Gravitropism

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Gravitropism

The ability of roots to perceive and respond to gravity has captured the interest of researchers ever since Darwin (1880) described the elimination of graviresponsiveness in primary roots by removing the root cap. The initial phase of gravitropism, that of perception, probably involves the root cap and, within the cap, the columella cells have been considered for some time to contain the gravity detecting mechanism. Although there is some controversy as to the exact mechanism, the popular model describes the settling of cell components with sufficient mass (e.g., amyloplasts or nucleus) on the bottom of the cells to trigger a pressure-sensitive effect from the ER or plasmelemma. This would ultimately result in the well-documented response phase, the asymmetric elongation of cells in the area 2-4 mm behind the cap.

If the cap is responsible for graviperception, a signal of some sort has to travel from the cap to the zone of elongaton. Results of experiments performed in the mid to late 70's suggested the signal involved a growth inhibitor influencing a reduction in growth rate on the lower side of the root. At first ABA was implicated as the inhibitor but subsequent research during the early 80's seem to rule out this possibility.

During this same period a lot of attention was given to the possible role of IAA as well as calcium to signal transduction. For instance, a correlation was shown between gravicurvature and the asymmetric efflux of acid in the elongating zone. Auxin transport inhibitors abolished this. As for calcium, it has been found roots oriented horizontally resulted in a Ca movement from the upper to lower sides of the cap. Application of Ca chelators eliminated root graviresponsiveness. Furthermore, inhibitors of auxin transport not only abolished gravitropic response in the elongating zone but blocked Ca movement in the root cap as well.

It was, therefore, suggested that pressure on the ER results in release of Ca on the lower side of the columella cells which, in turn, activates cell wall Ca pumps on that side, allowing Ca movement and subsequent concentration on the lower side of the cap and a coupled triggering of auxin along a similar asymmetrical gradient.

This model, however, became subject to question when cytochemical staining experiments showed a lack of any up-down calcium gradient in the columella of either vertical or horizontal roots and when it appeared that calcium moved acropetally within the columella. It was, therefore, difficult to explain the asymmetric basipetal movement of IAA in terms of calcium efflux from the symplast of the columella cells.

This set the stage for the research conducted during the past five years concerning the role of calcium and auxin in gravitropism.

Randy Moore has been a remarkably prolific worker in this area. Several papers have been published, but one particular report (1) demonstrated several results that appear to support the part of Moore's own model of root gravitropism that involves the apoplastic movement of Ca.

Moore placed roots from Zea mays as well as an ageotropic variety horizontally and allowed them to curve downward for 3 hours, during which time he placed agar receiver blocks along the upper and lower sides of the tips of the roots. Also, roots were used whose growth was slowed down by either cold treatment or application of CoCl to test the movement of Ca independent of gravicurvature. Vertical roots were used for controls. The blocks were then assayed by an atomic absorption spectrophotometer.

In the samples from the horizontal geotropic roots the concentration of Ca collected from the blocks placed on the upper side was significantly lower (almost 4X) than that collected from the lower blocks, and also lower in concentration than those solutions collected from the vertical roots. There was no significant difference in concentration between the upper and lower sides of the ageotropic roots, a condition similar to that of the vertical roots. This suggested to Moore the downward movement of Ca is important for graviresponsiveness.

The roots treated for slow growth concentrated more Ca in the lower than the upper side. This gradient formed before curvature occurred, indicating the gradient is not a result of curvature.

These results appear to be consistent with those of earlier experiments and with Moore's model in suggesting that: (1) Ca moves to the lower side of the root cap when the root is undergoing gravitropic response, (2) calcium is present in, and can be collected from, the mucilage (3) this movement of Ca in the mucilage is gravity-induced, and (4) since the ageotropic roots had no Ca gradient in the mucilage, the downward movement of calcium in mucilage is an important element in gravitropic response.

This particular experiment in and of itself does not present a totally convincing case concerning the role of gravity and Ca loading of the mucilage from the apoplast that Moore suggests. The Ca, for instance, may be actively moved symplastically by the cortical cells and the Ca gradient in the mucilage simply an efflux "residue" from the process. This distinction may be important in determining the mechanism in signal transduction. However, Moore cites an earlier report by himself utilizing an electron probe that indicated a lack of any endogenous calcium gradient across the cortex of the elongating zone in graviresponding Zea mays roots. Furthermore, removal of mucilage or of the epidermis (which produces the mucilage layer) had resulted in elimination of gravitropic response. These results provide a strong case for the role of the epidermis as well as for the gravity-induced downward movement of Ca from the apoplast to the mucilage layer in connection with the gravitropic reponse. Moore further suggests basipetal movement of Ca across the elongating zone and a subsequent induction of a growth regulator (such as auxin) along a gradient, but this particular experiment has nothing to directly contribute to that idea.

This matter was dealt with by Hasenstein and Evans (2). The study involved an exhaustive battery of experiments involving the effect of several cations and chelators on the polarity of movement of auxin in both intact and decapped root segments. The experiment most relevant to this paper involved donor and receiver agar blocks placed on either the tip or base of the segments. Labelled IAA was loaded into the donor blocks while different cations (here we are interested mostly in Ca2+ and Al3+) were applied to the blocks at the apical end.

After recording and comparing the amount of labelled IAA in the receiver blocks it was found that the cations significantly affected auxin transport, particularly the polarity of hormone movement. Compared to controls, calcium dramatically promoted basipetal over acropetal auxin movement, while aluminum strongly enhanced acropetal movement more than basipetal. Furthermore, the calcium chelator EGTA reversed the effect of calcium by decreasing basipetal movement of auxin.

Another result was that decapping roots caused auxin transport to change from basipetal to acropetal -- without influence of applied cations. This phenomenon may help to clear up some earlier confusion caused by experiments that seemed to show auxin transport was primarily acropetal, since those particular experiments were done with decapped roots. Furthermore, application of Ca to the cut surface of decapped roots promoted acropetal auxin movement. This further exemplifies the importance of the role of the cap in influencing the direction of auxin transport - at least in Zea mays .

Since it has been shown many times that root tips produce a curvature toward the side of calcium applied to the root cap, Havenstein and Evans proposed that it is the effect on polar movement of auxin that explains the link between Ca gradients in the tip and curvature in the elongating region below the tip. This is assuming, of course, that both Ca and auxin concentrations are correspondingly asymmetrical during the gravitropic response period -- a fact that is yet to be proven.

The fact that Al produced the opposite effect on polar movement, i.e., enhanced acropetal movement, prompted the researchers to apply labelled IAA to the basal cut surface and Al3+ in one side of the cap and measure the amount of labelled IAA that showed up on the elongating zone. It was found the auxin ended up concentrated on the side away from the applied Al3+.

It had been shown in earlier experiments that in the root tip auxin travels acropetally through the stele but exhibits basipetal movement through tissue peripheral to the stele. Evans and Hasenstein propose a model that describes a gravity-induced movement of Ca toward the lower side of the root cap as well as an asymmetric Ca-induced movement of auxin into a basipetal auxin stream in such tissue as the epidermis. The fact that auxin transport is acropetal in decapped roots is consistent with this idea as well as supporting the direct role of the cap in auxin distribution in the growing region during graviresponse.

An idea on how calcium may affect auxin transport is offered. In a report published by the same researchers (3) it was shown that certain cations can mimic the curvature induction exhibited by Ca but that this activity is eliminated by calmodulin inhibitors. It had also been found by other workers that CaM can be bound by cations at sites other than those bound by Ca, thus modifying the affects of CaM. Al, for example, was found to bind to CaM and thereby inhibit its activation by Ca. This may explain, in part, the effects of Al opposite to those of Ca. Calmodulin, therefore, may be a link between Ca and auxin transport, by promoting the extrusion of Ca into the apoplast and/or promoting basipetal auxin transport by activating auxin pumps or effecting polar membrane permeability.

The importance of the role of CaM and Ca was further demonstrated by Thomas Bjorkman and A. Leopold. In 1987 they published the results of the use of a bioelectric current to detect gravity sensing in the root (4). They first tested auxin. Although earlier experiments utilizing auxin transport inhibitors resulted in the cessation of gravitropism in Zea mays roots, it could not be shown if the inhibitors affected the initial perception of gravity by the root or some other subsequent step in the gravitropic mechanism. In other words, it was not clear if auxin itself had a contributing role in the sensing of gravity. By applying an auxin inhibitor to horizontally-oriented roots and at the same time testing to see if the sensing-induced current was inhibited as well, Bjorkman and Leopold were able to show that auxin transport was not part of the sensing process by noting that a change in current density occured during the first hour when auxin transport was totally inhibited and, in fact, continued into the subsequent period (after 1 hr.) when the inhibitor's effects wore off. Two different auxin-transport inhibitors, TIBA and NPA, gave similar results.

Since it had been difficult to isolate events in the sensing region without coupling to growth response events it had also been impossible to say for certain if redistribution of certain elements such as Ca and auxin are direct or secondary events in the gravitropic mechanism.

Another part of this experiment involved testing the effects of applying CaM inhibitors. CaM is involved in many varied aspects of metabolic activity, therefore the fact that CaM inhibitors negatively affect graviture (as reported by earlier research) did not necessarily narrow down the role of CaM. In contrast to the results of the auxin tests no significant change in current was detected when the CaM inhibitors were applied. This suggests CaM is a component of gravity sensing rather than simply involved in the growth mechanism. If this is true, Ca movement must also be involved since Ca is directly associated with CaM. It must also be kept in mind that CaM (as well as Ca) also may be involved in other phases of gravitropism, such as in transport of the signal or the response mechanism.

Bjorkman published a separate paper (5) at the same time explaining the theory behind the technique as well as reporting the results of tests performed on Zea may root caps. A preliminary current profile (to determine initial current distribution) was first prepared, and then the current density was recorded in three regions: the elongating zone, columella and tip of the cap. The results did record a density change in the columella cells, particularly on the upper side, at a distinctly regular interval proceeding gravistimulation (3.5 min.). Measurements taken at the root tip and elongating zone revealed a constant current for at least 20 minutes. This indicates gravity sensing occurs in the columella region only, in agreement with the proposed model.

It should be noted that there is no certainty here as to the exact cause of current density change. The possibility of interference or outright induction of current production in the tissue by the apparatus itself cannot be entirely ruled out. Furthermore, no adequate explanation was given as to why the measurements of current change on the lower side of the columella were not very distinct (all reported measurements with the inhibitors were consequently done on the upper side)

One could easily be swayed into agreement with the part of the model proposed by Moore and others that describes the movement of auxin as being stimulated by Ca and that auxin's presumed asymmetric distribution is somehow a result of the asymmetric release of Ca in the root cap. The link between Ca and auxin is the least understood part of the mechanism, however, and many interpretations of experimental results attempting to elucidate the matter are possible. A paper published by Migliaccio and Galston (6) describes a battery of experiments whose results call for reappraisal of this idea. Excised pea epicotyls were abraded to increase permeability and were pre-loaded with 3H-IAA or 45Ca2+ for one hour after which they were incubated in a solution containing one of various auxin or calcium inhibitors and then placed horizontally for 90 minutes. The epidermis was then stripped from the top and bottom sides and measured for radioactivity.

Normally in the horizontal shoot IAA will concentrate more on the lower side while Ca collects on the upper side. The controls (without inhibitors) reflected this. When Ca transport inhibitors (such as Nitrendipine, Nisoldipine and Bay K 8644) were used there was no appreciable difference in the asymmetries of either calcium or IAA. That is to say, the concentrations were the same as those of the controls. Furthermore, curvature measurements were similar. The application of IAA inhibitors (such as TIBA and 9-HFCA), however, significantly affected the asymmetries of IAA as well as calcium. IAA became more concentrated on the upper side while Ca concentrations were greater on the lower side. Gravicurvature was virtually nullified.

Migliaccio and Galston interpreted this to mean that calcium asymmetry during normal gravitropic response may be the result of IAA asymmetry rather than the reverse. To demonstrate one way this might work the workers exposed 45Ca2+-loaded epicotyls to varous pH's in the form of localized applications of filter papers soaked in buffers as well as total immersion of the epicotyls into buffered solution and then measured the 45Ca2+ uptake in the solution. The latter experiment resulted in data that showed a linear relationship between external pH and calcium release.

The localized pH applications produced the same results, while application of pH 3 to one side and pH 7 to the opposite side resulted in calcium loss only on the acidic side. No 45Ca2+ redistribution was apparent in the epicotyls. 50% of the Ca loss occured within 15 min. It was also shown that the introduction of IAA or fusicoccin, both proton efflux activators, rapidly release Ca from the tissue while cycloheximide retarded such release. Migliaccio and Galston took this all to be an indication that the Ca that is redistributed during the gravitropic event comes largely from the apoplast.

The researchers further concluded the role of Ca is secondary to that of IAA. IAA may be influencing the redistribution of Ca through proton release and growth in the lower side of the shoot. Since earlier experiments have recorded an increase in proton extrusion on the elongating side in Zea mays roots and sunflower stems during gravitropic response it is possible that the observed Ca release during this event is a secondary event. The researchers did not offer specific proof that Ca asymmetry follows H+ asymmetry but they did cite papers that reported an 8 to 10 minute lag before IAA-induced acidification compared with a 10 minute lag before appearance of calcium asymmetry.

There are some limitations inherent in this paper. The first obvious consideration is that these experiments involved pea epicotyls whereas all the previous research cited was done with Zea mays primary roots. There are some similarities, however, since in both cases the kinetics of the gravitropic response reportedly are alike, and since Ca is always redistributed on the side of slow growth whereas auxin is always on the lower sides (although auxin induces two opposite effects on stem and root cells). Also, it should be kept in mind we are considering the linkage of auxin and calcium as it applies to the sequence of events in the transduction stage rather than, say, the role of auxin in the response phase or calcium in the perception phase; it is not unreasonable to suggest this particular process is similar for both roots and stems.

Some questions are raised by the results of the experiments themselves. Migliaccio and Galston's main argument is that IAA must be involved in primary response activity rather than Ca since IAA transport inhibitors affect asymmetric distribution not only of IAA but also of Ca, while Ca inhibitors did not affect the redistribution of either. As stated, this is true, but the data also show that the affect of TIBA and 9-HFCA was to reverse the % of IAA and calcium found on the lower and upper side as compared to the control. No possible explanations were offered of this phenomenon even though one might expect homogenous distribution. Furthermore, the stems exhibited little or no curvature in this state. The authors state the lack of inversion of curvature accompanying an inversion of IAA and calcium may point to a discrepancy in Cholodny-Went theory. One may make the same judgement of this experiment. The inhibitors or the preparation protocal may have triggered secondary effects or otherwise obscurred the data, for example. It has been shown that excising tissure may result in wound-reactions that alter ion balances in the cells.

Otherwise, the results are provocative. This paper serves to remind us that any assumptions involving the sequence of cause and effect must be carefully inspected before constructing a model to explain all the processes of gravitropism. It should be pointed out that this experiment does not rule out the possibility that Ca acts as a second messenger during the perception phase in the symplast (as the Bjorkman experiment suggests)

Most of the earlier experiments with Ca before 1987 dealt with responses to exogenous Ca. Unfortunately, little could be said of what happens to endogenous Ca within the cells. Many of the techniques previously used had inherent problems that made it difficult to localize and quantify the ion in situ at the subcellular level. For example, there is the possibility of inducing wound-reaction artifacts and the difficulty of accurately measuring specific compartments such as columella cytoplasm. In 1987 R. Moore and others used energy dispersive X-ray microanalysis (EDS) as a means to solve most of these problems. This involves freezing the samples, sectioning them ultrathin on a cryomicrotome, freeze-drying the samples, transferring them onto a film and probing the samples under a scanning electron microscope equipped with an X-ray analysis system. The freezing process reduces the exposure to aqueous liquids that may cause ion movement, while the thin sections allow more accurate ion measurements in subcellular compartments under SEM without confusing the X-ray signal from underlying compartments.

According to his first report (6), Moore discovered several other ions, especially P and Na, also redistribute asymmetrically in both the root cap and elongating zone (but not to the extent of Ca). However, since exogenous applications of these ions does not induce curvature it is assumed they are an indirect consequence of gravitropic response. For instance, those that accumulate on the upper side may be a consequence of more rapid growth in that region. Calcium, on the other hand, appeared to play a direct role in graviresponse, but only in specific tissues. The results of the tests for this element indicated to Moore that "epidermal cells are developmentally and physiologically different from cortical cells, and may indeed by the responsive cells for differential growth associated with root gravicurvature".

In 1989 Moore published another paper (8) describing similar experiments and the same technique to compare the endogenous movement of calcium alone in the walls and cytoplasm of the epidermal cells as well as the mucilage -- all in the elongation zone. Samples were taken from Zea mays seedlings oriented vertically or horizontally. Gravitropic response usually begins 30 after stimulation, so measurements were taken at 30 and 90 minutes. The results of these measurements in the elongation zone were similar to those reported for the root cap in the 1987 paper, and are given in Table 1 (p. 20).

After gravistimulation epidermal cells showed dramatic increase on Ca concentrations in the lower side of the root. Mucilage (a product of the epidermis) on the lower side of the root also showed a similar increase in Ca. In contrast, cortical cells showed little difference between the vertical controls and gravity stimulated roots in Ca concentration in the root. In fact, the mild redistribution that did occur was loaded toward the upper side. This may be connected with the demands of the growth process occuring on that side. The cytoplasm in the cortical cells had no detectable concentrations at any time. On the other hand, the cytoplasm of the epidermal cells, which also contained no detectable amount of Ca in the vertical roots did gain significant increase of the ion, but only on the lower side of the root.

These results indicate a downward apoplastic movement of Ca, perhaps along an electrochemical gradient (as suggested by Moore), in epidermal but not cortical cells proceeding gravistimulation. The changes coincided with the timing of gravitropic response which suggests linkage between the two events. The cytoplasm of the epidermal cells contained no detectable amount of ion either in the vertical roots or on the upper side of the gravity- stimulated root. However, a moderate amount was detected on the lower side of the stimulated root. According to Moore this could be the result of either of two events: (1) the accumulation of Ca in the apoplast in this area influencing cytoplasmic uptake, or (2) the "gravity-triggered release of Ca sequestered in these cells". If the first possiblility were the case, exogenous application of Ca to the root would be expected to result in curvature. However, experiments involving the application of 45Ca2+ to the mucilage at the top of the root resulted in accumulation of the ion at the lower side in the mucilage unaccompanied by any curvature. This indicates the increase in Ca inside the cytoplasm in the lower epidermal cells may be involved directly in graviresponse in both the elongation zone and root cap. This is consistent with earlier experiments which showed the removal of the epidermis on the upper and lower side eliminates gravitropic response. This suggests a link between this event and hormone activity. However, since detachment of the root tip results in the abolishment of graviresponse while application of Ca to the elongation zone alone results in no response it is evident that either the Ca found in the elongation zone has little to do with the polar transport of auxin (but is perhaps a part of the response mechanism), or a continuous gradient of Ca on the lower side from the tip to the growing region is coupled with signal transduction.

It has already been reasonably established that the cap is the site of perception. Roots will not respond to stimulation without the presence of a functional and intact cap. The recent experiments involving current density change indicate the primary perception event occurs in the cap region, particularly the columella, and includes a direct involvement by CaM, which is reported to be at high levels in root caps.

Soon after gravity stimulation, events involving Ca are triggered in both the root cap and the growing region. The recent experiments cited clearly indicate a Ca gradient is induced by gravity, that this gradient is reflected in the mucilage, and that both conditions are important in gravitropic response since ageotropic roots have no Ca gradient and since the removal of mucilage eliminates the response in normal roots.

Since the perception and response events are located in two different areas some type of signal is required. Ca is not involved as the signal itself since application of Ca to the growing region does not result in a response. The signal does involve a growth inhibitor since early experiments have shown that roots with half the cap removed have slowed growth on the side of the remaining cap portion. This inhibitor is probably IAA since: (1) artificially induced asymmetries of auxin at the root tip induce curvature, (2) this curvature correlates positively with the development of asymmetric acid efflux in the elongation zone and (3) this curvature as well as proton efflux can be eliminated by auxin transport inhibitors.

Auxin, however, travels basipetaly in the stele but acropetaly through tissue peripheral to the stele in the root tip. The report by Hasenstein and Evans indicates Ca in the root cap promotes basipetal over acropetal auxin movement and that CaM may be involved in this process. If so, it is not unreasonable to assume Ca activates CaM which in turn, metabolically facilitates the basipetal routing of auxin. It is not clear if Ca may be active in two separate roles. In other words, what is the role of Ca in the columella cells during graviperception as compared to the movement of Ca associated with the lower epidermal cells and mucilage as it applies to the rerouting of auxin?

The experiments by Moore, Cameron and Smith indicate a downward, apoplastic movement of Ca associated with the stimulation of epidermal cells occurs across the entire region involved in graviresponse -- that is, a similar movement along the cap as well as the elongation zone. It is the elongating region, of course, that is the site of reponse and, as mentioned before, this probably involves auxin. It is reasonable to assume that there is a link between asymmetric Ca and hormone activity in the elongation zone, but the sequence and mechanism of cause and effect is not clearly established. The experiments by Migliaccio and Galston introduce the possibility that the asymmetry of Ca may be a result rather than the cause of auxin asymmetry. Auxin-mediated efflux of acid in the apoplast, for instance, may induce the movement of Ca. It is not unreasonable to question if the asymmetrical movement of Ca in the growing region is an event distinct from that in the root cap. Auxin may induce Ca movement in the elongation zone, for instance, but the asymmetrical movement of auxin may itself be the result of Ca movement in the cap epidermis or columella. The latter event could involve activation of CaM resulting in a turning on of auxin pumps or a change in polar membrane permeability. This would be consistent with the results of Hasenstein and Evans which show that application of a Ca chelator at the root tip decreases basipetal auxin movement. If however, auxin is the primary mover in the cap but affects Ca movement as it moves into the growing region, this would explain early experiments in which auxin transport inhibitors also block Ca movement. Moore assumes Ca is responsible for the asymmetric movement of both regions and offers a model which describes the role of Ca either as a sink for auxin, resulting in the hormone's accumulation on the lower side of the root, or as a sensitizer of the tissues to auxin. The latter possibility may explain why an auxin gradient in the growing region has not been sufficiently demonstrated, since an asymmetry would not necessarily be required.

Clearly, much work is left to be done. Ca appears to be involved in all phases of gravitropic response and the possibility has emerged that separate and distinct events involving Ca activity are associated with each of these stages. Further research into cell wall biochemistry might clarify the issue. It is yet to be determined, for instance, what occurs in the walls of cells in the different regions immediatly preceeding graviperception. Are calcium and/or auxin pumps turned on? If so, in what specific areas of the root and in what sequence? The location and isolation of Ca and auxin pumps and subsequent manipulation of these pumps may facilitate this research.

Timing of events is important. Exactly when does auxin begin moving asymmetrically (if it is asymmetrical at all), and how does this compare with the appearance of Ca in the columella, the cap epidermis and the epidermis of the elongation zone? Moore's experiments that measured endogenous amounts of Ca in the cap and elongation zone during graviresponse have been helpful, but the same must be done for auxin to clearly understand the pathway. If possible, finer time intervals must be employed in similar experiments involving both auxin and Ca to aid in tracing specific events as well as the link between auxin and Ca movements in the different regions.

An experiment is needed to determine if Ca increases sensitivity or otherwise act as a sink in the cells of the growing region. This would help to determine if gradients are a result or a cause of auxin movement -- at least in the growing region.

Further experiments are needed to elucidate the mechanism by which auxin normally leaves the stele in vertical roots (e.g., by diffusion or active transport). This information could be used for comparison to events after gravistimulation. The fact that Ca promotes acropetal auxin movement but has little effect on basipetal movement (as reported by Hasenstein and Evans) is curious. This may suggest something else besides Ca must be required for basipetal movement.

If any of these experiments could have been done someone probably would have implemented them; such research may not occur for quite a while due to the inherent difficulties involved. In the meantime, the same experiments performed by Migliaccio and Galston on pea shoots could be administered to roots to test their hypothesis that Ca movement is a result of auxin movement, specifically in the elongation zone. Also, it has been reported that Michael Evans used radioimmunoassay and enzyme activity assays to successfully detect CaM concentration in the root tip apoplast. Perhaps in conjunction with the application of CaM inhibitors it can be determined how or if CaM is operating in the epidermal cells of both the cap and elongation zone. Darwin once professed, "There is no structure in plants more wonderful, as far as its functions are concerned, than the tip of the radicle". Indeed, over a century later, the functions of the radicle have still captivated the wonder of all those who have cared to observe them.

(1) Randy Moore and W. Mark Fondren A Gradient of Endogenous Calcium Forms in Mucilage of Graviresponding Roots of Zea mays. Annals of Botany 61, 113-116 (1988).

(2) Karl Hasenstein and Michael Evans Effects of Cations on Hormone Transport in Primary Roots of Zea mays. Plant Physiology 86, 890-894. (1988).

(3) Karl Hasenstein, Randy Moore, et. al. Comparative Effectiveness of Metal Ions in Inducing Curvature of Primary Roots of Zea mays. Plant Physiology 86, 885-889 (1988).

(4) Thomas Bjorkman and A. Leopold Effect of Inhibitors of Auxin Transport and of Calmodulin on a Gravisensing-Dependent Current in Maize Roots. Plant Physiology 84, 8847-850 (1987).

(5) Thomas Bjorkman and A. Leopold An Electric Current Associated with Gravity Sensing in Maize Roots. Plant Physiology 84, 841-846 (1987).

(6) Fernando Migliaccio and Arthur Galston On the Nature and Origin of the Calcium Asymmetry Arising during Gravitropic Response in Etiolated Pea Epicotyls. Plant Physiology 85, 542-547 (1987).

(7) Randy Moore, Ivan Cameron, et. al. The Locations and Amounts of Endogenous Ions and Elements in the Cap and Elongating Zone of Horizontally Oriented Rooots of Zea mays L.: An Electron-probe EDS Study. Annals of Botany 59, 667-677 (1987).

(8) Randy Moore, Ivan Cameron, and Nancy Smith Movement of Endogenous Calcium in the Elongating Zone of Graviresponding Roots of Zea mays. Annals of Botany 63, 589-593 (1989).