Isoforms of the apical na-k-2cl transporter in murine thick
ascending limb. i: molecular characterization and intra-renal
localization.
Mount, David B., Allan Baekgaard, Amy E. Hall, Consuelo Plata, Jason
Xu, David R. Beier, Gerardo Gamba and Steven C. Hebert.
Nephrology Division, Vanderbilt University, MCN S-3223, Nashville,
TN 37232,
APStracts 5:0199F, 1998.
We have identified several alternatively spliced cDNAs encoding mBSC1,
an apical bumetanide-sensitive Na+-K+-2Cl- cotransporter from mouse
kidney. Two full-length clones were isolated, designated C4 and C9,
predicting proteins of 770 and 1095 amino acids respectively. The C4
isoforms are generated by utilization of an alternative poly
-adenylation site located within the intron between exons 16 and 17 of
the mBSC1 gene on chromosome 2; the resultant transcripts predict a
truncated C-terminus ending in a unique 55 amino acid sequence. The
predicted C4 and C9 C-termini differ in the distribution of putative
phosphorylation sites for both PKA and PKC. Independent splicing
events involve three previously described cassette exons, which are
predicted to encode most of the second transmembrane domain. A total
of six different isoforms are expressed, generated by the
combinatorial association of three cassette exons and two alternative
3( ends. C9-specific and C4-specific antibodies detect proteins of
[sim]150 kDa and 120 kDa, respectively, in mouse kidney.
Immunofluorescence and immunohistochemistry indicate expression of
both C-terminal isoforms within the thick ascending limb. However,
staining with the C4 antibody is more heterogeneous, with a decreased
proportion of positive cells in the cortical thick ascending limb.
Functional expression in Xenopus oocytes indicates a dominant
negative function for C4 isoforms (companion paper, Plata et al.),
and the differential expression of these isoforms may contribute to
functional heterogeneity of Na+-K+-2Cl- cotransport in mouse TAL.
Received 15 June 1998; accepted in final form 30 October 1998.
APS Manuscript Number F148-8.
Article publication pending Am. J. Physiol. (Renal Physiology).
ISSN 1080-4757 Copyright 1998 The American Physiological Society.
Published in APStracts on 10 November 1998