Iron Absorption by Enzymically Isolated Leaf Cells'

The rate of Fe absorption by cells enzymically isolated from tobacco leaves is corre'ated with the age of the leaves from which the cells are derived4 The cells obtained from younger leaves absorb Fe more rapidly than those from older ones. Ca inhibits Fe and Mn absorption. Fe and Mn are mutually antagonitic in their absorption by leaf cells. Ca enhances the inhibition of Mn absorption by Fe, but reduces the inhibition of Fe aibsorption by Mn. The affinity constant for Fe absorption by leaf cells is low. The chelate EDDHA (ethylenediamine di (o-hydroxyph-nylacetate) comp)eLitively inhibits Fe absorption. Ioni uptake by variotus plant parts, incltlding leaves, hias been the sul)ject of muclh researclh in recent y-ears (9,13, 18, 19, 22). Howvever, absorption of ions by leaves has received scant attention colimpared to roots. It has been slhowvni that leaves and leaf tis tuCs actively accumulate iolns fromi the ambient solution (7, 19). Living cells have been isolated enzy1mcalllv fromii leaves, stems anid roots (7,23) and the nmechanisms of ion uptake lhave been sttudied at tlle celluilar level (7, 11). Fur.hermore, it has been demonstrated that lighlt, succinate and bicarbonate as a source of CO.! for phlotosynitlhesis promote Rb and pho lphate ul)take by leaf cells. Metabolic inhibi.ors like NaN, and DNP inhibit ioll uptake by cells (7). Absorption of Fe by tobacco leaf cells has been slhoNvii to be metabolic (11). Stuldies of some factors affeciting Fe absorption by enzymically isolatcd leaf cells are reported lierein. Materials and Methods Plant AMaterinls. Tobacco (ATicotiana tabacutit L., Bturlev type) vas growvn in a greenhlotuse and cells isolated from leaves of different positions on the plant wvere tised to sttudy the effects of leaf age. All other experimeicnits vere carried out with cells or discs obtained fromii fuilly expan(de(d green leaves. Procedure for Isolationz of Cclls. The proceduire wvas essentiallv the samne as that described by Zaitlin (23) and has been outlinied in detail (7, 11). Leaves ' Aichigan Agricultural Experiment Station Journal Article No. 4867. 21PIresenit address: Biology Division, Bhabha Atomic Research Centre, Bombay 85, India. wvere sliced into 2 to 3 mm2 sections, and treated with enzyme solution. The isolation mixture contained nmineral salts and organic constituents (16). Pectinase (1 % w/v) was added and the pH adjusted to 6. Isolated cells were washed with cold 0.3 M sucrose and suspended in it at 3' until used. For eaclh experiment, cells were freshly isolated and used within 48 hr. Procedutre for lncubating Leaf Cells and Leaf Discs. The inicuibation mixture contained per 10 ml, 50 mg dry weight equivalent of cells, 1170 ,umoles sucrose, 100 jutmoles tris-maleate (pH 6.4) and 5 /An-oles "9Fe labeled FeSO4 or "4Mn labeled MnSO4 unless otlherwise indicated. The incubation medium for studies of ion uptake by leaf discs was of the same composition as above, but a volume of 20 ml instead of 10 ml was used. It contained besides, 4 leaf discs (0.8 cm2) weighing about 100 mg, and 10 ,umoles of isotopically labeled FeSO4, MnSO4, ZnSO0 cr RbCl. The flasks containing the incubation miixture were slhaken in a reciprocating waterbatlh in the lighlt (ca. 500 ft-c) at 20 ± 2°. M1casuireinent of Ion Uptake. At the end of the absorption period, cells in 2 ml of the incubation miixture were collected on weighed filter paper under mild suction. washed twice with non-labeled 0.1 niM soltution of FeSO4 or MnSO4 and finally with 0.3 M sucrose. Leaf discs were removed from the incubation meditum and rinsed in non-labeled 0.1 mM solution of the respective salt for 2 min and finally in deionized water. Air dry samples were radioassayed in a gamma well scintillation counter. Significant differences between triplicate samples were establislied statistically (2). The data presented in the table and figtures were derived from separate experiments and are comparable within an experiment but not between experiments. 1457 www.plantphysiol.org on August 29, 2017 Published by Downloaded from Copyright © 1969 American Society of Plant Biologists. All rights reserved.