Calcium Movement, Graviresponsiveness and the Structure of Columella Cells and Columella Tissues in Roots of Allium cepa L.

Roots of Allium cepa L. cv. Yellow are differentially responsiveto gravity. Long (e.g. 40 mm) roots are strongly graviresponsive,while short (e.g. 4 mm) roots are minimally responsive to gravity.Although columella cells of graviresponsive roots are largerthan those of nongraviresponsive roots, they partition theirvolumes to cellular organelles similarly. The movement of amyloplastsand nuclei in columella cells of horizontally-oriented rootscorrelates positively with the onset of gravicurvature. Furthermore,there is no significant difference in the rates of organellarredistribution when graviresponsive and nongraviresponsive rootsare oriented horizontally. The more pronounced graviresponsivenessof longer roots correlates positively with (1) their caps being9.6 times more voluminous, (2) their columella tissues being42 times more voluminous, (3) their caps having 15 times morecolumella cells, and (4) their columella tissues having relativevolumes 4·4 times larger than those of shorter, nongraviresponsiveroots. Graviresponsive roots that are oriented horizontallyare characterized by a strongly polar movement of ⁴⁵Ca²⁺ acrossthe root tip from the upper to the lower side, while similarlyoriented nongraviresponsive roots exhibit only a minimal polartransport of ⁴⁵Ca²⁺. These results indicate that the differentialgraviresponsiveness of roots of A. cepa is probably not dueto either (1) ultrastructural differences in their columellacells, or (2) differences in the rates of organellar redistributionwhen roots are oriented horizontally. Rather, these resultsindicate that graviresponsiveness may require an extensive columellatissue, which, in turn, may be necessary for polar movementof ⁴⁵Ca²⁺ across the root tip. Allium cepa, onion, root, columella tissue, columella cell, gravitropism, calcium, ultrastructure     

CALCIUM MOVEMENT,GRAVIRESPONSIVENESS, AND THE STRUCTURE OF COLUMELLA CELLS IN PRIMARY ROOTS OF AMYLOMAIZE MUTANTS OF ZEA MAYS

Primary roots of Zea mays cv. Amylomaize were less graviresponsive than primary roots of the wild-type Calumet cultivar. There were no significant differences in: 1) the partitioning of volume to organelles in columella cells, 2) the size or density of amyloplasts, or 3) rates and overall patterns of organelle redistribution in horizontally-oriented roots of the two cultivars. Amyloplasts and nuclei were the only organelles whose movement correlated positively with the onset of root gravicurvature. However, the onset of gravicurvature was not directly proportional to the average sedimentation rate of amyloplasts, since amyloplasts sedimented at equal rates in columella cells of both cultivars despite their differences in root gravicurvature. The more graviresponsive roots of Calumet seedlings were characterized by a more strongly polar movement of ⁴⁵Ca²⁺ from the upper to lower sides of their root tips than the less graviresponsive roots of Amylomaize seedlings. These results suggest that the decreased graviresponsiveness of horizontally-oriented roots of Amylomaize seedlings may be due to a delay in or decreased ability for polar transport of calcium rather than to smaller, more slowly sedimenting amyloplasts as has been suggested for their less graviresponsive coleoptiles.     

Movement of calcium across tips of primary and lateral roots of Phaseolus vulgaris

Calcium (Ca) movement across tips of primary and lateral roots of Phaseolus vulgaris was determined by applying 45Ca2+ to one side of the root and collecting radioactivity in an agar receiver block on the opposite side of the root. The ratios of cpm in receiver blocks on the bottom of primary roots : cpm in receiver blocks on the top of the primary roots were 1.87 and 2.47 after 1 and 2 hr, respectively. This polar transport of Ca across tips of primary roots correlated positively with a graviculture of 43 degrees after 2 hr. The ratio of cpm in receiver blocks on the bottom of lateral roots : cpm in receiver blocks on the top of lateral roots was 1.20 after 2 hr. The decreased polar movement of Ca across tips of lateral roots correlated positively with lateral roots being nongraviresponsive. These data 1) support the suggestion that gravistimulation induces polar movement Ca toward the lower side of tips of primary roots, and 2) suggest that the reduced polar movement of Ca across tips of lateral roots may be involved in uncoupling gravistimulation from gravicurvature in lateral roots.     

A morphometric analysis of the redistribution of organelles in columella cells in primary roots of normal seedlings and agravitropic mutants of Hordeum vulgare

The redistribution of organelles in columella cells of horizontally-oriented roots of Hordeum vulgare was quantified in order to determine what structural changes in graviperceptive (i.e., columella) cells are associated with the onset of the root gravicurvature. The sedimentation of amyloplasts is the only major change in cellular structure that correlates positively with the onset of root gravicurvature, which begins within 15 min after re-orientation. There is no consistent contact between sedimented amyloplasts and any other organelles. Nuclei are restricted to the proximal ends of columella cells in vertically-oriented roots, and remain there throughout gravicurvature after roots are oriented horizontally. Root gravicurvature does not involve significant changes in (1) the volume of columella cells, (2) the relative or absolute volumes of organelles in columella cells, or (3) the distribution of endoplasmic reticulum (ER). The size, number and sedimentation rates of amyloplasts in columella cells of non-graviresponsive roots of mutant seedlings are not significantly different from those of graviresponsive roots of normal seedlings. Similarly, there is no significant difference in (1) cellular volume, (2) distribution or surface area of ER, (3) patterns or rates of organelle redistribution in horizontally-oriented roots, (4) relative or absolute volumes of organelles in columella cells of graviresponsive and non-graviresponsive roots. These results suggest that the lack of graviresponsiveness by roots of mutant seedlings is probably not due to either (1) structural differences in columella cells, or (2) differences in patterns or rates of organelle redistribution as compared to that characteristic of graviresponsive roots. Thus, the basis of non-graviresponsiveness in this mutant is probably different from other agravitropic mutants so far studied.     

Inhibition of gravitropism in primary roots of Zea mays by chloramphenicol

Primary roots of Zea mays seedlings germinated and grown in 0.1 mM chloramphenicol (CMP) were significantly less graviresponsive than primary roots of seedlings germinated and grown in distilled water. Elongation rates of roots treated with CMP were significantly greater than those grown in distilled water. Caps of control and CMP-treated roots possessed extensive columella tissues comprised of cells containing numerous sedimented amyloplasts. These results indicate that the reduced graviresponsiveness of CMP-treated roots is not due to reduced rates of elongation, the absence of the presumed gravireceptors (i.e., amyloplasts in columella cells), or reduced amounts of columella tissue. These results are consistent with CMP altering the production and/or transport of effectors that mediate gravitropism.     

Graviresponsiveness and columella cell structure in primary and secondary roots of Ricinus communis

In order to determine what structural changes are associated with the onset of graviresponsiveness by plant roots, we have monitored the quantitative ultrastructures of columella (i.e., graviperceptive) cells in primary and secondary roots of Ricinus communis. The relative volumes of cellular components in lateral (i.e., minimally graviresponsive) roots were not significantly different from those of primary roots. The relative volumes of cellular components in secondary roots growing laterally were not significantly different from those of graviresponsive secondary roots. Therefore, the onset of graviresponsiveness by secondary roots of R. communis is not correlated with changes in organellar concentrations in columella cells. These results are discussed relative to a model for the differential graviresponsiveness of plant roots.     

Cytochemical localization of calcium in cap cells of primary roots of Zea mays L

The distribution of calcium (Ca) in caps of vertically- and horizontally-oriented roots of Zea mays was monitored to determine its possible role in root graviresponsiveness. A modification of the antimonate precipitation procedure was used to localize Ca in situ. In vertically-oriented roots, the presumed graviperceptive (i.e., columella) cells were characterized by minimal and symmetric staining of the plasmalemma and mitochondria. No precipitate was present in plasmodesmata or cell walls. Within 5 min after horizontal reorientation, staining was associated with the portion of the cell wall adjacent to the distal end of the cell. This asymmetric staining persisted throughout the onset of gravicurvature. No staining of lateral cell walls of columella cells was observed at any stage of gravicurvature, suggesting that a lateral flow of Ca through the columella tissue of horizontally-oriented roots does not occur. The outermost peripheral cells of roots oriented horizontally and vertically secrete Ca through plasmodesmata-like structures in their cell walls. These results are discussed relative to proposed roles of root-cap Ca in root gravicurvature.     

A Morphometric Analysis of the Redistribution of Organelles in Columella Cells in Primary Roots of Normal Seedlings and Agravitropic Mutants of Hordeum vulgare

Moore, R. 1985. A morphometric analysis of the redistributionof organellcs in columella cells in primary roots of normalseedlings and agravitropic mutants of Hordeum vulgare.—J.exp. Bot. 36:1275–1286. The redistribution of organeUes m columella cells of horizontally-orientedroots of Hordeum vulgare was quantified in order to determinewhat structural changes in graviperceptive (i.e, columella)cells are associated with the onset of root gravicurvature.The sedimentation of amyloplasts is the only major change incellular structure that correlates positively with the onsetof root gravicurvature, which begins within 15 min after re-orientation.There is no consistent contact between sedimented amyloplastsand any other organelles. Nuclei are restricted to the proximalends of columella cells in vertically-oriented roots, and remainthere throughout gravicurvature after roots are oriented horizontally.Root gravicurvature does not involve significant changes in(1) the volume of columella cells, (2) the relative or absolutevolumes of organelles in columella cells, or (3) the distributionof endoplasmic reticulum (ER). The size, number and sedimentationrates of amyloplasts in columella cells of non-graviresponsiveroots of mutant seedlings are not significantly different fromthose of graviresponsive roots of normal seedlings. Similarly,there is no significant difference in (1) cellular volume, (2)distribution or surface area of ER, (3) patterns or rates oforganelle redistribution in horizontally-oriented roots, or(4) relative or absolute volumes of organelles in columellacells of graviresponsive and non-graviresponsive roots. Theseresults suggest that the lack of gravi-responsiveness by rootsof mutant seedlings is probably not due to either (1) structuraldifferences in columella cells, or (2) differences in patternsor rates of organelle redistribution as compared to that characteristicof graviresponsive roots. Thus, the basis of non-graviresponsivenessin this mutant is probably different from other agravitropicmutants so far studied. Key words: Agravitropic mutant, barley, columella cell, gravitropism (root), Hordeum vulgare, ultrastructure     

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     

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.     

Polar Transport of Auxin across Gravistimulated Roots of Maize and Its Enhancement by Calcium

The effect of Ca on the polar movement of [³H]indoleacetic acid ([³H] IAA) in gravistimulated roots was examined using 3-day-old seedlings of maize (Zea mays L.). Transport of label was measured by placing an agar donor block containing [³H]IAA on one side of the elongation zone and measuring movement of label across the root into an agar receiver block on the opposite side. In vertically oriented roots, movement of label across the elongation zone into the receiver was slight and was not enhanced by incorporating 10 millimolar CaCl₂ into the receiver block. In horizontally oriented roots, movement of label across the root was readily detectable and movement to a receiver on the bottom was about 3-fold greater than movement in the opposite direction. This polarity was abolished in roots from which the caps were removed prior to gravistimulation. When CaCl₂ was incorporated into the receivers, movement of label across horizontally oriented intact roots was increased about 3-fold in both the downward and upward direction. The ability of Ca to enhance the movement of label from [³H]IAA increased with increasing Ca concentration in the receiver up to 5 to 10 millimolar CaCl₂. With the inclusion of CaCl₂ in the receiver blocks, gravity-induced polar movement of label into receiver blocks from applied [³H]IAA was detectable within 30 minutes, and asymmetric distribution of label within the tissue was detectable within 20 minutes. The results indicate that gravistimulation induces a physiological asymmetry in the auxin transport system of maize roots and that Ca increases the total transport of auxin across the root.     

Graviresponsiveness and the Development of Columella Tissue in Primary and Lateral Roots of Ricinus communis

Half-tipped primary and lateral roots of Ricinus communis cv Hale bend toward the side of the root on which the intact half-tip remains. Therefore, the minimal graviresponsiveness of lateral roots is not due to the inability of their caps to produce growth effectors (presumably inhibitors). The columella tissues of primary (i.e. graviresponsive) roots are (a) 4.30 times longer, (b) 2.95 times wider, (c) 37.4 times more voluminous, and (d) composed of 17.2 times more cells than those of lateral roots. The onset of positive gravitropism by lateral roots is positively correlated with a (a) 2.99-fold increase in length, (b) 2.63-fold increase in width, and (c) 20.7-fold increase in volume of their columella tissues. We propose that the minimal graviresponsiveness of lateral roots is due to the small size of their columella tissues, which results in their caps being unable to (a) establish a concentration gradient of the effector sufficient to induce gravicurvature and (b) produce as much of the effector as caps of graviresponsive roots.     

The effects of asymmetric calcium application on primary root curvature of ageotropum pea mutant.

Recent studies indicate that roots of ageotropum seedlings can be used to study the hydrotropic response of roots independent of physiological events related to the gravity response of roots. There is evidence that Ca2+ ions are important in both the gravitropic and hydrotropic response of roots. In this study, we have compared three fully graviresponsive pea cultivars and the ageotropum mutant with regard to: 1) general root anatomy, 2) the effects of unilateral Ca application to both the root cap and DEZ region on root curvature, and 4) effects of unilateral application of EGTA to the DEZ region.     

Dimensions of Root Caps and Columella Tissues of Primary Roots of Ricinus communis Characterized by Differing Degrees of Graviresponsiveness

Primary roots of Ricinus communis having large caps and columellatissues are more graviresponsive than primary roots with smallcaps and columella tissues. The increased graviresponsivenessof roots with larger caps correlates positively with their columellatissues having larger length: width ratios than less graviresponsiveroots having smaller caps. Roots with wider tips typically aremore graviresponsive and have more extensive columellas thanroots with thinner tips. However, the size of the columellatissue correlates positively with graviresponsiveness, irrespectiveof the width of the root tip. These results indicate that differingdimensions of the columella tissue may be the basis for thediffering graviresponses of primary roots of R. communis. Root gravitropism, columella, root cap, primary root, Ricinus communis, castor bean     

Inhibition of polar calcium movement and gravitropism in roots treated with auxin-transport inhibitors

Primary roots of maize (Zea mays L.) and pea (Pisum sativum L.) exhibit strong positive gravitropism. In both species, gravistimulation induces polar movement of calcium across the root tip from the upper side to the lower side. Roots of onion (Allium cepa L.) are not responsive to gravity and gravistimulation induces little or no polar movement of calcium across the root tip. Treatment of maize or pea roots with inhibitors of auxin transport (morphactin, naphthylphthalamic acid, 2,3,5-triiodobenzoic acid) prevents both gravitropism and gravity-induced polar movement of calcium across the root tip. The results indicate that calcium movement and auxin movement are closely linked in roots and that gravity-induced redistribution of calcium across the root cap may play an important role in the development of gravitropic curvature.Abbreviations 9-HFCA 9-hydroxyfluorenecarboxylic acid - NPA naphthylphthalamic acid - TIBA 2,3,5-triiodobenzoic acid - IAA indole-3-acetic acid     

Gravity-Induced Polar Transport of Calcium across Root Tips of Maize

Calcium movement across primary roots of maize (Zea mays, L.) was determined by application of ⁴⁵Ca²⁺ to one side of the root and collection of radioactivity in an agar receiver block on the opposite side. Ca movement across the root tip was found to be at least 20 times greater than movement across the elongation zone. The rapid movement of Ca across the tip was severely inhibited in roots from which the root cap had been removed. Ca movement across the tip was also strongly retarded in roots pretreated with 2,4-dinitrophenol or potassium cyanide. Orientation of roots horizontally had no effect on Ca movement across the elongation zone but caused a strong asymmetry in the pattern of Ca movement across the tip. In gravistimulated roots, the movement of Ca from top to bottom increased while movement from bottom to top decreased. The data indicate that gravistimulation induces polar movement of Ca toward the lower side of the root cap. An earlier report (Lee, Mulkey, Evans 1983 Science 220: 1375-1376) from this laboratory showed that artificial establishment of calcium gradients at the root tip can cause gravitropic-like curvature. Together, the two studies indicate that Ca plays a key role in linking gravistimulation to the gravitropic growth response in roots.     

Influence of microgravity on root-cap regeneration and the structure of columella cells in Zea mays

We launched imbibed seeds and seedlings of Zea mays into outer space aboard the space shuttle Columbia to determine the influence of microgravity on 1) root-cap regeneration, and 2) the distribution of amyloplasts and endoplasmic reticulum (ER) in the putative statocytes (i.e., columella cells) of roots. Decapped roots grown on Earth completely regenerated their caps within 4.8 days after decapping, while those grown in microgravity did not regenerate caps. In Earth-grown seedlings, the ER was localized primarily along the periphery of columella cells, and amyloplasts sedimented in response to gravity to the lower sides of the cells. Seeds germinated on Earth and subsequently launched into outer space had a distribution of ER in columella cells similar to that of Earth-grown controls, but amyloplasts were distributed throughout the cells. Seeds germinated in outer space were characterized by the presence of spherical and ellipsoidal masses of ER and randomly distributed amyloplasts in their columella cells. These results indicate that 1) gravity is necessary for regeneration of the root cap, 2) columella cells can maintain their characteristic distribution of ER in microgravity only if they are exposed previously to gravity, and 3) gravity is necessary to distribute the ER in columella cells of this cultivar of Z. mays.     

Root Gravitropism in a Cultivar of Zea mays Whose Columella Cells Contain Starch-deficient Amyloplasts

Starch occupies 4.2 per cent of the volume of plastids in calyptrogencells in primary roots of Zea mays L. cv. vp-7 wild type. Plastidsin calyptrogen cells are distributed randomly around large,centrally located nuclei. The differentiation of calyptrogencells into columella cells is characterized by cellular enlargementand the sedimentation of plastids to the bottom of the cells.Although sedimented plastids in columella cells do not containsignificantly more starch than those in calyptrogen cells, primaryroots are graviresponsive. The onset of root gravicurvatureis not associated with a significant change in the distributionof plastids in columella cells. These results indicate thatin this cultivar of Z. mays (1) the sedimentation of plastidsin columella cells is not based upon their increased densityresulting from increased starch content alone, (2) starch-ladenamyloplasts need not be present in columella cells for rootsto be graviresponsive, and (3) the onset of root gravicurvaturedoes not require a major redistribution of plastids in columellacells. Columella cell, gravitropism (root), plastids, root cap, Zea mays     

Species differences in ligand specificity of auxin-controlled elongation and auxin transport: comparing Zea and Vigna

The plant hormone auxin affects cell elongation in both roots and shoots. In roots, the predominant action of auxin is to inhibit cell elongation while in shoots auxin, at normal physiological levels, stimulates elongation. The question of whether the primary receptor for auxin is the same in roots and shoots has not been resolved. In addition to its action on cell elongation in roots and shoots, auxin is transported in a polar fashion in both organs. Although auxin transport is well characterized in both roots and shoots, there is relatively little information on the connection, if any, between auxin transport and its action on elongation. In particular, it is not clear whether the protein mediating polar auxin movement is separate from the protein mediating auxin action on cell elongation or whether these two processes might be mediated by one and the same receptor. We examined the identity of the auxin growth receptor in roots and shoots by comparing the response of roots and shoots of the grass Zea mays L. and the legume Vigna mungo L. to indole-3-acetic acid, 2-naphthoxyacetic acid, 4,6-dichloroindoleacetic acid, and 4,7-dichloroindoleacetic acid. We also studied whether or not a single protein might mediate both auxin transport and auxin action by comparing the polar transport of indole-3-acetic acid and 2-naphthoxyacetic acid through segments from Vigna hypocotyls and maize coleoptiles. For all of the assays performed (root elongation, shoot elongation, and polar transport) the action and transport of the auxin derivatives was much greater in the dicots than in the grass species. The preservation of ligand specificity between roots and shoots and the parallels in ligand specificity between auxin transport and auxin action on growth are consistent with the hypothesis that the auxin receptor is the same in roots and shoots and that this protein may mediate auxin efflux as well as auxin action in both organ types.     

The Possible Involvement of Root-cap Mucilage in Gravitropism and Calcium Movement Across Root Tips of Allium cepa L

Roots of Allium cepa L. grown in aerated water elongate rapidly,but are not graviresponsive. These roots (1) possess extensivecolumella tissues comprised of cells containing numerous sedimentedamyloplasts, (2) lack mucilage on their tips, and (3) are characterizedby a weakly polar movement of calcium (Ca) across their tips.Placing roots in humid air correlates positively with the (1)onset of gravicurvature, (2) appearance of mucilage on tipsof the roots, and (3) onset of the ability to transport Ca polarlyto the lower side of the root tip. Gravicurvature of roots previouslysubmerged in aerated water is more rapid when roots are orientedvertically for 1–2 h in humid air prior to being orientedhorizontally. The more rapid gravicurvature of these roots correlatespositively with the accumulation of mucilage at the tips ofroots during the time the roots are oriented vertically. Therefore,the onset of gravicurvature and the ability of roots to transportCa to the lower sides of their tips correlate positively withthe presence of mucilage at their tips. These results suggestthat mucilage may be important for the transport of Ca acrossroot caps. Allium cepa, root gravitropism, root mucilage, calcium, onion