The Effect of Light Intensity and Relative Humidity on ...

Journal ofExperimental Botany, Vol. 31, No. 125, pp. 1621-1630, December 1980

The Effect of Light Intensity and Relative Humidity on Growth Rate and Root Respiration ofPlantago lanceolata and Zea mays

Department of Plant Physiology, University ofGroningen, P.O. Box 14, 9750 AA Haren (Gn.), The Netherlands Received 19 July 1979

ABSTRACT Plants of Plantago lanceolata L. and Zea mays L., cv. 'Campo' were grown at two levels of light intensity. Especially in the roots, the rate of dry matter accumulation decreased at low light intensity. The carbohydrate content of both roots and shoots of P. lanceolata was not affected by light intensity. The relative contribution of SHAM'-sensitive respiration, the alternative chain, to total root respiration of both P. lanceolata and Z. mays, was not affected by light intensity during the daytime. The alternative pathway was somewhat decreased at the end of the dark period, but not in the root tips (0-5 mm) where it still contributed 56% in respiration. It was, therefore, concluded that photosynthesis is not a major factor in regulation of root growth in the species investigated. To see whether the effect of light intensity on root growth rate was via transpiration, plants of Z. mays were grown at different air humidities. Both high humidity and low light intensity affected the root morphology in such a way that the distance between the apex and the first laterals on the primary root axis increased. It is suggested that this effect on root morphology is due to transpiration and the subsequent removal of root-produced inhibitors of lateral root growth; although light intensity also affected the rate of dry matter accumulation of roots and the rate was not affected by the humidity of the air. It is, therefore, concluded that the effect of light intensity on the rate of dry matter accumulation of roots of Z. mays is not via an effect on transpiration.

INTRODUCTION

High light intensities, below an optimum, stimulate the rate of dry matter production on higher plants. Light intensity affects the growth rate of roots generally more than that of shoots (Brouwer, 1963). It has been suggested that this growth-stimulating effect of light intensity is due to its effect on photosynthesis. Growth of the roots, being organs further removed from the 'source' than the shoots, would be stimulated due to a better supply of carbohydrates under conditions of high light intensity as compared with conditions of low light intensity (Brouwer, 1963). The following reports are all in accordance with the concept that root growth is carbohydrate-limited. Some metabolic activities of the root, such as CO 2 1

SHAM = salicylhydroxamic acid.

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HANS LAMBERS AND FREEK POSTHUMUS

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Lambers and Posthumus—Light Intensity and Root Growth


production and nitrate uptake (Koster, 1973), maintenance of the metabolically derived transmembrane potential (Hatrick and Bowling, 1973), and increase in root length (Bunce, 1978; Richardson, 1953) decrease upon transfer of plants to darkness or to low light conditions or upon ringing the stem phloem. Other reports suggest that plant growth is only carbohydrate-limited under specific environmental conditions. Hofstra and Hesketh (1975) observed that CO 2 enrichment of the atmosphere stimulated the growth rate of soybean at near optimum growth temperatures. Ito (1973) and Madsen (1976) studied the effect of CO 2 enrichment on growth and a number of physiological processes in tomato plants. They found an increase in carbohydrate content, growth, and fruit yield, in combination with a change in morphological characteristics. Carbon dioxide enrichment of the atmosphere did not increase growth rate in either Lemna (Miiller, Feller, and Ehriszmann, 1977) or Glycine max at suboptimal temperatures (Hofstra and Hesketh, 1975), although it did increase their carbohydrate content. There are also reports that suggest that root metabolism is not carbohydratelimited. Veen (1977) found the same rates of potassium uptake and respiration in roots of Zea mays during the light and dark period and Doddema (1978) observed the same rate of nitrate uptake in roots of Arabidopsis thaliana in light and darkness. Root respiration of several species investigated by us is also not affected by excision of the shoot: it is constant during several hours (unpublished data). In a previous paper (Lambers, 19796) it was concluded that light regulates the growth rate of several plant species via a 'light factor', produced in the shoot under light conditions and which is distinct from carbohydrates. The reason for the introduction of this idea was that under conditions of light-limited growth the relative contribution of an alternative non-phosphorylative electron transport chain was the same as that of plants grown under conditions of higher light intensities. Evidence was presented (Lambers, 1979a, b, 1980; Lambers and Van de Dijk, 1979) that assimilates are only 'wasted' via the alternative chain when they are not required for growth and storage. In view of the different interpretation on how light intensity regulates root growth, it was decided to seek further information on the existence and the nature of the 'light factor'. Plants ofPlantago lanceolata and Zea mays were grown under conditions of both high and low light intensity and the rates of dry matter accumulation and of root respiration and the contribution of the alternative oxidative pathway to root respiration were measured. The content of ethanol-soluble and -insoluble carbohydrates was determined in roots and shoots of Plantago lanceolata. It was hypothesized that the 'light factor' regulated root growth rate. Several processes in a plant, apart from photosynthesis, may depend on light intensity, e.g. transpiration. Leaf area and light intensity are equally important in regulation of adventitious root growth rate in leaf cuttings of Phaseolus vulgaris (Oppenoorth and Lambers, unpublished data). Since the adventitious roots of P. vulgaris contained a large quantity of carbohydrates, it was concluded that photosynthesis was not a major factor regulating adventitious root growth. Transpiration might affect root growth by removal of inhibitors that are produced in the roots and further transported by the transpiration stream. Since some root-produced growth

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inhibitors (e.g. cytokinins and abscisic acid) are known to affect root morphology (Bottger, 1974, 1978; Torrey, 1976), root length and the distribution of dry matter over the root system (primary root, secondary roots = side branches) were also measured. As root morphology of Zea mays can be investigated more readily than that of Plantago lanceolata, the former species was used in experiments in which the interaction of root morphology with the shoot environment was studied. MATERIALS AND METHODS

Respiration measurements These were carried out polarographically as described by Lambers and Steingrover (1978a). Excised but intact roots of sets of four plants {Zea) or 12 plants (Plantago) were used. After measurement of the rate of respiration in the absence of inhibitors, the culture solution in the vessel in which the measurement was carried out was replaced by a culture solution in which 25 mM SHAM (pH 6), an inhibitor of the alternative respiratory pathway, was present. The culture solution did not contain iron. Except for the absence of iron, the culture solution during respiration was the same as during growth of the plants. Carbohydrate analyses Carbohydrates were extracted from dried and pulverized material as described by Lambers, Steingrover, and Smakman (1978).

RESULTS

Experiments with Plantago lanceolata Figure 1 shows that the growth rate of both shoots and roots of Plantago lanceolata decreased upon transfer of the plants to conditions of low light intensity. Root growth was more decreased by low light intensity than shoot growth, yielding an increased shoot/root ratio under conditions of low light intensity (Fig. 2), in accordance with results of Brouwer (1967). The decreases in relative growth rate with time correlated with the commencement of mutual shading (cf. Lambers, 19796). Figure 3 shows that the content of soluble sugars in roots and shoots of plants grown at low light intensity was not significantly lower than that of plants grown under conditions of high light intensity. Starch content of the roots was also the same under both conditions; high light intensity: 55 ± 10 mg starch g"1 dry wt. {n = 8), low light intensity: 60 ± 8 mg starch g"1 dry wt. (n = 10). These results


Growth conditions Plantago lanceolata L. was germinated as described by Lambers and Steingrover (19786). Seedlings of c. 2 weeks old were transferred into a culture solution of 1 strength of that described by Hoagland and Snijder (1933). The plants were grown in a growth chamber at constant temperature (18 °C). Relative humidity was c. 65%. Light intensity was 60 W m"2 (high light intensity) or 10 W m~2 (low light intensity) (white fluorescent lamps, Sylvania and incandescent lamps, Philips 50 W in a ratio of 4:5). Day length was 12 h. Seeds of Zea mays L. cv. Campo were soaked in tap water for c. 6 h and then germinated in the dark on moistened filter paper, at 25 °C. After 4 d the seedlings were transferred into a culture solution of ^ strength of that described by Hoagland and Snijder (1933). The plants were grown in a growth chamber at constant temperature (25 °C). Light intensity was 63 W m~2 (high light intensity) or 20 W m~2 (low light intensity) supplied by Philips HPL lamps provided with a water cuvette. Day length was 12 h. Relative humidity was as indicated in the legends to the figures.

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Lambers and Posthumus—Light Intensity and Root Growth

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suggest that root growth of plants at low light intensity was not limited by carbohydrate supply. Figure 4 shows the rate of root respiration of P. lanceolata, grown at high or low light intensity. Respiration decreased with age of the plants. The decrease was slower at low light intensity, which was possibly related to differences in root

40

50 60 Age (days) FIG. 2. Time course of the shoot/root ratio in P. lanceolata, grown at high (O) and at low (#) light intensity. For further information, see the legend to Fig. 1.


50 Age (days) Fio. 1. Accumulation of dry matter in roots (A and • ) and shoots (O and • ) of P. lanceolata, grown at a high light intensity (O and A) and at a low light intensity ( • and A). Days were counted from the day of germination. Each symbol is the mean of at least two determinations of 12 plants. Time of transfer to low light intensity is indicated by the arrow (s.d. was c. 10%). 30


50

60

Age (days) Fio. 3. Time course of the content of soluble carbohydrates in shoots (A) and roots (B) of P. lanceolata, grown at a high (O) and at a low ( # ) light intensity. Each symbol represents one determination with 12 plants. For further information, see the legend to Fig. 1.

weight (Fig. 1). The degree of inhibition by SHAM was the same in roots of plants grown at both light intensities; root respiration was inhibited by 53 ± 13% (« = 19) at high light intensity and 51 ± 13% (n — 18) at low light intensity. Therefore, these results also indicate that carbohydrate supply did not limit the rate of dry matter production of the roots of plants grown at low light intensity. Experiments with Zea mays Figure 5 A shows the effect of light intensity during growth on the rate of dry matter production of roots of Z. mays. Low light intensity only slightly reduced the elongation rate of the primary root axis, but the morphology of the root system was

40

50

60

Age (days) Fio. 4. Time course of the rate of root respiration of P. lanceolata, grown at a high (O) and at a low ( # ) light intensity. Each symbol is the mean of two or three independent determinations of 12 plants each. For further information, see the legend to Fig. 1.


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Lambers and Posthumus-Light Intensity and Root Growth :

A

•

/

on

30

v-

OJ

3

I

Fio. 5. Growth of Z. mays at two different light intensities, A. Accumulation of dry matter of the roots, B. Increase in length of the apical part ( O and 9 ) and length of the basal part (D and • ) of the roots, c. Increase in weight of the apical part (O and • ) and weight of the basal part (D and • ) . (Closed symbols: low light intensity; open symbols: high light intensity). The symbols in A and c are the mean of two or three determinations of four plants each. The symbols in B represent the mean of six to eight independent determinations of one root system each. The relative humidity ofthe air was c. 80%. Days were counted from the day of germination.

strongly affected (Fig. 5B). Accumulation of dry matter in the laterals and in the primary root axis decreased at low light intensity (Fig. 5 c). Since about 80% of the root weight was accounted for by the laterals, this indicates that growth of the secondary roots was inhibited under low light conditions.

10

15

10 15 Age (days)

10

15

FIG. 6. Growth of Z. mays at relative humidity of 95% (A and A), 65% (D and • ) , and 50% (O and • ) . A. Accumulation of dry matter of the roots, B. Increase in length of the apicaJ part (closed symbols) and length of the basal part (open symbols) of the roots, c. Increase in weight of the apical part (closed symbols) and weight of the basal part (open symbols) of the roots. For further explanation, see the legend to Fig. 5.


10 15 Age (days)


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1. Growth and root respiration ofZ. mays grown at high light intensity and at different humidities TABLE

RGR = relative growth rate; AC = the activity of the alternative pathway as a percentage of total root respiration. RGR (mg g"1 d"1) AC (%)

Relative humidity (%)

Shoot

Root

50 65 80 95

180 190 190 190

150 110 140 140

56±ll(n=13) 56 ± 1 4 ( n = 16) 53 ± 8 ( n = 1 7 ) 51 ± 8 (n= 12)

Figure 6B shows that a high air humidity increased the distance between the root apex and the site where the first laterals appeared ('apical part')- Roots were longer when the plants were grown in high humidity than in low humidity. However, if roots of the same weight were compared, total root length did not significantly differ. Figure 6 c shows the distribution of root dry matter over the apical part and the basal part (which includes the laterals). A comparison of roots of the same weight shows that the weight of the basal part (including the laterals) was not significantly affected by humidity, although the length of this zone was twice as long at 50 and 65% r.h. This was at variance with the results of plants grown at different light intensities, where the length of the basal part and its weight were both reduced by low light intensity (Fig. 5). The relative contribution of the alternative pathway to total root respiration in the daytime was the same in all treatments (Table 1), suggesting that carbohydrates did not limit dry matter production of Z. mays under any of the experimental conditions. In the last 2 h of the dark period the activity of the alternative chain decreased to 21 + 13% (« = 8). However, the alternative pathway contributed 56 ± 15% (n = 7) in respiration of the root tips (0-5 mm), even at the end of the dark period. This is in agreement with our unpublished results showing that the alternative chain is more active in root tips than in the basal parts of the roots. A combination of cyanide (0-2 mM) and SHAM (25 mM) invariably gave an inhibition of respiration of c. 85%.


Figure 6A, B, and c shows the effect of the humidity of the air on growth and morphology of roots of Z. mays. Shoot weight of the plants was significantly lower at 95% r.h., shoot weight of the plants at 50, 65 and 80% r.h. was the same. Low humidity decreased the final yield of dry matter of the roots (Fig. 6 A). However, the relative growth rate of both shoots and roots of Z. mays was not significantly different at the different humidities (Table 1), indicating that differences in final weight were due to differences at the beginning of the experimental period. These initial differences in weight are likely to be caused by adaptation to the low humidity of the air, as demonstrated by the number of stomata per cm2 leaf area, which was strongly reduced at 65 and 50% r.h., compared with 95% r.h.

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DISCUSSION SHAM and other substituted hydroxamic acids have been used as specific inhibitors of the alternative respiratory path (Schonbaum, Bonner, Storey, and Bahr, 1971) in studies on mitochondrial respiration (e.g. Bahr and Bonner, 1973; Van der Plas, Schoenmaker, and Gerbrandy, 1977). More recently these inhibitors have also been used in in vivo studies on root respiration (e.g. Van der Plas et al., 1977; Lambers and Smakman, 1978). Although the effects of substituted hydroxamic acids on respiration of intact tissue should be interpreted with care (Parrish and Leopold, 1978) it appears that sufficient information is now available to use these inhibitors in a study on the contribution of the alternative pathway in respiration of roots (for a more extensive discussion of this matter: see Lambers, 1980). SHAM at 25 mM gave less than 10% inhibition of respiration of shoots of Senecio (Lambers, Noord, and Posthumus, 1979) and of nodulated roots of Lupinus albus (Lambers, Layzell, and Pate, 1980). Since a combination of 25 mM SHAM with cyanide (0-2 mM) gave at least 80% inhibition of respiration of the above tissues, this low inhibition was not due to lack of penetration and indicates that the side effects of this inhibitor are irrelevant in short term studies on respiration. In potato slices, inhibition of the alternative pathway did not stimulate the cytochrome pathway in vivo (Theologis and Laties, 1978). Since a combination of cyanide (0-1 or 0-2 mM) and SHAM (25 mM) inhibits respiration of various tissues (e.g. Senecio shoots: Lambers et al, 1979; Hypochaeris roots: Lambers and Van de Dijk, 1979; Lupinus roots, both N2-fixing and NO3-fed: Lambers et al, 19806) by 80-90%, the present SHAM concentration is sufficiently high for investigation of the alternative pathway in vivo. Accumulation of dry matter in roots of both P. lanceolata and Z. mays decreased at low light intensity. The data on carbohydrate content do not support the hypothesis that carbohydrate supply limits root growth at low light intensity. Since the alternative pathway in roots of higher plants appears to be operative only where more carbohydrates are translocated to them than can be used for such processes as structural growth, storage, energy production, and osmoregulation (Lambers, Blacquiere, and Stuiver, 1980a; Lambers; 1980 and references cited therein), the present data on the alternative pathway indicate that carbohydrate supply does not limit root growth at low light intensity. The possibility that growth of meristematic zones of the roots was carbohydrate-limited can be excluded since the concentrations of carbohydrates in these zones are high (Wanner, 1952; Rogozinska, Bryan, and Whaley, 1965) and since the alternative pathway is more active in root tips than in the basal parts of the roots (maize roots: Lambers and Hoogenboom, unpublished; pea roots: Van Mil, to be published), even at the end of the dark period in roots of plants grown at low light intensity. Thus, the present results suggest the existence of a 'light factor', distinct from carbohydrate supply to the roots (Lambers, 1979Z?). This 'light factor' enabled the plants to adjust root growth rate to match the supply of assimilates available and prevented growth becoming directly carbohydrate-limited. When plants with the same root weight were compared with respect to their morphology those grown at 65 and 59% r.h. had primary axes of approximately

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ACKNOWLEDGEMENTS

This research was supported in part by the Foundation for Fundamental Biological Research (BION) which is subsidized by the Netherlands Organization for the Advancement of Pure Research (Z.W.O.). Grassland species research group Publ. No. 17. The authors thank Prof. Dr. R. Brouwer, Dr. J. J. Hofstra, and Prof. Dr. Ir. P. J. C. Kuiper for the critical reading of the manuscript. LITERATURE CITED T, and BONNER, W. D. JR., 1973. /. biol. Chem. 248,3441-5.

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1978. Z. PflPhysiol. 86,283-6. 1963./. Inst. Bodemsch. 213,31-9. 1967. Angew. Bot. 41,244-54. BUNCE, J. A., 1978./. exp. Bot. 29,595-601. DODDEMA, H., 1978. Thesis, University of Groningen. HATRICK, A. A. and BOWLING, D. J. F., 1973./. exp. Bot. 24,607-13. HOAGLAND, D. R., and SNUDER, W. C , 1933. Proc. Am. Soc. hort. Scl. 30,288-96. BROUWER, R.,

HOFSTRA, G., and HESKETH, J. D., 1975. In Environment and biological control of photosynthesis.

Ed. R. MarceUe. Dr. W. Junk B.V., The Hague. Pp. 71-80. ITO, T., 1973. The Transactions of the Faculty of Horticulture, Chiba University, Volume 7. KOSTER, A. L., 1973. Thesis, University of Leiden. LAMBERS, H., 1979a. Thesis, University of Groningen. ISBN 90 9000082 8. 19796. Physiologia PI. 46,194-202. 1980. Plant, CellEnvir. (in press).


the same length as those grown at 95% r.h. However, the distance between the apex and the site where the first laterals appeared was twice as long in plants grown at 95% r.h. than in plants grown at 65 and 50% r.h. Root axes of plants grown at different light intensities also had the same length. The root morphology induced by high humidity was also found at low light intensity. It was, therefore, concluded that the root morphology might be affected by the rate of the removal of root-produced inhibitors of lateral root growth by the transpiration stream. However, despite the above described similarities in root morphology of plants grown at high humidity and at low light intensity, there was a significant difference in the rate of dry matter accumulation in the laterals in the two treatments. Moreover, the rate of dry matter accumulation (relative growth rate) of roots of Z. mays at the four air humidities was the same, although their morphology was different. It is concluded that a decreased rate of transpiration affected root morphology only, without affecting the rate of dry matter accumulation of roots. Thus, the present results suggest that light does not affect the rate of dry matter accumulation of roots of Z. mays via its effect on transpiration and the subsequent removal of root-produced inhibitors of lateral growth. The results might be explained by the transport of a growth stimulator that is affected by light intensity. A high light intensity might increase the transport of the 'light factor' either by affecting its rate of production or by affecting its rate of transport (cotransport with assimilates?). Although no conclusive statement can be made on the nature of the 'light factor', results of MacDowall (1972) on the effect of light and CO 2 on the rate of dry matter production in wheat plants favour the hypothesis of a distinct light factor and not the hypothesis of cotransport with assimilates.

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- BLACQUIERE, T., and STUTVER, C. E. E., 1980a. Abstr., FESPP Meeting, Santiago de Compostela, Spain. • LAYZELL, D. B., and PATE, J. S., 19806. Physiologia PI. (in press). • NOORD, R., and POSTHUMUS, F., 1979. Ibid. 45,351-6.

• and SMAKMAN, G., 1978. Ibid. 42, 163-6. - and STEINGROVER, E., 1978a. Ibid. 42,179-84. 19786. Ibid. 43,219-24. • and SMAKMAN, G., 1978. Ibid. 43,219-24. • and VAN DE DUK, S. J., 1979. Ibid. 45,235-9. MACDOWALL, F. D. H. 1972. Can. J. Bot. 50, 883-9.

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ROGOZINSKA, J. H., BRYAN, P. A., and WHALEY, W. G., 1965. Phytochemistry,

4,919-24.

SCHONBAUM, G. R., BONNER, W. D. JR., STOREY, B., and BAHR, J. T., 1971. PI. Physiol,

Lancaster,

47,124-8. THEOLOGIS, A., and LATIES, G. G., 1978. Ibid. 62,232-7.

TORREY, J. G., .1976. A. Rev. PI. Physiol. 27,435-60. VAN DER PLAS, L. H. W., SCHOENMAKER, G. S., and GERBRANDY, S. J., 1977. PI. Sci. Lett. 8,

31-3. VEEN, B. W., 1977. / . exp. Bot. 28,1389-98.

WANNER, H., 1952. Ber. schweiz. bot. Ges. 62, 205-17.


RICHARDSON, S. D., 1953. Proc. K. ned. Acad. Wet. C56,185-253. PARJUSH, D. J., and LEOPOLD, A. C , 1978. PL Physiol. Lancaster, 62,470-72.