Functional expression of GFP-linked human heart sodium channel (hH1)
and subcellular localization of the a subunit in HEK293 cells and
dog cardiac myocytes.
Recent evidence suggests that biosynthesis of the human heart Na$^+$
channel (hH1) protein is rapidly modulated by sympathetic interventions.
However, data regarding the intracellular processing of hH1 in vivo
are lacking. In this study we sought to establish a model that would
allow us to study the subcellular localization of hH1 protein. Such
a model could eventually help us to better understand the trafficking
of hH1 in vivo and its potential role in cardiac conduction. We labeled
the C-terminus of hH1 with the green fluorescent protein (GFP) and
compared the expression of this construct (hH1-GFP) and hH1 in transfected
HEK293 cells. Fusion of GFP to hH1 did not alter its electrophysiological
properties. Confocal microscopy revealed that hH1-GFP was highly
expressed in intracellular membrane structures. Immuno-electronmicrographs
showed that transfection of hH1-GFP and hH1 induced proliferation
of three types of endoplasmic reticulum (ER) membranes to accommodate
the heterologously expressed proteins. Labeling with specific markers
for the ER and the Golgi apparatus indicated that the intracellular
channels are almost exclusively retained within the ER. Immunocytochemical
labeling of the Na$^+$ channel in dog cardiomyocytes showed strong
fluorescence in the perinuclear region of the cells, a result consistent
with our findings in HEK293 cells. We propose that the ER may serve
as a reservoir for the cardiac Na$^+$ channels and that the transport
from the ER to the Golgi apparatus is among the rate-limiting steps
for sarcolemmal expression of Na$^+$ channels.
%0 Journal Article
%1 Zimm_2002_1
%A Zimmer, T.
%A Biskup, C.
%A Dugarmaa, S.
%A Vogel, F.
%A Steinbis, M.
%A B�hle, T.
%A Wu, Y. S.
%A Dumaine, R.
%A Benndorf, K.
%D 2002
%J J. Membr. Biol.
%K Animals; Apparatus, Biological; Cells, Channels, Cultured; Dogs; Electrophysiology; Endoplasmic Fluorescent Fusion Golgi Green Humans; Intracellular Luminescent Membranes, Models, Myocardium, Protein Proteins, Proteins; Recombinant Reticulum, Sodium Subunits; Transport; cytology/metabolism; drug effects/metabolism/physiology; genetics/metabolism/pharmacology genetics/metabolism; genetics; metabolism;
%N 1
%P 1--12
%R 10.1007/s00232-001-0130-1
%T Functional expression of GFP-linked human heart sodium channel (hH1)
and subcellular localization of the a subunit in HEK293 cells and
dog cardiac myocytes.
%U http://dx.doi.org/10.1007/s00232-001-0130-1
%V 186
%X Recent evidence suggests that biosynthesis of the human heart Na$^+$
channel (hH1) protein is rapidly modulated by sympathetic interventions.
However, data regarding the intracellular processing of hH1 in vivo
are lacking. In this study we sought to establish a model that would
allow us to study the subcellular localization of hH1 protein. Such
a model could eventually help us to better understand the trafficking
of hH1 in vivo and its potential role in cardiac conduction. We labeled
the C-terminus of hH1 with the green fluorescent protein (GFP) and
compared the expression of this construct (hH1-GFP) and hH1 in transfected
HEK293 cells. Fusion of GFP to hH1 did not alter its electrophysiological
properties. Confocal microscopy revealed that hH1-GFP was highly
expressed in intracellular membrane structures. Immuno-electronmicrographs
showed that transfection of hH1-GFP and hH1 induced proliferation
of three types of endoplasmic reticulum (ER) membranes to accommodate
the heterologously expressed proteins. Labeling with specific markers
for the ER and the Golgi apparatus indicated that the intracellular
channels are almost exclusively retained within the ER. Immunocytochemical
labeling of the Na$^+$ channel in dog cardiomyocytes showed strong
fluorescence in the perinuclear region of the cells, a result consistent
with our findings in HEK293 cells. We propose that the ER may serve
as a reservoir for the cardiac Na$^+$ channels and that the transport
from the ER to the Golgi apparatus is among the rate-limiting steps
for sarcolemmal expression of Na$^+$ channels.
@article{Zimm_2002_1,
abstract = {Recent evidence suggests that biosynthesis of the human heart {N}a$^{+}$
channel (hH1) protein is rapidly modulated by sympathetic interventions.
However, data regarding the intracellular processing of hH1 in vivo
are lacking. In this study we sought to establish a model that would
allow us to study the subcellular localization of hH1 protein. Such
a model could eventually help us to better understand the trafficking
of hH1 in vivo and its potential role in cardiac conduction. We labeled
the C-terminus of hH1 with the green fluorescent protein (GFP) and
compared the expression of this construct (hH1-GFP) and hH1 in transfected
HEK293 cells. Fusion of GFP to hH1 did not alter its electrophysiological
properties. Confocal microscopy revealed that hH1-GFP was highly
expressed in intracellular membrane structures. Immuno-electronmicrographs
showed that transfection of hH1-GFP and hH1 induced proliferation
of three types of endoplasmic reticulum (ER) membranes to accommodate
the heterologously expressed proteins. Labeling with specific markers
for the ER and the Golgi apparatus indicated that the intracellular
channels are almost exclusively retained within the ER. Immunocytochemical
labeling of the {N}a$^{+}$ channel in dog cardiomyocytes showed strong
fluorescence in the perinuclear region of the cells, a result consistent
with our findings in HEK293 cells. We propose that the ER may serve
as a reservoir for the cardiac {N}a$^{+}$ channels and that the transport
from the ER to the Golgi apparatus is among the rate-limiting steps
for sarcolemmal expression of {N}a$^{+}$ channels.},
added-at = {2009-06-03T11:20:58.000+0200},
author = {Zimmer, T. and Biskup, C. and Dugarmaa, S. and Vogel, F. and Steinbis, M. and B�hle, T. and Wu, Y. S. and Dumaine, R. and Benndorf, K.},
biburl = {https://www.bibsonomy.org/bibtex/2324a3ec7af13204e893a8249b0e7af30/hake},
description = {The whole bibliography file I use.},
doi = {10.1007/s00232-001-0130-1},
file = {Zimm_2002_1.pdf:Zimm_2002_1.pdf:PDF},
institution = {Institute of Physiology II, Friedrich Schiller University Jena, Teichgraben
8, 07740 Jena, Germany. tzim@mti-n.uni-jena.de},
interhash = {3407333f4c95be54f561a5b9b1c85e96},
intrahash = {324a3ec7af13204e893a8249b0e7af30},
journal = {J. Membr. Biol.},
keywords = {Animals; Apparatus, Biological; Cells, Channels, Cultured; Dogs; Electrophysiology; Endoplasmic Fluorescent Fusion Golgi Green Humans; Intracellular Luminescent Membranes, Models, Myocardium, Protein Proteins, Proteins; Recombinant Reticulum, Sodium Subunits; Transport; cytology/metabolism; drug effects/metabolism/physiology; genetics/metabolism/pharmacology genetics/metabolism; genetics; metabolism;},
month = Mar,
number = 1,
pages = {1--12},
pdf = {Zimm_2002_1.pdf},
pmid = {11891584},
timestamp = {2009-06-03T11:21:39.000+0200},
title = {Functional expression of GFP-linked human heart sodium channel (hH1)
and subcellular localization of the a subunit in HEK293 cells and
dog cardiac myocytes.},
url = {http://dx.doi.org/10.1007/s00232-001-0130-1},
volume = 186,
year = 2002
}