American Journal of Botany 89(3): 375±382. 2002. INVITED SPECIAL PAPER

THIGMO RESPONSES IN PLANTS AND FUNGI1

MORDECAI J. JAFFE,A.CARL LEOPOLD,2 AND RICHARD C. STAPLES

Boyce Thompson Institute, Cornell University, Tower Road, Ithaca, New York 14853 USA

Thigmo mechanisms are adaptations that permit a plant to alter growth rates, change morphology, produce tropisms, avoid barriers, control germination, cling to supporting structures, infect a host plant, facilitate pollination, expedite the movement of pollen, spores, or seeds, and capture prey. Through these varied functions, plant thigmo systems have evolved impressive controls of cell differenti- ation, localized growth rates, regulated synthesis of novel products, and some elegant traps and projectile systems. For most thigmo events, there will be a dependence upon transmission of a signal from the cell wall through the plasmalemma and into the cytoplasm. We propose the possible involvement of integrin-like proteins, Hechtian strands, and cytoskeletal structures as possible transduction components. Many thigmo mechanisms may use some modi®cation of the calcium/calmodulin signal transduction system, though the details of transduction systems are still poorly understood. While transmission of thigmo signals to remote parts of a plant is associated with the development of action potentials, hormones may also play a role. Thigmo mechanisms have facilitated an enormous array of plant and fungal adaptations that make major contributions to their success despite their relatively sessile or immobile states.

Key words: mechanosensor; sensing; thigmomorphogenesis; thigmotactic; thigmotropism.

The relative immobility of plants as compared with animals We expected there to be commonalities among the thigmo has naturally provoked a dependence upon their ability to responses with regard to perception, transduction, and re- sense and respond to subtle environmental signals. Among sponse mechanisms. The evidence for such commonalties is these, the adaptations of plants to touch, i.e., thigmo responses, scattered at the present time, but an overview of some re- have become an elaborate and often elegant series of mechan- peating themes from relevant experimental studies may be ical signalling systems. Thigmo responses are found over the helpful in developing a general scheme for thigmo-regulated entire range of the plant kingdom, from fungi to algae, mosses, mechanisms. We have undertaken this review to point out the ferns, and higher plants. In this review we attempt to bring commonalities and to offer at the end a unifying mechanism. together an assemblage of thigmo responses under four cate- gories and then consider the mechanisms that may initiate THIGMOTACTIC, THIGMOMORPHOGENETIC, AND them. THIGMOTROPISTIC RESPONSES A thigmo stimulus may be quite subtle. For example, in handling experimental plant materials, just the act of moving In many instances, a thigmo stimulus leads to changes in a potted plant from the greenhouse to the laboratory may ex- plant movement. For example, swimming movements reverse pose the plant to suf®cient mechanical perturbation to cause direction of the alga Euglena when it runs into an obstacle changes in the morphology and growth rate of the plant (Jaffe, (Jennings, 1906). Similar movement adaptations occur with 1985). Indeed, moving the plant can result in inhibition of swimming Chilomonas and Cryptomonas cells (Nageli, 1860) growth rate, and touching the plant can result both in alter- and in the motile spores of many algae (Strasburger, 1878). ations of phloem cell structure (Jaffe and Leopold, 1984) and Changes in both growth rates and morphological differen- differentiation of stem pith (Jaffe and Lineberry, 1988). tiation, due to thigmomorphogenesis, are illustrated by thigmo Responses to mechanical perturbation can occur with im- responses of numerous fungi. The ridge pattern on the surface pressive rapidity; for example, touching the leaf of Mimosa of a leaf can induce a directional orientation of mycelial will cause leaf folding beginning within 1 s (Jaffe, 1985); dis- growth (thigmotropism) (Johnson, 1934; Hoch et al., 1987). turbing the trigger hairs on a Venus ¯ytrap leaf will cause the In another example, in certain species of the rust fungi, sensing trap to close within 1 s; and touching the trigger hairs in a the stomatal ridge will actually induce an alteration of cell Utricularia bladder cause the trap to close within an estimated differentiation (thigmomorphogenesis). Thus germ tube 0.01 s (Jaffe, 1985; Simons, 1992). growth over a stomatal ridge can alter growth rates and induce Although the initial thigmo stimulus is received by the cell appressorium differentiation in various species of rust fungi, wall, the magnitude of the deformation may be very slight as in Puccinia or Uromyces (Allen, 1923; Wynn, 1976), or indeed. For example, localized pressure can elicit the trans- bring about encystment of hop downy mildew zoospores (Dea- location of the nucleus and cytoplasm, the intracellular gen- con and Donaldson, 1993). Again, in the bean rust fungus eration of reactive oxygen species, and the transcription of (Uromyces appendiculatus), the stimulus for appressorium dif- defense-related genes (Gus-Mayer et al., 1998). Furthermore, ferentiation is induced when the germ tube grows over a tiny these tiny perturbations can be transduced into enormous stomatal ridge on the epidermis of the host plant, causing a changes in morphology, growth rates, and metabolic products, termination of elongation of the sporeling germ tube, followed as for example in thigmo growth and tropistic responses, thig- by activation of cell division and development of the appres- mo secretions, thigmo traps, and thigmo projectiles as re- sorium (Allen, 1923; Maheshwari, Hildebrandt, and Allen, viewed below. 1967). The mechanical nature of the thigmo stimulus is evi- dent in the fact that cellular differentiation can be stimulated 1 Manuscript received 21 August 2001; revision accepted 30 October 2001. by contact of the germ tube with a plastic impression of the 2 Author for reprint requests (e-mail: [email protected]). leaf stomatal surface. In some other cases, differentiation of 375 376 AMERICAN JOURNAL OF BOTANY [Vol. 89 zoospores at stomatal locations may also involve a cooperative water loss from one side of the pulvinus, and closing of the action with some chemical signals emerging from the leaf sto- leaf, a constriction in the vascular strand serving as a hinge ma (Royle and Thomas, 1973). (Esau, 1965). The thigmo stimulation of Mimosa leaf move- A dramatic example of thigmomorphogenesis is observed ment involves an action potential, which occurs ®rst in the in the growth of a Monstera vine. On the ground, the seedling stimulated lea¯et and then advances to each of the other pul- grows initially in a tropistic manner toward a dark object (sko- vini above and below it (Sibaoka, 1966; Pickard, 1971). The totropism); when it touches a tree, the thigmo stimulus chang- movement of the signal has been suggested to involve the es from skototropic to phototropic, and it climbs the tree ad- movement of a water-soluble substance (Ricca, 1916), which hering to the tree's bark. At the same time the leaf form is could be hormonal (Umrath and Thaler, 1980). thigmomorphogenetically altered from small nearly circular leaves to large ®nely divided leaves as it climbs the tree (Buss- THIGMO SECRETIONS man, 1939). Many plant adaptations to physical perturbations involve the Thigmomorphogenesis is involved in numerous types of ad- synthesis or secretion of adhesives. Examples among the fungi aptation to stress. These include adaptation to wind, drought, are the spores of Magnaporthe, which respond to the sensing frost, and infection (Aist, 1977; Pressman et al., 1983; Te- of landing on a surface by secreting an adhesive that holds the lewski and Jaffe, 1986a, b; Gilroy, Read, and Trewavas, 1990; spore at the landing site while the spore germinates (Hamer et Coops and Van der Velde, 1996). However, thigmomorpho- al., 1988). In some instances the secretion of the adhesive is genetic responses may involve different regulatory systems. necessary for spore germination (Shaw and Hoch, 1999). A For example, suppressed elongation in response to touch may variant of this requirement occurs in spores of Chochliobolus, involve an inhibition of auxin effects in Bryonia dioica (Boyer, which secrete adhesive only after experiencing both the con- 1967), or by an increase in abscisic acid content in Phaseolus tact and a wetting (Braun and Howard, 1994). A parallel in- vulgaris (Erner and Jaffe, 1982), or a biosynthesis of ethylene stance is the secretion of vitronectin-like molecules by Fucus in Pinus taeda (Telewski and Jaffe, 1986c). zygotes in attaching to a solid surface (Wagner, Brian, and Some of the most common thigmo responses are the tro- Quatrano, 1992). pistic responses. Thigmotropism most often involves unilateral In higher plants, the secretion of adhesives provides for at- growth inhibitions (Jaffe, 1976b), as in the enhancement of tachment to trees or rocks by epiphytes, lichens, and some tropistic bending of styles, ¯owers, or petals toward visiting climbing vines and roots. The attachment of vines to walls pollinators (Jaffe, Gibson, and Biro, 1977; Faegri and van der involves the formation of stunted roots, which become hold- Pijl, 1979). The ¯owers of the orchid Melampyrum sense vi- fasts, secreting strong adhesive substances (Darwin, 1881; brations of a visiting bee and respond by releasing a shower Dyer, 1979). Such adhesive, vitronectin-like proteins are of pollen (Meidell, 1945). Likewise, in the ¯owers of Pedi- formed at areas where the cell wall contacts a solid surface. cularis, a visiting bee may shake the stamens by wing vibra- Proteinaceous adhesives are involved in the sticking of pol- tions and collect the falling pollen on its back (Macior, 1969). len to the stigma of some ¯owers (Lolle and Pruitt, 1999). The thigmonastic coiling of tendrils following a mechanical When compatible pollen touches the stigma of ¯owers of bean perturbation occurs in at least two phases in pea, an initial and several other legumes, and ¯owers of Arabidopsis, the rapid nastic bending, which is predominantly a hydraulic-driv- stigma secretes an adhesive substance that binds the pollen. en contraction on the ventral side of the tendril (Jaffe and The stigma then creates a channel of lipidic substances that Galston, 1968), and a subsequent growth on the dorsal side facilitate the growth of the pollen tube (Sanders et al., 1991; (Jaffe and Galston, 1966). It has been suggested that the sens- Lolle and Pruitt, 1999). Another instance of thigmo-secretory ing of the touch by tendrils may be achieved by small tactile activity is the case of formation of mucilage on the leaf-hairs protuberances on the ventral side that have particularly thin of insect-trapping plants such as sundews and butterworts. As epidermal walls (Tronchet, 1964). an insect enters the leaf-trap, its movements constitute the thig- The thigmonastic twining of the stems or roots of strangler mo stimulus causing the trap to produce more mucilage, in- ®gs (Ficus costaricensis) deserves mention here. The seedling creasing the immobilization of the insect (Lloyd, 1942). starts as an epiphyte on a host tree. As it develops, the root of the ®g plant winds down around the target tree, and it im- THIGMO TRAPS poses an increasing pressure on the tree through the differen- tiation of increasing amounts of cortical tissue on the side Insect traps occur principally in sunny, moist environments away from the target tree. As this pressure against the tree of extreme nitrogen de®ciency (Givnish et al., 1984), such as increases, the twining ®g ultimately kills the host tree and the in sphagnum bogs where the acidic pH limits the degradation ®g becomes free-standing (Putz and Holbrook, 1989). of organic matter. Among carnivorous plants there are ®ve Perhaps the most heralded thigmo responses are the nastic types of thigmo traps. The sundews (Drosera) form rosettes leaf-folding movements in response to touch. A mechanical of green or red spoon-shaped leaves with tentacles on the up- perturbation of Mimosa pudica leaves results in the dramatic per surface. Each tentacle secretes globules of mucilage, which folding and lowering of the leaves. Such nastic leaf move- glisten in the sunshine and attract small insects. The thigmo ments occur also in Cassia sensitivum and some species of stimulus generated by an insect landing and struggling with Oxalis (Umrath, 1958). While this thigmo response is very the sticky mucilage results in several responses: a bending of dramatic, its physiological function is very much in doubt. The the tentacles toward the captured prey, an increased secretion leaf-folding movements in sensitive plants have both structural of mucilage, a closure movement of the leaf, and subsequently and physiological similarities to the diurnal folding of leaves the secretion of digesting enzymes by the leaf (Lloyd, 1942). that occurs commonly in legumes. Both movements involve The butterworts (Pinguicula) generate an insect trap involv- hydraulic shifts in glandular organs, the pulvini, at the base of ing the formation of two types of glandular tentacles: stalked leaves or lea¯ets. Structurally, the folding movement involves glands that produce mucilage and sessile glands that produce March 2002] JAFFE ET AL.ÐTHIGMO RESPONSES 377 digestive juices. The thigmo stimulus of an insect landing re- patiens) is a common example. As the fruit matures, the outer sults in a gradual rolling of the leaf serving to engulf the insect shell develops lines of cell separation, which, when combined as it is digested (Darwin, 1881; Lloyd, 1942). An impressive with a partial drying of the tissues, constitute a poised spring. adaptation of the butterworts is that during the winter season Thigmo stimulation (touching) will cause the fruit to explode, when insects would not be expected to visit, the secretions do and the seeds will be thrown for a meter or more from the not occur, even if given a thigmo stimulation (M. J. Jaffe, mother plant. This type of seed projection is very common, personal observation). occurring for example in Geranium, manioc, and in broom and The venus ¯ytrap (Dionaea) operates on a different basis. gorse bushes (Simons, 1992). Instead of secreting mucilage, the photosynthetic leaves de- Exploding ¯owers are another type of thigmo system. In velop a trap involving a row of spines on each margin and a many legumes, such as alfalfa (Medicago), when a ¯ower is cluster of trigger-hairs at the center of the leaf. Thigmo stim- being visited by a pollinating insect, its entry spreads some of ulation by a landing insect or spider causes a very rapid clo- the ¯ower parts apart, causing the ¯ower to ``explode,'' sud- sure of the two lobes of the leaf; the spines then restrain the denly exposing its anthers or stigmata (Faegri and van der Pijl, prey while digestive juices are secreted. An impressive feature 1979). of this thigmo trap is the rapidity of the trap closure, viz. A more aggressive strategy is taken by some orchids such shutting like a castle's portcullis, it closes within a second as the twayblade (Listera). As a bee enters the ¯ower or hovers (Brown, 1916). close to the ¯ower, the thigmo perturbation of wing vibration Traps are not restricted to land plants. The aquatic Aldro- causes the ¯ower to ¯ing a pollinium at the bee, along with a vanda is a tiny aquatic version of the trap mechanism of Dion- quick-drying adhesive that attaches the pollinium to the face aea (Darwin, 1881; Lloyd, 1942). The hydrophyte bladder- of the bee (Faegri and van der Pijl, 1979). wort (Utricularia) presents an even more rapid trapping action. This aquatic plant forms a bladder, with long guide-hairs ad- NATURE OF THE THIGMO STIMULUS joining the small entrance. Swimming daphnia or crustaceans can be steered by the guide hairs into the entrance, where The perturbation needed for thigmo stimuli varies consid- trigger hairs sense their presence. The thigmo stimulus on the erably, although the range is frequently in the vicinity of 1 g trigger activates a rapid sucking of water (along with the prey) or less (Darwin, 1891; Simons, 1992). During the sensing of into the trap by an expanding action of the bladder, after which leaf-surface irregularities by pathogenic fungi, a ridge of 0.2 a trapdoor closes over the entrance. The rapidity of this trap ␮m is suf®cient to reorient the growth direction of a uredio- action is impressive, estimated at Ͻ0.01 sec (Fineran, 1985). spore germ tube (Hoch and Staples, 1987; Hoch et al., 1993). The soil fungus Haptoglossa has evolved another trap sys- Phycomyces sporangiophores are stimulated to bend by a 0.5- tem. The fungus forms a trap as a circle of three cells, arranged ␮m movement (Dennison and Roth, 1967). For a higher plant in the manner of a lasso. Attracted by chemical pheromones, such as Drosera, the magnitude of touch perturbation needed a nematode enters and touches the trap, and the three cells for response is estimated to be ϳ0.3 mg (Darwin, 1881). Such exercise a very rapid hydraulic swelling, trapping the prey in very low thresholds suggest that there must be sensitive sys- the lasso. After the trap has been sprung, movements by the tems to substantially amplify the signal. Environmental fea- nematode create a further thigmo stimulus for the actual ®ring tures may also be involved because the pea tendril is sensitive of a needle-like gland that injects the fungus into the prey, to pH, such that exposing it to a low-pH solution will enhance starting the digestive process (Robb and Barron, 1982). The responsiveness to touch (Jaffe and Galston, 1966), and blue fungus then grows further into the nematode body and com- light is a corequisite for tendril coiling (Jaffe and Galston, pletes the digestion of the trapped nematode (Lee, Vaughan, 1966; Shotwell and Jaffe, 1979). and Durschner-Pelz, 1992). In at least some cases, the chem- For fungal spores, the hardness of the landing site in¯uences ical sensing of the nematodes stimulates the fungus to form the response in terms of secretion and/or germination. For ex- more traps (Robb and Barron, 1982). ample, anthracnose fungi are able to form appressoria on a hard glass surface, and spores of several species of anthracnose THIGMO PROJECTILES fungi germinate poorly or not at all when suspended in liquids. There is often a requirement also for the surface to be hydro- A number of species in the plant kingdom have developed phobic (Nicholson and Epstein, 1991; Terhune and Hoch, thigmo-triggered projectiles. Mention has already been made 1993; Hoch, Bojko, Comeau et al., 1995). Thus hard surfaces that Sphagnum spores are thrown in response to thigmo stim- such as polystyrene, Te¯on, polycarbonate, collodion, or glass ulation by rain. A mechanism reminiscent of the Haptoglossa treated with organic silanes (compounds of carbon and sili- injection device has been developed by stinging nettles (Ur- cone) will all serve to stimulate germination. tica). The stinging device is located in a series of glands It is common for calcium ions to play a role in the mediation around the edges of the leaf with a sting mechanism inside of thigmo signals (Hoch and Staples, 1987; van der Luit et al., (Esau, 1965). The gland contains a pouch of toxin, a mixture 1999). For example, pretreatment of the surface with calcium of histamine and acetylcholine (Feldberg, 1950), and a ®ne ions induces both attachment and germination of pycnidios- hollow needle, which points outward with a calci®ed seal at pores of Phyllosticta ampelicida (Shaw and Hoch, 2000), and the extended end. Upon being touched, the calci®ed seal Royle and Thomas (1973) have suggested that calcium signals breaks off, and the bared needle readily penetrates the in- are involved in the encystment of the downy mildew fungi. truder's skin and introduces the sting, serving to repel herbi- Some higher plants have developed receptors for thigmo vores or wandering humans. signals. For example, the pea tendril has a specialized surface A more dynamic projectile action is found in numerous on the concave side, and when one draws a ®ber across the ferns and higher plants that actually throw their spores or seeds sensing pad, one can feel that there are ridges present (M. J. into the air. The well-known jewel-weed or touch-me-not (Im- Jaffe, personal observation). Grape tendrils have specialized 378 AMERICAN JOURNAL OF BOTANY [Vol. 89 epidermal cells in which the surface cell walls are very thin, and Heath, 1994), and higher plants, e.g., Arabidopsis (Canut a quality that may increase the sensitivity for thigmo stimuli. et al., 1998; Galaud et al., 1998). Indeed, genes have been The tendrils of Eccromonocarpus have surface cells that bulge cloned from Saccharomyces cerevisiae (Hostetter et al., 1995), out hemispherically, and Tronchet (1964) has claimed to be Candida albicans (Gale et al., 1996), and Arabidopsis thaliana able to see plasma membrane invagination in these surface (Galaud et al., 1998; Laval et al., 1999; Nagpal and Quatrano, cells after thigmo stimulation. 1999) that have partial sequence similarity to the animal in- tegrins. A PROPOSAL FOR A THIGMO RECEPTOR Animal integrins mediate signalling as well as anchorage MECHANISM (Hynes, 1992; Sastry and Horwitz, 1993), and in the fungus Saprolegnia ferax, the distribution of an integrin-like protein How is a touch stimulus to the cell wall transduced into correlates with stretch-activated calcium channels (Garrill, intracellular responses, and how may the signal bring about Lew, and Heath, 1992; Garrill et al., 2001; Levina, Lew, and responses at remote sites? Any thigmo event in plants and Heath, 2001). Channel function (Garrill et al., 2001) and an fungi involves a perturbation at the cell wall. We know that integrin±extracellular matrix interaction (Bachewich and at least some thigmo signals are picked up through patches of Heath, 1993) are both required for growth in Saprolegnia, sug- sensitized cells, in which the surfaces of exposed cell walls gesting that they may be interdependent. In characean inter- are slightly corrugated (e.g., pea tendrils). In other instances node cells of the alga Chara, stretch-activated calcium chan- there are sensing hairs that serve as thigmo sensors (e.g., Utri- nels and an integrin-like protein are co-distributed, both are cularia, Dionaea) and in which the cell walls are presumably required for gravisensing (Wayne, Staves, and Leopold, 1992; bent by mechanical force (Lloyd, 1942; Simons, 1992). In the Staves, Wayne, and Leopold, 1992), and again the signal trans- thigmo systems of ®lamentous fungi, the hyphal growing duction chain linking the cell wall to the plasmalemma of the points usually serve as sensors (Hoch et al., 1987; Gow, 1994; cells apparently consists of integrin-like proteins. These nu- CorreÃa and Hoch, 1995; Jelitto, Page, and Read, 1995). merous reports of integrin-like proteins being involved in Whereas perception of thigmo stimuli can be presumed to stretch regulation of calcium channels may well relate to the originate at the exposed cell wall, a basic uncertainty is how transduction of thigmo events. the signal is transmitted to the interior of a cell. Despite the apparent presence of integrin-like proteins in the Peptides containing the integrin-binding sequence, RGD plasmalemma of some fungi and higher plants, the exact struc- (Arg-Gly-Asp), applied to walled cells interfere with a wide ture of these RGD-binding proteins in plants may differ from range of physiological events including gravisensing (Wayne, that of the animal integrins. The latter are responsible for com- Staves, and Leopold, 1992), development of sun¯ower pro- munication with an entirely different kind of extracellular ma- toplasts (Barthou et al., 1998), expression of plant defense re- trix than the rigid cell wall typical of plants and fungi. The sponses during host penetration by a rust fungus (Mellersh and integrin-like protein, Int1p, from Candida albicans has limited Heath, 2001), and accumulation of an antimicrobial phyto- sequence similarity to the ␣-integrins (Gale et al., 1998), and alexin in pea epicotyls (Kiba et al., 1998). In a preliminary the sequence of an Arabidopsis protein deduced from a cDNA experiment, RGD peptides were found to retard the rate of clone (Laval et al., 1999) had restricted similarity to integrin- lea¯et closure involved by touch in Mimosa pedica to about like proteins from Drosophila (Yee and Hynes, 1993) and hu- one-third of the control rate (M. J. Jaffe, personal observation). mans (Argraves et al., 1995). It seems quite possible that the Among the fungi, treatment with RGD peptides interferes with thigmo receptor proteins of fungi and higher plants may be- the association of the cell wall and plasmalemma in Saproleg- long to a family of signal receptors that are quite different in nia ferax (Kaminskyj and Heath, 1994), and in the bean rust structure from the animal integrins, but which share the com- fungus, Uromyces appendiculatus, RGD peptides interfere mon feature of binding and responding to RGD ligands. In- with development of the appressorium (CorreÃa, Staples, and deed, the genes ␣-Int1, responsible for adhesion in C. albicans Hoch, 1996). As demonstrated for onion and Arabidopsis (Gale et al., 1996), and AtELP1, a member of a multigenic (Canut et al., 1998; Laval et al., 1999; Nagpal and Quatrano, family composed of at least six water-de®cit-response genes 1999), bean (Garcia-GoÂmez et al., 2000), Saprolegnia ferax in Arabidopsis (Laval, Chabannes, CarrieÁre et al., 1999), are (Kaminskyj and Heath, 1994), and Candida albicans (Gale et plasma membrane receptors containing domains speci®c to an- al., 1996), proteins that bind the RGD sequence are located in imal integrins (Laval et al., 1999). However, they are other- the plasmalemma of both fungi and higher plants. Such evi- wise quite different in sequence from typical animal integrin dence strongly suggests a role for integrin-like proteins in the genes. communication of a thigmo signal from the wall to the pro- Recently, Lang-Pauluzzi and Gunning (2000) have de- toplasm. scribed the existence of Hechtian strands, i.e., strands linking Knowledge of integrins in animal cells is extensive now, the cell wall to the plasmalemma. These strands, which be- and integrins have been recognized as a widely expressed fam- come visible during plasmolysis (Hecht, 1912), can contain ily of cell surface adhesion receptors since 1987 (Hynes, actin micro®laments and microtubules (Lang-Pauluzzi and 1987). These proteins bind RGD sequences. They transfer ex- Gunning, 2000). They have been proposed to have a role in ternal signals from the extra-cellular matrix (ECM), across the cell±cell communication and signal transduction routes in plasmalemma to the cytoplasm, and have important roles in plant cells (Zandomeni and Schopfer, 1994; Reuzeau et al., cell-cell adhesion events (Hynes, 1992). Recent reviews in- 1997; Reuzeau, McNally, and Pickard, 1997; Canut et al., clude those by Hynes (1999), Hynes and Zhao (2000), and 1998; Glass, Jacobson, and Shiu, 2000), a proposal supported Calderwood, Shattil, and Ginsberg (2000). by the fact that exposure of walled cells to RGD-containing Proteins apparently related to the integrins have been dem- peptides causes a loss of Hechtian strands accompanied by loss onstrated in the plasma membranes of fungi, e.g., Candida of resistance to pathogens (Canut et al., 1998; Kiba et al., albicans (Gale et al., 1996) and Saprolegnia ferax (Kaminskyj 1998; Mellersh and Heath, 2001) and loss of the signal trans- March 2002] JAFFE ET AL.ÐTHIGMO RESPONSES 379 mission pathway between the cell wall and plasmalemma (Kiba et al., 1998). Both integrins and Hechtian strands are considered to be linked to cortical microtubules, which in turn can regulate numerous cellular functions including ATPase, metabolism, and growth itself (Nick, 1999). A number of proteins have been shown to bind to both cell wall and plasma membrane and have the potential to signal directly between the compartments. These proteins include the arabinogalactans, cellulose synthases, and wall-associated ki- nases (Anderson et al., 2001). However, it has not been shown yet that RGD sequences can disrupt the association of the pro- teins with either the cell wall or plasmalemma as is true for Hechtian strands. It should be noted, however, that wall-as- sociated kinases are induced by pathogen infection and wounding and have roles in cell growth and cell wall expan- sion (Anderson et al., 2001). The general occurrence of changes in calcium ions with signal stimulations has led to the appellation of Caϩ2 as a sig- nalling molecule in plants (Toriyama and Jaffe, 1972; Trewa- vas and Knight, 1994; Berridge, Lipp, and Bootman, 2000). The role of calcium in thigmo responses has been widely noted and indicates a common role for calcium in the transduction of the signal in the cytoplasm or nucleus (Toriyama and Jaffe, 1972; Trewavas and Knight, 1994; Legue et al., 1997). The intimate connection between Caϩ2 and calmodulin has been noted in many instances, and Braam and Davis (1990) have shown that three of four genes involved in thigmo responses in Arabidopsis that serve as calmodulin regulators are quickly induced by touch. The calmodulin involved in the thigmo re- sponse is principally located in the nucleus (van der Luit et al., 1999). Indeed, inhibitors of calmodulin have been shown Fig. 1. Proposed model of components of the thigmo transduction system to suppress thigmo responses (Jones and Mitchell, 1989), an from cell wall into the cytoplasm. Pressure against the cell wall causes dis- placement of the plasma membrane via Hechtian strands and integrin-like effect that may be related to the dissociation or rearrangement linkages, thus repositioning the microtubules. At the same time, ion channels of microtubules (Braam and Davis, 1990). are opened, altering the intracellular Caϩϩ concentration and inducing action The changes in cytoplasmic Caϩ2 occur in some instances potentials. through opening of calcium channels in the plasma membrane (Ding and Pickard, 1993), or in some instances the rise in ϩ2 ϩ2 cytoplasmic Ca may result from release of Ca from intra- be one of the cell's immediate responses to shaking, perhaps cellular pools (Knight, Smith, and Trewavas, 1992; Kluesener in conjunction with rapid ¯uxes of calcium ions (Braam and et al., 1995), perhaps via phosphoinositides released at the Davis, 1990). plasma membrane (Drobak, 1993). A signalling propagation So, a tentative model for thigmo responses (Fig. 1) would mechanism that is common to all thigmo systems involves begin with a perturbation of the cell wall followed by signal changes in electrical resistance associated with development transduction to the plasmalemma via Hechtian strands involv- of an electrical potential across the plasmalemma (Sibaoka, ing integrin-like proteins that can be disrupted by the RGD 1966; Pickard, 1971; Jaffe, 1976a). The participation of an motif. These proteins appear to form a continuum from the electrical potential in thigmo responses is most vividly evi- cell wall to the cytoskeleton (Wyatt and Carpita, 1993). Clear- denced in the leaf movement of Mimosa, where a perturbation ly, thigmo signals received at the cell wall are transmitted leads to an action potential that moves from leaf to leaf, through the plasma membrane, probably opening cation- and spreading the thigmo response systemically. The rise time for stretch-activated channels and may involve MAP kinases. The the bioelectrical potential relates to the particular thigmo re- perturbation may thus alter both membrane and cytoskeletal sponse. Thus the carnivorous and sensitive plants may show structures and lead ®nally to alterations in gene expression. an electric potential rise time averaging ϳ0.9 sec, whereas the Whether the RGD-sensitive Hectian strands serve the function rise time for thigmomorphogenetic changes averages ϳ25 sec visualized here for the reception of touch signals and whether (Jaffe, 1976a). Action potentials change the ionic balance in they would be the only means of communication that enable the cytoplasm, have been shown to induce some gene tran- response to thigmo signals (Kohorn, 2000) will remain am- scription (Stankovic and Davies, 1998), and have the capabil- biguous until a proper experimental foundation has been pro- ity of altering a broad range of cellular activities. vided. Mitogen-activated protein (MAP) kinases may also be in- volved in mechanosensing in plants. BoÈgre et al. (1996) have LITERATURE CITED reported that a protein, p44MMK4, was activated within 1 min after alfalfa leaves were mechanically stimulated for 2 sec. The AIST, J. R. 1977. Mechanically induced wall appositions of plant cells can activation was transient and disappeared after 10 min. The prevent penetration by a parasitic fungus. Science 197: 568±571. authors suggested that activation of the MMK4 pathway must ALLEN, R. F. 1923. A cytological study of infection of Baart and Kanred 380 AMERICAN JOURNAL OF BOTANY [Vol. 89

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