ACCELERATED PUBLICATION
Oxygen Sensing and HIF-1 Activation Does Not Require an
Active Mitochondrial Respiratory Chain Electron-transfer Pathway*
Vickram
Srinivas
,
Irene
Leshchinsky,
Nianli
Sang,
Michael P.
King§,
Alex
Minchenko, and
Jaime
Caro
From the Department of Medicine, Cardeza Foundation for Hematologic
Research and § Department of Biochemistry and Molecular
Pharmacology, Jefferson Medical College of Thomas Jefferson University,
Philadelphia, Pennsylvania 19107-5099
Received for publication, April 9, 2001
 |
ABSTRACT |
Hypoxia induces the
stabilization and transcriptional activation of the hypoxia-inducible
factor 1
(HIF-1
) protein, the regulatory member of the HIF-1
complex. The molecular mechanisms that are responsible for oxygen
sensing and the downstream pathways utilized by the hypoxic signal are
still poorly understood. One hypothesis for oxygen sensing has
postulated that reactive oxygen species generated at mitochondrial
complex III are the initiators of the hypoxic signal. Here we
find that mitochondrial DNA-less (
o) cells
have a normal response to hypoxia, measured at the level of HIF-1
protein stabilization, nuclear translocation, and its transcriptional activation activity. Furthermore, overexpression of
catalase, either in the mitochondria or in the cytosol, fails to modify
the hypoxia response indicating that hydrogen peroxide is not a
signaling molecule in the hypoxic signaling cascade that culminates
with HIF-1 activation.
 |
INTRODUCTION |
In multicellular organisms, oxygen, as the final acceptor of
electrons in the respiratory chain, is an absolute requirement for
life. Hypoxia, the decrease in oxygen supply to tissues, determines a
series of metabolic and systemic adaptations that tend to overcome the
detrimental effect of the lack of oxygen. Central to this adaptation is
the regulation of hypoxia-responsive genes that control multiple
cellular and systemic functions (reviewed in Ref. 1). Most of these
genes are regulated by a common mechanism of transcriptional activation
that involves the hypoxia-inducible factor 1 (HIF-1)1 complex (2). The
HIF-1 complex is composed of two subunits. HIF-1
and HIF-1
,
members of the helix loop helix-PAS family of transcription
factors. The function of the HIF-1 complex is primarily regulated by
the abundance of the HIF-1
subunit. Although HIF-1
is
constitutively expressed in normoxic cells, the HIF-1
subunit is
only detectable in hypoxic cells. Under normoxic conditions, the
HIF-1
protein is ubiquitinated and rapidly degraded by the proteasomal system (3, 4). The ubiquitination process depends on its
interaction with the von Hippel Lindau protein (VHL) that acts as a
ubiquitin protein ligase (5-7). Hypoxia, transition metals, and iron
chelators inhibit this degradation process and allow for HIF-1
accumulation and formation of the transcriptionally active complex.
Significantly, HIF-1
responses are very rapid and occur at levels of
hypoxia that are well within the physiological range (8). Furthermore,
the induction of HIF-1
during hypoxia is practically instantaneous,
with protein HIF-1
detected in the nucleus as early as 2 min after
exposure to low oxygen (9).
Despite the advances in the area of gene regulation by hypoxia, the
molecular mechanisms that are responsible for oxygen sensing and the
downstream pathways utilized by the hypoxic signal are still poorly
understood. In biological systems, oxygen reactivity is usually
associated with metals, most notably iron, as it intervenes in two
types of reactions in the following ways: (a) as an acceptor of electrons in redox reactions, or (b) in liganding to heme
groups. In the latter case, as it happens in its interactions with
hemoglobin, there is no transfer of electrons. This last property
suggested to Goldberg et al. (10) that oxygen sensing
in vertebrates may utilize a rapidly turning over heme-containing
protein, such that oxygen-dependent changes in conformation
would initiate the hypoxia response. However, inhibitors of heme
synthesis had no significant affect on hypoxia response (11, 12). Other
models for oxygen sensing are based on electron transfer or redox
reactions. In redox reactions, oxygen acts as the acceptor of electrons
and may result in generation of reactive oxygen species (ROS). Based on
the stimulatory effect of iron chelators on HIF-1
activation, Srinivas et al. (11) proposed that iron may be interacting
directly, or through an intermediate iron-binding protein, with
HIF-1
by inducing localized oxidative reactions that would act as a
signal for degradation. However, no such protein has yet been found
(11). Acker and colleagues (13, 14) have postulated that a low output NADPH oxidase, similar to the one present in neutrophils, could act as
an oxygen sensor. In this model, hydrogen peroxide would be
continuously generated by the oxidase in an
oxygen-dependent manner and would have a continuous
negative tonic effect on HIF-1
survival. During hypoxia, the
decrease in peroxide production would result in HIF-l
accumulation.
However, evidence against this model is the finding that diphenylene
iodonium (DPI), an NADPH oxidase inhibitor, decreases rather than
stimulates the hypoxia response (15). Chandel et al. (16)
have proposed another redox model of oxygen sensing based on the
production of ROS by the mitochondria. These investigators have
reported that during hypoxia there is an increase in superoxide
production at the level of complex III of the respiratory electron
transfer chain. The increase in ROS production would be proportional to
the degree of hypoxia and would be, following dismutation to hydrogen
peroxide, the starting point of the hypoxic signal. Experimental
support for their model is the unresponsiveness to hypoxia of cells
lacking mitochondrial DNA,
o cells. Furthermore,
blocking of electron transfer at complex I by rotenone resulted in
inhibition of the hypoxia response. However, other studies had found
that inhibitors of cytochrome c oxidase, like cyanide and
azide, had no effect on the hypoxia response and suggested that the
mitochondria was not involved in oxygen sensing. The results by Chandel
et al. (16) prompted us to re-evaluate the role of
mitochondria in HIF-1
activation. Our results indicate that the
integrity of the mitochondrial respiratory chain is not necessary for
hypoxia response. Furthermore, they show that hydrogen peroxide is not
an intermediary molecule involved in oxygen sensing.
 |
MATERIALS AND METHODS |
Cell Lines and Culture Conditions--
HeLa cells were obtained
from ATCC and grown in high glucose DMEM (Mediatech, Herndon, VA)
supplemented with 10% fetal bovine serum, penicillin, and
streptomycin. HepG2 cells and its derivative catalase overexpressing
cells Mc5 (mitochondrial localization) and C33 (cytoplasmic
localization) were obtained from Dr. Cederbaum (Mt. Sinai School of
Medicine, New York, NY) and grown in minimal essential medium
(Mediatech, Herndon, VA) supplemented with 10% fetal bovine serum
(Hyclone, Logan, UT), penicillin (100 units/ml), and streptomycin (100 µg/ml) (17). Mitochondrial DNA-less (
o cells) were
prepared by prolonged exposure to ethidium bromide as described by King
and Attardi (18) and grown in DMEM supplemented with 5% or 15% fetal
bovine serum, penicillin, streptomycin, sodium pyruvate (1 mM), and uridine (50 µg/ml). Cells were maintained at
saturation humidity at 37 °C in 5% CO2, 95% air. For
hypoxic stimulation, the cultures were flushed in a modular incubator (Billups-Rothenburg, Del Mar, CA) with a gas mixture of 0.5%
O2, 5% CO2, and nitrogen was balanced. Cells
were transfected with LipofectAMINE Plus (Life Technologies,
Inc.) as described previously (19). Expression vectors used in
the functional Luciferase assays have been described previously
(19).
Nuclear Extracts, Electrophoretic Mobility Shift Assay, and
Immunoblot Analysis--
Nuclear extracts were prepared from normal or
treated cells as described previously (19). Electrophoretic mobility
shift assay was performed by incubating 10 µg of nuclear extract with 32P-labeled double stranded oligonucleotide probe from the
human HIF-1 Epo enhancer as described (4). For immunoblot analysis, equal amounts of nuclear extracts (10-20 µg of protein) were
fractionated by SDS-polyacrylamide gel electrophoresis and transferred
to polyvinylidene difluoride membranes prior to detection using a
stabilized alkaline phosphatase system (Promega, Madison, WI) (19).
Monoclonal anti-HIF-1
antibody was purchased from Transduction
Laboratories (Lexington, KY), and alkaline phosphatase-conjugated
anti-mouse secondary antibody was purchased from Southern
Biotechnology (Birmingham, AL).
Indirect Immunofluorescence Microscopy--
Cells growing on
glass slides were fixed with 4% paraformaldehyde in PBS (pH 8.0) for
10 min and then washed with PBS three times. Subsequently, they were
permeabilized with 0.5% Triton X-100 in PBS for 5 min and rinsed again
three times with PBS. After blocking nonspecific binding with 10%
fetal calf serum in PBS for 30 min, the cells were incubated with a
monoclonal anti-HIF-1
antibody (Novus Biologicals, Littleton, CO).
Primary antibody was detected by incubation with an fluorescein
isothiocyanate-conjugated rabbit anti-mouse antibody (Southern
Biotechnology, Birmingham, AL) at room temperature for 30 min.
Following extensive washing in PBS and mounting in Miowol solution
(Calbiochem), the cells were visualized and photographed utilizing a
fluorescence microscope (Nikon).
RNA Isolation and Ribonuclease Protection Assays--
Total RNA
was isolated with the Trizol reagent (Life Technologies, Inc.)
according to the manufacturer's instructions. RNase protection assays
was performed utilizing a 32P-labeled single stranded
vascular endothelial growth factor (VEGF) probe as described
previously (20).
 |
RESULTS |
Activation of HIF-1
by Hypoxia in
o
Cells--
Mitochondria are the center of oxidative phosphorylation, a
process by which ATP is formed as a result of the transfer of electrons
from NADH or FADH2 to O2 by a series of
electron carriers. The mammalian mitochondrial DNA encodes thirteen
proteins, all subunits of the respiratory chain enzymes, including
critical catalytic subunits for complex I (NADH dehydrogenase), complex III (bcl complex), complex IV (cytochrome c oxidase), and
ATP synthetase. Elimination of mitochondrial DNA from cells by
prolonged ethidium bromide treatment results in the absence of
respiratory chain electron transfer and mitochondrial ATP production by
oxidative phosphorylation (18). Cells lacking mitochondrial DNA can
survive, however, when provided with external sources of pyrimidines
and pyruvate (18). We utilized two independently generated
o cell lines, one derived from the human osteosarcoma
cells (143B206) (18) and one from a human fibroblast cell line (GM 701)
(21). The
o cells and the corresponding parental cell
lines were subjected to hypoxia (0.5% O2) for 4-6 h, and
the nuclear extracts were analyzed for HIF-1
expression by Western
blots and for HIF-1 complex formation by electrophoretic mobility shift
analysis. As shown in Fig. 1,
a and b, exposure to hypoxia and desferrioxamine results in increased expression of HIF-1
protein in both
o cell lines, their parental wild types, and HeLa cells.
Similarly, hypoxia, desferrioxamine, and cobalt induced the formation
of the HIF-1 DNA binding complexes, as measured by gel shift assays (Fig. 1c). Furthermore, rotenone, an inhibitor of
mitochondrial complex I, had no effect on the hypoxia responses (not
shown).

View larger version (52K):
[in this window]
[in a new window]
|
Fig. 1.
Expression of HIF-1a in
o cells.
Parental (WT) and o cells were exposed to
hypoxia (Hx; 0.5% O2), desferrioxamine
(Df; 130 µM), or cobalt (Co; 100 µM) for 4-6 h, and nuclear extracts were analyzed by
Western blots (a) and (b) or electrophoretic
mobility shift assays (c). N, normoxia.
|
|
The activation of HIF-1
expression by hypoxia in
o
cells was confirmed by immunofluorescence and functional studies. Fig. 2a shows the induction of
nuclear fluorescence, detected with an anti-HIF-1
monoclonal
antibody, in parental 143B (wild type) and
o 143B206
cells exposed to hypoxia (2 and 5) or treated with desferrioxamine (3 and 6). To test whether the transcriptional activation of HIF-1
by
hypoxia was conserved in
o cells, we utilized an
expression vector containing the transcriptionally active C-terminal
domain of HIF-1
protein (amino acids 740 to 826) fused to the
DNA-binding domain of the yeast transactivator GAL-4 (19).
Cotransfection of this plasmid with a luciferase reporter containing a
GAL-4 DNA-binding site showed that hypoxia induced the
transactivational activity of HIF-1
, both in
o and
wild type cells (Fig. 2b). Furthermore, confirmation of the hypoxia responsiveness of
o cells was obtained by the
hypoxic activation of the VEGF gene as shown in the RNase protection
assay in Fig. 2c.

View larger version (67K):
[in this window]
[in a new window]
|
Fig. 2.
Functional activity of
HIF-1 in
o cells. a,
nuclear translocation. Wild type (WT) and o
cells were maintained in normoxia (1 and 4) or
exposed to hypoxia (2 and 5) or desferrioxamine
(3 and 6) for 2-4 h and immunostained with an
anti HIF-1 monoclonal antibody. b, transcriptional
activity. Wild type and o cells were transfected with a
GAL-4-HIF-1 fusion (amino acids 740-826) and a GAL-4-DNA-binding
domain luciferase expression plasmid and maintained in normoxia
(N) or exposed to hypoxia (Hx). Results are
expressed as the mean of arbitrary luciferase units per µg of
protein ± S.D. c, RNase protection assay for Northern
blot VEGF expression in o cells exposed to hypoxia
(Hx) or desferrioxamine (Df).
|
|
Effect of Overexpression of Catalase on HIF-1
Expression--
Most of the currently proposed redox models for oxygen
sensing suggest that H2O2 is the intermediary
signal molecule in the hypoxia response. To evaluate the role of
hydrogen peroxide in oxygen sensing, we utilized cell lines that
overexpress the enzyme catalase in either the mitochondrial or
cytosolic compartments. These cell lines were produced by introducing a
catalase cDNA or a catalase cDNA with a superoxide dismutase
mitochondrial leader sequence into a human hepatoma cell line (HepG2
cells). Published results (17) have shown that these cell lines are
more resistant to H2O2-, menadione-, or
antimycin A-induced apoptosis as compared with cells transfected with
the vector alone. After exposure to hypoxia and desferrioxamine of
control and catalase-overexpressing cells, the induction of HIF-1
was measured by Western blots and electrophoretic motility shift
assays. Fig. 3a shows that
overexpression of catalase, either in the cytosol (C33 cells) or in the
mitochondria (Mc5 cells), does not affect the response to hypoxia or
change the basal expression of HIF-1
protein. Similar results were
obtained using the gel shift assay (not shown). The effect of catalase overexpression on the transcriptional activity of HIF-1
was
evaluated by using the GAL-4-HIF-1
fusion system, as
described above. Fig. 3b indicates that overexpression of
catalase does not affect the transcriptional response to hypoxia (Fig.
3d).

View larger version (26K):
[in this window]
[in a new window]
|
Fig. 3.
Hypoxia responses in catalase overexpressing
cells. a, expression of HIF-1 . Control HepG2 cells
or cells overexpressing catalase in the cytosol (C33) or in
the mitochondria (MC5) were exposed to normoxia
(N), desferrioxamine (Df), or hypoxia
(Hx) for 2-4 h, and nuclear extracts were subjected to
Western blot analysis. b, HIF-1 transcriptional activity.
Control and catalase overexpressing cells were transfected, as
described in Fig. 2, and exposed to normoxia (N) or hypoxia
(Hx) for 4 h. Results are expressed as arbitrary
luciferase units per µg of protein ± S.D.
|
|
 |
DISCUSSION |
Early work showing that cyanide and sodium azide do not affect the
hypoxia response suggested that the mitochondrial respiratory chain was
not involved in oxygen sensing (22). However, Chandel et al.
(16) have proposed that an increase in superoxide production at the
level of mitochondrial complex III (quinol-cytochrome c oxidoreductase) is the initial event that triggers the hypoxia response
(16). Superoxide is produced during normal mitochondrial function, and
it is rapidly converted to H2O2, a readily
diffusible molecule (23). A compelling argument for their hypothesis
was the finding that
o cells, which lack mitochondrial
DNA, have an impaired response to hypoxia. In the work presented here,
we have utilized two independent human
o cell lines
created by prolonged exposure to low doses of ethidium bromide. This
treatment results in inhibition of mitochondrial DNA replication
without affecting the nuclear DNA (18). As mitochondrial DNA encodes
essential subunits of the respiratory chain and ATP synthetase,
o cells have no mitochondrial electron transfer and are
completely dependent on glycolysis for the generation of ATP.
Furthermore, these cells depend on exogenous sources of pyrimidines
(pyrimidine auxotrophy). The need for pyrimidine is explained by the
fact that the enzyme dihydrooratate dehydrogenase is located in the mitochondrial inner membrane and requires mitochondrial electron transfer for normal function. In the presence of pyruvate and pyrimidines, these cell lines can maintain almost normal cell growth
rates. One of the cell lines (143B206) was derived from an osteosarcoma
cell line and the other (701-2c) from an SV-40 T-antigen-transformed
fibroblast cell line (18, 21). Oxygen consumption in the
o cells was found to be less than 5% of the control
cells, and no cytochrome c oxidase activity was detected
(24). The
o status of both cell lines was confirmed by
their inability to grow in pyruvate- and pyrimidine-deficient media and
by DNA hybridization studies showing no traces of mitochondrial DNA.
Our results with the
o cells indicate that an active
mitochondrial respiratory electron transport chain is not an absolute
requirement for the activation of HIF-1 protein in response to hypoxia.
Furthermore, we find that in both cell lines the functional activity of
the HIF-1 complex, its nuclear translocation, and its hypoxia-induced
transactivation activity are preserved, as well. The differences
between our results and those of Chandel et al. (16) are
unclear and may derive from differences in the cell lines or the
procedures used to establish the
o cells. It should be
noted that in the preparation of
o cells, Chandel
et al. (16) utilized a selection that included growth in the
presence of rotenone and antimycin. It is not known whether this
selection step may affect the properties of the cells.
Electron transfer reactions (redox reactions) appear to be an important
component of the mechanisms of oxygen sensing. Acker (13) and Fandrey
et al. (14) have postulated that a low output NADPH oxidase,
similar to the one present in granulocytes, may initiate the hypoxic
response. In their model, the continuous production of superoxide by
this NADPH oxidase would act as an active negative signal to induce
HIF-1
degradation. During hypoxia, the decrease in hydrogen peroxide
would result in increased HIF-1
stability. However, Gleadle et
al. (15) observed that DPI, a flavoprotein inhibitor whose targets
include NADPH oxidases, decreases, rather than enhances, the hypoxia
response (15). Dismutation of superoxide, either spontaneously or
through superoxide dismutase, results in the production of hydrogen
peroxide. This compound is quite soluble and diffusible and has been
postulated as a likely intermediate of the hypoxic signal. Reaction of
H2O2 with iron (Fenton reaction) results in the
generation of hydroxyradicals, which are highly reactive molecules. To
test the role of H2O2 as a signal molecule, we
utilized cell lines that overexpress the enzyme catalase either in the
cytosol or in the mitochondria. These cell lines have been shown to
decompose H2O2 very effectively and to provide
resistance to the apoptotic effects of menadione or antimycin (17).
Overexpression of catalase, either in the cytosol or in the
mitochondria, did not affect the normoxic levels of HIF-1
protein or
its response to hypoxic stimulation. These results cast doubt on the
role of H2O2 as a signal molecule in the
hypoxic response, either as a negative signal, as postulated by Acker
(13) and Fandrey et al. (14), or as a positive signal, as
postulated by Chandel et al. (16).
The strong stimulatory effect of the iron (III) chelator
desferrioxamine in HIF-1
activation suggests that an iron-containing protein(s) is most likely involved in oxygen sensing. In general, iron
(III)-chelating agents are not active against heme-containing enzymes
or iron-sulfur cluster proteins, although they do interfere with
enzymes containing mono-iron or bi-iron centers coordinated to oxygen
ligands, including amino acids hydroxylases, lipoxygenases, and
ribonucleotide reductase (25). As shown by the studies presented here,
the putative oxygen sensor is independent of the mitochondrial respiratory chain activity, and it does not utilize
H2O2 as a signal molecule.
 |
ACKNOWLEDGEMENTS |
We thank Dr. Cederbaum for generously
providing the catalase overexpressing cells. We acknowledge the support
of and thank our secretary, Rosemarie Silvano, and our graphic
illustrator, Drew Likens.
 |
FOOTNOTES |
*
This work was supported in part by Grants 0060194U (to
V. S.) and 9950122N (to J. C.) from the American Heart Association and by National Institutes of Health Grant 1RO1 CA89212-02 (to J. C.).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.
To whom correspondence should be addressed: Cardeza
Foundation for Hematologic Research, 1015 Walnut St., Philadelphia, PA 19107-5099. Tel.: 215-955-5118; Fax: 215-923-3836; E-mail:
Vickram.Srinivas@mail.tju.edu.
Published, JBC Papers in Press, May 7, 2001, DOI 10.1074/jbc.C100177200
 |
ABBREVIATIONS |
The abbreviations used are:
HIF, hypoxia-inducible factor, DPI, diphenylene iodonium, ROS, reactive
oxygen species;
DMEM, Dulbecco's modified Eagle's medium;
PBS, phosphate-buffered saline;
VEGF, vascular endothelial growth
factor.
 |
REFERENCES |
1.
|
Wenger, R. H.
(2000)
J. Exp. Biol.
203,
1253-1263[Abstract/Free Full Text]
|
2.
|
Wang, G. L.,
Jiang, B.-H.,
Rue, E. A.,
and Semenza, G. L.
(1995)
Proc. Natl. Acad. Sci. U. S. A.
92,
5510-5514[Abstract]
|
3.
|
Huang, L. E.,
Gu, J.,
Schau, M.,
and Bunn, H. E.
(1998)
Proc. Natl. Acad. Sci. U. S. A.
95,
7987-7992[Abstract/Free Full Text]
|
4.
|
Salceda, S.,
and Caro, J.
(1997)
J. Biol. Chem.
272,
22642-22647[Abstract/Free Full Text]
|
5.
|
Maxwell, P. H.,
Wiesener, M. S.,
Chang, G. W.,
Clifford, S. C.,
Vaux, E. C.,
Cockman, M. E.,
Wykoff, C. C.,
Pugh, C. W.,
Maher, E. R.,
Ratcliffe, S. C.,
and Vaux, E. C.
(1999)
Nature
399,
271-275[CrossRef][Medline]
[Order article via Infotrieve]
|
6.
|
Tanimoto, K.,
Makino, Y.,
Pereira, T.,
and Poellinger, L.
(2000)
EMBO J.
19,
4298-4309[Abstract/Free Full Text]
|
7.
|
Ohh, M.,
Park, C. W.,
Ivan, M.,
Hoffman, M. A.,
Kim, T. Y.,
Huang, L. E.,
Pavletich, N.,
Chau, V.,
and Kaelin, W. G.
(2000)
Nat. Cell Biol.
2,
423-427[CrossRef][Medline]
[Order article via Infotrieve]
|
8.
|
Jiang, B.-H.,
Semenza, G. L.,
Bauer, C.,
and Marti, H. H.
(1996)
Am. J. Physiol.
271,
C1172-C1180[Abstract/Free Full Text]
|
9.
|
Jewell, U. R.,
Kvietikova, I.,
Scheid, A.,
Bauer, C.,
Wenger, R. H.,
and Gassman, M.
(2001)
FASEB
15,
1312-1314[Abstract/Free Full Text]
|
10.
|
Goldberg, M. A.,
Dunning, S. P.,
and Bunn, H. F.
(1988)
Science
242,
1412-1415[Medline]
[Order article via Infotrieve]
|
11.
|
Srinivas, V.,
Zhu, X.,
Salceda, S.,
Nakamura, R.,
and Caro, J.
(1998)
J. Biol. Chem.
273,
18019-18022[Abstract/Free Full Text].; correction 274, 1180, 1999
|
12.
|
Horiguchi, H.,
and Bunn, H. F.
(2000)
Biochim. Biophys. Acta
1495,
231-236[Medline]
[Order article via Infotrieve]
|
13.
|
Acker, H.
(1994)
Ann. N. Y. Acad. Sci.
718,
1-10
|
14.
|
Fandrey, J.,
Frere, S.,
and Jelkmann, W.
(1994)
Biochem. J.
303,
507-510[Medline]
[Order article via Infotrieve]
|
15.
|
Gleadle, J. M.,
Ebert, B. L.,
and Ratcliffe, P. J.
(1995)
Eur. J. Biochem.
234,
92-99[Abstract]
|
16.
|
Chandel, N. S.,
McClintock, D. S.,
Feliciano, C. E.,
Wood, T. M.,
Melendez, J. A.,
Rodriguez, A. M.,
and Schumacker, P. T.
(2000)
J. Biol. Chem.
275,
25130-25138[Abstract/Free Full Text]
|
17.
|
Bai, J.,
Rodriguez, A. M.,
Melendez, J. A.,
and Cederbaum, A. I.
(1999)
J. Biol. Chem.
274,
26217-26224[Abstract/Free Full Text]
|
18.
|
King, M. P.,
and Attardi, G.
(1996)
Methods Enzymol.
264,
304-313[Medline]
[Order article via Infotrieve]
|
19.
|
Srinivas, V.,
Zhang, L.-P.,
Zhu, X.-H.,
and Caro, J.
(1999)
Biochem. Biophy. Res. Commnun.
260,
557-561[CrossRef]
|
20.
|
Minchenko, A.,
Bauer, T.,
Salceda, S.,
and Caro, J.
(1994)
Lab. Invest.
71,
374-379[Medline]
[Order article via Infotrieve]
|
21.
|
Jacobson, M. D.,
Burne, J. F.,
King, M. P.,
Miyashita, T.,
Reed, J. C.,
and Raff, M. C.
(1993)
Nature
361,
365-369[CrossRef][Medline]
[Order article via Infotrieve]
|
22.
|
Necas, E.,
and Thorling, E. B.
(1972)
Am. J. Physiol.
222,
1187-1190[Free Full Text]
|
23.
|
Han, D.,
Williams, E.,
and Cadenas, E.
(2001)
Biochem. J.
353,
411-416[CrossRef][Medline]
[Order article via Infotrieve]
|
24.
|
King, M. P.,
and Attardi, G.
(1989)
Science
246,
500-503[Medline]
[Order article via Infotrieve]
|
25.
|
Liu, Z. D.,
Lockwood, M.,
Rose, S.,
Theobald, A. E.,
and Hider, R. C.
(2001)
Biochem. Pharmacol.
61,
285-290[CrossRef][Medline]
[Order article via Infotrieve]
|
Copyright © 2001 by The American Society for Biochemistry and Molecular Biology, Inc.