1Department of Medicine, Uniformed Services University, Bethesda 20814; 2Department of Medicine, University of Maryland, Baltimore, Maryland 21201; and 3Department of Natural Environment Sciences, Kyoto University, Kyoto 606-01, Japan
Submitted 19 November 2002 ; accepted in final form 2 April 2003
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ABSTRACT |
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Madin-Darby canine kidney cells; N-acetylcysteine; catalase
All mammalian cells have inward Na+ and outward K+ gradients. Maintenance of these two ion gradients relies on the Na,K-ATPase that acts through coupled active transport of Na+ and K+ across the plasma membrane. In cell culture media, a decrease in K+ concentration <1 mM inhibits the activity of the Na,KATPase. In most mammalian cells, the Na,K-ATPase represents the only pathway to expel intracellular Na+ and uptake extracellular K+. Any malfunctioning or inhibition of the Na,K-ATPase will result in decreased cellular Na+ extrusion and K+ uptake. Under conditions of continuous passive leaks, there will be a net gain of intracellular Na+ and a net loss of intracellular K+, thus creating a decrease in the Na+ and K+ gradients. Cells respond to this stress by upregulation of the Na,K-ATPase. This process is constituted by increases in 1) Na,K-ATPase mRNA abundance within the first few hours, 2) Na,K-ATPase protein synthesis and abundance within 8 to 24 h, and 3) the function of the enzyme within 20 to 24 h (3, 12, 15, 16). A reduction of serum K+ concentration to about 3 mM by restriction of K+ intake also stimulates Na,K-ATPase activity with coordinated increases in the mRNA and protein abundances in the medullary collecting duct of rats (4, 6, 9, 13). However, the K+ concentration used in vitro is much lower than that used in vivo. A similar sequence of reactions is also induced when cells are exposed to a sublethal concentration of ouabain, a specific inhibitor of the enzyme (17, 18). The effect of low K+ on Na,K-ATPase activity is abolished when the Na+ concentration is reduced (2). Subsequent studies demonstrate that veratridine or monensin, used to increase Na+ entry, produces a similar effect on the Na,K-ATPase as does low K+ (15, 23, 26). The critical role of Na+ in the effect of low K+, therefore, has been established.
Although the effect of low K+ on the Na,K-ATPase has been
repeatedly demonstrated both in vitro and in vivo, the signaling pathway that
transduces the effect of low K+ remains largely unknown. Reactive
oxygen species (ROS) are generated as byproducts of cellular metabolism. Over
the past decade, ROS have emerged as an important integral component of
membrane receptor signaling. ROS fulfill important prerequisites for
intracellular messenger molecules; they are easily synthesized, highly
diffusible, easily degraded, and ubiquitously present within all types of
cells. Although ROS can be generated from a variety of sources, the ROS acting
as messengers are usually produced from NADPH oxidase, a flavoprotein similar
to the phagocytic NADPH oxidase. This enzyme produces ROS with rapid kinetics
of activation and inactivation. The rapid kinetics allows a tight up- and
downregulation of intracellular ROS levels within the short time required for
transduction of signals (5).
ROS have been demonstrated to be involved in the signaling pathways
originating from growth receptors, cytokine receptors, receptor
serine/threonine kinases, G protein-coupled receptors, and ion channel-linked
receptors (5). In rat
cardiomyocytes, the partial inhibition of the Na,KATPase by a nontoxic
concentration of ouabain causes a rapid generation of ROS that is prevented by
preexposure of the cells to the antioxidants N-acetylcysteine (NAC)
or vitamin E. In parallel, these antioxidants also block or attenuate the
ouabain-induced expression of genes that are related to cardiac hypertrophy
like skeletal -actin and atrial natriuretic factor
(28). In Madin-Darby canine
kidney (MDCK) cells, low K+-induced biosynthesis of the Na,K-ATPase
is dependent on the intracellular iron activity that is important to the
generation of ROS and is inhibited by catalase
(30). In the present study, we
examined whether the stimulation of the Na,K-ATPase by low K+
required ROS, using MDCK cells as a model.
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MATERIALS AND METHODS |
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Cell culture. MDCK cells were purchased from the American Type Culture Collection (Manassas, VA). The cells were kept in Dulbecco's modified essential medium (DMEM) with 10% fetal bovine serum at 37°C in an atmosphere containing 5% CO2. The cells were collected after exposure to trypsin for 10 min and placed down at a high density. After incubation for 24 h, the cells became confluent and were then treated with either control or low-K+ medium for appropriate periods of time. The control medium contained K+-free DMEM (JRH Biosciences, Lenexa, KS) plus 7.5% horse serum and 2.5% fetal bovine serum that had been dialyzed against K+-free DMEM or Ca2+, K+-free Hanks' solution (150 mM NaCl, 0.5 mM MgCl2, 0.2 mM Na2HPO4, 0.4 mM NaH2PO4) and was supplemented with 5.25 mM K+. All low-K+ media were identical to the control medium except some K+ was substituted by Na+ (3). In most of the experiments, the concentration of K+ in the low-K+ group was 0.1 mM, except for in the concentration-response study in which serially reduced K+ concentrations were used. The antibiotic gentamycin (25 µg/ml; GIBCO BRL, Grand Island, NY) was included in all culture media.
Ouabain binding assay. The cells were placed down at 6 x 104 cells/well in a 96-well plate. After treatments, the confluent cells were washed twice with Ca2+, K+-free Hanks' solution and incubated in the same solution plus 2 mM EGTA at 37°C for 15 min to disrupt tight junctions of the cells. Next, the cells were incubated in ouabain binding media at 37°C for an additional 15 min, washed four times with ice-cold Ca2+, K+-free Hanks' solution, and solubilized in 0.4 N NaOH. Radioactivity remaining in the cells was quantified by scintillation counting. The total binding medium contained 4 x 10-7 M [3H]ouabain in Ca2+, K+-free Hanks' solution. This concentration of ouabain was demonstrated to be high enough to saturate binding sites (data not shown). Nonspecific binding was measured in the presence of 10-4 M unlabeled ouabain. Nonspecific binding was <2% of total binding. Specific ouabain binding was defined as the difference between total binding and nonspecific binding. Binding assays done in transwells were essentially the same as those in plastic dishes, except the cells were incubated at room temperature, and 2 mM CaCl2 was included in both binding media and washing solution to preserve the tight junctions of the epithelial cells. When apical binding assays were performed, the medium from the basolateral side was checked for radioactivity to ensure no leakage of [3H]ouabain and vice versa.
Rb uptake assay. The cells were placed down at 5 x 105 cells/well in a 24-well plate. 86Rb was used as a cognate for measuring K+ uptake. After the tight junctions were disrupted, the cells were washed twice with Ca2+-free Hanks' solution containing a suboptimal K+ concentration (2 mM KCl) and incubated with the same solution (total uptake) or plus 0.1 mM ouabain (background uptake) at 22°C for 30 min to saturate ouabain binding sites. The uptake was measured by adding 86Rb and incubating for 15 min at 22°C. This period of incubation was within the linear uptake range (data not shown). Rb+/K+ transport mediated by the Na,K-ATPase is the difference between total and background uptakes.
Na,K-ATPase activity assay. The cells were placed down at 5 x 105 cells/well in a 24-well plate. After treatments, the confluent cells were washed twice with a hypotonic solution (1 mM MgCl2, 0.25 mM EDTA, 0.1% bovine serum albumin, and 1 mM imidazole, pH 7.4) and then incubated with the assay buffer (130 mM NaCl, 20 mM KCl, 4 mM MgCl2, 1 mM EGTA, 3 mM NaAzid, 0.1% saponin, and 30 mM imidazole-HCl, pH 7.4) or plus 10-4 M ouabain in background assays at 37°C for 20 min. Reactions were started by adding 3 mM ATP and incubated at 37°C for an additional 45 min. The release of inorganic phosphate (Pi) by the enzyme was within the linear range during the incubation period (data not shown). The liberated Pi was measured as absorption at 850 nm by the method of Baginski et al. (1). The Na,K-ATPase activity was defined as the difference between the total ATPase activity and the background ATPase activity and accounted for about 35% of the total ATPase activity in most cases.
ROS measurement. Separation of confluent monolayers of MDCK cells requires prolonged exposure to trypsin, which causes damage to the cells. Thus, for these experiments, cell suspensions trypsinized from plates of subconfluent cultures with 1 x 106 cells/assay were used. The cells were preloaded with 20 µM freshly prepared 2'7'-dichlorofluorescin diacetate (DCF-DA) in phenol red-free DMEM in some cases also with antioxidants at 37°C for 30 min, washed with 0.9% NaCl to remove K+, and then treated with phenol red-free control or low-K+ medium. After treatments, the cells were directly analyzed by flow cytometry with excitation at 475 nm and emission at 525 nm, using software System II (Coulter, Miami, FL). The gate was appropriately set to distinguish oxidant-stressed cells from non-oxidant-stressed cells. The reading in the first control was arbitrarily assigned a value of 100%. The rest of the data were normalized to this value (33). ROS signals were also acquired by fluorescence microscopy (Zeiss EL-Einsatz) with the software IntelligentImaging.
Western analysis. The cells were placed down at 8 x
105 cells/well in a 12-well plate. After treatments, the confluent
cells were rinsed with ice-cold phosphate-buffered saline (PBS) and scraped
with a rubber policeman in a loading buffer supplemented with 5%
-mercaptoethanol and 0.1 mg/ml phenylmethysulfonyl fluoride (PMSF) and
0.04 µg/ml aprotinin. Samples were loaded into 10% sodium dodecyl
sulfate-polyacrylamide gel in 12 µg/lane, resolved by electrophoresis, and
electrophoretically blotted onto PVDF membranes. The membranes were first
hybridized with primary antibodies and then with horseradish
peroxidase-conjugated anti-mouse or anti-rabbit IgG secondary antibody
(Sigma). Antibody binding was visualized by the enhanced chemiluminescence
method (Amersham, UK) (32) and
semi-quantified by the software GelPro. The antibody against the
1-subunit of the Na+,K+-ATPase was a
generous gift from Dr. Thomas A. Pressley (Texas Tech University Health
Sciences Center) (14).
Antibodies against
1-subunit and GAPDH were purchased from
Upstate Biotechnology (Lake Placid, NY) and RDI Research Diagnostics
(Flanders, NJ), respectively.
Transfection and chloramphenicol acetyl transferase assay. The cells were placed down at 6 x 105 cells/60 mm dish overnight and then transfected with chloramphenicol acetyl transferase (CAT) expression plasmid constructs mixed with a vector pCB6 that confers G418 resistance. LipofectAMINE was used according to the manufacturer's instructions (GIBCO BRL), and the cells were selected in growth medium supplemented with 600 µg/ml G418. For CAT assays, the cells were placed down at 8 x 105/well in a 12-well plate. After treatments, the cells were transferred into an Eppendorff tube with the lysis buffer included in the assay kit. A freezing and thawing method was used to rupture the cells. CAT assays were performed according to the manufacturer's procedures (Promega, Madison, WI).
Statistical analyses. Data are expressed as means ± SE. Statistical analyses were performed using analysis of variance (ANOVA) or Student t-test as appropriate. Post hoc comparisons were made by Dunnett's test. P < 0.05 was considered significant.
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RESULTS |
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Low K+ stimulates ROS activity. Low K+ also increased the intracellular ROS activity in a dose- and time-dependent manner as determined by DCF-DA-based flow cytometry (Fig. 2). Low K+ rapidly increased ROS activity, and the effect of low K+ reached the peak within 10 min. Catalase inhibits Cr(VI)-induced generation of ROS and activation of p53 in a human lung epithelial cell line (24). Catalase or NAC, another well-known antioxidant, blocked the effect of low K+ on ROS production (Fig. 2B). Low-K+-induced ROS activity was also suppressed by 5 µM diphenylene iodonium, an inhibitor of the NADPH oxidase (Fig. 3).
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ROS mediate the effect of low K+ on the
Na,KATPase. The increased ROS activity induced by low K+ that
occurs at early stages implies that ROS may be required for the stimulation of
the Na,K-ATPase gene expression. To address this issue, the cells were first
stably transfected with expression constructs in which the reporter gene CAT
was under the control of the avian Na,K-ATPase -subunit 1.9-kb and
900-bp 5'-flanking regions that have a negative regulatory element
(31). Because the CAT activity
in the mixture of transfectants was not detectable, subcloning was performed,
and positive clones were identified by CAT activity. Low K+
increased the CAT expression in both constructs. Catalase or NAC inhibited the
effect of low K+ but had no significant effect on basal CAT
activity (Fig. 4). To determine
whether the increased CAT activity induced by low K+ was mediated
through releasing the repressive effect or a direct stimulation of the
promoter, the cells were transfected with a CAT expression construct that is
directed by a 96-bp promoter fragment that is located immediately upstream to
the coding region and has no negative regulatory element
(31). Because the mixture of
G418-resistant cells displayed CAT activity, CAT assays were conducted
directly in the pool of the G418-resistant cells without subcloning. Low
K+ also augmented the CAT activity expressed by this construct,
implying that the effect of low K+ is due to the direct stimulation
of the promoter activity. More importantly, both catalase and NAC abolished
the low-K+-induced CAT activity, which suggests that ROS mediate
the stimulation of low K+ on the
-subunit promoter
(Fig. 4).
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Low K+ elevates the 1- and
-subunit mRNA
levels associated with increased protein abundance of the enzyme
(3,
16). To test whether the
antioxidants also suppressed the effect of low K+ at the protein
level, Western analyses were employed first to determine the linear
relationship between protein loading and the chemiluminescence signal.
Figure 5, A and
B, shows that the amount of protein loading between 6 and
15 µg was within the linear range of both
1- and
1-subunit detections. Next, the same analyses were used to
examine the effect of antioxidants. As shown in
Fig. 5, C and
D, catalase or NAC inhibited low-K+-induced
increases in
1- and
1-subunit protein
abundance. The
1-subunit showed a bigger increase than the
1-subunit. It is possible that the
1-subunit is more responsive than the
1-subunit. The
1-subunit also exhibited a
greater response to hyperoxia than the
1-subunit in MDCK
cells (25). In a low
electrical field (20 volts), the
1-antibody recognized four
closely migrated faint bands that most probably result from different stages
of glycosylation (22). In a
high electrical field (40 volts), only one band was detected as shown in
Fig. 5. Inhibition of
low-K+-induced increases in protein abundance by the antioxidants
would be expected to result in inhibition of low-K+-induced
increases in cell surface presence of Na,K-ATPase molecules. To test this
possibility, the effect of low K+ on ouabain binding sites was
examined in the presence of catalase or NAC. Catalase at 500 U/ml or NAC at 35
mM almost completely inhibited the effect of low K+, whereas
catalase or NAC had no significant effect under the control conditions
(Fig. 6, A and
B). NAC at 35 mM did not induce cell death. Similarly,
diphenylene iodonium also abolished the effects of low K+ on
ouabain binding sites (Fig.
6C).
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DISCUSSION |
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By CAT activity assays, Yu et al.
(31) have identified that the
avian -subunit gene contains a repressive region located upstream to
the promoter region. Gel retardation assays reveal the existence of a set of
proteins binding to this repressive region, although the binding amount of
each individual protein varies in different types of cells
(31). Low K+ not
only increased CAT activity directed by the avian 1.9-kb and 900-bp
5'-flanking regions that have the negative regulatory element, but also
the CAT expression controlled by the 96-bp promoter region that does not
contain the negative regulatory element. More importantly, both catalase and
NAC abrogated the effect of low K+ in every construct
(Fig. 4). These data suggest
that the stimulatory effect of low K+ on the
-subunit gene
expression results from the direct stimulation of the promoter activity
instead of from lifting repressors and that ROS mediate this stimulation.
Substantial evidence suggests that the transient production of ROS is an
important signaling event that mediates a variety of agonist-induced gene
expressions, most of which are related to cell hypertrophy and proliferation.
ROS stimulate the promoter activity of a gene directly or through other
signaling cascades, like protein tyrosine phosphorylation and MAP kinases
(5). The effect of low
K+ is mitogenic (3).
The present finding is within the purview of this paradigm.
However, how ROS stimulate the -subunit promoter activity remains
unknown. ROS regulate activities of several transcription factors, most
notably nuclear factor-
B (NF-
B) and activator protein-1 (AP-1).
Although the 5'-flanking region of the
-subunit gene has six AP-1
and AP-2 binding sites, none of these is located in this 96-bp region. Rather,
this region is nucleotide GC-rich with five putative Sp-1 targeted sequences
(31). The Sp-1 binding site is
present in both
- and
-subunit promoters across isoforms and
species (10,
20,
21). In rat cardiac myocytes,
the increased
1-subunit gene transcription by low
K+ is mediated by increasing Sp-1/Sp-3 binding to the promoter
region (34). Sp-1 is known to
be modulated by the redox state. Moreover, upregulation of the
1-subunit of the Na,KATPase by hyperoxia in MDCK cells is
mediated by stimulation of Sp-1/Sp-3 activity
(25). It is plausible to
speculate that low K+ increases the activity of ROS that enhance
the binding of Sp-1/Sp-3 to the promoter, thereby increasing transcription of
the Na,K-ATPase gene.
The discovery of the inhibitory effect of catalase and NAC on low-K+-induced increases in Na,K-ATPase protein abundance and ouabain binding sites is somewhat surprising. In contrast to ARL15 cells, a rat hepatoma cell line, in which low K+ induces a constant elevation of the Na,K-ATPase mRNA level (16), in MDCK cells, a previous report (3), in addition to our own studies (data not shown), reveals that low K+ only transiently increases the Na,K-ATPase mRNA abundance. The return of the mRNA abundance to the basal level within 3 h of exposure to low-K+ medium implies that transcriptional regulation may not be significant in the effect of low K+ on the Na,K-ATPase protein abundance. However, the inhibition of the Na,KATPase protein abundance by the antioxidants argues against this possibility. In outer medullary kidney tubules or chick skeletal muscle, ouabain- or veratridine-induced increases in Na,K-ATPase activity is constituted by two steps: 1) increased protein synthesis at an early stage and 2) decreased protein degradation at a late stage (18, 26). It is not clear whether ROS are also involved in stabilizing the Na,K-ATPase protein.
Low K+ not only stimulates the biosynthesis of the Na,K-ATPase but also induces general protein synthesis (3, 16, 29). The Na,K-ATPase mediates a ouabain-induced mitogenic effect in cardiac myocytes by interacting with neighboring membrane proteins like Src and epidermal growth factor receptors, which send messages to nuclei through organized cytosolic cascades of signaling events. Among those signaling events, ROS and intracellular Ca2+ are essential second messengers (27). Whether the Na,K-ATPase also acts as a signal transducer that mediates the pleiotropic effect of low K+ remains to be determined.
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DISCLOSURES |
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ACKNOWLEDGMENTS |
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FOOTNOTES |
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The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
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REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() |
---|
2. Boardman L, Huett M, Lamb JF, Newton JP, and Polson JM. Evidence for the genetic control of the sodium pump density in HeLa cells. J Physiol 241: 771794, 1974.[ISI][Medline]
3. Bowen JW and
McDonough A. Pretranslational regulation of Na,K-ATPase in cultured canine
kidney cells by low K+. Am J Physiol Cell
Physiol 252:
C179C189, 1987.
4. Buffin-Meyer B, Verbavatz JM, Cheval L, Marsy S, Younes-Ibrahim M, Le Moal C, and Doucet A. Regulation of Na+, K+-ATPase in the rat outer medullary collecting duct during potassium depletion. J Am Soc Nephrol 9: 538550, 1998.[Abstract]
5. Finkel T. Redox-dependent signal transduction. FEBS Lett 476: 5254, 2000.[ISI][Medline]
6. Hayashi M and
Katz AI. The kidney in potassium depletion. I.
Na+-K+-ATPase activity and [3H]ouabain binding in MCT.
Am J Physiol Renal Fluid Electrolyte Physiol
252: F437F446,
1987.
7. Heaton JH and Gelehrter TD. Derepression of amino acid transport by amino acid starvation in rat hepatoma cells. J Biol Chem 252: 29002907, 1977.[Abstract]
8. Hume SP and Lamb JF. Proceedings: effect of growth in various concentrations of amino acids on the properties of the A-mediated amino acid uptake system in cultured cells. J Physiol 239: 46P47P, 1974.[Medline]
9. Imbert-Teboul M, Doucet A, Marsy S, and Sianne-Perez S.
Alteration of enzymatic activity along rat collecting tubule in potassium
depletion. Am J Physiol Renal Fluid Electrolyte
Physiol 253:
F408F417, 1987.
10. Ikeda K, Nagano
K, and Kawakami K. Anomalous interaction of Sp1 and specific binding of an
E-box-binding protein with the regulatory elements of the Na,K-ATPase
2 subunit gene promoter. Eur J
Biochem 218:
195204, 1993.[Abstract]
11. Kletzien RF and Perdue JF. Induction of sugar transport in chick embryo fibroblasts by hexose starvation. Evidence for transcriptional regulation of transport. J Biol Chem 250: 593600, 1975.[Abstract]
12. Lescale-Matys L, Hensley CB, Crnkovic-Markovic R, Putman DS, and
McDonough AA. Low K+ increases Na,KATPase abundance in
LLC-PK1/CL4 cells by differentially increasing , and not
,
subunit mRNA. J Biol Chem 265:
1793517940, 1990.
13. McDonough AA,
Magyar CE, and Komatsu Y. Expression of
Na+-K+-ATPase - and
-subunits along rat
nephron: isoform specificity and response to hypokalemia. Am J
Physiol Cell Physiol 267:
C901C908, 1994.
14. Pressley TA. Phylogenetic conservation of isoform-specific
regions within -subunit of Na+-K+-ATPase.
Am J Physiol Cell Physiol 262:
C743C751, 1992.
15. Pressley TA, Haber RS, Loeb JN, Edelman LS, and Ismail-Beigi F. Stimulation of Na,K-activated adenosine triphosphatase and active transport by low external K+ in a rat liver cell line. J Gen Physiol 87: 591606, 1986.[Abstract]
16. Pressley TA,
Ismail-Beigi F, Gick GG, and Edelman IS. Increased abundance of
Na,K-ATPase mRNA in response to low external K. Am J Physiol Cell
Physiol 255:
C252C260, 1988.
17. Rayson BM.
Calcium: a mediator of the cellular response to chronic
Na+/K+-ATPase inhibition. J Biol
Chem 268:
88518854, 1993.
18. Rayson BM.
Rates of synthesis and degradation of Na+-K+-ATPase
during chronic ouabain treatment. Am J Physiol Cell
Physiol 256:
C75C80, 1989.
19. Salter DW and Cook JS. Reversible independent alterations in glucose transport and metabolism in cultured human cells deprived of glucose. J Cell Physiol 89: 143155, 1976.[ISI][Medline]
20. Shull MM, Pugh
DG, and Lingrel JB. The human Na, KATPase 1 gene:
characterization of the 5'-flanking region and identification of a
restriction fragment length polymorphism. Genomics
6: 451460,
1990.[ISI][Medline]
21. Takeyasu K,
Hamrick M, Barnstein AM, and Fambrough DM. Structural analysis and
expression of a chromosomal gene encoding an avian
Na+/K+-ATPase . Biochim Biophys
Acta 1172:
212216, 1993.[ISI][Medline]
22. Tamkun MM and
Fambrough DM. The Na+/K+-ATPase of chick sensory
neurons. Studies on biosynthesis and intracellular transport. J
Biol Chem 261:
10091019, 1986.
23. Taormino JP and
Fambrough DM. Pre-translational regulation of the
Na+/K+-ATPase in response to demand for ion transport in
cultured chicken skeletal muscle. J Biol Chem
265: 41164123,
1990.
24. Wang S, Leonard
SS, Ye J, Ding M, and Shi X. The role of hydroxyl radical as a messenger
in Cr(VI)-induced p53 activation. Am J Physiol Cell
Physiol 279:
C868C875, 2000.
25. Wendt CH, Gick
G, Sharma R, Zhuang Y, Deng W, and Ingbar DH. Up-regulation of Na,K-ATPase
1 transcription by hyperoxia is mediated by SP1/SP3 binding.
J Biol Chem 275:
4139641404, 2000.
26. Wolitzky BA and
Fambrough DM. Regulation of the Na+,K+-ATPase in
cultured chick skeletal muscle modulation of expression by the demand for ion
transport. J Biol Chem 261:
99909999, 1986.
27. Xie Z and
Askari A. Na+/K+-ATPase as a signal transducer.
Eur J Biochem 269:
24342439, 2002.
28. Xie Z,
Kometiani P, Liu J, Li J, Shapiro JI, and Askari A. Intracellular reactive
oxygen species mediate the linkage of Na+/K+-ATPase to
hypertrophy and its marker genes in cardiac myocytes. J Biol
Chem 274:
1932319328, 1999.
29. Xie Z, Liu J, Malhotra D, Sheridan T, Periyasamy SM, and Shapiro JI. Effects of hypokalemia on cardiac growth. Ren Fail 22: 561572, 2000.[ISI][Medline]
30. Yin W, Jiang G, Takeyasu K, and Zhou X. Stimulation of Na,K-ATPase by low potassium is dependent on transferrin. J Membr Biol. In press.
31. Yu HY,
Nettikadan S, Fambrough DM, and Takeyasu K. Negative transcriptional
regulation of the chicken Na+/K+-ATPase
1-subunit gene. Biochim Biophys Acta
1309: 239252,
1996.[ISI][Medline]
32. Zhou X and Fambrough DM. Expression of the avian Na,KATPase subunits in Dictyostelium discoideum. J Membr Biol 167: 1924, 1999.[ISI][Medline]
33. Zhou X, Zhao A,
Goping G, and Hirszel P. Gliotoxin-induced cytotoxicity proceeds via
apoptosis and is mediated by caspases and reactive oxygen species in LLC-PK1
cells. Toxicol Sci 54:
194202, 2000.
34. Zhuang Y, Wendt
C, and Gick G. Regulation of Na,K-ATPase 1 subunit gene
transcription by low external potassium in cardiac myocytes. Role of Sp1 AND
Sp3. J Biol Chem 275:
2417324184, 2000.