Reconstitution of calyculin-inhibited k-cl cotransport in dog
erythrocyte ghosts by exogenous protein phosphatase type 1.
Dunham, Thomas Krarup Philip B.
Department of Biology, Syracuse University, 130 College Place,
Syracuse, New York 13244-1220
APStracts 2:0348C, 1995.
Osmotic swelling of dog and other mammalian erythrocytes activates Cl
-dependent K transport, K-Cl cotransport. This activation can be
abolished by nanomolar concentrations of calyculin, a potent
inhibitor of serine/threonine protein phosphatases. Therefore K-Cl
cotransport is probably activated by dephosphorylation by a type 1
and/or type 2A phosphatase. This was tested directly by incorporating
exogenous protein phosphatases into resealed ghosts made from dog
erythrocytes previously exposed to calyculin. K-Cl cotransport was
nearly completely inhibited in the ghosts. Incorporation of PP-1
reconstituted K-Cl cotransport. Maximal reconstitution was up to 90%
of the control flux in the ghosts and 0.1 units of PP-1 per ml of
lysate gave half-maximal reconstitution of cotransport. In contrast,
PP-2A had no effect. This result with PP-1 provides direct evidence
that K-Cl cotransport is activated by PP-1 in dog erythrocytes. Half
-maximal activation of K-Cl cotransport required 180 molecules of PP-1
per ghost.
Received 26 June 1995; accepted in final form 22 September 1995.
APS Manuscript Number C372-5.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 6 November 95