The Root Cap: Cell Dynamics, Cell Differentiation and Cap Function

The root cap is a universal feature of angiosperm, gymnosperm, and pteridophyte roots. Besides providing protection against abrasive damage to the root tip, the root cap is also involved in the simultaneous perception of a number of signals – pressure, moisture, gravity, and perhaps others – that modulate growth in the main body of the root. These signals, which originate in the external environment, are transduced by the cap and are then transported from the cap to the root. Root gravitropism is one much studied response to an external signal. In the present paper, consideration is given to the structure of the root cap and, in particular, to how the meristematic initial cells of both the central cap columella and the lateral portion of the cap which surrounds the columella are organized in relation to the production of new cells. The subsequent differentiation and development of these cells is associated with their displacement through the cap and their eventual release, as border cells, from the cap periphery. Mutations, particularly in Arabidopsis, are increasingly playing a part in defining not only the pattern of genetic activity within different cells of the cap but also in revealing how the corresponding wild-type proteins relate to the range of functions of the cap. Notable in this respect have been analyses of the early events of root gravitropism. The ability to image auxin and auxin permeases within the cap and elsewhere in the root has also extended our understanding of this growth response. Images of auxin distribution may, in addition, help extend ideas concerning the positional controls of cell division and cell differentiation within the cap. However, firm information relating to these controls is scarce, though there are intriguing suggestions of some kind of physiological link between the border cells surrounding the cap and mitotic activity in the cap meristem. Open questions concern the structure and functional interrelationships between the root and the cap which surmounts it, and also the means by which the cap transduces the environmental signals that are of critical importance for the growth of the individual roots, and collectively for the shaping of the root system. Current address (Peter W. Barlow): School of Biological Sciences, University of Bristol, Woodland Road, Bristol BS8 1UG, UK     

Mapping the Functional Roles of Cap Cells in the Response of Arabidopsis Primary Roots to Gravity

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.     

Complex physiological and molecular processes underlying root gravitropism

Gravitropism allows plant organs to guide their growth in relation to the gravity vector. For most roots, this response to gravity allows downward growth into soil where water and nutrients are available for plant growth and development. The primary site for gravity sensing in roots includes the root cap and appears to involve the sedimentation of amyloplasts within the columella cells. This process triggers a signal transduction pathway that promotes both an acidification of the wall around the columella cells, an alkalinization of the columella cytoplasm, and the development of a lateral polarity across the root cap that allows for the establishment of a lateral auxin gradient. This gradient is then transmitted to the elongation zones where it triggers a differential cellular elongation on opposite flanks of the central elongation zone, responsible for part of the gravitropic curvature. Recent findings also suggest the involvement of a secondary site/mechanism of gravity sensing for gravitropism in roots, and the possibility that the early phases of graviresponse, which involve differential elongation on opposite flanks of the distal elongation zone, might be independent of this auxin gradient. This review discusses our current understanding of the molecular and physiological mechanisms underlying these various phases of the gravitropic response in roots.     

Negative gravitropic response of roots directs auxin flow to control root gravitropism

Root tip is capable of sensing and adjusting its growth direction in response to gravity, a phenomenon known as root gravitropism. Previously, we have shown that negative gravitropic response of roots (NGR) is essential for the positive gravitropic response of roots. Here, we show that NGR, a plasma membrane protein specifically expressed in root columella and lateral root cap cells, controls the positive root gravitropic response by regulating auxin efflux carrier localization in columella cells and the direction of lateral auxin flow in response to gravity. Pharmacological and genetic studies show that the negative root gravitropic response of the ngr mutants depends on polar auxin transport in the root elongation zone. Cell biology studies further demonstrate that polar localization of the auxin efflux carrier PIN3 in root columella cells and asymmetric lateral auxin flow in the root tip in response to gravistimulation is reversed in the atngr1;2;3 triple mutant. Furthermore, simultaneous mutations of three PIN genes expressed in root columella cells impaired the negative root gravitropic response of the atngr1;2;3 triple mutant. Our work revealed a critical role of NGR in root gravitropic response and provided an insight of the early events and molecular basis of the positive root gravitropism.     

Root gravitropism requires lateral root cap and epidermal cells for transport and response to a mobile auxin signal

Re-orientation of Arabidopsis seedlings induces a rapid, asymmetric release of the growth regulator auxin from gravity-sensing columella cells at the root apex. The resulting lateral auxin gradient is hypothesized to drive differential cell expansion in elongation-zone tissues. We mapped those root tissues that function to transport or respond to auxin during a gravitropic response. Targeted expression of the auxin influx facilitator AUX1 demonstrated that root gravitropism requires auxin to be transported via the lateral root cap to all elongating epidermal cells. A three-dimensional model of the root elongation zone predicted that AUX1 causes the majority of auxin to accumulate in the epidermis. Selectively disrupting the auxin responsiveness of expanding epidermal cells by expressing a mutant form of the AUX/IAA17 protein, axr3-1, abolished root gravitropism. We conclude that gravitropic curvature in Arabidopsis roots is primarily driven by the differential expansion of epidermal cells in response to an influx-carrier-dependent auxin gradient.     

Gravity signal transduction in primary roots

AIMS: The molecular mechanisms that correlate with gravity perception and signal transduction in the tip of angiosperm primary roots are discussed. SCOPE: Gravity provides a cue for downward orientation of plant roots, allowing anchorage of the plant and uptake of the water and nutrients needed for growth and development. Root gravitropism involves a succession of physiological steps: gravity perception and signal transduction (mainly mediated by the columella cells of the root cap); signal transmission to the elongation zone; and curvature response. Interesting new insights into gravity perception and signal transduction within the root tip have accumulated recently by use of a wide range of experimental approaches in physiology, biochemistry, genetics, genomics, proteomics and cell biology. The data suggest a network of signal transduction pathways leading to a lateral redistribution of auxin across the root cap and a possible involvement of cytokinin in initial phases of gravicurvature. CONCLUSION: These new discoveries illustrate the complexity of a highly redundant gravity-signalling process in roots, and help to elucidate the global mechanisms that govern auxin transport and morphogenetic regulation in roots.     

ALTERED RESPONSE TO GRAVITY is a peripheral membrane protein that modulates gravity-induced cytoplasmic alkalinization and lateral auxin transport in plant statocytes

ARG1 (ALTERED RESPONSE TO GRAVITY) is required for normal root and hypocotyl gravitropism. Here, we show that targeting ARG1 to the gravity-perceiving cells of roots or hypocotyls is sufficient to rescue the gravitropic defects in the corresponding organs of arg1-2 null mutants. The cytosolic alkalinization of root cap columella cells that normally occurs very rapidly upon gravistimulation is lacking in arg1-2 mutants. Additionally, vertically grown arg1-2 roots appear to accumulate a greater amount of auxin in an expanded domain of the root cap compared with the wild type, and no detectable lateral auxin gradient develops across mutant root caps in response to gravistimulation. We also demonstrate that ARG1 is a peripheral membrane protein that may share some subcellular compartments in the vesicular trafficking pathway with PIN auxin efflux carriers. These data support our hypothesis that ARG1 is involved early in gravitropic signal transduction within the gravity-perceiving cells, where it influences pH changes and auxin distribution. We propose that ARG1 affects the localization and/or activity of PIN or other proteins involved in lateral auxin transport.     

How roots perceive and respond to gravity

Graviperception by plant roots is believed to occur via the sedimentation of amyloplasts in columella cells of the root cap. This physical stimulus results in an accumulation of calcium on the lower side of the cap, which in turn induces gravicurvature. In this paper we present a model for root gravitropism integrating gravity-induced changes in electrical potential, cytochemical localization of calcium in cells of gravistimulated roots, and the interdependence of calcium and auxin movement. Key features of the model are that 1) gravity-induced redistribution of calcium is an early event in the transduction mechanism, and 2) apoplastic movement of calcium through the root-cap mucilage may be an important component of the pathway for calcium movement.     

Happy end in sight after 70 years of controversy

Differential auxin transport from the columella to lateral root cap cells as a result of root gravitropism has recently been shown using a GFP-based auxin biosensor. Together with the recent finding of gravity-dependent localization of an auxin efflux carrier, these results strongly support the Cholodny-Went hypothesis for the tropic response, which has been disputed for 70 years.     

Cell Displacement Through the Columella of the Root Cap of Zea mays L

Exposing roots of Zea mays to a solution of caffeine for 1 hinduces a small population of binucleate cells in the meristem.The progress of the binucleate cell population was then followed,in time, as it was displaced along the length of the cap columella.Since this method of marking cells seems to have no effect onthe subsequent pattern of cell proliferation in the cap meristem,the movement of the binucleate cells through the cap is inferredto be similar to the movement of cells in an undisturbed cap.The binculeate cells that persist in the cap are believed tobe cells that were engaged in their final mitosis at the timeof the caffeine treatment, so the time that it takes for themto appear at the edge of the cap is a measure of the periodfor which a cell is contained in the non–dividing portionof the tissue before being lost from the cap surface. In rootsof Zea grown at 22 °C cells take about 7 days to reach thetip of the cap columella and about 2 to 3 days to reach theflanks of the cap following their displacement from the capmeristem. Zea mays, root cap, cell displacement, binucleate cells     

Root Graviresponsiveness and Cellular Differentiation in Wild-type and a Starchless Mutant of Arabidopsis thaliana

Primary roots of a starchless mutant of Arabidopsis thalianaL. are strongly graviresponsive despite lacking amyloplastsin their columella cells. The ultrastructures of calyptrogenand peripheral cells in wild-type as compared to mutant seedlingsare not significantly different. The largest difference in cellulardifferentiation in caps of mutant and wild-type roots is therelative volume of plastids in columella cells. Plastids occupy12.3% of the volume of columella cells in wild-type seedlings,but only 3.69% of columella cells in mutant seedlings. Theseresults indicate that: (1) amyloplasts and starch are not necessaryfor root graviresponsiveness; (2) the increase in relative volumeof plastids that usually accompanies differentiation of columellacells is not necessary for root graviresponsiveness; and (3)the absence of starch and amyloplasts does not affect the structureof calyptrogen (i.e. meristematic) and secretory (i.e. peripheral)cells in root caps. These results are discussed relative toproposed models for root gravitropism. Arabidopsis thaliana, gravitropism (root), plastids, root cap, stereology, ultrastructure     

Root gravitropism: a complex response to a simple stimulus?

Roots avoid depleting their immediate environment of essential nutrients by continuous growth. Root growth is directed by environmental cues, including gravity. Gravity sensing occurs mainly in the columella cells of the root cap. Upon reorientation within the gravity field, the root-cap amyloplasts sediment, generating a physiological signal that promotes the development of a curvature at the root elongation zones. Recent molecular genetic studies in Arabidopsis have allowed the identification of genes that play important roles in root gravitropism. Among them, the ARG1 gene encodes a DnaJ-like protein involved in gravity signal transduction, whereas the AUX1 and AGR1 genes encode proteins involved in polar auxin transport. These studies have important implications for understanding the intra- and inter-cellular signaling processes that underlie root gravitropism.     

Recovery of gravitropic response during regeneration of root caps does not require developed columella cells and sedimentation of amyloplasts

Correlations between regeneration of the root cap and recovery of a gravitropic response were studied using primary roots of Phaseolus vulgaris. After removal of various lengths of the root tip a gravistimulus was continuously given to the root. The statistical analysis of data showed that recovery of the gravitropic response was gradually delayed as the length of the tips removed increased. This suggested that the columella cells of the root cap were involved in gravitropism. When the root cap was completely removed, the roots did not respond to gravistimuli for the first 15 h and began to reorient their growth direction at 20 h. At this time, the columella cells had just begun to regenerate and had immature amyloplasts which did not sufficiently form a sediment. These results suggest that other systems of perception exist in plant cells in addition to the amyloplast-based model of graviperception.     

How do Arabidopsis Roots Differentiate Hydrotropism from Gravitropism?

Root hydrotropism is a response to moisture gradients, which is considered to be important for drought avoidance. Recent reevaluation of root hydrotropism has emphasised the dominating effect of root gravitropism on it. It has been suggested that amyloplast dynamics inside columella cells and auxin regulation play roles in this interacting mechanism, even though the existence of distinct pathways of two tropisms derived from different stimuli remained unclear. We have recently found two factors that separate the mechanism of hydrotropism from that of gravitropism in Arabidopsis seedling roots. One is the difference in the mode of auxin-mediated growth regulation between two tropisms, and the other is the identification of gene indispensable only for root hydrotropism. Here we summarize the recent progress on root hydrotropism research and discuss the remaining and emerging issues.Key Words: auxin, gravitropism, hydrotropism, root, MIZU-KUSSEI1 (MIZ1)     

Early development and gravitropic response of lateral roots in Arabidopsis thaliana

Root system architecture plays an important role in determining nutrient and water acquisition and is modulated by endogenous and environmental factors, resulting in considerable developmental plasticity. The orientation of primary root growth in response to gravity (gravitropism) has been studied extensively, but little is known about the behaviour of lateral roots in response to this signal. Here, we analysed the response of lateral roots to gravity and, consistently with previous observations, we showed that gravitropism was acquired slowly after emergence. Using a lateral root induction system, we studied the kinetics for the appearance of statoliths, phloem connections and auxin transporter gene expression patterns. We found that statoliths could not be detected until 1 day after emergence, whereas the gravitropic curvature of the lateral root started earlier. Auxin transporters modulate auxin distribution in primary root gravitropism. We found differences regarding PIN3 and AUX1 expression patterns between the lateral root and the primary root apices. Especially PIN3, which is involved in primary root gravitropism, was not expressed in the lateral root columella. Our work revealed new developmental transitions occurring in lateral roots after emergence, and auxin transporter expression patterns that might explain the specific response of lateral roots to gravity.     

ARL2, ARG1 and PIN3 define a gravity signal transduction pathway in root statocytes

ALTERED RESPONSE TO GRAVITY1 (ARG1) and its paralog ARG1-LIKE2 (ARL2) are J-domain proteins that are required for normal root and hypocotyl gravitropism. In this paper, we show that both ARL2 and ARG1 function in a gravity signal transduction pathway with PIN3, an auxin efflux facilitator that is expressed in the statocytes. In gravi-stimulated roots, PIN3 relocalizes to the lower side of statocytes, a process that is thought to, in part, drive the asymmetrical redistribution of auxin toward the lower flank of the root. We show that ARL2 and ARG1 are required for PIN3 relocalization and asymmetrical distribution of auxin upon gravi-stimulation. ARL2 is expressed specifically in the root statocytes, where it localizes to the plasma membrane. Upon ectopic expression, ARL2 is also found at the cell plate of dividing cells during cytokinesis, an area of intense membrane dynamics. Mutations in ARL2 and ARG1 also result in auxin-related expansion of the root cap columella, consistent with a role for ARL2 and ARG1 in regulating auxin flux through the root tip. Together these data suggest that ARL2 and ARG1 functionally link gravity sensation in the statocytes to auxin redistribution through the root cap.     

Role of cytokinin in the regulation of root gravitropism

The models explaining root gravitropism propose that the growth response of plants to gravity is regulated by asymmetric distribution of auxin (indole-3-acetic acid, IAA). Since cytokinin has a negative regulatory role in root growth, we suspected that it might function as an inhibitor of tropic root elongation during gravity response. Therefore, we examined the free-bioactive-cytokinin-dependent ARR5::GUS expression pattern in root tips of transformants of Arabidopsis thaliana (L.) Heynh., visualized high cytokinin concentrations in the root cap with specific monoclonal antibodies, and complemented the analyses by external application of cytokinin. Our findings show that mainly the statocytes of the cap produce cytokinin, which may contribute to the regulation of root gravitropism. The homogenous symmetric expression of the cytokinin-responsive promoter in vertical root caps rapidly changed within less than 30 min of gravistimulation into an asymmetrical activation pattern, visualized as a lateral, distinctly stained, concentrated spot on the new lower root side of the cap cells. This asymmetric cytokinin distribution obviously caused initiation of a downward curvature near the root apex during the early rapid phase of gravity response, by inhibiting elongation at the lower side and promoting growth at the upper side of the distal elongation zone closely behind the root cap. Exogenous cytokinin applied to vertical roots induced root bending towards the application site, confirming the suspected inhibitory effect of cytokinin in root gravitropism. Our results suggest that the early root graviresponse is controlled by cytokinin. We conclude that both cytokinin and auxin are key hormones that regulate root gravitropism.Electronic Supplementary Material Supplementary material is available in the online version of this article at http://dx.doi.org/10.1007/s00425-004-1381-8     

Negative phototropism of rice root and its influencing factors

Some characteristics of the rice (Oryza sativa L.) root were found in the experiment of unilaterally irradiating the roots which were planted in water: (i) All the seminal roots, adventitious roots and their branched roots bent away from light, and their curvatures ranged from 25° to 60°. The curvature of adventitious root of the higher node was often larger than that of the lower node, and even larger than that of the seminal root, (ii) The negative phototropic bending of the rice root was mainly due to the larger growth increment of root-tip cells of the irradiated side compared with that of the shaded side, (iii) Root cap was the site of light perception. If root cap was shaded while the root was irradiated the root showed no negative phototropism, and the root lost the characteristic of negative phototropism when root cap was divested. Rice root could resume the characteristic of negative phototropism when the new root cap grew up, if the original cells of root cap were well protected while root cap was divested, (iv) The growth increment and curvature of rice root were both influenced by light intensity. Within the range of 0–100 μmol · m² -s⁻¹, the increasing of light intensity resulted in the decreasing of the growth increment and the increasing of the curvature of rice root, (v) The growth increment and the curvature reached the maximum at 30°C with the temperature treatment of 10–40°C. (vi) Blue-violet light could prominently induce the negative phototropism of rice root, while red light had no such effect. (vii) The auxin (IAA) in the solution, as a very prominent influencing factor, inhibited the growth, the negative phototropism and the gravitropism of rice root when the concentration of IAA increased. The response of negative phototropism of rice root disappeared when the concentration of IAA was above 10 mg · L⁻¹     

The Locations and Amounts of Endogenous lons and Elements in the Cap and Elongating Zone of Horizontally Oriented Roots of Zea mays L: An Electron-probe EDS Study

We used quantitative electron-probe energy-dispersive x-raymicroanalysis to localize endogenous Na, Cl, K, P, S, Mg andCa in cryofixed and freeze-dried cryosections of the cap (i.e.the putative site of graviperception) and elongating zone (i.e.site of gravicurvature) of horizontally oriented roots of Zeamays. Ca, Na, Cl, K and Mg accumulate along the lower side ofcaps of horizontally oriented roots. The most dramatic asymmetriesof these ions occur in the apoplast, especially the mucilage.We could not detect any significant differences in the concentrationsof these ions in the central cytoplasm of columella cells alongthe upper and lower sides of caps of horizontally-oriented roots.However, the increased amounts of Na, Cl, K and Mg in the longitudinalwalls of columella cells along the lower side of the cap suggestthat these ions may move down through the columella tissue ofhorizontally-oriented roots. Ca also accumulates (largely inthe mucilage) along the lower side of the elongating zone ofhorizontally-oriented roots, while Na, P, Cl and K tend to accumulatealong the upper side of the elongating zone. Of these ions,only K increases in concentration in the cytoplasm and longitudinalwalls of cortical cells in the upper vs lower sides of the elongatingzone. These results indicate that (1) gravity-induced asymmetriesof ions differ significantly in the cap and elongating zoneof graviresponding roots, (2) Ca accumulates along the lowerside of the cap and elongating zone of graviresponding roots,(3) increased growth of the upper side of the elongating zoneof horizontally-oriented roots correlates positively with increasedamounts of K in the cytoplasm and longitudinal walls of corticalcells, and (4) the apoplast (especially the mucilage) may bean important component of the pathway via which ions move ingraviresponding rots of Zea mays. These results are discussedrelative to mechanisms for graviperception and gravicurvatureof roots. Corn, gravitropism (root), ions, x-ray microanalysis, Zea mays     

Growth and gravitropism of corn roots in solution

Growth and early gravitropic responses of corn roots in solution have been studied using time-lapse photography. Aeration was required for both root growth and gravitropism. The optimum pH for gravitropism was in the range 5 to 6. The bending response seemed to be greater for roots in non-buffered solution than in buffered solution. Fastest growth and maximum curvature occurred with about 0.2 mol m⁻³ Ca²⁺. Under some conditions, the gravitropic response started with apparently negligible time delay after the start of the gravitropic stimulus. This may denote graviperception in or near the elongation zone itself. This mechanism for early but relatively weak gravitropism may help to explain a variety of gravitropic responses such as the ‘early wrong way’ curvature, and the behaviour of roots whose columella cells lack amyloplasts. More rapid bending appears to start at about 20 min, which is consistent with observations on roots in humid air and with the accepted statolith model of perception in the root cap.