The cap is widely accepted to be the site of gravity sensing in roots because removal of the cap abolishes root curvature. Circumstantial evidence favors the columella cells as the gravisensory cells because amyloplasts (and often other cellular components) are polarized with respect to the gravity vector. However, there has been no functional confirmation of their role. To address this problem, we used laser ablation to remove defined cells in the cap of Arabidopsis primary roots and quantified the response of the roots to gravity using three parameters: time course of curvature, presentation time, and deviation from vertical growth. Ablation of the peripheral cap cells and tip cells did not alter root curvature. Ablation of the innermost columella cells caused the strongest inhibitory effect on root curvature without affecting growth rates. Many of these roots deviated significantly from vertical growth and had a presentation time 6-fold longer than the controls. Among the two inner columella stories, the central cells of story 2 contributed the most to root gravitropism. These cells also exhibited the largest amyloplast sedimentation velocities. Therefore, these results are consistent with the starch-statolith sedimentation hypothesis for gravity sensing.Root gravitropism consists of gravity perception, signal transduction, and the growth response, which is manifested by differential growth across the elongation zone. Gravity perception in roots is generally believed to occur in the root cap (Sack, 1991), but it has been argued that perception might also be possible in other regions of the root (Poff and Martin, 1989). Previous attempts to prove that the root cap is the site of gravity perception involved surgical removal of the whole cap (Juniper et al., 1966) or small portions of the cap (Younis, 1954; Konings, 1968). These early studies implied a major role for the cap in root gravitropism, particularly the cells in the columella, the central region of the cap (Konings, 1968). In addition to its proposed role in gravity perception, the root cap has also been suggested as the site of initial development of auxin asymmetry, which is transmitted to the elongation zone during the gravity response (Konings, 1967(Konings, , 1968 Moore and Evans, 1986; Hasenstein and Evans, 1988). Recently, it was shown that the apoplastic pH differences that form between the upper and lower flanks of gravistimulated roots can be abolished by surgically removing the cap (Monshausen et al., 1996). This provides evidence that a signal originating from the cap is transmitted to the elongation zone during gravistimulation.Despite the wealth of information derived from the surgical approach, this method has one disadvantage: it does not allow the precise removal of defined cap cells (i.e. removing only columella cells versus removing only peripheral cap cells). The surgical approach was modified by Konings (1968) such that only small portions of pea root caps were removed prior to gravistimulation. Progressive elimination of the c...

Although the columella cells of the root cap have been identified as the site of gravity perception, the cellular events that mediate gravity signaling remain poorly understood. To determine if cytoplasmic and/or wall pH mediates the initial stages of root gravitropism, we combined a novel cell wall pH sensor (a cellulose binding domain peptide-Oregon green conjugate) and a cytoplasmic pH sensor (plants expressing pH-sensitive green fluorescent protein) to monitor pH dynamics throughout the graviresponding Arabidopsis root. The root cap apoplast acidified from pH 5.5 to 4.5 within 2 min of gravistimulation. Concomitantly, cytoplasmic pH increased in columella cells from 7.2 to 7.6 but was unchanged elsewhere in the root. These changes in cap pH preceded detectable tropic growth or growth-related pH changes in the elongation zone cell wall by 10 min. Altering the gravity-related columella cytoplasmic pH shift with caged protons delayed the gravitropic response. Together, these results suggest that alterations in root cap pH likely are involved in the initial events that mediate root gravity perception or signal transduction.

Touch and gravity are two of the many stimuli that plants must integrate to generate an appropriate growth response. Due to the mechanical nature of both of these signals, shared signal transduction elements could well form the basis of the cross-talk between these two sensory systems. However, touch stimulation must elicit signaling events across the plasma membrane whereas gravity sensing is thought to represent transformation of an internal force, amyloplast sedimentation, to signal transduction events. In addition, factors such as turgor pressure and presence of the cell wall may also place unique constraints on these plant mechanosensory systems. Even so, the candidate signal transduction elements in both plant touch and gravity sensing, changes in Ca2+, pH and membrane potential, do mirror the known ionic basis of signaling in animal mechanosensory cells. Distinct spatial and temporal signatures of Ca2+ ions may encode information about the different mechanosignaling stimuli. Signals such as Ca2+ waves or action potentials may also rapidly transfer information perceived in one cell throughout a tissue or organ leading to the systemic reactions characteristic of plant touch and gravity responses. Longer-term growth responses are likely sustained via changes in gene expression and asymmetries in compounds such as inositol-1,4,5-triphosphate (IP3) and calmodulin. Thus, it seems likely that plant mechanoperception involves both spatial and temporal encoding of information at all levels, from the cell to the whole plant. Defining this patterning will be a critical step towards understanding how plants integrate information from multiple mechanical stimuli to an appropriate growth response.

Although the columella cells of the root cap have been identified as the site of gravity perception, the cellular events that mediate gravity signaling remain poorly understood. To determine if cytoplasmic and/or wall pH mediates the initial stages of root gravitropism, we combined a novel cell wall pH sensor (a cellulose binding domain peptide-Oregon green conjugate) and a cytoplasmic pH sensor (plants expressing pH-sensitive green fluorescent protein) to monitor pH dynamics throughout the graviresponding Arabidopsis root. The root cap apoplast acidified from pH 5.5 to 4.5 within 2 min of gravistimulation. Concomitantly, cytoplasmic pH increased in columella cells from 7.2 to 7.6 but was unchanged elsewhere in the root. These changes in cap pH preceded detectable tropic growth or growth-related pH changes in the elongation zone cell wall by 10 min. Altering the gravity-related columella cytoplasmic pH shift with caged protons delayed the gravitropic response. Together, these results suggest that alterations in root cap pH likely are involved in the initial events that mediate root gravity perception or signal transduction. INTRODUCTIONRoot gravitropism requires a coordination and interaction of cells responsible for gravity perception, signal transduction, signal transmission, and the growth response. The gravity perception step occurs in the columella cells of the root cap (Sack, 1997;Blancaflor et al., 1998), whereas the growth response (i.e., curvature) is initiated in the distal elongation zone (DEZ) and fully expressed in the central elongation zone (CEZ) (Ishikawa et al., 1991;Ishikawa and Evans, 1993). Thus, for curvature to occur, a signal must move from the gravity-sensing columella cells to the graviresponding cells of the elongation zone. Although there is a wealth of literature implicating the sedimentation of amyloplasts in the columella cells as one of the initial gravity-sensing events (Sack, 1997) and auxin as a factor mediating the growth response (reviewed by Chen et al., 1999;Rosen et al., 1999), the nature of the signal transduction events in the root cap that lead to the growth response have remained elusive.The molecular mechanisms that mediate the growth response in the elongating cells of the root also are largely unknown. Indeed, growth control in different regions of the root may be very different. The DEZ is defined as being centered on the region of the root elongation zone showing 30% maximal growth rate, and the CEZ is defined as being centered on the region of maximal growth rate (Ishikawa and Evans, 1993). The current evidence suggests that the growth of the CEZ, but not the DEZ, is regulated in part by auxin (Ishikawa and Evans, 1993;Evans et al., 1994;Mullen et al., 1998) and likely is mediated, at least in part, via acid growth phenomena (Edwards and Scott, 1974;Evans, 1976;O'Neill and Scott, 1983;Collings et al., 1992; Taylor et al., 1996;Büntemeyer et al., 1998;Felle, 1998;Peters and Felle, 1999). For example, both auxins and auxin antagonists cause root growth and the pH o...

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