Isoforms of the apical na-k-2cl cotransporter in murine thick
ascending limb. ii: functional characterization and mechanism of
activation by cyclic-amp.
Plata, Consuelo, David B. Mount, Verena Rubio, Steven C. Hebert and
Gerardo Gamba.
Molecular Physiology Unit. Department of Nephrology and Mineral
Metabolism, Instituto Nacional de la Nutrici[acute]on Salvador
Zubir[acute]an and Department of Medicine, Instituto de
Investigaciones Biom[acute]edicas, National University of Mexico,
Mexico City CP 14000, Mexico and *Division of Nephrology, Department
of Medicine, Vanderbilt University Medical Center, Nashville, TN
37232
APStracts 5:0198F, 1998.
The functional properties of alternatively spliced isoforms of the
mouse apical Na+-K+-2Cl- cotransporter (mBSC1) were examined, using
expression in Xenopus oocytes and measurement of 22Na+ or 86Rb+
uptake. A total of six isoforms, generated by the combinatorial
association of three 5( exon cassettes (A, B, and F) with two
alternative 3( ends, are expressed in mouse thick ascending limb
(TAL) (companion paper, Mount, et al.). The two 3( ends predict C
-terminal cytoplasmic domains of 129 amino acids (the _C4_ C-terminus)
and 457 amino acids (the _C9_ terminus). The three C9 isoforms
(mBSC1-A9/F9/B9) all express Na+-K+-2Cl- cotransport activity,
whereas C4 isoforms are non-functional in Xenopus oocytes. Activation
or inhibition of protein kinase A (PKA) does not affect the activity
of the C9 isoforms. The co-injection of mBSC1-A4 with mBSC1-F9
reduces tracer uptake, as compared with mBSC1-F9 alone, an effect of
C4 isoforms that is partially reversed by the addition of cAMP-IBMX
to the uptake medium. The inhibitory effect of C4 isoforms is a dose
-dependent function of the alternatively spliced C-terminus. Isoforms
with a C4 C-terminus thus exert a dominant-negative effect on Na+-K+
-2Cl- cotransport, a property that is reversed by the activation of
PKA. This interaction between co-expressed C-terminal isoforms of
mBSC1 may account for the regulation of Na+-K+-2Cl- cotransport in
the mouse TAL by hormones that generate cAMP.
Received 15 June 1998; accepted in final form 30 October 1998.
APS Manuscript Number F149-8.
Article publication pending Am. J. Physiol. (Renal Physiology).
ISSN 1080-4757 Copyright 1998 The American Physiological Society.
Published in APStracts on 10 November 1998