Recent literature focuses on the phenomenal desiccation tolerance (although this is only relevant for the eurypoikilohydrous organisms) and wonders whether it is a primitive strategy (Oliver et al. 2005) or how it can be genetically traced down (Illing et al. 2005) or whether it constrains growth and competetivity (Alpert 2006). Today, 74 pteridophyte and 145 angio-sperm poikilohydrous species have been investigated with respect to desiccation tolerance (see Proctor and Pence 2002).
The first, but not the only, stress during desiccation is the lack of water itself, which imposes dramatic structural and physiological changes on the tissues of poikilohydrous organisms. Some poikilohydrous plants exhibit a remarkable tolerance to intense desiccation. The moss T. ruralis survived a water content as low as 0.008%, which is equivalent to —6000 bars (Schonbeck and Bewley 1981a). Desiccation tolerance of eurypoikilohydrous autotrophs can be defined as the capacity to withstand equilibrium with a relative humidity less than 20% (Lange 1953, Biebl 1962, Bertsch 1966b, Gaff 1986). The lowest relative water contents tolerated by vascular plants were reported to be between 1.4% and 8.4%, which is equivalent to 4% and 15% of their dry weight (Kaiser et al. 1985) (see Table 2.1 for a detailed list). Detached leaves of resurrection plants proved to be much less tolerant to severe water loss (Gaff 1980). Table 2.1 also shows tolerance to extended periods of anabiosis. Nostoc may tolerate 5 years of desiccation, but other cyanobacteria do not survive desiccation at all (Biebl 1962, Scherer et al. 1986). Most lichens, bryophytes (Lange 1953, Biebl 1962, Proctor 1982), and poikilohydrous vascular plants (Hallam and Gaff 1978, Lazarides 1992) survive dry periods that last for a few months. Although Marchantiales were reported to be exposed to 6-8 months of drought in their habitat in Namibia (Volk 1979), such extremely long periods of desiccation are usually rare for this kind of bryophyte. Several vascular resurrection plants have been shown to tolerate air-dry periods of 2-5 years (Hickel 1967, Gaff and Ellis 1974, Gaff 1977), and some lichens can become photosynthetically active again after a period of 10 years frozen in the dry state (Larson 1988).
Bryophytes comprise taxa that have an intrinsic capacity of desiccation tolerance, as well as taxa that need acclimation (Proctor and Tuba 2002). Although the so-called xerophytic moss species such as Syntrichia ruralis, Rhacomitrium canescens, Neckera crispa, and others always immediately survive extreme desiccation, mesophytic and even hygrophytic species such as Bryum caespititium, Plagiothecium platyphyllum, Pohlia elongata, or Mnium seligeri become extremely desiccation tolerant only if they were pretreated at a relative humidity of
96% for 24 h (Abel 1956, Biebl 1962). Most remarkable is that the water moss Fontinalis squamosa is as tolerant as a xerophytic moss, whereas Fontinalis antipyretica is drought-sensitive even when pretreated at 96% relative humidity. The intrinsic desiccation tolerance of eurypoikilohydrous bryophytes can be modified by the rapidity of desiccation processes (Gaff 1980). Desiccation tolerance was observed to vary seasonally in many bryophytes (Dircksen 1964, Dilks and Proctor 1976a,b), in ferns (Kappen 1964), and also in Borya nitida (Gaff 1980) as they can acclimate to frost desiccation in winter or to summer drought. Several bryophytes (Dicranum scoparium and Mnium punctatum) and ferns (Asplenium spp. and Polypodium vulgare) gained an extremely high desiccation tolerance in winter (Dircksen 1964, Kappen 1964). Like seeds, air-dry lichens (Lange 1953), bryophytes (Hosokawa and Kubota 1957, Gaff 1980, Proctor 1990), and vascular resurrection plants can persist longer if stored at humidities lower than 30% relative humidity (Leopold 1990). This was explained by the fact that intermediate-to-low water contents allow some enzyme activity and lead to respiratory carbon loss, destructive processes, and infections (Gaff and Churchill 1976 [B. nitida], Proctor 1982). Most of our current knowledge of the effects of environmental conditions before and during desiccation on the tolerance to desiccation and related stresses comes from experiments under controlled conditions.
More important for eurypoikilohydrous autotrophs is their capacity to withstand repeated changes between dry and moist states. Lichens in deserts and the Mediterranean regions, for instance, oscillate regularly between periods of a few hours of activity and anabiosis for the rest of the day (Kappen et al. 1979, Redon and Lange 1983, Kappen 1988, Lange et al. 1991, Sancho et al. 1997). Similarly, it is typical of several poikilohydrous phanerogams such as Chamaegigas intrepidus living in ephemeral rock pools (Woitke et al. 2004) and changing repeatedly between hydrated and dried state within one season. The capacity to tolerate several changes between dry and wet states was tested experimentally for different moss species by Dilks and Proctor (1976a, 1979). Tortula ruralis, as an eurypoiki-lohydrous species, performed well during up to 63 changes within a period of 18 months, whereas Rhytidiadelphus loreus was killed when continually dry for 18 months or when the oscillation phase was 1 day wet/1 day dry, but it retained 50% of its normal net photosyn-thetic rate if the wet periods were longer (6 or 7 days) or the dry period was shorter. Mosses such as T. ruralis and the angiosperm B. nitida (Schonbeck and Bewley 1981b) were actually able to increase their desiccation tolerance if drying and rehydration were repeated. However, cultivation under moist conditions for 2 weeks can decrease the desiccation tolerance of most eurypoikilohydrous autotrophs (algae and lichens: Kappen 1973, Farrar 1976b; bryophytes: Schonbeck and Bewley 1981b, Hellwege et al. 1994; vascular plants: Gaff 1977, 1980). In contrast, continuous hydration over several days decreased the tolerance to desiccation in T. ruralis (Schonbeck and Bewley 1981b), a result also found for some lichens (Ahmadjian 1973). Apparently, in the absence of contrasting oscillations of moisture content, the algal partner of lichens grows excessively, altering its symbiotic relation with the mycobiont (Farrar 1976b). Repeated drying seems to be essential for the internal metabolic balance of lichens (McFarlane and Kershaw 1982), and also in poikilohydrous vascular plants, as indicated by the fact that cultivation of Myrothamnus flabellifolius is only successful if the plants dry occasionally (Puff 1978). For several desert cyanobacteria, hydration is a very rare event. Pleurococcoid green algae (e.g., Apatococcus lobatus) and many epilithic lichens are water-repellent (Bertsch 1966a). Species of the genera Chrysothrix, Lepraria, and Psilolechia growing under overhanging rocks never receive liquid water during their lifetime (Wirth 1987).
Cellular and Physiological Changes during Desiccation
Great attention has been given in the last 25 years to the investigation of the ultrastructural changes and the biochemical processes that take place during dehydration and rehydration of poikilohydrous autotrophs. Rather than providing a detailed account here, we refer the interested reader to some reviews (Gaff 1980, 1989, Bewley and Krochko 1982, Stewart 1989, Leopold 1990, Proctor 1990, Bewley and Oliver 1992, Ingram and Bartels 1996, Hartung et al. 1998, Bartels 2005, Rascio and Rocca 2005). Two main mechanisms are involved, one that downregulates the processes and structures, leading to desiccation tolerance, and another that contributes to full metabolic and structural recovery (Bernacchia et al. 1996).
Nonvascular autotrophs are desiccation tolerant if they can retain cellular integrity and limit cellular damage during drying. To accomplish this with bryophytes we summarize some salient facts from reviews by Bewley and Oliver (1992), Oliver and Bewley (1997), and Oliver et al. (2000, 2005). The capability of xeric bryophytes like Tortula ruralis to recover quickly even from extremely rapid desiccation indicates that their desiccation tolerance is intrinsic. Membrane structure does not suffer from drying, however, protein synthesis ceases rapidly during drying (Bewley and Krochko 1982). Specific protective substances or mechanisms are not apparent (Bewley and Oliver 1992, Oliver and Bewley 1997), as sugar content does not change significantly, and no membrane protective mechanism is detectable. Upon rehydra-tion a transient leakage indicates membrane phase transitions, and cells regain normal shape and structure within 24 h. During the first 2 h of rehydration an extensive alteration in gene expression indicates synthesis of proteins, the rehydrins. The rehydrins are late embryo abundant (LEA) or LEA-like proteins (see Section "Synthesis of Proteins and Protective Substances''), which may stabilize membranes lipids or help to enable quick lipid transport for reconstitution of eventually damaged membranes (Oliver 2005). They may be more important for the mesic, less-tolerant mosses and liverworts, where according to Bewley (1979) changes in membranes and metabolism during dehydration were observed. However, mesic bryopytes may increase tolerance by previous drying treatment or by addition of ABA. The presence of ABA was found in moss (e.g., Funaria hygrometrica: Werner et al. 1991) and in hepatic species (e.g., the europoikilohydric Exormotheca holstii: Hellwege et al. 1994, Hartung personal communication). Accumulation of ABA during drying and disappearance in wet culture and the capacity of inducing hardiness when applied to dehardened gametopytes reveal its role in the drought hardening process, except in T. ruralis where ABA was not detected (Reynolds and Bewley 1993a).
The mechanism of lichens for dealing with desiccation remains broadly unknown but is strongly related to their high content of polyols (Farrar 1976a). Membrane leakage in lichens as a consequence of repeated drying and wetting (Farrar 1976a) is harmless. The role of ABA, recently detected in lichens, is unclear, as this hormone is produced by the fungal biont and, as opposed to its activity in plants, increases as a response to water uptake (Dietz and Hartung 1998).
Pteridophytes have a performance, intermediate between bryophytes and angiosperms. For instance, as was shown with Polypodium virginianum (Oliver et al. 2000), they carry out synthesis of proteins upon dehydration (dydrins) as well as subsequent to rehydration (rehy-drins). Interestingly, the proteins synthesized during drying disappear rapidly upon rehydration, but later (after 24 h) some of these proteins increase again. In addition, immediately after rehydration novel polypeptides are synthesized, which are not exclusively related to the desiccation regime, and another set of new proteins is produced after 24 h. ABA is present in Polypodium but notably decreases during drying. Nonetheless, ABA application can induce synthesis of proteins similar to dehydrins and survival of otherwise letal-rapid desiccation (Reynolds and Bewley 1993a,b). From dehydrated microphylls of S. lepidophylla, an expressed gene sequence tag (EST) database was generated and compared with that of other, not poikilohydrous Selaginella species. The percentage of functional categories, which were disease/defense-related comprising induction of secondary metabolism, molecular chaperons, and LEA proteins, was significantly higher in the poikilohydrous Selaginella species (Iturriaga et al. 2006).
In S. lepidophylla, 74% of the activity of 9 enzymes of the carbohydrate metabolism was conserved during drying. The conservation of photosynthetic enzymes was lower than that for respiratory enzymes (Eickmeier 1986).
Based on the fact that most monocotyledonous poikilohydrous species show a dramatic color change when desiccated and most dicots do not, we can distinguish between poikilo-chlorophyllous and homoiochlorophyllous angiosperms. Comparative, mainly electron-microscopical studies illustrate these differences (Sherwin and Farrant 1996, 1998, Farrant 2000).
In the poikilochlorophyllous species, structural changes are considerable (see Gaff 1980, Hetherington and Smillie 1982, Tuba et al. 1996b, 1997), except in the nucleus (Barthley and Hallam 1979), containing a dense mass of chromatin (Hethrington and Smillie 1982). Ultrastructural changes in poikilochlorophyllous plants may be even greater than in desiccation-sensitive homoiohydrous species at a comparable dehydration level (Gaff 1989). Investigations with Xerophyta villosa and X. scabrida show the following: The polysome content rises significantly with water loss (X. villosa; Gaff 1989). Virtually all thylacoids and most of the carotenoids content are lost. Obviously, the destruction of the chlorophyll is structured, as grana are retained (Bartley and Hallam 1979). However, as most of the chlorophyll can be preserved in X. scabrida, if it is desiccated in darkness, Tuba et al. (1997) suggest that under natural conditions chlorophyll loss is due to photooxidation rather than metabolic destruction. Although most of their cristae disappear, mitochondria remain functional (see Hallam and Capicchianano 1974). A continuation of respiration still measurable below —3.2 MPa (Tuba et al. 1996b) in desiccated plants of X. scabrida suggests that energy is required for dismantling the thylacoids and the formation of the so-called desiccoplasts. Thus, rather than being deleterious, these organelle changes involve an organized remobilization of cell resources in these resurrection plants.
Structural changes by desiccation are usually small in the homoiochlorophyllous species, and thylacoid membranes and associated chlorophyll complexes apparently remain widely preserved (Owoseye and Sandford 1972, Hallam and Cappicchiano 1974, Gaff and McGregor 1979, Gaff 1989). However, changes in the chloroplast structure and loss of chlorophyll by 20% in Myrothamnus flabellifolius (Wellburn and Wellburn 1976, Farrant 2000) and by 40%-50% in Craterostigma wilmsii (Sherwin and Farrant 1998) were observed, which recovered within 24-45 h subsequent to rehydration. Kaiser and Heber (1981) and Schwab and Heber (1984) state that the lens shape of the chloroplasts permits dehydration without greater surface area reduction, and in vivo rupture of chloroplasts during desiccation is rarely observed. In dry Talbotia elegans, the mitochondria are reduced to membrane-bound sacks (Hallam and Gaff 1978). Transient membrane leakage has been reported for several species. Vacuoles, fragmented into numerous vesicles, become filled with a nonaqueous substance, obviously generating a backpressure for the desiccating cell (Farrant 2000) and lysosomes seem to be maintained intact (Gaff 1980, 1989, Hartung et al. 1998). As a consequence, repair processes may last for 2 days in small herbaceous species (Ramonda and Haberlea species: Gaff 1989, Markowska et al. 1995), and the extended time for recovery of the large fruticose M. flabellifolius is due to the restoration of its hydraulic system.
As a rule, the processes of desiccation and recovery take more time in the poikilochlor-ophyllous than in the homoiochlorophyllous species. Tuba et al. (1997) state that under field conditions X. scabrida may take more than 2 weeks until the plants are dry and inactive. Correspondingly, also repair processes are extended over periods of several days (X. viscosa 92-120 h: Sherwin and Farrant 1996, 1998). Poikilochlorophyllous performance is obviously a highly derived adaptation. It allows avoidance of stress from free radicals. In ecological terms, such species are adapted to sites where drought periods are extended over weeks or months, whereas homoiochorophyllous species easily manage in sites with frequent oscillations between dry and wet stages in shorter periods of time.
Obviously, a network of genes with presumably different functions is activated by water stress. Hartung et al. (1998) estimated that 800-3000 genes could be involved in the response of plants to desiccation. Poikilohydrous plants exhibit a great variety of down- and upregula-tion of cellular processes, which can be retained at very low water potentials (Leopold 1990). Particularly, genes that code for enzymes relevant to photosynthesis, both in vascular plants and in mosses (Ingram and Bartels 1996, Bernacchia et al. 1996, Oliver and Bewley 1997) were downregulated. In general, the decline of total protein is smaller than in drought-sensitive plants. Loss of water-insoluble proteins is common in resurrection plants, especially in the poikilochlorophyllous species, probably because of degradation of the lipoproteins of the membrane (Gaff 1980). The preservation of polysomes and of RNA may enable protein synthesis after drought (Bewley 1973). Many novel proteins (dehydrins) are synthesized during desiccation, most of which were considered specific to extremely desiccation-tolerant plants (Hallam and Luff 1980, Eickmeier 1988, Bartels et al. 1990, Piatkowski et al. 1990, Bartels et al. 1993, Kuang et al. 1995). Nevertheless, certain polypeptides, such as those found in desiccated Polypodium virginianum, are not exclusive to the desiccation regime (Reynolds and Bewley 1993b). The majority of the dehydrins belongs to LEA proteins, they are hydrophilic and resistant to denaturation, and typical of orthodox seeds. They are believed to protect desiccation-sensitive enzymes and to stabilize membranes during dehydration (Schneider et al. 1993, Bernacchia et al. 1996, Ingram and Bartels 1996, Bartels 2005). Proteins are necessary also during the rehydration phase. They can be gained by translation of already existing transcripts, as was shown for poikilohydrous species (Dace et al. 1998). Bartels and Salamini (2001) suggest that desiccation tolerance (of Craterostigma plantagineum) is in most cases not due to structural genes, unique to resurrection plants and could be present as well in desiccation-sensitive homoiohydrous plants. However, the latter may have less amounts of LEA proteins and the expression pattern may be different. Only one, a LEA-6 protein, was identified as typical exclusively for Xerophyta humilis and seeds (Illing et al. 2005).
In most resurrection plants, including the aquatic species Chamaegigas intrepidus, abscisic acid (ABA) is strongly accumulated and is involved in attaining desiccation tolerance and in stimulating the synthesis of dehydrins (Gaff 1980, 1989, Gaff and Loveys 1984, Reynolds and Bewley 1993a, Hellwege et al. 1994, Schiller et al. 1997). As was hypothesized by Bartels et al. (1990), Nelson et al. (1994), and Oliver and Bewley (1997), there is evidence now in vascular plants that ABA is necessary to induce the genes for desiccation tolerance. With experiments of mutants of C. plantagineum, the so-called CDT-1/2 gene family was shown to function by ABA signal transduction (Smith-Espinoza et al. 2005). Leaves of Myrothamnus flabellifolius and Borya nitida did not survive dehydration if they were dried so rapidly that ABA could not be accumulated (Gaff and Loveys 1984). Abscisic acid accumulation obviously can occur only in leaves attached to the whole plant (Hartung et al. 1998).
A common phenomenon in drought stress is the accumulation of organic compatible solutes because they stabilize proteins and membranes (Levitt 1980, Crowe and Crowe 1992). Lichens are permanently rich in sugar alcohols, which are assumed to be the basis of their remarkable desiccation tolerance (Kappen 1988). Cowan et al. (1979) have demonstrated that the synthesis of amino acids and sugar alcohols was active in lichens in equilibrium with humidities as low as 50%. In contrast, desiccation-tolerant bryophytes contain a low amount of sugars, mainly sucrose, and show no or very little increase in sugar content during drying (Bewley and Pacey 1978, Santarius 1994). Strong sugar accumulation, mainly sucrose, during desiccation has been demonstrated in seeds and many resurrection grasses, species of Ramonda, Haberlea, and Boea, and X. villosa (Kaiser et al. 1985, Scott 2000, Zirkovic et al. 2005). Other resurrection plants for example, of the genera Ceterach and Craterostigma already contain comparatively high amounts of sugar in the leaves when turgid
(Schwab and Gaff 1986). In these and other species (e.g., M. flabellifolius), sugar composition was observed to be changed during dehydration (Bianchi et al. 1991, Hartung et al. 1998). Unusual sugars such as stachyose that appear in the turgid leaves and roots are storage products, but they are converted into sucrose during the drying process (Bianchi et al. 1991, 1993, Albini et al. 1994, Heilmeier and Hartung 2001, Norwood et al. 2003). For instance, 2-octulose is typical for hydrated leaves of C. plantagineum and is converted into sucrose upon dehydration (Bartels and Salamini 2001). Thus, sucrose accumulation during desiccation is generally recruited from metabolizing storage carbohydrates rather than directly from photosynthesis.
Inositol, present in Xerophyta viscosa, may be an effective osmoprotectant (Mayee et al. 2005). Proline concentration in many plant species associated with water stress was comparatively high in Ceterach and Craterostigma, but did not significantly increase during dehydration (Schwab and Heber 1984). Application of proline had no effect on detached leaves of B. nitida and M. flabellifolius (Gaff 1980). In the latter species, polyphenols have been identified (Moore et al. 2005) that might be relevant to desiccation tolerance, as provenances from Namibia subjected to greater drought stress were genetically different from those in South Africa and contained more and different polyphenols (e.g., 3,4,5-tri-O-galloquinic acid).
One of the internal hazards of desiccation is the increase in oxidative processes, which occurs in plants exposed to a wide range of environmental stresses (Smirnoff 1995). An increase in or a high level of defense enzymes of the ascorbate/glutathione cycle was associated with the protection of the membrane lipids in Sporobolus stapfianus during drying (Sgherri et al. 1994a). Oxidized glutathione was much lower in slowly dried (unimpaired) samples than in rapidly (injured) dried samples of Boea hygroscopica (Sgherri et al. 1994b), and glutathione was shown to play the primary role in maintaining the sulfhydryl groups of thylacoid proteins in reduced state during desiccation (Navarri-Izzo et al. 1997). The reversible decrease in phenolic acids far below the level in the hydrated state, which is joined by a decrease in the enzyme ascorbate peroxidase (AP) while antioxidants were accumulated, indicates that Ramonda serbica leaves are able to keep up an antioxidative status when subjected to desiccation (Sgherri et al. 2004). Kranner and Grill (1997) postulate that glutathione reductase (GR) and glucose-6-phosphate dehydrogenase are needed for the reduction of desiccation-induced oxidized glutathione. It is suggested that this pathway provides the NADPH during the critical rehydration phase when photosynthesis is still inactive. Accordingly, when photosynthesis is recovered a decrease in antioxidants and production of reactive oxygen species was observed in a lichen species (Weissman et al. 2005). Antioxidants such as AP, GR, and SOD (superoxide dismutase) were increased but to various extents in subsequent phases of water loss in Craterostigma wilmsii, M. flabellifolius (mainly GR), and X. viscosa (Sherwin and Farrant 1998) and went down to normal level when the tissues were rehydrated. Anthocyanins recognized as antioxidants (Smirnoff 1993) were observed to increase in drying leaves of poikilochlorophyllous species such as Eragrostis nindensis (Van der Willigen et al. 2001) and particularly in Xerophyta humilis (Farrant 2000). In C. plantagineum, lipoxygenase, which catalyzes lipid peroxidation at membranes, becomes increasingly inhibited during drying (Smirnoff 1993). Similar processes may also operate in desiccation-tolerant bryophytes (Dhindsa and Matowe 1981, Seel et al. 1992a) in which lipid peroxidation during drought is low. Oxidative processes can take place both in the presence and the absence of light, and light can exacerbate oxidation. This situation— oxidative stress caused or accentuated by light—is discussed in the following section.
If plants absorb more light than required during photosynthesis, they are exposed to the risk of photooxidative destruction of their photosynthetic apparatus (Long et al. 1994).
Photooxidative stress can therefore be an important limiting factor for poikilohydrous autotrophs. Bryophytes and algae that are restricted to shady habitats were shown to have limited photoprotective capacities (Oquist and Fork 1982b). Negative effects of strong light were observed in hydrated lichens in the tropical, temperate (Coxson 1987a,b), and Mediterranean region (Manrique et al. 1993, Valladares et al. 1995). Nevertheless, tolerance to strong light can be enhanced by acclimation. Cyanobacterial mats taken from exposed habitats proved to be highly tolerant to high irradiance, whereas cyanobacteria from shaded sites were very sensitive (Luttge et al. 1995). Field studies have revealed, for instance, that the cyanobacterial lichen Peltigera rufescens was at least photoinhibited under certain conditions in winter (Leisner et al. 1996). On the other hand, cryptogam species in Antarctica such as Umbilicaria aprina, Leptogium puberulum, Xanthoria mawsonii, and Hennediella heimii were very resistant to the combination of low temperatures and high irradiance while the thallus was photosyn-thetically active (Schlensog et al. 1997, Kappen et al. 1998a, Pannewitz et al. 2003, 2006).
In hydrated autotrophs, photosynthetic productivity is maintained because only that part of the light energy that is in excess to that used for energy conservation is thermally dissipated by a mechanism that requires zeaxanthin, a carotenoid of the xanthophyll cycle, and the protonation of a special thylacoid protein (Niyogi 1999, Heber et al. 2006). Thermal energy dissipation should be in equilibrium in hydrated autotrophs with ongoing photosynthesis. This means that energy dissipation is in equilibrium with energy conservation based on charge separation, the production of a strong oxidant and a reductant in the reaction centers of PS II. If energy dissipation caused is speeded up (photostress), it would inhibit photosynthesis (Wiltens et al. 1978, Oquist and Fork 1982a, Demmig-Adams et al. 1990b). Downregulation of photosynthetic processes and the so-called dynamic or recoverable photoinhibition (i.e., inhibition of photosynthesis by light, but no damage) has been observed in a number of poikilohydrous plants, bryophytes, and lichens (e.g., Seel et al. 1992, Leisner et al. 1996, Ekmekci et al. 2005), and as a result, avoidance of photooxidation (Eickmeier et al. 1993, Valladares et al. 1995, Calatayud et al. 1997, Heber et al. 2000, 2001, Bukhov et al. 2001).
A prevention of photooxidative damage by drying may be apparent from the fact that isolated Trebouxia as well as green-algal lichens resisted photostress in the field by quick desiccation under high irradiances ((Oquist and Fork 1982b, Leisner et al. 1996). This would resemble in effect the strategy of poikilochlorophyllous plants that radically destruct the photosynthetic apparatus during desiccation (Smirnoff 1993). It was hypothesized that the photosynthetic apparatus of homoiochlorophyllous autotrophs cannot be affected by strong irradiance because it undergoes a functional dissociation between light harvesting complexes and photosystem II during desiccation (Bilger et al. 1989, Smirnoff 1993). However, water content has been proved to influence both dynamic and chronic photoinhibition of lichens (Valladares et al. 1995, Calatayud et al. 1997). Some air-dried lichens typical of shady habitats exhibited even damage after exposure to high light (Valladares et al. 1995, Gauslaa and Solhaug 1996, 1999, 2000, Gauslaa et al. 2001). In addition, stenopoikilohydrous mosses were more damaged by drying at high irradiance than at low irradiance (Seel et al. 1992a).
According to recent findings since Shuvalov and Heber (2003), it has become apparent that reaction centers are capable of charge separation even in the absence of water (Heber et al. 2006a). Thus, functional reaction centers would cause damaging oxidative reactions. A revised and more comprehensive approach to understanding photoprotection in desiccated autotrophs has recently come from Heber and coauthors (Heber et al. 2000, 2001, Heber and Shuvalov 2005, Kopecky et al. 2005, Heber et al. 2006a,b) who have demonstrated that more than one photoprotective mechanism of energy dissipation is active in lichens and bryophytes. Available evidence suggests that zeaxanthin-dependent energy dissipation remains active upon desiccation (Eickmeier et al. 1993, Kopecky et al. 2005, Georgieva et al. 2005), but it is not clear whether the zeaxanthin-dependent energy dissipation is fast enough to prevent charge separation in functional reaction centers particularly in lichens and xeric bryophytes.
A second protective mechanism was evident from the finding that functions of the reaction center of PS II can change upon desiccation (Heber et al. 2006a,b). In desiccated samples of the moss Rhytidiadelphus squarrosus, energy dissipation has been shown to occur in PS II reaction centers. In this case, a photoreaction is responsible for the formation of a quencher of fluorescence in the reaction center (Heber et al. 2006b). A third protective mechanism was apparent from the observation that light was not even necessary for the formation of a quencher during desiccation. After lichen thalli were carefully predarkened to avoid light activation of the mechanism of energy dissipation, fluorescence was quenched after desiccation took place in darkness. This reveals the activation of the third mechanism of thermal energy dissipation (Heber et al. 2006b, Heber personal communication). The latter two protective mechanisms were operating only under desiccation and ceased by rehydration. Lichens with cyanobacteria as photobionts lack the zeaxanthin-dependent thermal energy dissipation mechanism; however, other carotenoids may play a role at least in the hydrated state (Demmig-Adams et al. 1990b, Leisner et al. 1994, Lange et al. 1999). In the desiccated state, the desiccation-induced thermal energy dissipation mechanism (mechanism 3) may be operating (Heber, personal communication). As soon as water becomes available to the chloroplasts of homoiochlorophyllous autotrophs, photosynthetic water oxidation is resumed (Kopecky et al. 2005, Heber et al. 2006a).
In vascular plants, only the zeaxanthin-dependent dissipation mechanism is known to be protective also in the desiccated state. It also operates in homoiohydrous xerophytic plants (e.g., Nerium oleander. Demmig et al. 1988; Clusia spp. with CAM: Winter and Koniger 1989). In resurrection plants, it was shown with Selaginella lepidophylla (Casper et al. 1993, Eickmeier et al. 1993). Accumulation of zeaxanthin upon drying was reported for the poikilohydrous species Craterostigma Plantagineum (Amalillo and Bartels 2001) and Boea hygrometric (see Yang et al. 2003). In R. serbica, the photoprotection appeared to be achieved, when dry, by the zeaxanthin-dependent dissipation as well as by ascorbate and glutathione (Augusti et al. 2001). In the poikilochlorophyllous X. scabrida, 22% of the carotenoids were still preserved in the dry leaves when the photosynthetic apparatus was dismantled, but the carotenoids seemed to be protective or essential when the chloroplasts reorganized during rehydration (Tuba et al. 1993b).
Poikilohydrous autotrophs exhibit various photoprotective mechanisms in addition to the thermal energy dissipation via carotenoids. In the case of lichens, filtering or screening effects of the upper cortex formed by the mycobiont and of certain secondary compounds such as parietin have been shown to be potentially important in reducing the risk of photodamage of the photosynthetic units (Budel 1987, Solhaug and Gauslaa 1996, Kappen et al. 1998a, Gauslaa and Ustvedt 2003, Gauslaa and McEvoy 2005). Anthocyanins, which reflect photo-synthetically active light, may prevent excessive light-chlorophyll interaction. An accumulation of anthocyanins was observed in sun-exposed leaves of several resurrection plants, mainly on the abaxial leaf face, which is everted when the leaves are curled or folded (Farrant 2000). Leaf and shoot curling during drought, despite not effective in certain bryophytes (Seel et al. 1992a), can confer photoprotection simply by shadowing. For instance, leaf curling by everting the reflectant abaxial leaf surface was effective in Polypodium polypodioides, a species sensitive to strong light under water stress (Muslin and Homann 1992, Helseth and Fischer 2005). Additionally, photorespiration and light-activated photosynthetic enzyme activity were not affected by intense radiation in S. lepidophylla due to leaf curling (Lebkuecher and Eickmeier 1991, 1993). The protective role of curling is important for plants growing in open and exposed habitats and, although not yet thoroughly explored, it may be relevant also for lichens such as Parmelia convoluta and Chondropsis semiviridis, or for thallose desert liverworts all everting a whitish underside when dry (Lange et al. 1990a).
Another radiative stressor is UV B, particularly in polar and alpine regions; however, experiments proved that it never harmed lichens and mosses in open habitats (Lud et al. 2003,
Nybakken et al. 2004). Moreover, UV B (280-320 nm) was shown to be an essential requisite for the synthesis of sun-screening pigments (parietin, melanin) in lichens (Solhaug et al. 2003).
Desiccation Tolerance: An Old Heritage
Taking desiccation as a fundamental heritage, geneticists and molecular biologists were challenged to trace down this capability of plants to the beginning of plant evolution and to ask whether this is unique to poikilohydrous organisms and why it is absent in the vegetative parts of the majority of vascular plants and certain bryophytes, and, whether it is mono- or polyphyletically evolved, and, on which genes desiccation tolerance is located.
According to Oliver et al. (2000) recent synthetic phylogenetic studies confirm the idea that vegetative desiccation tolerance is primitively present in the bryophytes but was then lost in the evolution of vascular plants. It is hypothesized that desiccation tolerance was crucial for ancestral fresh-water autotrophs to live on land. The tolerance might have been lost on the way of a more complex (homoiohydrous) organization of the vascular plants, and independent evolution or re-evolution of desiccation tolerance has happened in the Selagi-nellales, leptosporangiate ferns, and the 10 families of angiosperms with poikilohydrous species. Evolutionary progress may be evident from the fact that pteridophytes, in common with bryophytes, are able to synthesize rehydrins but can also synthesize dehydrins, the only capability of angiosperms. Tracing possible pathways by identifying possible gene orthologs of LEA-like proteins that are synthesized upon dehydration provided a network of land plant phylogeny. Oliver et al. (2005) proved their earlier theory and extended it by discussing the fact that several genera of bryophytes (Haplomitrium, Sphagnum, Takakia, Tetraphis) lack the particular genes and are not desiccation tolerant either constitutively or due to loss of phenotype, which involves the possibility that precursors of these bryophytes also did not realize desiccation tolerance in the vegetative state. There was also negative evidence that ancestors of the hornworts and of all vascular plants were desiccation tolerant. Keeping up the idea that desiccation tolerance of spores may elucidate the problem, Illing et al. (2005) demonstrated by molecular responses a crucial role of orthodox seeds as the carriers of genes for desiccation tolerance, which may, or not, be expressed in the vegetative part of vascular plants. Analyzing sequences of genes of Arabidopsis seeds and a set of vegetative homoio- and poikilohydrous plant species with respect to certain LEA proteins, RNA transcripts for antioxidants that are expressed during desiccation, and in addition, sugar accumulation revealed the following results: Among the antioxidant activities only that with 1-cys perox-idase appeared to be a mechanism specific to desiccation tolerance of resurrection plants and seeds. Most of the antioxidants are activated also as a response to various other abiotic stresses. However, genes belonging to the LEA six superfamily are uniquely associated with dehydration stress in resurrection plants and in seeds. Moreover, sucrose accumulation upon drying seems to be a desiccation-tolerance specific mechanism of resurrection plants and orthodox seeds, albeit it was also observed in some recalcitrant seeds (Kermode and Finch-Savage 2002). As a general result, desiccation tolerance is not considered the result of parallel origins, and the loss of desiccation tolerance can be interpreted as the result of suppression of latent genes in the vegetative state of the plant. However, single gene loss in certain instances (Dickie and Pritchard 2002) cannot be excluded.
Generally speaking of desiccation tolerance suggests that it relates to a fundamental uniform mechanism; however, a differentiated view is obvious now. On the one hand, it has to be realized that the most primitive autotrophs, such as Cyanobacteria, are extremely desiccation tolerant, whether they live in water or in the porous space of rock. This indicates that desiccation tolerance is a very early trait in evolution. Moreover, the fact that organelles such as chloroplasts of spinach (Table 2.1) are desiccation tolerant and that vegetative tissues can be lyophilized without harm suggests that it is the way of withdrawing water that can be damaging but not the dehydration itself. Thus, it can be concluded that the capability of surviving dehydration must be basic and ubiquitous. On the other hand, phenotypic tolerance must have been acquired at various advanced stages of organismic complexity by evolving metabolic or structural protective mechanisms. These must be very specific, either because of a distinct combination or quantitatively selected ubiquitous "house-keeping" mechanisms (Illing et al. 2005), or because of inventing genetic structures that act species-specific, as most likely the CDT-1 gene in C. plantagineum (Furini et al. 1997, Bartels and Salamini 2001). So far it is reasonable to consider desiccation tolerance of vascular plants (and highly derived bryophytes) as modified (see Oliver and Bewley 1997) or secondary trait (Raven 1999).
Tolerance to Extreme Temperatures: A Property Linked to Poikilohydry
Plants capable of surviving desiccation may be tolerant also to other environmental stresses such as extreme temperatures. Some lichen species, for example, are extremely resistant to freezing both in hydrated and dehydrated states. The freezing tolerance of hydrated lichens can exceed by far the temperature stresses that occur in winter or in polar environments (Kappen 1973). Bryophytes are less freezing-tolerant, but many species are well adapted to live and persist in cold environments (Richardson 1981, Proctor 1982, Longton 1988b). Fern gametophytes and vascular resurrection plants from the temperate zone have a moderate (—9°C to — 18°C) freezing tolerance in winter (Kappen 1964, 1965). Most poikilohydrous vascular plants come from subtropical climates and are sensitive to freezing. For example, Borya nitida can only survive temperatures between — 1°C and —2°C in winter (Gaff and Churchill 1976), and poikilohydrous grass species do not survive temperatures below 0°C (Lazarides 1992). However, desiccated leaves, fronds, or other vegetative parts become highly tolerant to cold depending on the remaining water content. For instance, experiments with Ramonda myconi, Polypodium vulgare, Myrothamnus flabellifolius, and gametophytes of several fern species revealed that the tissues resisted — 196°C if they were desiccated to a relative water content of, for example, 6% (Kappen 1966, Vieweg and Ziegler 1969, Pence 2000). Thus, the tolerance of these vascular poikilohydrous plants to low temperatures was not different from that of dry algae, lichens, and bryophytes (Levitt 1980), which also indicates water content as a crucial factor in the freezing tolerance of organisms in general.
Heat tolerance of plants can also be increased if they are desiccated (Kappen 1981). This fact becomes ecologically relevant if we consider that dry plants can easily heat up to over 50°C in their natural habitats. Thallus temperatures in dry moss turfs and dry lichens can reach 60°C-70°C under field conditions (Lange 1953,1955, Richardson 1981, Proctor 1982, Kershaw 1985). A similarly high temperature was reported for desert soil around Selaginella lepidophylla (Eickmeier 1986). The temperature in the rock pools where Chamaegigas intrepidus lives can reach 41°C (Gaff and Giess 1986). Although some lichens such as Peltigera praetextata and Cladonia rangiferina did not survive temperatures higher than 35°C, even in the desiccated state (Tegler and Kershaw 1981,Gauslaa and Solhaug 1999, 2001), tolerance to 70°C and even to 115°C was recorded for many other dry lichen and bryophyte species (Lange 1953,1955, Proctor 1982, Meyer and Santarius 1998). Heat tolerance of bryophytes and vascular plant species varies with season. The maximal temperature tolerated in the turgescent state by the temporarily poikilohydrous fern P. vulgare and by R. myconi was highest in winter (approximately 48°C), and by decreasing water content, the heat tolerance could be increased to approximately 55°C (Kappen 1966, Figure 2.5). C. intrepidus was able to thrive at 60°C temperatures (Heil 1925), and dry leaves of M. flabellifolius were reported to have resisted at 80°C (Vieweg and Ziegler 1969). Eickmeier (1986) could demonstrate an increase in photosynthetic repair capacity of S. lepidophylla if dry plants were subjected to 45°C and 65°C, but he found also that desiccation tolerance decreased with increasing temperatures (25°C, 45°C, 65°C). However,
Water saturation deficit (%)
FIGURE 2.5 Shift of heat tolerance limit with increasing water saturation deficit in leaves of Ramonda myconi. (After Kappen, L., Flora, 156, 427, 1966.)
Water saturation deficit (%)
FIGURE 2.5 Shift of heat tolerance limit with increasing water saturation deficit in leaves of Ramonda myconi. (After Kappen, L., Flora, 156, 427, 1966.)
since the water content of these dry plants was not defined, it still remains unclear whether metabolic disturbance at the higher temperature was weakening the plants.
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