagery was not conclusive. This fluorescent Hsp12p construct was later employed in a novel application to sense the stress status of yeast, which bears future promise for use in an industrial setting.
- ItemOpen AccessMolecular and physiological study of water-deficit stress on selected Eragrostis species(2002) Ginbot, Zekarias Gebremedhin; Farrant, Jill M; Lindsey, George G; Brandt, Wolf FEragrostis nindensis and Eragrostis tef are wild and domestic grasses respectively that belong to the subfamily Eragrostideae. E. nindensis is desiccation tolerant while E. tef is desiccation sensitive. The responses of these plants to water-deficit stress were studied using molecular and physiological approaches. A cDNA library of E. nindensis was screened to identify differentially expressed genes during dehydration. Physiological studies included monitoring changes in photosynthesis, respiration, ultrastructure and membrane integrity of plants during dehydration and rehydration. The differential screening of the cDNA library, using a radio-labelled cDNA from hydrated and dehydrated leaves respectively, revealed two genes, referred as Nin-19 and Nin-44, that were differentially expressed in dehydrated leaves of E. nindensis. These genes were sequenced and partially characterized. Nin-19 did not show considerable identity with any known genes and was not studied any further. Nin-44 was identified as a dehydrin-like gene with approximately 99 % identity to seven water-deficit stress responsive genes on a section of about 60 bp near the 3' end. As its sequence was found to represent a partial insert size, two forward and reverse primers were designed to find the full length through RT-PCR. Despite repeated attempts, no products that could be used in subsequent procedures were achieved using this technique. Hence, further characterization of this gene also could not be performed and different approaches were suggested. The physiological studies showed that E. nindensis is desiccation tolerant but E. tefis not, the latter dying below RWC of about 33 %. Difference among plants in physiological responses became evident after 6 days of dehydration treatment, which resulted in a decline of RWC to 65%, 39% and 33% in E. nindensis, E.tif(R) (red-seeded) and E. tef (W) (white-seeded) plants respectively. A significant decrease in photosynthesis, transpiration, stomatal conductance, and an increase in electrolyte leakage occurred in all species after 6 days of dehydration, but leaves of E. tef (W) did not recover from this level of dehydration when watered. Instead, new leaves were observed to re-grow from the stem nodes. The leaves of red-seeded variety of E. tef did recover fully from RWC of 39 %. After a further 3 days dehydration both varieties of E. tef died. On the other hand, E. nindensis was found to survive extreme water-deficit (-10 % RWC tested here) and recovered full physiological activity when watered. The electrolyte leakage study on these plants indicated major injury on E. tef(W), being intermediate in E. tef(R) and very low in E. nindensis, which coincided with the trend of declining in RWC and other metabolic activities measured. The ultrastructural study on E. tef varieties also showed evidence of the damage caused by dehydration, but the difference among these species was not significant enough to indicate the level of susceptibility of the plants to dehydration damage. The study demonstrated that E. tef varieties are not drought tolerant and showed a considerable difference in their responses to water-deficit stress with each other and with respect to E. nindensis. However, E. tef (R) seems to have a better control over transpiration and some form of repair mechanism operational at least until dehydration to 39 % RWC. This is proposed to be a better performing cereal in conditions of water stress. On the other hand E. nindnsis did not suffer major injury from the dehydration treatment and confirmed to be desiccation tolerant.
- ItemOpen AccessAn overview of mechanisms of desiccation tolerance in selected angiosperm resurrection plants(Global Science Books, 2007) Farrant, Jill M; Brandt, Wolf B; Lindsey, George GThe vegetative tissues of resurrection plants, like seeds, can tolerate desiccation to 5% relative water content (RWC) for extended periods and yet resume full metabolic activity on re-watering. In this review we will illustrate how this is achieved in a variety of angiosperm resurrection plants, our studies ranging from the ecophysiological to the biochemical level. At the whole plant level, leaf folding and other anatomical changes serve to minimise light and mechanical stress associated with drying and rehydration. The mechanisms of cell wall folding are described for Craterostigma wilmsii and Myrothanmus flabellifolia. Free radicals, radical oxygen species (ROS) usually generated under water-deficit stress by photosynthesis, are minimised by either homoiochlorophylly (e.g. C. wilmsii and M. flabellifolia) or poikilochlorophylly (e.g. Xerophyta sp.). The antioxidant systems of these plants effectively deal with ROS generated by other metabolic processes. In addition to antioxidants common to most plants, resurrection plants also accumulate polyphenols such as 3, 4, 5 tri-O-galloylquinic acid in M. flabellifolia, and seed-associated antioxidants (e.g. 1-cys-peroxiredoxin and metallothionines) as effective ROS scavengers. Sucrose accumulates at low RWC, presumably protecting the sub-cellular milieu against desiccation-induced macromolecular denaturation.
- ItemRestrictedAn overview of the biology of the desiccation-tolerant resurrection plant Myrothamnus flabellifolia.(Oxford University Press, 2007) Moore, John P; Lindsey, George G; Farrant, Jill M; Brandt, Wolf FMyrothamnus flabellifolia (Welw.) is a relatively large resurrection plant, a woody shrub between 0.5 m and 1.5 m tall (Sherwin et al., 1998) that grows on rock inselbergs (Porembski and Barthlott, 2000) throughout southern Africa (Weimarck, 1936; Van Wyk et al., 1997; Glen et al., 1999). The plant was first recorded in 1859 by Friedrich Welwitsch, who named the plant Myrothamnus (myron meaning aromatic and thamnos meaning bush) flabellifolia (meaning fan-like leaves) (Puff, 1978a; Glen et al., 1999), the leaves having a balsamic-like odour (Puff, 1978a; Glen et al., 1999). Weiss (1906) was the first to note the ‘miraculous manner’ with which the desiccated plant revived when supplied with water (Fig. 1A, B). Myrothamnus flabellifolia occupies an important position in traditional African folklore and medicine (Watt and Breyer-Brandwijk, 1962; Hutchings, 1996; Van Wyk et al., 1997). The Zulu name for the plant is ‘uvukwaba- file’ (wakes from the dead). The reviving ability is believed to be passed on to the ill person during treatment (Hutchings, 1996; Van Wyk et al., 1997). The plant is a geophyte possessing an extensive root system which extends into the crevices of the rocky slopes upon which it grows (Child, 1960; Glen et al., 1999). Myrothamnus flabellifolia can dehydrate its vegetative tissue, in particular its leaves, to an air-dry state. In this state, the leaves and stem segments curl and change colour from green to dullbrown (Farrant et al., 1999; Glen et al., 1999). When water is provided to the roots the plant re-hydrates its desiccated tissue and returns to its original colour and shape (Glen et al., 1999; Farrant et al., 2003). Since the last review on M. flabellifolia was written many years ago (Puff, 1978a) and since considerable work has been published in the last decade, this review focuses on recent advances in the understanding of the physiology, biochemistry and chemistry of M. flabellifolia.
- ItemRestrictedThe predominant polyphenol in the leaves of the resurrection plant Myrothamnus flabellifolius, 3,4,5 tri-O-galloylquinic acid, protects membranes against desiccation and free radical-induced oxidation(Portland Press, 2005) Moore, John P; Westall, Kim L; Ravenscroft, Neil; Farrant, Jill M; Lindsey, George G; Brandt, Wolf FThe predominant (>90%) low-molecular-mass polyphenol was isolated from the leaves of the resurrection plant Myrothamnus flabellifolius and identified to be 3,4,5 tri-O-galloylquinic acid using 1 H and 13C one- and two-dimensional NMR spectroscopy. The structure was confirmed by mass spectrometric analysis. This compound was present at high concentrations, 44%(by weight) in hydrated leaves and 74% (by weight) in dehydrated leaves. Electron microscopy of leaf material fixed with glutaraldehyde and caffeine demonstrated that the polyphenols were localized in large vacuoles in both hydrated and dehydrated leaves. 3,4,5 Tri-O-galloylquinic acid was shown to stabilize an artificial membrane system, liposomes, against desiccation if the polyphenol concentration was between 1 and 2 µg/µg phospholipid. The phase transition of these liposomes observed at 46 ◦C was markedly diminished by the presence of 3,4,5 tri-O-galloylquinic acid, suggesting that the presence of the polyphenol maintained the membranes in the liquid crystalline phase at physiological temperatures. 3,4,5 Tri-O-galloylquinic acid was also shown to protect linoleic acid against free radical-induced oxidation.