Mechanism of swelling-activation of k-cl cotransport in inside-out
vesicles of lk sheep erythrocyte membranes.
Kelley, Scott J., and Philip B. Dunham.
Department of Biology, Syracuse University, Syracuse, NY 13244
APStracts 2:0371C, 1995.
Stimulation by swelling of K-Cl cotransport was studied in inside-out
vesicles (IOVs) made from membranes of LK sheep erythrocytes. The
purpose was to understand this stimulation in terms of the three
-state process proposed for regulation of the cotransporter (Dunham et
al. J. Gen. Physiol. 101: 733-765, 1993). The first step in this
process, A ---> B, is rate limiting and controlled by
transphosphorylation reactions. The second step, B ---> C, is
fast; its control is unknown. Predictions were that Jmax of
cotransport increases with A ---> B and K1/2 for K+ as a
substrate decreases with B ---> C. The hypothesis tested was
that most transporters in IOVs are in the B-state, and that swelling
activates cotransport in vesicles by the B ---> C conversion. In
accordance with the hypothesis, swelling should activate K+ influx
with no discernible delay. It did. K1/2 for K+ should decrease with
swelling, and Jmax should not change. K1/2 decreased 10-fold and Jmax
didn't change. Inhibitors of transphosphorylation, reactions of A --
-> B, should not affect K+ influx in IOVs, and did not. The
results support the hypothesis; swelling-activation of K+ influx in
IOVs corresponds to B ---> C. A mechanical change in the
membrane causes a specific change in the cotransporter, an increase
in apparent affinity for K+.
Received 23 May 1995; accepted in final form 5 October 1995.
APS Manuscript Number C295-5.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 6 November 95