PHYSIOLOGIA PLANTARUM 103: 437-441. 1998 Printed reland—al rights reseed Copyrgh © Physlogia Plantarum 1998 Minireview Heat-shock proteins and cross-tolerance in plants Adnan Sabehat, David Weiss and Susan Lurie Sabehat, A., Weiss, D. and Lurie, S. 1998. Heat-shock proteins and cross-tolerance in plants. ~” Physioi. Plant. 103: 437-441 Environmental stresses dramatically affect plant survival and productivity. Because plants are immobile, presumably different strategies are required for protection against transient stresses. Under stress, plants synthesize specific proteins, and their accumulation has a role in protecting the tissue ftom possible damage. An increasing ‘number of studies show the existence of eross-tolerance in plants: Exposure of tissue to moderate stress conditions often induces resistance to other stresses. Many varied ‘mechanisms explaining the phenomenon of eross-tolerance have been proposed, and they often, but not always, suggest that specific proteins are induced by one kind of stress and are involved in the protection against other kinds. Although various cross-protections have been demonstrated in a number of plants, a common mecha- nism has not been found. This review discusses heat-shock proteins and their possible roles in protecting the plant under heat and other stresses. Key words — Crosstolerance, cross-protection, heat-shock proteins, stress injures, stress proteins. 4, Sabehat and D. Weiss, The Kennedy-Leigh Centre for Horticultural Research, The Hebrew Univ. of Jerusalem, P.O. Box 12, Rehovot 76100, Israel; 8. Lurie (correspond- ing author, e-mail vilurie@voicani.agri-gov.il), Dept of Postharcest Science, Agricul- tural Research Organization, The Voleani Center, Bet Dagan 50250, Israel Introduction. Because higher plants are immobile, they may have a greater need of protection from transient stresses, such as high or low temperature, high or low pH, salinity, drought, oxidative conditions or heavy metals, which dramatically affect their survival and productivity. In many plants, gradual changes in the environmental conditions induce tolerance to the extreme situations. Stresses such as anaerobiosis (Sachs et al. 1980), water deficit (Rhodes and Matsuda 1976), heat shock (Key et al. 1981) and heavy metals (Orzech and Burke 1988) lead to a rapid shift from polysomes to monosomes, and to a low rate of total protein synthesis. However, while the synthesis of many pre-stress proteins is inhib- ited (Sachs et al. 1980, Key et al. 1981), the expression of other specific stress proteins is induced (Sachs and Ho 1986). Some of these newly-synthesized proteins are required for stress tolerance, and their accumulation has a role in protecting the tissue from possible damage Received 30 November, 1997; revised 30 January, 1998 Phyo. Pan. 10, 198 caused by these stresses (Sachs and Ho 1986). The alteration of protein level is a reflection of both tran- criptional and translational regulation (Sachs et al 1980, Key et al. 1981, Matters and Scandalios 1986, Sachs and Ho 1986). An increasing number of studies show the existence of cross-tolerance in plants: Exposure of tissue to mod- erate stress induces resistance to other stresses. This, cross-protection has been shown for different kinds of stress. Water stress confers cold hardiness in a variety of winter cereals (Siminovitch and Cloutier 1982, Cloutier and Andrews 1984) and induces chilling resis- tance in rice (Takahashi et al. 1994), Salt stress stimu- lates cold hardiness in potato and spinach seedlings (Ryu et al. 1995). Mechanical stress (brushing) in- creases chilling tolerance in tomato leaves (Keller and Steffen 1995), and ozone stress induces a defensive pathway against fungal attack in parsley (Kaltenbach et, al. 1994), Heat stress protects against heavy-metal toxi- city (Bonham-Smith et al. 1987, Orzech and Burke 4371988), increases salt resistance (Kuznetsov et al. 1993), induces water-stress tolerance (Bonham-Smith et al 1987) and reduces chilling injury in chilling-sensitive species, such as in tomato fruits, mung bean hypocotyls and cucumber cotyledons (Lafuente et al. 1991, Lurie and Klein 1991, Jennings and Saltveit 1994, Collins et al 1995, respectively). Abbreviations ~ LEA, late embryogenesis abundant; HSP, hneat-shock protein; smHSP, small heat-shock protein; Pf pathogen response, Cross-tolerance in plants: Protein synthesis and possible mechanisms Many varied mechanisms explaining the phenomenon of cross-tolerance have been proposed and they often, but not always, suggest that specific proteins are induced by one kind of stress and are involved in the protection against other kinds. Several cold-regulated proteins are homologous and have similar properties to those of drought- and ABA-induced proteins, such as LEAs, RABs, and dehydrins (Gilmour and ‘Thomashow 1991, Neven et al. 1993). The induction of chilling resistance by water stress involves the activation of drought-regulated genes (Takahashi et al. 1994), and salt stress stimulates cold hardiness by the activation of cold- and ABA-responsive genes (Ryu et al. 1995). Ozone and heavy-metal stress induce the synthesis of PR proteins and enhance plant resistance against fungal attack in parsley (Kaltenbach et al. 1994), Moderate heat shock induces the expression of heat-shock proteins (HSPs) in tomato and protects the fruits against chilling injury. It has been suggested that these HSPs are in- volved in the protection of the fruits against both high- and low-temperature damage (Sabehat et al. 1996) ‘One mechanism that may be involved in the resistance to many types of stress is the activity of the antioxidant pathway. The enzymes in this pathway include superox- ide dismutase, peroxidase and catalase, as well as those of the ascorbate-glutathione cycle. High activity levels of these enzymes have been found in response to heat, chilling, freezing, salt, drought and wounding, as well as, to oxidative stress (Foyer et al. 1994). The overproduc- tion of antioxidant enzymes under different kinds of stress may suggest that these proteins have a general role in the acquisition of tolerance by plants. ‘The interaction between heat, salt and heavy-metal stresses may, in some systems, involve the accumulation of low-molecular-mass compounds such as putrescine, ‘osmotin-like proteins, proline or linear polymers of glutathione (Bonham-Smith et al. 1987, Kuznetsov et al. 1993), Specific low-molecular-mass compounds accumu- late in a number of plants in response to a particular stress and it has been suggested that these compounds protect the tissue against another kind of stress. The function of these molecules in the acquisition of toler- 438 ance is still unknown, They do not normally accumulate in sufficient concentrations to be osmoprotectants, Although various cross-protections have been de- monstrated in a number of plants, a common mecha- nism has not been found. It may be that a plant can have a number of ways of dealing with a stress, using different stress proteins. Alternatively, different plants may have different mechanisms of defense against the same kind Of stress. What is still missing is an overall picture tying, the various studies into @ coherent whole. Heat-shock proteins and cross-tolerance Plants are sensitive to high and low temperatures, and both extremes inhibit photosynthesis, growth, pollina- tion, fruit set and fruit development (Vierling 1991). Plants from temperate zones are less sensitive to low than to high temperatures, whereas the opposite is true for plants originating from tropical or subtropical areas. The latter include tree crops such as mango, avocado and banana, as well as many vegetables, such as cucum- ber, pepper, squash and tomato. However, plants from both groups can be damaged by extended exposure to both high and low temperatures (Wang 1994). Plants, can be partially protected against extreme temperatures if conditions are changed gradually. Raising the temper- ature slowly to 38°C allows plants to acctimate and tolerate further increases to temperatures that are nor- mally lethal (Vierling 1991). Exposing plants to 15°C acclimates them to lower temperatures, which would normally cause chilling injuries (Wang 1994). Since these adaptations can be prevented by cycloheximide, it is reasonable to assume that de novo protein synthesis is required for the protection against both high- and low- temperature injuries (Vierling 1991), All organisms respond to high temperatures by in- ducing the synthesis of a small group of evolutionarily- conserved polypeptides known as HSPs. When plants ate exposed to high temperatures they synthesize both, high-molecular-mass HSPs (ranging from 60 to 110 kDa) and small HSPs (smHSPs), ranging from 15 to 45 kDa. Some plant HSPs are required for normal growth at the higher end of their normal growth-temperature range, whereas others help cells to withstand the ad- verse effect of extreme temperatures (Vierling 1991). The accumulation of HSPs is also induced by low temperatures. Recently, a class of proteins that accumu- late at low temperatures has been isolated and iden- tified as HSP70 and HSP90 (Neven et al. 1993, Krishna et al. 1995), The HSP70 family is known to have roles in processes of protein transport, folding, assembly and disassembly (Gething and Sambrook 1992). HSP90 proteins are known to have a general chaperone func- tion in vertebrate cells (Pratt 1993). The accumulation of two HSPs (104 and 109 kDa) has been detected during the acclimation of rice plants to low tempera- tures (Pareek et al. 1995) and the accumulation ofsmHSP transcripts has been observed during cold stor- age of potatoes (van Berkel et al. 1994). HSPs are also induced by other kinds of stress, such as drought, flooding, salinity and heavy metals (Nover et al. 1989, Anderson et al. 1994, Coca et al. 1994, Kiyosue et al 1994, Pareek et al. 1995). It is possible that a specific environmental stress induces the synthesis of specific HSPs which, with other siress proteins, either protect the plant from damage or help repair the damage caused by this stress, There is very likely some overlap- ping in function among the different HSPs, which may explain cross-tolerance Moderate heat shock has been found to protect against maay other kinds of stress in a number of species (Bonham-Smith ct al. 1987, Lurie and Klein 1991, Kuzznetsov ct al. 1993). Protection against chilling injury conferred by moderate heat shock in chilling-sen- sitive species has been investigated in several plants (Klein and Lurie 1991, Lafuente et al. 1991, Jennings and Sakvveit 1994, Collins et al. 1995). When mature green tomato frutis are stored at 2°C for 10 days, they develop chilling-injury symptoms and do not ripen when transferred to ambient temperature. However, if the fruits are first held at 38°C, they can be transferred to 2°C for several weeks without developing chilling- injury symptoms (Lurie and Klein 1991). These fruits then ripen normally under ambient temperatures. The same moderate heat shock can protect against a higher Heat shock 58°C Cold shock 2c Moderate heat shock 38°C High Cross protection Chilling temperature injury injury Fig. 1. Schematic illustration demonstrates eross-toleranee in toimato fruits, When tomato fruits are exposed 1 high (55°C) temperature for a few minutes or low (2°C) temperature for a few days they develop stress injuries. However, ifthe Frits are first held at a moderately high temperature (38°C), they di velop tolerance to both extremes. This moderately high’ tem- perature induces the synthesis of several HSPs. It ts suggested that these HSPs are involved in the mechanism of eross-pro- tection, Phil, Pat 103, 18 temperature stress and low-temperature injury (Fig. 1) Protection against chilling injury by high-temperature treatment has also been found in mung bean hypocotyls, (Collins et al. 1995) and in cucumber cotyledons and seeds (Lafuente et al, 1991, Jennings and Saltveit 1994) High temperatures induce the synthesis of several smHSPs in tomato fruits (Sabehat et al, 1996), These newly-synthesized smFSPS are also found in heated fruits after three weeks at 2°C. Loss of protection against low temperature is correlated with the disap- ppeatance of these proteins from the tissue. These results support the claim for a role played by HSPs in protec- tion against low-temperature damage. A similar correla- tion between expression of HSPs and tolerance to chilling injuries has been found in mung bean hypocotyls and cucumber cotyledons and seeds (La- fuente et al, 1991, Collins et al. 1995). To study the molecular mechanism involved in the acquisition of tolerance to chilling injuries by heat shock, we recently cloned several HSP genes that are expressed at low temperature in pre-heated fruits. Most of the cloned eDNAS belonged to tie smHSP group. One of the isolated clones, ficit2, encoding a heat- induced protein of 16.5 kDa, lacks the consensus se- quences of smHSPs, but has a putative trans-membrane hydrophobic sequence, suggesting membrane localiza- tion (Sabehat et al. 1998). One of the first symptoms of many kinds of stress including heat and chilling is a massive increase in ion and solute leakage caused by the loss of membrane components (Levitt 1980). Cooper and Ho (1987) identified several membrane-associated. simFSPs and HSP70 in maize, and suggested their potential role in maintaining normal membrane func- tions during heat stress. simFSP-enriched fractions from several plants provide thermo-stabilization in vitro to ‘membrane-associated proteins (Jinn et al. 1993). The HCIT2 protein may act to protect the cell membrane during both high- and low-temperature stresses. A study of the accumulation of fcit2 and smHSP transcripts in tomato fruits, following heat treatment and during storage at low temperature, showed the continued presence of these transcripts. This is in con- trast to the relatively rapid turnover of HSP transcripts if a tissue is removed from heat to 20 or 25°C. In the latter case, the transeripts disappear after 12 to 48h (Nover et al. 1989), [t may be that the low temperatur in slowing most cellular processes, also decreases the rate of mRNA turnover. It was found that the HSP transcripts present at low temperature are still active in synthesizing proteins (Sabehat et al. 1996). Several other studies have shown that specific smHISP genes are expressed at low temperatures. Class TI smHSP gene from heated tomato fruits is induced by high temperature and is also expressed when the heated fruits are transferred to low temperature (Kadyrzhanova et al. 1998), Similar results were found, 439