Planta 9 by Springer-Verlag 1979
Planta 146, 405-408 (1979)
Endogenous and Exogenous Auxin in the Control of Root Growth P.E. Pilet 1, M.C. Elliott 2, and M.M. Moloney 2 1 Institut de Biologie et de PhysiologicV4g4tales, Universit6 de Lausanne, 6 Place de la Riponne, CH-1005 Lausanne, Switzerland, and 2 School of Life Sciences, LeicesterPolytechnic,LeicesterLE1 9BH, U.K.
Abstract. The endogenous indol-3yl-acetic acid (IAA) of detipped apical segments from roots of maize (cv ORLA) was greatly reduced by an exodiffusion technique which depended upon the preferential acropetal transport of the phytohormone into buffered agar. When IAA was applied to the basal cut ends of freshly prepared root segments only growth inhibitions were demonstrable but after the endogenous auxin concentration had been reduced by the exodiffusion technique it became possible to stimulate growth by IAA application. The implications of the interaction between exogenous and endogenous IAA in the control of root segment growth are discussed with special reference to the role of endogenous IAA in the regulation of root growth and geotropism.
Key words: Auxin - Growth - Root growth - Zea.
Introduction
In recent years workers in several laboratories have carried out intensive investigations of the hormonal regulation of maize root growth and geotropism. Work on the endogenous growth inhibitors released from the root cap (Wilkins, 1977; Wain, 1977; Pilet, 1977) has tended to draw attention away from the classical concept which featured auxin as an essential regulator of root elongation. However, it has been clearly shown that IAA may affect the axial extension of roots (Pilet, 1961a; Street etal., 1967). Pilet (1961 b) noted that for very young roots a low concentration of IAA promoted growth, whereas, higher concentrations of IAA were growth inhibitory. Abbreviations: IAA=indol-3yl-acetic acid; GC-MS=gas chroma-
tography mass spectrometry
Growth of older roots was inhibited by all concentrations of IAA tested. By the use of classical techniques of auxin extraction and bioassay it was possible to show that growing roots contained auxin-like compounds (Pilet, 1961a; Street et al., 1967), and that the auxin content increased with age and finally reached a "supra-optimal" level such that application of exogenous auxin would cause an inhibition of growth (Pilet, 1961b). Recently, the use of the GCMS technique made it possible to demonstrate unequivocally that IAA was a natural auxin of maize roots (Elliott and Greenwood, 1974) and that it occurred at very high concentration in the root cap (Rivier and Pilet, 1974). IAA transport in the root is preferentially acropetal (Pilet, 1964, 1975; Scott and Wilkins, 1968) and occurs predominantly in the stele (Greenwood et al., 1973). In maize, the application of [5-3H] IAA to the cut mesocotyl surface or to the caryopses, led to the appearance of radioactivity in the root (Batra etal., 1975, and application to the shoot apices of intact maize seedlings resulted in radioactivity accumulating at the root tips (Martin et al., 1978). Although labeled IAA applied on the cap of the maize root possibly entered the tip, its basipetal movement was very limited (Pernet and Pilet, 1976). It seemed that IAA must be implicated in the regulation of root elongation (Pilet 1961b, 1975; Audus, 1975; Elliott, 1975, 1977). The aim of the experiments reported here was to determine the effects of endogenous and exogenous IAA on root elongation.
Design of the Experiments
Since IAA in roots is transported acropetally, it was possible to rapidly reduce the endogenous IAA concentration of detipped apical root segments by
0032-0935/79/0146/0405/$01.00
406
exodiffusion into buffered agar. The changes in endogenous IAA during the exodiffusion process were monitored by a highly specific and sensitive spectrophotofluorimetric method. Root segments with a range of different endogenous IAA contents were treated with exogenous IAA at the basal cut end and the elongation of the root segments was measured. Thus root growth could be considered in the context of changing endogenous and exogenous IAA concentrations. Figure 1 illustrates schematically the assays which were carried out. F r o m primary roots of maize (cv ORLA), 15.1 mm in length (A), apical segments 10 +0.5 mm long were prepared (B). 0.5 m m tips were removed from the segments (C), some of these detipped segments were used for IAA determination and the remainder were placed vertically (D) for 4 or 8 h with their apical cut surfaces (a) in contact with buffered agar (pa). Some of these segments (E) - the length (10) of which had been measured - were used for determination of their IAA content. The remaining segments (F) were inverted and covered on their morphologically basal cut ends (b) with buffered agar or buffered agar containing IAA at 10 9, 10 7 or 10- s mol din- 3. There after the length (14) of the segments (G) was measured and their relative elongation was calculated.
Materials and Methods The technique of preparation of primary roots has been described previously (Pilet, 1975, 1977). Zea mays L. cv O R L A 264 (Assoc. suisse des S~lectionneurs) was used. Selected caryopses were placed in darkness (22 ~ C) between plastic frames on moist paper towels, the roots elongated vertically. W h e n they reached 15+_3 m m in lenght, 10_+ 0.2 m m apical segments were prepared, detipped, and m o u n t e d in plastic frames (Fig. 2A). For the diffusion experiment, 1.5% Difco purified agar buffered at pH 6.1 with 0.35 mol d m 3 phosphate-citrate buffer was used, a n d all the preparative techniques were done in dim green light obtained by using a white-light source (Luxram incandescent lamp 6 V, 5 W) and a special filter in plexiglas, from R 6 h m and Haas G M B H Darmstadt, G e r m a n y (530 +_20 ran; 1.2 W cm-2). In the first set of assays, segments were kept vertical with the apical cut surface applied to the buffered agar (Fig. 2B). The frame was turned t h r o u g h 180 degrees and the segments, now inverted, had their basal cut ends in contact with buffered agar (control) or buffered agar containing I A A (Fig. 2C). For the IAA assays, 300 root segments (approximately 4 g fresh weight) were rapidly frozen in a beaker placed in a dry ice/acetone bath and then lyophilized (to yield approximately 300 m g o f lyophilized material - each batch was accurately weighed). The iyophitized material was g r o u n d dry with a small quantity o f acid-washed sand and then extracted for a total of 40 min with 6 • 70 cm 3 of cold (4 ~ C) redistilled methanol in a tissue grinder with a teflon plunger. The methanolic extracts were filtered, the filtrates combined and concentrated to a small volume under reduced pressure at 30 ~ C. The concentrated extract was purified by solvent partitioning in the m a n n e r described by Knegt and Bruinsma (1973). The purified extract contained pigments
P.E. Pilet et al. : Auxin and Root Growth which caused considerable quenching of fluorescencs, hence a further purification step was used before the spectrophotofluorimetric assay. The extract was passed through a 20.1.7 cm c o l u m n o f polyvinylpyrrolidone (Polyclar AT, G A F ) a n d eluted with 0.05 mol din- 3 potassium phosphate buffer p H 8. The first 150 cm 3 of eluate were collected, acidified to pH 3, and partitioned three times against equal volumes of diethyl ether, The ethereal solution was concentrated to a small volume under reduced pressure at 30 ~ C and the I A A content was determined using the spectrophotofluorimetric method of Knegt and Bruinsma (1973), modified by the use of methanol to stop the reaction. Losses during extraction were accounted for by isotope dilution. Extraction efficiencies were routinely 75-80%. Fluorescence measurements were performed on a Perkin-Elmer M P F - 4 3 A spectrophotofluorimeter. In all cases a complete fluorescence spectrum was taken around the fluorescence m a x i m u m (480 nm). Errors did not exceed + 8%.
Results and Discussion
In the first set of assays, the endogenous IAA content was determined for freshly prepared detipped root segments and for detipped segments kept for 4 h or 8 h in a vertical position with their apical cut ends in contact with buffered agar (Fig. 1 D and Fig. 2 B). Table 1 shows the change in the endogenous IAA content of the root segments (expressed in terms of ng IAA per segment and ~g IAA per g dry weight) with increasing time (from 0 to 8 h) of exodiffusion from the tip of the segment into the agar. These results confirm those briefly summarized in the Introduction of this paper on the IAA transport characteristics of maize root apices. Thus direct measurements of
E
c A
B
C
D IAA
G
Fig. 1 A - G . Diagram illustrating the principle of experiments. A Intact maize primary root. B Apical segment. C Detipped segment with the apical (a) and basal (b) cut ends marked. D Segment vertically maintained 4 or 8 h with its apical cut surface in contact with buffered agar (pa) : the arrow indicates the direction of preferential acropetaI transport of endogenous auxin. E Segment for which length (lo) was measured and then used for analysis of the endogenous auxin level. F Inverted segment kept vertical with its basal cut surface on buffered agar or buffered agar containing I A A at several concentrations; the arrow shows the direction of preferential acropetal movement of applied IAA. G Segment the length (14) of which was measured. Root segments not to scale
P.E. Pilet et al. : Auxin and Root Growth
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A Fig. 2A-C. Diagram showing the equipment used in treatment of apical maize root segments. A Plastic frames (J) with holes (h) and fixation feet (J)0. B Assay with root segments (rs) kept vertical for 4 or 8 h on the frame in a Petri dish (Pd), with their apical cut surfaces in contact with buffered agar (see Fig. 1D). C Assay with inverted segments maintained vertically for 4 h with their basal cut ends on buffered agar or buffered agar containing IAA (see Fig. 1 F)
20
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Fig. 4. Relation between the growth of apical maize root segments and their auxin level. Growth : relative elongation in % (see Fig. 3). Segments treated with IAA (see Fig. 1 F) at several concentrations (10- 9, 10 7 and 10- 5 mol dm- 3). Auxin level: relative endogenous IAA content in % (see Table 1 : ng per segment). Segments first vertically kept for 0, 4 or 8 h with their apical cut surfaces in contact with buffered (pH) 6.1) agar (see Figs. 1D and 2B)
concentration of IAA applied (10-s mol dm a) caused inhibition; and this was less than for segments which had undergone 4 h of exodiffusion (and had a higher endogenous IAA content). These results show clearly that endogenous and applied IAA cooperate in the regulation of root elongation. Figure 4 shows the way in which the growth of root segments containing different endogenous IAA levels changes in response to different applied IAA concentrations. The experiments described here provide further evidence that endogegenous auxin plays an essential role in the regulation of root elongation (Pilet, 1975; Audus, 1975; Elliott, 1975, 1977), and the results are in accord with our thesis that maize root elongation and georeaction are regulated by the interaction of acropetally moving IAA, derived partly from the shoot system, with growth inhibitors (probably including abscisic acid) transported in a basipetal direc-
References Audus, L.J. : Geotropism in roots. In: The Development and Function of Roots, pp. 32%363. Torrey, J.G., Clarkson, D.T., eds. London: Academic Press 1975 Batra, M.W., Edwards, K.L., Scott, T.K.: Auxin transport in roots: its characteristics and relationship to growth. In: The Development and Function of Roots, pp. 299-325. Torrey, J.G., Clarkson, D.T., eds. London: Academic Press 1975 Elliott, M.C. : Hormone interactions in regulation of root growth and geotropism. Plant Physiol. 56S, 39 (1975) Elliott, M.C. : Auxin and the regulation of root growth. In: Plant Growth Regulation, pp. 10(~108, Pilet, P.E., ed. Berlin, Heidelberg, New York: Springer 1977 Elliott, M.C., Greenwood, M.S.: Indol-3yl-acetic acid in roots of Zea mays. Phytochemistry 13, 239-241 (1974) Greenwood, M.S., Hillman, J.R., Shaw, S., Wilkins, M.B. : Localisation and identification of auxin in roots of Z e a mays. Planta 109, 369-374 (1973) Knegt, E., Bruinsma, J. : A rapid, sensitive and accurate determination of indolyl-3-acetic acid. Phytochemistry 12, 753-756 (1973) Martin, H.V., Elliott, M.C., Wangermann, E., Pilet, P.E.: Auxin gradient along the root of the maize seedlings. Planta 141, 179-181 (1978) Pernet, J.J., Pilet, P.E. : Indoleacetic acid movement in the root cap. Planta 128, 183-184 (1976) Pilet, P.E. : Auxins and the process of aging in root cells. In: Plant Growth Regulation, pp. 167-178, Wightmann, F., Setterfield, G., eds. Ames Iowa: The Iowa State University Press 1961 a Pilet, P.E.: L'action des auxines sur la croissance des cellules. In: Handbuch der Pflanzenphysiologie, XIV, pp. 784-806, Rubland, W., ed. Berlin, Heidelberg, New York: Springer 196Ib Pilet, P.E.: Auxin transport in roots. Nature 204, 561 562 (1964) Pilet, P.E. : Abscisic acid as a root growth inhibitor: physiological analyses. Planta 122, 299 302 (1975) Pilet, P.E. : Growth inhibitors in growing and geostimulated maize roots. In: Plant Growth Regulation, pp. 115-128, Pilet, P.E., ed. Berlin, Heidelberg, New York: Springer 1977 Rivier, L_, Pilet, P.E. : Indolyl-3-acetic acid in cap and apex of maize roots: identification and quantification by mass fragmentography. Planta 120, 107-112 (1974) Scott, T.K., Wilkins, M.B.: Auxin transport in roots. II. Polar flux of IAA in Zea roots. Planta 83, 232-334 (1968) Street, H.E., Bullen, P.M., Elliott, M.C. : The natural growth regulators of roots. In: Wachstums-Regulatoren bei Pflanzen, pp. 407-416, Libbert, E., ed. Rostock: Fischer 1967 Wain, R.L. : Root growth inhibitors. In: Plant Growth Regulation, pp. 109-114, PiIet, P.E., ed. Berlin, Heidelberg, New York: Springer 1977 Wilkins, M.B. : Geotropic response mechanism in roots and shoots. In: Plant Growth Regulation, pp. 199 207, Pilet, P.E., ed. Berlin, Heidelberg, New York: Springer 1977
Received 6 December 1978; accepted 16 May 1979
Planta 146, 405-408 (1979)
Endogenous and Exogenous Auxin in the Control of Root Growth P.E. Pilet 1, M.C. Elliott 2, and M.M. Moloney 2 1 Institut de Biologie et de PhysiologicV4g4tales, Universit6 de Lausanne, 6 Place de la Riponne, CH-1005 Lausanne, Switzerland, and 2 School of Life Sciences, LeicesterPolytechnic,LeicesterLE1 9BH, U.K.
Abstract. The endogenous indol-3yl-acetic acid (IAA) of detipped apical segments from roots of maize (cv ORLA) was greatly reduced by an exodiffusion technique which depended upon the preferential acropetal transport of the phytohormone into buffered agar. When IAA was applied to the basal cut ends of freshly prepared root segments only growth inhibitions were demonstrable but after the endogenous auxin concentration had been reduced by the exodiffusion technique it became possible to stimulate growth by IAA application. The implications of the interaction between exogenous and endogenous IAA in the control of root segment growth are discussed with special reference to the role of endogenous IAA in the regulation of root growth and geotropism.
Key words: Auxin - Growth - Root growth - Zea.
Introduction
In recent years workers in several laboratories have carried out intensive investigations of the hormonal regulation of maize root growth and geotropism. Work on the endogenous growth inhibitors released from the root cap (Wilkins, 1977; Wain, 1977; Pilet, 1977) has tended to draw attention away from the classical concept which featured auxin as an essential regulator of root elongation. However, it has been clearly shown that IAA may affect the axial extension of roots (Pilet, 1961a; Street etal., 1967). Pilet (1961 b) noted that for very young roots a low concentration of IAA promoted growth, whereas, higher concentrations of IAA were growth inhibitory. Abbreviations: IAA=indol-3yl-acetic acid; GC-MS=gas chroma-
tography mass spectrometry
Growth of older roots was inhibited by all concentrations of IAA tested. By the use of classical techniques of auxin extraction and bioassay it was possible to show that growing roots contained auxin-like compounds (Pilet, 1961a; Street et al., 1967), and that the auxin content increased with age and finally reached a "supra-optimal" level such that application of exogenous auxin would cause an inhibition of growth (Pilet, 1961b). Recently, the use of the GCMS technique made it possible to demonstrate unequivocally that IAA was a natural auxin of maize roots (Elliott and Greenwood, 1974) and that it occurred at very high concentration in the root cap (Rivier and Pilet, 1974). IAA transport in the root is preferentially acropetal (Pilet, 1964, 1975; Scott and Wilkins, 1968) and occurs predominantly in the stele (Greenwood et al., 1973). In maize, the application of [5-3H] IAA to the cut mesocotyl surface or to the caryopses, led to the appearance of radioactivity in the root (Batra etal., 1975, and application to the shoot apices of intact maize seedlings resulted in radioactivity accumulating at the root tips (Martin et al., 1978). Although labeled IAA applied on the cap of the maize root possibly entered the tip, its basipetal movement was very limited (Pernet and Pilet, 1976). It seemed that IAA must be implicated in the regulation of root elongation (Pilet 1961b, 1975; Audus, 1975; Elliott, 1975, 1977). The aim of the experiments reported here was to determine the effects of endogenous and exogenous IAA on root elongation.
Design of the Experiments
Since IAA in roots is transported acropetally, it was possible to rapidly reduce the endogenous IAA concentration of detipped apical root segments by
0032-0935/79/0146/0405/$01.00
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exodiffusion into buffered agar. The changes in endogenous IAA during the exodiffusion process were monitored by a highly specific and sensitive spectrophotofluorimetric method. Root segments with a range of different endogenous IAA contents were treated with exogenous IAA at the basal cut end and the elongation of the root segments was measured. Thus root growth could be considered in the context of changing endogenous and exogenous IAA concentrations. Figure 1 illustrates schematically the assays which were carried out. F r o m primary roots of maize (cv ORLA), 15.1 mm in length (A), apical segments 10 +0.5 mm long were prepared (B). 0.5 m m tips were removed from the segments (C), some of these detipped segments were used for IAA determination and the remainder were placed vertically (D) for 4 or 8 h with their apical cut surfaces (a) in contact with buffered agar (pa). Some of these segments (E) - the length (10) of which had been measured - were used for determination of their IAA content. The remaining segments (F) were inverted and covered on their morphologically basal cut ends (b) with buffered agar or buffered agar containing IAA at 10 9, 10 7 or 10- s mol din- 3. There after the length (14) of the segments (G) was measured and their relative elongation was calculated.
Materials and Methods The technique of preparation of primary roots has been described previously (Pilet, 1975, 1977). Zea mays L. cv O R L A 264 (Assoc. suisse des S~lectionneurs) was used. Selected caryopses were placed in darkness (22 ~ C) between plastic frames on moist paper towels, the roots elongated vertically. W h e n they reached 15+_3 m m in lenght, 10_+ 0.2 m m apical segments were prepared, detipped, and m o u n t e d in plastic frames (Fig. 2A). For the diffusion experiment, 1.5% Difco purified agar buffered at pH 6.1 with 0.35 mol d m 3 phosphate-citrate buffer was used, a n d all the preparative techniques were done in dim green light obtained by using a white-light source (Luxram incandescent lamp 6 V, 5 W) and a special filter in plexiglas, from R 6 h m and Haas G M B H Darmstadt, G e r m a n y (530 +_20 ran; 1.2 W cm-2). In the first set of assays, segments were kept vertical with the apical cut surface applied to the buffered agar (Fig. 2B). The frame was turned t h r o u g h 180 degrees and the segments, now inverted, had their basal cut ends in contact with buffered agar (control) or buffered agar containing I A A (Fig. 2C). For the IAA assays, 300 root segments (approximately 4 g fresh weight) were rapidly frozen in a beaker placed in a dry ice/acetone bath and then lyophilized (to yield approximately 300 m g o f lyophilized material - each batch was accurately weighed). The iyophitized material was g r o u n d dry with a small quantity o f acid-washed sand and then extracted for a total of 40 min with 6 • 70 cm 3 of cold (4 ~ C) redistilled methanol in a tissue grinder with a teflon plunger. The methanolic extracts were filtered, the filtrates combined and concentrated to a small volume under reduced pressure at 30 ~ C. The concentrated extract was purified by solvent partitioning in the m a n n e r described by Knegt and Bruinsma (1973). The purified extract contained pigments
P.E. Pilet et al. : Auxin and Root Growth which caused considerable quenching of fluorescencs, hence a further purification step was used before the spectrophotofluorimetric assay. The extract was passed through a 20.1.7 cm c o l u m n o f polyvinylpyrrolidone (Polyclar AT, G A F ) a n d eluted with 0.05 mol din- 3 potassium phosphate buffer p H 8. The first 150 cm 3 of eluate were collected, acidified to pH 3, and partitioned three times against equal volumes of diethyl ether, The ethereal solution was concentrated to a small volume under reduced pressure at 30 ~ C and the I A A content was determined using the spectrophotofluorimetric method of Knegt and Bruinsma (1973), modified by the use of methanol to stop the reaction. Losses during extraction were accounted for by isotope dilution. Extraction efficiencies were routinely 75-80%. Fluorescence measurements were performed on a Perkin-Elmer M P F - 4 3 A spectrophotofluorimeter. In all cases a complete fluorescence spectrum was taken around the fluorescence m a x i m u m (480 nm). Errors did not exceed + 8%.
Results and Discussion
In the first set of assays, the endogenous IAA content was determined for freshly prepared detipped root segments and for detipped segments kept for 4 h or 8 h in a vertical position with their apical cut ends in contact with buffered agar (Fig. 1 D and Fig. 2 B). Table 1 shows the change in the endogenous IAA content of the root segments (expressed in terms of ng IAA per segment and ~g IAA per g dry weight) with increasing time (from 0 to 8 h) of exodiffusion from the tip of the segment into the agar. These results confirm those briefly summarized in the Introduction of this paper on the IAA transport characteristics of maize root apices. Thus direct measurements of
E
c A
B
C
D IAA
G
Fig. 1 A - G . Diagram illustrating the principle of experiments. A Intact maize primary root. B Apical segment. C Detipped segment with the apical (a) and basal (b) cut ends marked. D Segment vertically maintained 4 or 8 h with its apical cut surface in contact with buffered agar (pa) : the arrow indicates the direction of preferential acropetaI transport of endogenous auxin. E Segment for which length (lo) was measured and then used for analysis of the endogenous auxin level. F Inverted segment kept vertical with its basal cut surface on buffered agar or buffered agar containing I A A at several concentrations; the arrow shows the direction of preferential acropetal movement of applied IAA. G Segment the length (14) of which was measured. Root segments not to scale
P.E. Pilet et al. : Auxin and Root Growth
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A Fig. 2A-C. Diagram showing the equipment used in treatment of apical maize root segments. A Plastic frames (J) with holes (h) and fixation feet (J)0. B Assay with root segments (rs) kept vertical for 4 or 8 h on the frame in a Petri dish (Pd), with their apical cut surfaces in contact with buffered agar (see Fig. 1D). C Assay with inverted segments maintained vertically for 4 h with their basal cut ends on buffered agar or buffered agar containing IAA (see Fig. 1 F)
20
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100 ( 4
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Fig. 4. Relation between the growth of apical maize root segments and their auxin level. Growth : relative elongation in % (see Fig. 3). Segments treated with IAA (see Fig. 1 F) at several concentrations (10- 9, 10 7 and 10- 5 mol dm- 3). Auxin level: relative endogenous IAA content in % (see Table 1 : ng per segment). Segments first vertically kept for 0, 4 or 8 h with their apical cut surfaces in contact with buffered (pH) 6.1) agar (see Figs. 1D and 2B)
concentration of IAA applied (10-s mol dm a) caused inhibition; and this was less than for segments which had undergone 4 h of exodiffusion (and had a higher endogenous IAA content). These results show clearly that endogenous and applied IAA cooperate in the regulation of root elongation. Figure 4 shows the way in which the growth of root segments containing different endogenous IAA levels changes in response to different applied IAA concentrations. The experiments described here provide further evidence that endogegenous auxin plays an essential role in the regulation of root elongation (Pilet, 1975; Audus, 1975; Elliott, 1975, 1977), and the results are in accord with our thesis that maize root elongation and georeaction are regulated by the interaction of acropetally moving IAA, derived partly from the shoot system, with growth inhibitors (probably including abscisic acid) transported in a basipetal direc-
References Audus, L.J. : Geotropism in roots. In: The Development and Function of Roots, pp. 32%363. Torrey, J.G., Clarkson, D.T., eds. London: Academic Press 1975 Batra, M.W., Edwards, K.L., Scott, T.K.: Auxin transport in roots: its characteristics and relationship to growth. In: The Development and Function of Roots, pp. 299-325. Torrey, J.G., Clarkson, D.T., eds. London: Academic Press 1975 Elliott, M.C. : Hormone interactions in regulation of root growth and geotropism. Plant Physiol. 56S, 39 (1975) Elliott, M.C. : Auxin and the regulation of root growth. In: Plant Growth Regulation, pp. 10(~108, Pilet, P.E., ed. Berlin, Heidelberg, New York: Springer 1977 Elliott, M.C., Greenwood, M.S.: Indol-3yl-acetic acid in roots of Zea mays. Phytochemistry 13, 239-241 (1974) Greenwood, M.S., Hillman, J.R., Shaw, S., Wilkins, M.B. : Localisation and identification of auxin in roots of Z e a mays. Planta 109, 369-374 (1973) Knegt, E., Bruinsma, J. : A rapid, sensitive and accurate determination of indolyl-3-acetic acid. Phytochemistry 12, 753-756 (1973) Martin, H.V., Elliott, M.C., Wangermann, E., Pilet, P.E.: Auxin gradient along the root of the maize seedlings. Planta 141, 179-181 (1978) Pernet, J.J., Pilet, P.E. : Indoleacetic acid movement in the root cap. Planta 128, 183-184 (1976) Pilet, P.E. : Auxins and the process of aging in root cells. In: Plant Growth Regulation, pp. 167-178, Wightmann, F., Setterfield, G., eds. Ames Iowa: The Iowa State University Press 1961 a Pilet, P.E.: L'action des auxines sur la croissance des cellules. In: Handbuch der Pflanzenphysiologie, XIV, pp. 784-806, Rubland, W., ed. Berlin, Heidelberg, New York: Springer 196Ib Pilet, P.E.: Auxin transport in roots. Nature 204, 561 562 (1964) Pilet, P.E. : Abscisic acid as a root growth inhibitor: physiological analyses. Planta 122, 299 302 (1975) Pilet, P.E. : Growth inhibitors in growing and geostimulated maize roots. In: Plant Growth Regulation, pp. 115-128, Pilet, P.E., ed. Berlin, Heidelberg, New York: Springer 1977 Rivier, L_, Pilet, P.E. : Indolyl-3-acetic acid in cap and apex of maize roots: identification and quantification by mass fragmentography. Planta 120, 107-112 (1974) Scott, T.K., Wilkins, M.B.: Auxin transport in roots. II. Polar flux of IAA in Zea roots. Planta 83, 232-334 (1968) Street, H.E., Bullen, P.M., Elliott, M.C. : The natural growth regulators of roots. In: Wachstums-Regulatoren bei Pflanzen, pp. 407-416, Libbert, E., ed. Rostock: Fischer 1967 Wain, R.L. : Root growth inhibitors. In: Plant Growth Regulation, pp. 109-114, PiIet, P.E., ed. Berlin, Heidelberg, New York: Springer 1977 Wilkins, M.B. : Geotropic response mechanism in roots and shoots. In: Plant Growth Regulation, pp. 199 207, Pilet, P.E., ed. Berlin, Heidelberg, New York: Springer 1977
Received 6 December 1978; accepted 16 May 1979
