PLANT PHYSIOLOGY , Vol 103, Issue 1 49-58, Copyright © 1993 by American Society of Plant Biologists
|
CELL BIOLOGY AND SIGNAL TRANSDUCTION |
Effect of Cutting on Solute Uptake by Plasma Membrane Vesicles from Sugar Beet (Beta vulgaris L.) Leaves
S. Sakr, R. Lemoine, C. Gaillard and S. Delrot
Laboratoire de Physiologie et Biochimie Vegetales, Unite Associee Centre National de la Recherche Scientifique 574, Universite de Poitiers, 25 rue du Faubourg Saint-Cyprien, 86000 Poitiers, France
The uptake of sucrose, 3-O-methylglucose (3-O-MeG), and valine were studied
in discs and in purified plasma membrane vesicles (PMV) prepared from sugar
beet (Beta vulgaris L.) exporting leaves. The uptake capacities of freshly
excised leaf discs were compared with the uptake in discs that had been
floated for 12 h on a simple medium (aging) and with discs excised from
leaves that had been cut from the plant 12 h before the experiments
(cutting). After cutting, sucrose uptake amounted to twice the uptake
measured in fresh discs, whereas the uptake of 3-O-MeG and valine remained
unaffected. In aged leaf discs, there was a general stimulation of uptake,
which represented 400, 300, and 400% of the uptake measured in fresh discs
for sucrose, 3-O-MeG, and valine, respectively. Sucrose uptake in fresh
discs was sensitive to N-ethylmaleimide (NEM), to
p-chloromercuribenzenesulfonic acid (PCMBS), and to mersalyl acid (MA).
Although cutting induced the appearance of a sucrose uptake system that is
poorly sensitive to NEM but sensitive to PCMBS and MA, aging induced the
development of an uptake system that is sensitive to NEM but poorly
sensitive to PCMBS and MA. Autoradiographs of discs fed with [14C]sucrose
show that cutting resulted in an increase of vein labeling with little
effect in the mesophyll, whereas aging induced an increase of labeling
located mainly in the mesophyll. The data show that cutting is sufficient
to induce dramatic and selective changes in the uptake properties of leaf
tissues and that the effects of cutting and aging on the uptake of organic
solutes are clearly different. Parallel experiments were run with purified
PMV prepared from fresh and cut leaves. The uptake of sugars and amino
acids was studied after imposition of an artificial proton motive force
(pmf). Comparison of the uptake properties of PMV and of leaf tissues
indicate that the recovery of the sucrose uptake system in PMV is better
than the recovery of the hexose and of the valine uptake systems. As
observed with the leaf discs, cutting induced a 2-fold increase of the
initial rate of sucrose uptake in PMV but did not affect the uptake of
valine and 3-O-MeG. Cutting induced an increase of both Vmax and Km of the
sucrose transport system in PMV. Measurements of the pmf imposed on the
vesicles indicated that the increase of sucrose uptake induced by cutting
was not due to a better integrity of the vesicles. Hexoses did not compete
with sucrose for uptake in PMV from fresh and cut leaves, and maltose was a
stronger inhibitor of sucrose uptake in PMV from cut leaves than in PMV
from fresh leaves. The sensitivity of sucrose uptake to NEM, PCMBS, and MA
in PMV from fresh and cut leaves paralleled that described above for the
corresponding leaf discs. These data show that (a) the changes induced by
cutting on sucrose uptake by leaf discs are due to membrane phenomena and
not to the metabolism of sucrose; (b) the study of sucrose uptake with PMV
gives a good account of the physiological situation; and (c) the specific
effects induced by cutting on the sucrose uptake system are not lost during
the preparation of the PMV.
