cell wall adaptations to multiple environmental stresses ...cell wall adaptations to multiple...

Journal of Experimental Botany, Vol. 51, No. 344, pp. 595–603, March 2000

Cell wall adaptations to multiple environmental stressesin maize roots

Birgit Degenhardt and Hartmut Gimmler1

Julius-von-Sachs-Institut fur Biowissenschaften, Lehrstuhl Botanik I, Universitat Wurzburg, Germany

Received 19 July 1999; Accepted 10 October 1999

Abstract shown the alterations of root architecture and structureinduced by a variety of stressful conditions (Neumann

A municipal solid-waste bottom slag was used to growet al., 1994; Shannon et al., 1994; Tsegaye and Mullins,

maize plants under various abiotic stresses (high pH,1994; Plaut et al., 1997). For instance, it is known that

high salt and high heavy metal content) and to analyse plants respond to salt stress by a reduction in root growththe structural and chemical adaptations of the cell and length. A decrease in the diameter of wheat rootswalls of various root tissues. When compared with and of the transectional area of cortical cells was observedroots of control plants, more intensive wall thickenings with heavy metal treatment (Setia and Bala, 1994).were detected in the inner tangential wall of the endo- Besides changes in external root morphology there aredermis. In addition, phi thickenings in the rhizodermis also alterations in the fine structure of the root. It hasin the oldest part of the seminal root were induced been shown (Shannon et al., 1994) that salinity promoteswhen plants were grown in the slag. The role of the suberization of the hypodermis and endodermis and thatphi thickenings may not be a barrier for solutes as an the Casparian strip is developed closer to the root tipapoplastic dye could freely diffuse through them. The than in non saline roots. In cotton seedling roots, thechemical composition of cell walls from endodermis formation of an exodermis can be induced by salinityand hypodermis was analysed. Slag-grown plants had (Reinhardt and Rost, 1995). In addition, some planthigher amounts of lignin in endodermal cell walls when species develop cell wall thickenings in different rootcompared to control plants and a higher proportion of tissues. These thickenings are modifications of the mid-H-type lignin in the cell walls of the hypodermis. portion of the radial cell walls and are therefore calledFinally, the amount of aliphatic suberin in both endo- phi thickenings. They can either form a uniseriate or aand hypodermal cell walls was not affected by growing multiseriate layer (Guttenberg, 1968; Peterson et al., 1981;the plants on slag. The role of these changes in relation Eschrich, 1995). Phi thickenings are formed on theto the increase in mechanical strengthening of the walls of certain cell layers in the root cortex of severalroot is discussed. species of gymnosperms (Taxaceae, Podocarpaceae,

Cupressaceae) (Guttenberg, 1968; Haas et al., 1976) andKey words: Endodermis, exodermis, heavy metals, lignin, a few families of angiosperms (Rosaceae, Geraniaceae,phi thickenings. Berberidaceae, Sapindaceae) (Peterson et al., 1981;

Praktikakis et al., 1998). Thickenings can also be formedin the hypodermis (e.g. geranium, Haas et al., 1976). AllIntroductionspecies reported in the literature are dicotyledons.

Plants have developed a variety of strategies and mechan- However, the occurrence of phi thickenings in roots ofisms to react to any change of their environment. monocotyledons has not yet been documented.Anatomical and physiological adaptations allow better Bulk density, porosity and mechanical impedance aregrowth under unfavourable conditions. The root system other factors influencing root growth and extensionis particularly affected by these changes because the root (Bennie, 1996). Increased mechanical impedance slowsis that part of the plant which is in direct contact to the root extension and can alter cell size and cell number in

the cortex (Goss and Russell, 1980).contaminated soil substrate. A number of studies have

1 To whom correspondence should be addressed. Fax: +49 931 888 6158. E-mail: [email protected]

© Oxford University Press 2000

596 Degenhardt and Gimmler

Soil solutionIn this study, two different soil substrates were com-Soil solutions were obtained by negative pressure filtration withpared for their effect on the growth of corn roots: gardensuction cups (ceramics suction cups, Oekos, Gottingen,mould was used as a control substrate and MSW (muni-Germany).cipal solid waste) bottom incinerator slag was used as a

contaminated substrate including different sources of Microscopic techniquesstress like salinity, high pH, heavy metals, and high For all microscopic investigations sections were made in themechanical impedance. MSW incinerator slag is used in upper third of the primary roots. By reason of different rootGermany in accordance with German law as support and growth of slag and control plants, the whole root was divided

into three parts. For slag roots, the upper third is 1–5 cm fromconstruction material (LAGA, 1994). In general, it isthe root base, for control roots 1–15 cm. Before use, rootused for roads, banks or parking lots as a 30–50 cm thicksegments were fixed for 24 h in a phosphate buffer (10 mM,top layer. The environmental compatibility of this applica- pH 7.4) containing 3.7% (w/w) formaldehyde. Sections of 20 mm

tion is under discussion. were cut at −25 °C using a cryomicrotome (Cryostat H 500 M;The aim of this study was to investigate how tissues in Microm) and examined by an Axioplan microscope (Zeiss,

Oberkochen, Germany) equipped with an Osram HBO 50 Wcorn roots were affected by the presence of various sourcesmercury lamp and Zeiss filter sets (exciter filter 365 nm,of stress. Therefore, not only the development and struc-dichroitic mirror FT 395, barrier filter LP 395).ture of the root, but also particular components of the

cell wall were examined. Tracer application

To test the permeability properties of the phi thickenings, aberberine–thiocyanate tracer procedure was applied (Enstoneand Peterson, 1992). The chemicals necessary for the experimentMaterials and methodswere obtained from Sigma (Deisenhofen, Germany). Themovement of the fluorescent dye was observed on sectionsPlant materialunder blue light using an axioplan microscope (Carl Zeiss,Seeds of corn (Zea mays L. cv. Garant FAO 240) were obtainedOberkochen, Germany).from Asgrow GmbH, Bruchsal, Germany. Seeds were imbibed

The Casparian bands of endodermis and hypodermis werein aerated water overnight and planted in pots (90 mm diameter)visualized by treating root sections with 0.1% berberinefilled with soil. Plants were grown in a greenhouse (20 °C,hemisulphate for 1 h, rinsing the tissue several times with water16/8 h, light/dark) for 20 d, and watered daily with tap water.and transferring to a 0.5% toluidine blue staining solution for30 min. Toluidine blue is used instead of aniline blue. The tissue

Soil substrates was rinsed once again with distilled water, then placed on slidesin mounting medium (0.1% FeCl3 in 50% glycerol ) and viewedProcessed MSW slag, obtained from a MSW slag processingunder violet fluorescence excitation (Brundrett et al., 1988).plant Wurzburg (CC Reststoff GmbH, Germany), was used as

a substrate expected to induce various kinds of stresses (alkalineIsolation of hypodermal and endodermal cell wallsand salt stress, heavy metal stress) on plant roots.

In order to distinguish the alkaline stress from the salt stress The isolation and purification procedures of cell walls of theof the slag, the salt burden of the slag was removed by washing hypodermis and endodermis have been described in detailwith a 3-fold volume of water. For this purpose a defined previously (Schreiber et al., 1994). Roots were cut into segmentsvolume of slag was put into an Erlenmeyer flask, the same of approximately 5 cm length using a razor blade. Rootvolume of distilled water was added and the mixture was segments were incubated at room temperature in a citric buffershaken for 1 h. Then the mixture was allowed to settle, filtered solution (10−2 M, pH 3) containing cellulase (Onozuka R-10,and the solid was resuspended with fresh water. This procedure Serva, Heidelberg) and pectinase (Macerozyme R-10, Serva,was repeated three times. The supernatant was discarded, the Heidelberg). Sodium azide was added to prevent microbialslag was dried at 105 °C and stored until use. growth. After 10–15 d of enzymatic digestion, hypodermal and

A garden mould composed of organic soil, peat and sand in endodermal cell walls were separated by means of two forcepsa ratio of 452:1, by vol. was used as a reference soil. Before use and a binocular microscope (SZ 30, Olympus, Hamburg).the organic soil was steam-heated (20 min, 90 °C) to kill fungiand weed seeds. Determination of lignin and suberin

Thioacidolysis: To detect the biopolymer lignin, isolated cellElement analysis wall material was subjected to the thioacidolysis-procedure

(according to Lapierre et al., 1991). Cell wall isolates wereThe elemental composition of the municipal incinerator slagstirred in a mixture of BF3 etherate (Merck, Darmstadt,and the garden mould was determined after homogenization ofGermany), ethanethiol (Fluka, Neu-Ulm) and dioxane in argonsamples and pressurized extraction in ultra grade 65% HNO3 atmosphere at 100 °C for 4 h. The reaction mixture was allowed(170 °C, 10 h) by ICP emission spectroscopy (ICP Jobin 70to cool, diluted with 2 ml water and extracted three times withplus, ISA GmbH, Grasbrunn, Germany).3 ml of CHCl3 containing 20 mg dotriacontane (Fluka) asinternal standard. The combined organic phases were dried

Soil physical properties with Na2SO4.Mechanical impedance was determined by a penetrometer (Ele,Great Britain). For determination of the bulk density (mass of Transesterification: The degradation of cell wall material for

detecting suberin was carried out according to the transes-dried soil per unit volume) the soil substrates were dried at105 °C, placed in a calibrated glass cylinder and weighed. terification method (Kolattukudy and Agrawal, 1974). 0.5–1 mg

Corn roots and environmental stress 597

Table 2. Heavy metal content of garden mould and slagof isolated cell wall material was added to 1 ml of a 10%BF3/methanol solution (Fluka). The reaction mixture was

Element Content (mg kg−1)heated to 70 °C for 24 h. After cooling, the solution wasremoved from the cell wall sample and collected. The cell walls

Garden mould Slagwere washed three times with 1 ml CHCl3 containing 20 mgdotriacontane (Fluka) as internal standard. The chloroformic

Cd 0.2 10and methanolic solutions were combined and washed twice with Cr 19–45 320–5202 ml saturated sodium chloride solution. The organic phase was Cu 13–30 1260–3600separated and dried with Na2SO4. Ni 11–20 190–600

Pb 26–35 800–1600Zn 97–170 3260–5900Gas chromatography and mass spectrometry

Gas chromatographic analysis and mass spectrometric identi-fication of the reaction products obtained by thioacidolysis andtransesterification were performed as described in detail previ- (60–180-fold), cadmium (50-fold), lead (25–50-fold),ously (Zeier and Schreiber, 1997). Separation and quantitative and zinc (25–50-fold)). A closer estimation of the phyto-sample analysis were achieved by a gas chromatograph (HP

toxicity of this substrate is to assay the availability of5890 Series II gas chromatograph, Hewlett-Packard, California,these elements in the soil solution. In Table 3 the chemicalUSA) equipped with a flame ionization detector. Qualitative

sample analysis was carried out on a gas chromatograph (HP composition of soil solutions obtained by suction cups is5890 Series II gas chromatograph, Hewlett-Packard) coupled shown. The solutions exhibited strong alkaline pH valueswith a quadrupole mass selective detector (HP 5971 mass for slag and washed slag while the pH of the soil solutionselective detector, Hewlett-Packard). Before injecting, samples

of garden mould was only slightly alkaline. Even thoughwere derivatized using BSTFA (N,N-bis-trimethylsilyl-the amount of P and Fe was high in slag, these elementstrifluoroacetamide, Machery-Nagel, Duren).were hardly detectable in the soil solution, due to the

Analysis of amino acids insolubility of phosphate and iron compounds at alkalineCell wall isolates were hydrolysed with 1 ml 6 N HCl (130 °C, pH. Furthermore, the electric conductivity of the slag soil24 h, argon atmosphere). After filtration the hydrochloric acid solution indicated a 10 times higher concentration ofwas removed (20 mbar, 40 °C), the residue washed twice with

solutes than in the reference soil solution. The main1 ml distilled water and the aquatic solvent was evaporateddifference between slag and washed slag was the loweragain. Then 200 ml sample buffer (0.1 M lithium citrate,

68.5 mM citric acid, 20 mM bis-(2-hydroxy-ethyl )-sulphide, content of soluble salts (alkali-chloride and -sulphate) inpH 2.2) was added. Amino acids were analysed by HPLC washed slag. Details about the chemical composition of(LC 5001 Biotronic, Eppendorf, Hamburg). the soil solution of slag and other properties of MSW

Results Table 3. Chemical composition of the soil solutions obtained bysuction cups

Characterization of the soil substratesContent in substrate (mg l−1)MSW bottom slag compared to garden mould was charac-

terized by its high Ca, Fe, Na, Cl, S, and B content Garden mould Slag washed Slag(Table 1), its high amount of heavy metals (Table 2) and

pH 7.4–7.9 8.1–8.4 8.4–9.0its low N content (Table 1). When compared to garden Conductivity 0.931–2.48 3.3–6.7 21.4–33.3mould the metals were 25–200-fold enriched (copper (mS cm−1),

20 °CAl (mg l−1) 0.02–0.06 n.d.a 0.30–0.95

Table 1. Major elements of MSW incinerator slag, washed slag B (mg l−1) 0.22–0.47 1.88–2.71 2.8–3.9and garden mould Ca (mg l−1) 81.2–170.7 636–829 881–1142

Fe (mg l−1) 0.043–0.122 n.d.–0.029 n.d.–0.03Element Content (g kg−1) K (mg l−1) 98.9–322 160–302 1304–2117

Mg (mg l−1) 17.3–34.3 17.4–20.6 74.8–96.9Garden mould Washed slag Slag Mn (mg l−1) 0.0025–0.0197 0.048 0.013–0.037

Na (mg l−1) 24.2–77.4 683–1150 3716–6337Cd (mg l−1) n.d.–0.008 n.d.–0.021 n.d.–0.04Si 280–355 138–165 138–165

Ca 15–17 69.8–106 99–126 Cr (mg l−1) n.d. 0.0146–0.0324 0.22–0.47Cu (mg l−1) 0.038–0.111 0.07–0.097 0.36–0.49Fe 12–23 76.9–96.5 69–100

Na 0.2 3.23–4.24 7.7–15 Ni (mg l−1) n.d. n.d. n.d.Pb (mg l−1) n.d. n.d. n.d.Mg 4.2–5.3 6.03–10.4 9.4–13

K 5.8–7.1 2.59–3.73 5.1–10 Zn (mg l−1) 0.073–0.80 0.03–0.099 0.0025–0.043Cl− (mM) 2.64–6.08 9.7–55.5 82–204Cl 0.1 n.m.a 5

P 0.87–1.4 2.2–3.3 2.2–5.1 NH+4 (mM ) 16.5–34.2 13.9–25.3 21.5–130NO−3 (mM) 4.8–10.98 0.4–0.5 n.d.S 0.36–0.75 3.7–5.4 6.3–8.1

N 1.1–5.2 0.02–0.17 0.037–0.2 PO3−4 (mM) 0.3–1.6 n.d.–0.04 n.d.–0.4SO2−4 (mM) 5.4–6.5 19.5–23.2 26.4–37.6B 0.02 0.2 0.2

an.d.: Not detectable.an.m., Not measured.


slag are listed (Fuchs et al., 1997). MSW slag is a rather failed (not shown). Also hydroponic and aeroponic cul-tures of Zea mays did not induce phi thickenings in theinhomogeneous substrate ( large differences in size of soil

particles, 40% of slag particles are larger than 7 mm in rhizodermis (Freundl et al., 2000).To get more information about the importance of cellcomparison to 15% of the control soil ). For a description

of a soil substrate some physical properties should be wall thickenings, the permeability properties were testedby application the apoplastic tracer berberine hemisulph-mentioned as well. Compared to garden mould

(0.07 MPa) the mechanical impedance of slag is increased ate (Enstone and Peterson, 1992). Berberine moves radi-ally into the root and forms crystals with potassiumabout a factor of three (0.2 MPa). Bulk soil density of

slag (1.14 g cm−3) was higher than that of garden mould thiocyanate. As shown in Fig. 1E the phi thickenings(0.92 g cm3). were penetrated, but the inward movement of the dye

was stopped at the exodermis. The cell walls with phiModifications in root architecture and structure thickenings were permeable for the apoplastic dye in

contrast to the exodermal Casparian band. Figure 1FThe effect of culture in slag was first a reduction of rootillustrates the occurrence of the Casparian band in thegrowth. Root length was reduced by 50% by cultivationexodermis and endodermis and reveals that maize is anon slag compared to garden mould (Table 4). However,exodermal species (Enstone and Peterson, 1992).the root5shoot ratio was increased for slag plants by a

factor of 2. Root diameter was also affected. Roots grownin MSW slag were 14% thicker. Microscopic examination Chemical composition of cell wall isolatesrevealed that the increase of the root diameter was due

Lignin: Quantitative chromatographic results revealedto an increased thickness of the cortex (Table 4).that isolated cell walls of the endodermal layer of slag-In roots of slag-grown plants intensive U-shaped thick-cultivated plants contained higher amounts of lignin thanenings of the inner tangential wall of the endodermiscell walls of control roots (Table 5). By contrast, roots(Fig. 1A) were observed. In the tertiary state of develop-grown in prewashed slag did not contain higher concen-ment endodermal cells deposit lignified cell wall materialtrations of lignin. Furthermore, variations in the ligninonto the suberin lamella. These deposits were pronouncedcomponents were studied. The biopolymer lignin is com-more intensive in slag-cultivated roots (0.5±0.08 mm)posed of the cinnamic acid derivates p-coumaryl, coniferylthan in control roots (0.32±0.04 mm) (Fig. 1B). Anotherand sinapyl alcohols, which differ in the extent of methox-phenomenon detected during the slag-cultivation of cornylation. According to the aromatic nucleus in the polymerwere cell wall modifications like phi thickenings in thethe terms guaiacyl (G), syringyl (S) and p-hydroxyphenylradial cell walls of the rhizodermis (Fig. 1C, D). These(H) are used. The monomeric composition of ligninphi thickenings were present only on slag culture. Theshowed a difference both between cell walls of the endo-development of phi thickenings during these experimentaldermis and hypodermis and between the cell wall materialconditions was variable, both longitudinal and tangential.at different culture conditions. Although there was noThickenings were not formed continuously from the tipeffect of the growth substrate on the total lignin contentto the base, but occurred only in the upper third, theof hypodermal cell walls (Table 5) there were variationsoldest part of the root. In addition, if present, phiin the proportions of the monomeric units of lignin. Thethickenings did not occur continuously in the rhizodermalproportion of the H-monomer in hypodermal cell wallsperiphery of the root. Rather intermissions in thickeningwas much higher than in endodermal tissue (Fig. 2A, B).formation were observed. Between 6 and 50 adjacentFurthermore, growth on slag induced a higher proportionrhizodermal cells formed clusters equipped with phi thick-of H-units in hypodermal cell walls than in the controlenings. Attempts to induce phi thickenings by treatingones. In addition, hydroponically grown maize showedroots with 100–200 mM NaCl in hydroponic culture orproportions of H-units even lower than garden mouldwith sand as carrier material or by growing them in glass(Fig. 2A). In endodermal cell walls the proportion ofballotini of varying size to simulate mechanical impedanceG-units were increased and the proportion of the S-unitswere decreased by growth on slag. G-units were the

Table 4. Growth parameters of shoot and root of corn grown on abundant monomer in the endodermis. In the hypodermisslag or garden mould; values represent means±SD, n=10 the proportion of the three monomers is more

homogeneous.Garden mould Slag

Seminal root length (cm) 63.2±5.8 35.2±4.2 Amino acids: Hypodermal cell walls in roots of slag-Root fresh weight (g) 2.60±0.67 1.6±0.54

treated maize plants exhibited a nearly 3-fold higherShoot fresh weight (g) 6.33±0.63 1.8±0.53Root5shoot ratio 0.41 0.92 amount of p-hydroxyproline than the roots of the controlRoot diameter (mm) 1.16±0.11 1.35±0.16 plants (Table 6). The amount of the amino acids threon-Diameter of the central cylinder (mm) 0.55±0.02 0.55±0.03

ine, proline and histidine were also increased in roots of


Fig. 1. Fluorescent microscopic investigation of root cross-sections of primary roots of 20-d-old maize plants. (A–D, G) Autofluorescence (FT 395)of unstained sections, made by the aid of a microtome. (A) Cross-section of the root of a slag plant showing massive tertiary wall thickenings ofthe endodermis. (B) Cross-section of a similar aged root of a control plant. The U-shaped thickenings of the endodermis are significantly smallercompared to the slag-treated root ones. (C) Cross-section of the root of a slag plant showing wall modifications in the radial cell walls of therhizodermis (100×). (D) Cross-section as in (C), 1000×. (E) Freehand cross-section through the root of corn grown on slag, treated with berberinehemisulphate and potassium thiocyanate (FT 460). Arrowheads indicate that the dye penetrated the wall thickenings, whereas the inward movementwas stopped at the Casparian band of the hypodermis (see F). (F) Cross-section of a control root treated with berberine hemisulphate,counterstained with toluidine blue and viewed with violet illumination. The Casparian bands of the endodermis and hypodermis were obvious whilethe fluorescence of the lamellar suberin and lignin is quenched. (G) Cross-section of a control root showing the lack of phi thickenings.


Table 5. Amount of lignin of hypodermal (HCW) and endodermal(ECW) cell walls obtained by thioacidolysis

Values in mg lignin mg−1 dry weight. Mean values and SD from threeindependent experiments.

Lignin content (mg mg−1)

Garden mould Slag washed Slag

HCW 26.8±9.0 24.5±0.37 24.4±2.2ECW 20.0±5.6 21.85±5 27.7±5.3

Fig. 3. Sum of aliphatic suberin monomers obtained after transes-terification of hypodermal (HCW ) and endodermal (ECW ) cell wallisolates. Values are means of three replicates, bars indicate SD.

slag-cultivated maize plants. Finally, the sum of alldetected amino acids in hypodermal cell wall material ofslag-cultivated roots was higher than in control roots.

Hydrophobic compounds: There was a significant higheramount of suberin in the cell wall of the hypodermis thanin that of the endodermis (Fig. 3). However, there wasno significant increase in the total amount of hypodermalsuberin when the plants were grown on slag. It was also

Fig. 2. Monomeric units of hypodermal (A) and endodermal (B) cellwalls obtained after thioacidolysis; H ( p-hydroxyphenyl ), G (guaiacyl ),S (syringyl ). Mean values of three independent experiments, barsindicate SD. (Data of hydroponics were taken from Zeier, 1998).

Table 6. Amino acid composition of acid hydrolysates fromhypodermal cell walls in mg amino acid per mg dry weight; valuesrepresent means±SD, n=3

Amino acid Content (mg mg−1 DW )

Garden mould Slag

0.15±0.04 0.47±0.054HydroxyprolineProline 0.88±0.15 1.30±0.14Histidine 0.26±0.035 0.36±0.03Threonine 0.89±0.10 0.92±0.075Sum (of all 15.5±1.9 18.63±1.51

Fig. 4. Composition of aliphatic suberin components (sum=100%) ofdetectedhypodermal (A) and endodermal (B) cell walls obtained afteramino acids)transesterification. Values are means of three replicates, bars indicate SD.


tested, whether growth of corn on slag would alter the length and the increase in root diameter are typicalcomposition of the suberin fraction. Figure 4 demon- morphological changes for mechanically impeded rootsstrates that this was not the case. The proportion of (Bennie, 1996). Morphological and anatomical changesv-hydroxyacids (C16–C30), dicarboxylic acids (C16–C26), of roots of young maize were also observed when thecarboxcylic acids (C16–C26), alcohols (C16–C24), and seedlings had to penetrate soil of high mechanical imped-2-hydroxyacids (C16–C26) were roughly similar in hypod- ance (Hartung et al., 1994). Salinity stress also causes aermis and endodermis and within a given cell wall type thickening of the root. The increased root to shoot ratiobetween the various treatments. corresponds to the fact, that plants invest more energy in

root than in shoot growth in order to increase absorptionof ions when they suffer nutrient deficiency, here N and P.Discussion

Even though Zea mays is an intensively studied mat-Soil substrates erial, nothing else has been reported in the literature

about the presence of cell wall modifications like phiWhen MSW slag is used as soil substrate, plants experi-thickenings in this plant to date. However, phi thickeningsence various stresses: salinity, nutrient deficiency, heavyhave already been described for several other species (e.g.metal toxicity, alkaline pH, and compaction. Highpelargonium, apple, Pyrus) as ordinary structures in cellamounts of soluble salts, mainly alkali chloride andwalls of root tissue. They have been described for varies-sulphate salts are expected to cause considerable saltroot tissues, but never for the rhizodermis. At least, nostress to plants (up to−0.5 MPa) while the low N contentspecial conditions were reported to be necessary to induceinduces nitrogen deficiency. The low nitrogen content ofthem. However, a phi layer in Ceratonia siliqua L. hasslag is due to its origin by burning of waste at 800–been described, which was observed when plants were1000 °C. The main difference between crude slag andgrown in soil but not in perlite (Praktikakis et al., 1998).prewashed slag is the amount of salt. When slag is washedIn the case of Zea mays, phi thickenings were onlywith a 3-fold volume of water, most of the salt burden isdetected in slag culture. Therefore the properties of theremoved because of the high solubility of alkali salts,soil substrate were an important parameter for the pres-mainly chlorides and sulphates.ence or absence of such cell wall thickenings. The vari-The high pH of the soil solution of slag, which causesation in the frequency of phi thickenings may be attributedalkaline stress, produces additional nutrition deficiency.to the inhomogeneity of the soil substrate. MSW slag isAlthough the elements P and Fe were present in highera heterogeneous substrate. Before use, rough particlesconcentration in slag than in garden mould, at alkaline(particle size>2 cm) were removed, but the bulk fractionpH they are scarcely available to the plant. Another stressstill contains fragments of glass or metals and the particlearises from the heavy metal content. Even though thesize varies considerably. Thus, the bulk soil density couldamount of heavy metals in slag is very high, the availabil-

ity for plants is reduced due to the alkaline pH and the differ locally. Therefore, roots could be influenced onlychemical binding form of the metals (sequential extraction by stages in their extension. The exact reason for theprocedure according to Tessier et al., 1979; not shown). presence of phi thickenings in maize grown on slag is notThus, the contribution of heavy metals to the total stress clear, but a relationship between the supporting role ofon slag-grown roots may be low. the wall thickenings and the mechanical impedance of the

Furthermore, plant growth was influenced by the com- soil substrate is very likely. Furthermore, the fact thatpaction of the soil. Slag exhibits a stronger mechanical the thickenings do not function as a barrier for theimpedance for the root system to penetrate into the soil apoplastic movement of solutes and that they are notthan the control substrate. It was reported that the present around the whole root circumference suggest thatextension rate of the root was reduced when an external they have no relevance as a barrier for solutes in thepressure of about 100 kPa was applied (Goss and Russell, rhizodermis. This is a further argument for the mechanical1980). Other authors published values about 1.5 MPa as supporting role, as suggested by the literaturemoderate impedance. However, a lower impedance mayalso impede root growth when soil aeration is low

Chemical composition of cell wall isolates(Vepraskas, 1994). Typical bulk soil density of heavilycompacted soils is about 1.5 g cm−3 (Iijima and Kono, The chemical composition of hypodermal and endo-1992). Comparing with these data from literature the dermal cell walls were compared with respect to themechanical impedance of MSW slag could be valued as amount of lignin, amino acids of cell wall proteins andrather moderate and not severe. hydrophobic compounds, because knowledge of the

chemical composition of cell wall material can be usefullyRoot structure used to interpret the function of structural changes.

Referring to the hypothesis of mechanical adaptation ofBy cultivation of corn in slag, various architectural andstructural changes were observed. The decrease in root roots grown in slag, the higher amount of lignin of the


endodermis is a further argument. The incrustation of endodermis emphasizes the role of the hypodermis as aphysiological barrier for hydrophilic compounds.the cell walls with lignin is reported to provide mechanical

Finally the results of investigations so far revealed thatstability for root architecture. The tight binding of ligninalterations induced by cultivation in MSW slag were ofto the cellulose-fibrils gives high static properties to thea structural nature. The roots responded to cultivation incell walls (Richter, 1996).slag with mechanical strengthening. In future, efforts willDiscussing the differences in lignin composition,be exercised to find further physiological adaptations.H-enriched lignin is proposed to be stress lignin (MontiesSome work dealing with peroxidase activity which isand Chalet, 1992) or lignin associated with compressioninvolved in modifying cell wall components and detoxi-wood as in gymnosperms (Campbell and Sederoff, 1996).fication of highly reactive molecules is in progress.A higher proportion of H-units produces a more con-

densed lignin because of more intermonomeric C–C-linkages. This functional adaptation of the hypodermis

Acknowledgementscould be related to the fact that the hypodermis is theexternal sealing tissue of the root, which is in direct This investigation was supported by a grant from thecontact with the surrounding rhizosphere and its micro- Graduiertenkolleg ‘Pflanze im Spannungsfeld zwischen

Nahrstoffangebot, Klimastreb und Schadstoffbelastung’ (Borganisms. This fact supports the hypothesis of the contri-Degenhardt) and the SFB 251 (TP A2, H Gimmler). We arebution of the hydroxyphenyl component to the hardeningindebted to Lukas Schreiber for offering the opportunity forof the cell walls.fluorescence microscopic investigations and to Jurgen Zeier for

Cell walls may contain both structural proteins and supporting gas chromatographic and mass spectrometricenzymes. The amino acid composition of these peptides analysis.can be analysed after acid hydrolysis. After the pretreat-ment of the cell wall material as carried out in thisinvestigation, proteins with a structural function are Referencesassumed to remain in the cell wall because soluble or

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cell wall adaptations to multiple environmental stresses ...cell wall adaptations to multiple...

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