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Development of Cell Wall Ingrowths in Aging Slices of Radish (Raphanus sativus L.)
Short Communication
Development of Cell Wall Ingrowths in Aging Slices of Radish (Raphanus sativus 1.) DAISY PEREZ-RODRIGUEZ
1
)
Ecole Normale Superieure et Universite P. et M. Curie, Paris, France Received December 1, 1980 . Accepted January 30, 1981
Summary During aging of radish slices, some xylem parenchyma cells in contact with the incubation medium develop local wall thickenings. These protuberances are morphologically similar to wall ingrowths of transfer cells but they differ by their heterogeneous appearance and irregular polysaccharidic content. Key words: Raphanus sativus, transfer cell, aging, xylem parenchyma.
Introduction The elaborate wall-membrane apparatus characteristic of transfer cells is one of several specializations related to intensive trans-plasma membrane transport in plants. In these cells, the inner surface of the cell wall develops irregular, more or less numerous protuberances lined by a large surfa;ce area of plasma membrane. The amplification of surface area and the geometry of the wall labyrinth indicate special transport properties. Since they were described for the first time by Gunning et a1. (1968), transfer cells were found in many anatomical locations in plants and formation of the wall apparatus appeared to be inducible in many instances (see Gunning, 1977, for a review). For instance such apparatus is known to develop in cells cultured in suspension (Davey and Street, 1971). On the other hand no induction of transfer cells was reported in organ slices during aging phenomena (see Barckhausen, 1978). 1) Address to which reprint requests should be sent: Laboratoire de Botanique, 24 rue Lhomond 75231 Paris Cedex OS. Author's address: Universidad Simon Bolivar, Departamento de Biologia de Organismos, Apartado Postal 80. 659, Caracas, Venezuela.
Z. Pflanzenphysiol. Bd. 102. S.279-282. 1981.
280
DAISY PEREZ-RODRIGUEZ
It is generally thought that in this case surface cells become either suberized or lignified (Kahl, 1973). Development of cell wall in growths, however, was observed during our study of lignification processes in aging radish slices. Material and Methods 2 mmm-thick slices were cut from sterilized hypocotyl tubers of radish. Some were used directly, the others were washed (<< aged ») in sterile water with agitation up to 72 h. For some experiments, indole-acetic acid (5 mg / l), gibberellin (0.1 mg / l) or zeatin (0.1 mg/I) were added to the washing solution. Small pieces of tissues were cut from the slices at 0, 24, 48 or 72 h. They were fixed in 4 % glutaraldehyde buffered at pH 7.4 for 1 h 30, rinsed in buffer and postfixed in buffered 1 % osmic acid for 1 h. Specimens were dehydrated through a graded series of alcohol and propylene oxide and embedded in araldite. Ultrathin sections were either viewed unstained or contrasted by 1 % potassium permanganate or treated by P ATAg technique (Thiery, 1967) to v isualize polysaccharides with vic-glycol groups. For lignin visualization some pieces were fixed in 15 Ofo alcohol and treated with ethanolamine and silver nitrate before embedding (Cop pick and Fowler, 1939; for details on this technique, see Czaninski, 1979).
Results and Discussion The storage tissue in radish root tubers is constituted by xylem parenchyma 'cells. Their walls are thin and unlignified and they do not present any protuberances. During aging, cell wall lignification occurs in many parenchyma cells along slice surfaces. It begins in cells surrounding vascular strands and then spreads (Perez-Rodriguez, 1979). In some cells, however, another phenomenon can be observed. After a 24 h- or 48 h-incubation, plasmalemma infoldings appear in connexion with local accumulMions of polysaccharidic (PATAg-positive) material along the inner surface of the cell wall (Fig. 1). At the same time, a number of Golgi vesicles with PATAg-positive content may be seen in the cytoplasm or fusing with the plasmalemma (Fig. 1 and 2). The localized polysaccharide deposits then develop into local wall in growths. They are present in slices aged on distilled water as well as in slices incubated in presence of growth substances (Fig. 3 to 6). These wall protuberances appear heterogeneous, with local deposits of osmiophilic material while P A TAg tests show that some regions of the protuberances are devoid of polysaccharides with free vic-glycol groups (Fig. 5 and 6). No lignified Fig. 1 and 2: 48 h aging on water. P ATAg test. Dictyosomes (D) produce vesicles (V) containing polysaccharides and eventually fusing with the plasmalemma (black arrow) and producing plasmalemma infoldings (white arrows). Fig. 1 X40,000; Fig. 2 X 60,000. Fig. 3 and 4: 72 h aging either on zeatin (fig. 3) or IAA (Fig. 4) . Contrast KMn0 4 • Walls in growths (1) with osmiophilic, dense regions. W: normal cell wall. Fig. 3 X 20,000; Fig. 4 X 30,000. Fig. 5 and 6: 72 h aging either on gibberellin (fig. 5) or water (fig. 6). PATAg test. Heterogeneous deposit of polysaccharides in the wall ingrowths. Fig. 5 X 11,000; Fig. 6 X 14,000.
z.
Pjlanzenphysiol. Bd. 102. S.279-282. 1981.
Cell wall in growths in aging slices
281
Z. PJlanzenphysiol. Bd. 102. S.279-282. 1981.
282
DAISY PEREZ-RODRIGUEZ
wall ingrowths were observed under our experimental conditions even after a 72 hincubation. Morphologically, the irregular wall thickenings which develop during aging in radish slices appear somewhat similar to the wall apparatus of transfer cells. But structurally they are different. In radish slices, wall thickenings represent local masses of deposit and are heterogeneous while the typical wall protuberances of normal transfer cells are generally more homogeneous, although varying from a very open appearance to quite densely packed fibrils and matrix material (Czaninski, 1977; Gunning, 1977). Highly heterogeneous wall protuberances were described in callus tissue of Andrographis paniculata but at the time were not related to transfer cell wall thickenings (Bowes and Butcher, 1967; Bowes, 1969). On the other hand, the slightly heterogeneous wall invaginations observed in cells cultured in suspension were compared to transfer cell wall apparatus (Davey and Street, 1971). That cultured tissue fragments or cultured cells can develop wall ingrowths somewhat similar to those of 'transfer cells is clearly related to the need of intensive absorption from ·the medium. The same is probably true for aging slices although in this case the incubation medium does not contain any nutrients. In radish slices, lignification processes start in superficial cells 24 h. after the beginning of the incubation period and specialized portions of the cell wall such as those described here are then necessary to maintain exchanges with the surrounding medium . That this phenomena was not recognized in other aging materials may be due to the fact tha.t aging experime11lts rarely extend beyond 24 h. Acknowledgements A grant to the laboratory from the CNRS (LA nO 311) is gratefully aknowledged.
References BARCKHAUSEN, R.: In : G. KAHL (Ed.): Biochemistry of wounded plant tissues, pp. 1-42 (1978). BOWES, B. G.: The New Phytol. 68, 619-626 (1969). BOWES, B. G . and D. N . BUTCHER: Z. Pflanzenphysiol. 58, 86-89 (1967). COPPICK, S. and W. F. FOWLER, Jr.: Paper Trade J. 81 186, (Tappi section, 135-140), (1939). CZANINSKI, Y.: Biologie Cellulaire 29, 221-224 (1977). - Biologie Cellulaire 35, 97-102 (1979). DAVEY, M. R. and H. E. STREET: J. Exper. Bot. 22, 90-95 (1971). GUNNING, B. E. S. : Science Progress, London, 64, 539-568 (1977). GUNNING, B. E. S., J. S. PATE, and L. G. BRIARTY: J. Cell BioI. 37, C7-C12 (1968). KAHL, G.: Bot. Rev. 39, 274- 299 (1973). PEREZ-RODRIGUEZ, D.: These de 3eme cycle, Paris (1979). THIERY, J. P.: J. Microscopie 6, 987-1018 (1967) .
Z. PJlanzenphysiol. Ed. 102. S.279-282 . 1981.
Development of Cell Wall Ingrowths in Aging Slices of Radish (Raphanus sativus 1.) DAISY PEREZ-RODRIGUEZ
1
)
Ecole Normale Superieure et Universite P. et M. Curie, Paris, France Received December 1, 1980 . Accepted January 30, 1981
Summary During aging of radish slices, some xylem parenchyma cells in contact with the incubation medium develop local wall thickenings. These protuberances are morphologically similar to wall ingrowths of transfer cells but they differ by their heterogeneous appearance and irregular polysaccharidic content. Key words: Raphanus sativus, transfer cell, aging, xylem parenchyma.
Introduction The elaborate wall-membrane apparatus characteristic of transfer cells is one of several specializations related to intensive trans-plasma membrane transport in plants. In these cells, the inner surface of the cell wall develops irregular, more or less numerous protuberances lined by a large surfa;ce area of plasma membrane. The amplification of surface area and the geometry of the wall labyrinth indicate special transport properties. Since they were described for the first time by Gunning et a1. (1968), transfer cells were found in many anatomical locations in plants and formation of the wall apparatus appeared to be inducible in many instances (see Gunning, 1977, for a review). For instance such apparatus is known to develop in cells cultured in suspension (Davey and Street, 1971). On the other hand no induction of transfer cells was reported in organ slices during aging phenomena (see Barckhausen, 1978). 1) Address to which reprint requests should be sent: Laboratoire de Botanique, 24 rue Lhomond 75231 Paris Cedex OS. Author's address: Universidad Simon Bolivar, Departamento de Biologia de Organismos, Apartado Postal 80. 659, Caracas, Venezuela.
Z. Pflanzenphysiol. Bd. 102. S.279-282. 1981.
280
DAISY PEREZ-RODRIGUEZ
It is generally thought that in this case surface cells become either suberized or lignified (Kahl, 1973). Development of cell wall in growths, however, was observed during our study of lignification processes in aging radish slices. Material and Methods 2 mmm-thick slices were cut from sterilized hypocotyl tubers of radish. Some were used directly, the others were washed (<< aged ») in sterile water with agitation up to 72 h. For some experiments, indole-acetic acid (5 mg / l), gibberellin (0.1 mg / l) or zeatin (0.1 mg/I) were added to the washing solution. Small pieces of tissues were cut from the slices at 0, 24, 48 or 72 h. They were fixed in 4 % glutaraldehyde buffered at pH 7.4 for 1 h 30, rinsed in buffer and postfixed in buffered 1 % osmic acid for 1 h. Specimens were dehydrated through a graded series of alcohol and propylene oxide and embedded in araldite. Ultrathin sections were either viewed unstained or contrasted by 1 % potassium permanganate or treated by P ATAg technique (Thiery, 1967) to v isualize polysaccharides with vic-glycol groups. For lignin visualization some pieces were fixed in 15 Ofo alcohol and treated with ethanolamine and silver nitrate before embedding (Cop pick and Fowler, 1939; for details on this technique, see Czaninski, 1979).
Results and Discussion The storage tissue in radish root tubers is constituted by xylem parenchyma 'cells. Their walls are thin and unlignified and they do not present any protuberances. During aging, cell wall lignification occurs in many parenchyma cells along slice surfaces. It begins in cells surrounding vascular strands and then spreads (Perez-Rodriguez, 1979). In some cells, however, another phenomenon can be observed. After a 24 h- or 48 h-incubation, plasmalemma infoldings appear in connexion with local accumulMions of polysaccharidic (PATAg-positive) material along the inner surface of the cell wall (Fig. 1). At the same time, a number of Golgi vesicles with PATAg-positive content may be seen in the cytoplasm or fusing with the plasmalemma (Fig. 1 and 2). The localized polysaccharide deposits then develop into local wall in growths. They are present in slices aged on distilled water as well as in slices incubated in presence of growth substances (Fig. 3 to 6). These wall protuberances appear heterogeneous, with local deposits of osmiophilic material while P A TAg tests show that some regions of the protuberances are devoid of polysaccharides with free vic-glycol groups (Fig. 5 and 6). No lignified Fig. 1 and 2: 48 h aging on water. P ATAg test. Dictyosomes (D) produce vesicles (V) containing polysaccharides and eventually fusing with the plasmalemma (black arrow) and producing plasmalemma infoldings (white arrows). Fig. 1 X40,000; Fig. 2 X 60,000. Fig. 3 and 4: 72 h aging either on zeatin (fig. 3) or IAA (Fig. 4) . Contrast KMn0 4 • Walls in growths (1) with osmiophilic, dense regions. W: normal cell wall. Fig. 3 X 20,000; Fig. 4 X 30,000. Fig. 5 and 6: 72 h aging either on gibberellin (fig. 5) or water (fig. 6). PATAg test. Heterogeneous deposit of polysaccharides in the wall ingrowths. Fig. 5 X 11,000; Fig. 6 X 14,000.
z.
Pjlanzenphysiol. Bd. 102. S.279-282. 1981.
Cell wall in growths in aging slices
281
Z. PJlanzenphysiol. Bd. 102. S.279-282. 1981.
282
DAISY PEREZ-RODRIGUEZ
wall ingrowths were observed under our experimental conditions even after a 72 hincubation. Morphologically, the irregular wall thickenings which develop during aging in radish slices appear somewhat similar to the wall apparatus of transfer cells. But structurally they are different. In radish slices, wall thickenings represent local masses of deposit and are heterogeneous while the typical wall protuberances of normal transfer cells are generally more homogeneous, although varying from a very open appearance to quite densely packed fibrils and matrix material (Czaninski, 1977; Gunning, 1977). Highly heterogeneous wall protuberances were described in callus tissue of Andrographis paniculata but at the time were not related to transfer cell wall thickenings (Bowes and Butcher, 1967; Bowes, 1969). On the other hand, the slightly heterogeneous wall invaginations observed in cells cultured in suspension were compared to transfer cell wall apparatus (Davey and Street, 1971). That cultured tissue fragments or cultured cells can develop wall ingrowths somewhat similar to those of 'transfer cells is clearly related to the need of intensive absorption from ·the medium. The same is probably true for aging slices although in this case the incubation medium does not contain any nutrients. In radish slices, lignification processes start in superficial cells 24 h. after the beginning of the incubation period and specialized portions of the cell wall such as those described here are then necessary to maintain exchanges with the surrounding medium . That this phenomena was not recognized in other aging materials may be due to the fact tha.t aging experime11lts rarely extend beyond 24 h. Acknowledgements A grant to the laboratory from the CNRS (LA nO 311) is gratefully aknowledged.
References BARCKHAUSEN, R.: In : G. KAHL (Ed.): Biochemistry of wounded plant tissues, pp. 1-42 (1978). BOWES, B. G.: The New Phytol. 68, 619-626 (1969). BOWES, B. G . and D. N . BUTCHER: Z. Pflanzenphysiol. 58, 86-89 (1967). COPPICK, S. and W. F. FOWLER, Jr.: Paper Trade J. 81 186, (Tappi section, 135-140), (1939). CZANINSKI, Y.: Biologie Cellulaire 29, 221-224 (1977). - Biologie Cellulaire 35, 97-102 (1979). DAVEY, M. R. and H. E. STREET: J. Exper. Bot. 22, 90-95 (1971). GUNNING, B. E. S. : Science Progress, London, 64, 539-568 (1977). GUNNING, B. E. S., J. S. PATE, and L. G. BRIARTY: J. Cell BioI. 37, C7-C12 (1968). KAHL, G.: Bot. Rev. 39, 274- 299 (1973). PEREZ-RODRIGUEZ, D.: These de 3eme cycle, Paris (1979). THIERY, J. P.: J. Microscopie 6, 987-1018 (1967) .
Z. PJlanzenphysiol. Ed. 102. S.279-282 . 1981.