Main conclusion Provides a first comprehensive review of integrated physiological and molecular a... more Main conclusion Provides a first comprehensive review of integrated physiological and molecular aspects of desiccation tolerance Xerophyta viscosa. A synopsis of biotechnological studies being undertaken to improve drought tolerance in maize is given. Xerophyta viscosa (Baker) is a monocotyledonous resurrection plant from the family Vellociacea that occurs in summer-rainfall areas of South Africa, Lesotho and Swaziland. It inhabits rocky terrain in exposed grasslands and frequently experiences periods of water deficit. Being a resurrection plant it tolerates the loss of 95 % of total cellular water, regaining full metabolic competency within 3 days of rehydration. In this paper, we review some of the molecular and physiological adaptations that occur during various stages of dehydration of X. viscosa, these being functionally grouped into early and late responses, which might be relevant to the attainment of desiccation tolerance. During early drying (to 55 % RWC) photosynthesis is shut down, there is increased presence and activity of housekeeping antioxidants and a redirection of metabolism to the increased formation of sucrose and raffinose family oligosaccharides. Other metabolic shifts suggest water replacement in vacuoles proposed to facilitate mechanical stabilization. Some regulatory processes observed include increased presence of a linker histone H1 variant, a Type 2C protein phosphatase, a calmodulin-and an ERD15-like protein. During the late stages of drying (to 10 % RWC) there was increased expression of several proteins involved in signal transduction, and retroelements speculated to be instrumental in gene silencing. There was induction of antioxidants not typically found in desiccation-sensitive systems, classical stress-associated proteins (HSP and LEAs), proteins involved in structural stabilization and those associated with changes in various metabolite pools during drying. Metabolites accumulated in this stage are proposed, inter alia, to facilitate subcellular stabilization by vitrification process which can include glass-and ionic liquid formation. Keywords Physiology Á Proteome Á Resurrection plant Á Transcriptome Á Vegetative desiccation tolerance Abbreviation ERD (LRD) Early (Late) response to desiccation
Provides a first comprehensive review of integrated physiological and molecular aspects of desicc... more Provides a first comprehensive review of integrated physiological and molecular aspects of desiccation tolerance Xerophyta viscosa . A synopsis of biotechnological studies being undertaken to improve drought tolerance in maize is given. Xerophyta viscosa (Baker) is a monocotyledonous resurrection plant from the family Vellociacea that occurs in summer-rainfall areas of South Africa, Lesotho and Swaziland. It inhabits rocky terrain in exposed grasslands and frequently experiences periods of water deficit. Being a resurrection plant it tolerates the loss of 95 % of total cellular water, regaining full metabolic competency within 3 days of rehydration. In this paper, we review some of the molecular and physiological adaptations that occur during various stages of dehydration of X. viscosa, these being functionally grouped into early and late responses, which might be relevant to the attainment of desiccation tolerance. During early drying (to 55 % RWC) photosynthesis is shut down, there is increased presence and activity of housekeeping antioxidants and a redirection of metabolism to the increased formation of sucrose and raffinose family oligosaccharides. Other metabolic shifts suggest water replacement in vacuoles proposed to facilitate mechanical stabilization. Some regulatory processes observed include increased presence of a linker histone H1 variant, a Type 2C protein phosphatase, a calmodulin- and an ERD15-like protein. During the late stages of drying (to 10 % RWC) there was increased expression of several proteins involved in signal transduction, and retroelements speculated to be instrumental in gene silencing. There was induction of antioxidants not typically found in desiccation-sensitive systems, classical stress-associated proteins (HSP and LEAs), proteins involved in structural stabilization and those associated with changes in various metabolite pools during drying. Metabolites accumulated in this stage are proposed, inter alia, to facilitate subcellular stabilization by vitrification process which can include glass- and ionic liquid formation.
Previous studies report that leaf tensile strength (TS) of the desiccation tolerant (resurrection... more Previous studies report that leaf tensile strength (TS) of the desiccation tolerant (resurrection) grass Eragrostis nindensis does not change on drying, but increases in dried desiccation sensitive Eragrostis species. In this paper we tested whether unchanging TS on dehydration is a common feature among 4 resurrection species, Craterostigma wilmsii, Sporobolus stapfianus, Xerophyta humilis and Xerophyta schlecteri, and how this might relate to leaf structure and mechanisms of protection against mechanical stress of drying. Desiccation sensitive controls were Zea mays and Arabidopsis thaliana. Light and transmission electron microscopy of leaves was performed to determine lignification and the nature of subcellular mechanical stabilization. There was a positive correlation between % lignin/unit cross-sectional area and TS of hydrated leaves. Only the grass, S. stapfianus, did not change TS when naturally dried. All others increased in TS when naturally dried, but there was variation among them when flash dried. In S. stapfianus, mechanical stabilization was by both wall folding (mesophyll) and vacuole packaging (bundle sheath) as reported for E. nindensis. This combination may account, in part, for unchanging TS during drying and may be a feature of resurrection grasses. We conclude that leaf tensile properties differ among resurrection plants and are not necessarily affected by protection mechanisms associated with mechanical stress.
the work reported in this paper, and the relationship among desiccation sensitivity of axes, chan... more the work reported in this paper, and the relationship among desiccation sensitivity of axes, changes in cell ultrastructure of axes and activities of superoxide dismutase (SOD), ascorbate peroxidase (APX), catalase (CAT), glutathione reductase (GR) and dehydroascorbate reductase (DHAR), and the content of TBA (thiobarbituric acid)-reactive products in axes and washed mitochondria were studied. Desiccation tolerance of axes was gradually lost with dehydration, and desiccation tolerance of epicotyls was larger than that of radicles. The seed water content at which 50% of epicotyl and radical were killed by dehydration was about 0.58 and 0.66 g/g (dry weight base, dwb), respectively. At this water content level, cell ultrastructure of axes show damage. All of the epicotyls and radicals were killed by dehydration at 0.26 and 0.48 g/g (dwb) water content levels, respectively. Cytochrome c oxidase (CCO) activities of washed mitochondria from axes increased during the initial phase of dehydration, and then notably decreased. Latencies of CCO activities of washed mitochondria were about 93%-97%. Activities of SOD, APX, CAT, GR, DHAR of axes and washed mitochondria increased at initial phase of dehydration, and then decreased rapidly. These five enzyme activities were more sensitive to dehydration in washed mitochondria than in the axes, and the content of TBA-reactive products in axes and washed mitochondria dramatically increased with dehydration of axes. These results showed that A. toxicaria seed is a typical recalcitrant seed; that loss of desiccation tolerance in axes was a quantitative feature; and that antioxidant enzyme activities in mitochondria could be one of the more sensitive parameters for study of seed recalcitrance.
At harvest, sunflower (Helianthus annuus L.) seeds are dormant and unable to germinate at tempera... more At harvest, sunflower (Helianthus annuus L.) seeds are dormant and unable to germinate at temperatures below 15°C. Seed storage in the dry state, known as after-ripening, is associated with an alleviation of embryonic dormancy allowing subsequent germination at suboptimal temperatures. To identify the process by which dormancy is broken during after-ripening, we focused on the role of reactive oxygen species (ROS) in this phenomenon. After-ripening entailed a progressive accumulation of ROS, namely superoxide anions and hydrogen peroxide, in cells of embryonic axes. This accumulation, which was investigated at the cellular level by electron microscopy, occurred concomitantly with lipid peroxidation and oxidation (carbonylation) of specific embryo proteins. Incubation of dormant seeds for 3 h in the presence of hydrogen cyanide (a compound that breaks dormancy) or methylviologen (a ROS-generating compound) also released dormancy and caused the oxidation of a specific set of embryo proteins. From these observations, we propose a novel mechanism for seed dormancy alleviation. This mechanism involves ROS production and targeted changes in protein carbonylation patterns.
As there is limited information on the mechanisms of vegetative desiccation tolerance in pteridop... more As there is limited information on the mechanisms of vegetative desiccation tolerance in pteridophytes, we undertook a comprehensive anatomical, ultrastructural, physiological and biochemical study on the fern Mohria caffrorum. Our data show that this species is desiccation-tolerant during the dry season, and desiccation-sensitive in the rainy season. This system allows the verification of protection mechanisms by comparison of tolerant and sensitive tissues of the same species at the same developmental age. Tolerant fronds acquire protection mechanisms during drying that are mostly similar to those reported for angiosperms. These include: (i) chlorophyll masking by abaxial scales and frond curling; (ii) increased antioxidant capacity that is maintained in dry tissues; (iii) mechanical stabilization of vacuoles in the dry state; (iv) de novo production of heat stable proteins (at least one identified as a putative chaperonin); (v) accummulation of protective carbohydrates (sucrose, raffinose family oligosaccharides and cyclitols). This study has implications for the biotechnological production of drought-tolerant crops, and allows speculation on the evolution of vegetative desiccation tolerance.
is a resurrection species that is thought to rely primarily on the protection of cellular compone... more is a resurrection species that is thought to rely primarily on the protection of cellular components during drying to survive desiccation. The time taken for this protection to be instituted is thought to preclude recovery after rapid drying. Thus the response of C. wilmsii plants to rapid dehydration was investigated. The effect of rapid drying on sucrose accumulation was determined and the cellular ultrastructure was investigated during natural (slow) and rapid dehydration and on subsequent rehydration. The dependence of naturally and rapidly dried C. wilmsii on de novo transcription and translation during and after rehydration was determined by examining quantum ef®ciency, changes in photosynthetic pigments and subcellular organization of excised leaves with rehydration in water and using the metabolic inhibitors, distamycin A and cycloheximide. Slowly dried C. wilmsii required no new transcription or translation during rehydration in order to recover. With rapid dehydration, cells showed ultrastructural damage, which indicated that at least some protective mechanisms were affected (as evidenced by a reduced accumulation of sucrose). C. wilmsii was able to limit the damage and recover upon rehydration in water, but rapidly dried plants did not survive if mRNA or protein synthesis was inhibited by distamycin A or cycloheximide, respectively. This demonstrates an induction of repair mechanisms during rehydration, which enables recovery from rapid drying. Thus, although C. wilmsii does rely almost entirely on protection during natural drying, it apparently has the ability to repair if protection is inadequate and damage is incurred.
De-etiolation of dark-grown seedlings is a commonly used experimental system to study the mechani... more De-etiolation of dark-grown seedlings is a commonly used experimental system to study the mechanisms of chloroplast biogenesis, including the stacking of thylakoid membranes into grana, the response of the nuclear-chloroplast transcriptome to light, and the ordered synthesis and assembly of photosystem II (PSII). Here, we present the xeroplast to chloroplast transition during rehydration of the resurrection plant Xerophyta humilis as a novel system for studying chloroplast biogenesis, and investigate the role of light in this process. Xeroplasts are characterized by the presence of numerous large and small membrane-bound vesicles and the complete absence of thylakoid membranes. While the initial assembly of stromal thylakoid membranes occurs independently of light, the formation of grana is light dependent. Recovery of photosynthetic activity is rapid in plants rehydrated in the light and correlates with the light-dependent synthesis of the D1 protein, but does not require de novo chlorophyll biosynthesis. Light-dependent synthesis of the chlorophyll-binding protein Lhcb2 and digalactosyldiacylglycerol synthase 1 correlated with the formation of grana and with the increased PSII activity. Our results suggest that the molecular mechanisms underlying photomorphogenic development may also function in desiccation tolerance in poikilochlorophyllous resurrection plants.
At harvest, sunflower (Helianthus annuus L.) seeds are dormant and unable to germinate at tempera... more At harvest, sunflower (Helianthus annuus L.) seeds are dormant and unable to germinate at temperatures below 15°C. Seed storage in the dry state, known as after-ripening, is associated with an alleviation of embryonic dormancy allowing subsequent germination at suboptimal temperatures. To identify the process by which dormancy is broken during after-ripening, we focused on the role of reactive oxygen species (ROS) in this phenomenon. After-ripening entailed a progressive accumulation of ROS, namely superoxide anions and hydrogen peroxide, in cells of embryonic axes. This accumulation, which was investigated at the cellular level by electron microscopy, occurred concomitantly with lipid peroxidation and oxidation (carbonylation) of specific embryo proteins. Incubation of dormant seeds for 3 h in the presence of hydrogen cyanide (a compound that breaks dormancy) or methylviologen (a ROS-generating compound) also released dormancy and caused the oxidation of a specific set of embryo proteins. From these observations, we propose a novel mechanism for seed dormancy alleviation. This mechanism involves ROS production and targeted changes in protein carbonylation patterns.
The effect of drying rate on the survival of three angiosperm resurrection plants, Craterostigma ... more The effect of drying rate on the survival of three angiosperm resurrection plants, Craterostigma wilmsii (homoiochlorophyllous), Xerophyta humilis (poikilochlorophyllous) and Myrothamnus flabellifolius (homoiochlorophyllous) was examined. All species survived slow drying, but only C. wilmsii was able to survive rapid drying. C. wilmsii was rapidly able to induce protection mechanisms such as folding of cell walls to prevent mechanical stress and curling of leaves to minimize light stress, and thus survived fast drying. Rapid drying of X. humilis and M. flabellifolius appeared to allow insufficient time for complete induction of protection mechanisms. In X. humilis, there was incomplete replacement of water in vacuoles, the photosynthetic apparatus was not dismantled, plasma membrane disruption occurred and quantum efficiency of photosystem II (F V \F M ) did not recover on rehydration. Rapidly dried leaves of M. flabellifolius did not fold tightly against the stem and F V \F M did not recover. Ultrastructural studies showed that subcellular damage incurred during drying was exacerbated on rehydration. The three species co-occur in environments in which they experience high desiccation pressures. C. wilmsii has few features to retard water loss and thus the ability for rapid induction of subcellular protection is vital to survival. X. humilis and M. flabellifolius are able to retard water loss and protection is acquired relatively slowly.
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