 |
|  |
Work
in the Caplan laboratory is focused on understanding how membrane proteins
are sorted to the appropriate cell surface domains of polarized epithelial
cells. One of the proteins whose trafficking we study is the Na,K-ATPase,
or sodium pump, which generates the ion gradients responsible for most
fluid and electrolyte transport processes in the kidney. The Na,K-ATPase
must be restricted to the basolateral surfaces of renal tubule epithelial
cells. Much remains to be learned about the partner proteins and trafficking
pathways that determine the sodium pump’s subcellular distribution
and modulate its activity. We have adapted a novel labeling methodology
to investigate the attributes of temporally defined cohorts of Na,K-ATPase.
We can observe directly the trafficking itinerary pursued by newly synthesized
Na,K-ATPase and isolate newly synthesized Na,K-ATPase in association
with its collections of partner proteins. We find that the basolateral
delivery of newly synthesized Na,K-ATPase occurs via a pathway distinct
from that pursued by other basolateral membrane proteins. We have also
detected interactions between the Na,K-ATPase a-subunit and a collection
of novel partner proteins that may govern the pump’s trafficking
properties. Thus, we have developed tools that permit us to evaluate
the trafficking pathways and partner proteins that govern the post-synthetic
sorting and regulation of the epithelial Na,K-ATPase.
We also study a common genetic disease that dramatically alters the
structure and function of polarized epithelial cells. In Autosomal Dominant
Polycystic Kidney Disease (ADPKD) the normal architecture of the kidney
tubules is replaced by large fluid filled cysts, which can ultimately
result in renal failure. ADPKD is caused by mutations in the PKD1 or
PKD2 genes, which encode the polycystin-1 and polycystin-2 proteins,
respectively. Both of these proteins are targeted to cilia in polarized
epithelial cells. We have found that polycystin-1 undergoes such a proteolytic
cleavage that releases its C-terminal tail (CTT), which enters the nucleus
and initiates signaling processes. The cleavage occurs in vivo in association
with alterations in mechanical stimuli that may be communicated by signaling
through the cilium. The C-terminal tail fragment of polycystin-1 participates
in a complex with ß-catenin and acts to profoundly inhibit canonical
ß-catenin-dependent Wnt signaling. The polycystin-1 C-terminal
tail fragment also appears to modulate gene expression, and may induce
expression of cilia-related proteins in renal epithelial cells. We find
that all of the signal transduction machinery found in the cilia of
olfactory epithelial cells is present in renal epithelial cells. Our
data suggest that olfactory receptors and proteins involved in olfactory
signal transduction may play a role in regulating renal flow or transport
in response to chemosensory cues.
Figure
caption:
Polarized
pig kidney epithelial cells have been transfected with a cDNA encoding
a chimeric ion pump composed of portions of the Na,K-ATPase and the
gastric H,K-ATPase. The chimera and the endogenous population of Na,K-ATPase
were localized by immunfluorescence confocal microscopy. The distribution
of the chimera (red staining) is restricted to the apical membranes
of these epithelial cells (shown in cross-section), whereas the Na,K-ATPase
(green staining) is limited to the basolateral surfaces. Despite ~85%
amino acid sequence identity, therefore, the chimera and the Na,K-ATPase
are differentially sorted by these cells.
Selected
publications:
Click
 for PDF
Chauvet,
V., X. Tian, H. Husson , D.H. Grimm , T. Wang , T. Hieseberger, P. Igarashi,
A.M. Bennett , O. Ibraghimov-Beskrovnaya , S. Somlo and M.J. Caplan.
Mechanical stimuli induce the cleavage and nuclear translocation of
the polycystin-1 C-terminus. J. Clin. Invest. 114: 1433-1443, 2004.
Zhang,
L., J. Li, L.H. Young and M.J. Caplan. AMP-activated protein kinase
regulates the assembly of epithelial tight junctions. Proc. Nat. Acad.
Sci. (USA) 103: 17272-17277, 2006.
Kamsteeg,
E.J., A.S. Duffield, I.B.M. Konings, J. Spencer, P. Pagel, P.M.T. Deen
and M. J. Caplan. MAL decreases the internalization of the aquaporin-2
water channel. Proc. Nat. Acad. Sci. (USA) 104:16696-16701, 2007.
Lal,
M., X. Song, J.L. Pluznick, V. Di Giovanni, D.M. Merrick, N.D. Rosenblum,
V. Chauvet, C.J. Gottardi, Y. Pei and M.J. Caplan. Polycystin-1 C-terminal
Tail Associates with
ß-catenin and Inhibits Canonical Wnt Signaling. Hum. Mol. Gen.
17:3105-3117, 2008.
Pluznick,
J.L., D.J. Zou, X. Zhang, Q. Yan, D.J. Rodriguez-Gil, C. Eisner, E.
Wells, C.A. Greer, T. Wang, S. Firestein, J. Schnermann, and M.J. Caplan.
Functional expression of the olfactory signaling system in the kidney.
Proc. Nat. Acad. Sci., 106:2059-2064 , 2009.
Farr,
G.A., M. Hull, I.S. Mellman and M.J. Caplan. Membrane Proteins Follow
Multiple Trafficking Pathways to the Basolateral Cell Surface in Polarized
Epithelial Cells. J. Cell Biol., 186:269-282, 2009.
michael.caplan@yale.edu
|
|