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Polyamines: New Cues in Cellular Signal
Transduction
Uriel Bachrach, Yong-Chun Wang, and Amalia Tabib
The naturally occurring polyamines putrescine, spermidine, and spermine are involved in signal
transduction. This has been demonstrated by using inhibitors for polyamine biosynthesis (such as
D-difluoromethylornithine) or adding polyamines to cultured cells. Different polyamines, preferentially activated protein kinases (tyrosine kinases and MAP kinases), stimulated the
expression of nuclear protooncogenes (myc, jun, and fos).
T
he distance between the cellular outer membrane and the
nucleus seems to be tiny, only 20 Pm. Yet that minute distance encompasses a major mystery: how do the cells of
higher organisms respond to the many growth signals that they
receive from the environment? The answer is crucial to underU. Bachrach, Y.-C. Wang, and A. Tabib are in the Department of Molecular Biology, Hebrew University-Hadassah Medical School, 91120
Jerusalem, Israel.
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News Physiol. Sci. • Volume 16 • June 2001
standing such long-standing questions in cell biology as what
causes cancer and how cellular growth is initiated or arrested.
It has been well established that growth factors (mitogens) bind
to specific receptors located on the cellular membrane. The
mitogen-receptor complexes then trigger a cascade of events,
including the activation of Ras by converting it from the GDP
bound form to the GTP form. The activation of protein kinases
is considered to be the next step in signal transduction, and we
now recognize its critical role in the regulation of cell growth
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Ca2+ influx and fill SR Ca2+ stores. In relation to abnormal
pacemaker function, drugs or disease processes that elevate SR
Ca2+ may act via low-voltage-activated SR Ca2+ release to
abnormally enhance atrial pacemaker automaticity. Abnormal
pacemaker activity may therefore arise from Ca2+-mediated
mechanisms that need not invoke delayed afterdepolarizations
and triggered activity, which require spontaneous diastolic
Ca2+ release from an SR overloaded with Ca2+. Because latent
atrial pacemakers are more dependent on low-voltage-activated SR Ca2+ release than primary pacemakers, they may be
more susceptible to Ca2+-mediated dysrhythmic activities.
Moreover, diastolic SR Ca2+ release may play a more important
role in generating activity in pacemaker cells lacking If. An
understanding of the different mechanisms underlying atrial
pacemaker activity may provide insight into the etiology and
ultimate prevention of certain types of atrial arrhythmias.
kinase to the other and how the phosphorylation reaction can
be controlled.
Polyamines and proliferation processes
and development. The extracellular signal-regulated kinases
(ERKs), also referred to as mitogen-activated protein kinases
(MAPKs), play a number of important roles in signal transduction in eukaryotic cells (see Fig. 1). They are phosphorylated by
MAPK/ERK kinases (MEKs), which are, in turn, activated by
Raf. During the immediate-early responses of mammalian cells
to mitogens, histone H3 is rapidly phosphorylated by Rsk-2, a
member of the p90 family. p90 Proteins are translocated into
the nucleus on phosphorylation by MAPKs, where changes in
chromatin structure take place and transcription factors are
synthesized. The mutated Ras has been defined as an oncogene and has been associated with various malignancies (9).
Another oncogene, Src, is also activated by mitogens and is
induced during malignant transformation. It may be seen (Fig.
1) that the cascade of events leading to enhanced growth by
Src follows a different pathway. The various kinases, as well as
the membrane-bound Ras, have been studied in detail, yet little is known as to how information is transmitted from one
FIGURE 2. Structure and biosynthesis of the naturally occurring polyamines.
News Physiol. Sci. • Volume 15 • June 2001
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FIGURE 1. Signal transduction from the cellular membrane to its nucleus.
Mitogens or growth factors are bound to specific receptors located on the
membrane. The mitogen-receptor complexes then trigger several cascades of
events. One of them includes tyrosine kinases, which activates Ras. This protooncogene then activates mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase (ERK) kinases (MEKs) via Raf. ERKs, which play
important roles in signal transduction, are then phosphorylated by MEK. From
there, signals are transduced to the nucleus, where histone H3 (His) is phosphorylated and thereby changes the structure of chromatin. Nuclear oncogenes, which may serve as transcription factors, are also activated by phosphorylation. CREB is a cAMP binding site located on the ornithine decarboxylase (ODC) gene. Another oncogene, Src, is also induced by mitogens,
leading to the activation of another pathway. PKB and PKC, protein kinases B
and C; SOS, son-of-sevenless protein; PI3K, phosphitidylinositol 3-kinase;
FRAP, FKBP-rapamycin-associated protein; FKBP, FK506 binding protein,
CREB, cAMP-responsive element binding protein.
The naturally occurring polyamines (the diamine putrescine,
the triamine spermidine, and the tetra-amine spermine) are
ubiquitous polycations (Fig. 2). They are present in all prokaryotic and eukaryotic cells thus far studied. They stabilize nucleic
acids and stimulate their replication (6). Polyamines are therefore essential for growth processes, and they have also been
associated with carcinogenesis. Their assay in biological fluids
has been used for cancer diagnosis and for monitoring anticancer treatment (10). The intracellular concentration of
polyamines can be regulated by 1) biosynthesis, 2) uptake, 3)
oxidation, and 4) acetylation.
Biosynthesis. The rate-limiting step in the biosynthesis of
polyamines is the conversion of ornithine into putrescine (Fig.
2). This step is catalyzed by ornithine decarboxylase (ODC;
E.C.4.1.1.17), which has also been defined as an oncogene (1).
ODC has an extremely short half-life (1525 min in eukaryotic
cells), and its synthesis is induced by hormones (10). Inhibition
of ODC by specific inhibitors such as D-difluoromethylornithine (DFMO) results in inhibition of malignant growth,
abortion, and prevention of parasitic growth (6).
Uptake. In addition to biosynthesis, cellular polyamine concentrations can be modulated by uptake. The transport of
polyamines by yeast and bacterial cells has been studied
extensively, and transporters have been isolated and cloned
(6). The rate of polyamine uptake by bacteria is energy dependent and is a function of external pH and exogenous amine
concentrations. The transporters can be either membrane-associated proteins or contain transmembrane-spanning segments,
some of which bind ATP (6).
It is evident from Fig. 2 that putrescine can be converted into
spermidine or spermine. This occurs when cells are saturated
with putrescine. In our studies, cells were starved before the
addition of the polyamines, polyamines did not reach saturation, and therefore the interconversion was minimal.
Polyamines and protein kinases
FIGURE 3. Effect of spermidine on signal transduction events. Spermidine,
which is formed from the diamine putrescine, stimulates the phosphorylation
of tyrosine kinases and ERK1/2 (p42 and p44). Spermidine also activates the
nuclear oncogene c-myc, which may be regarded as a transcription factor. PD98059 inhibits MEK1/2, whereas D-difluoromethylornithine (DFMO) inhibits
the phosphorylation of ERK1/2 and the expression of c-myc.
Oxidation. Oxidation and/or excretion are the main
processes leading to the reduction in cellular polyamine levels.
Polyamines and diamines can be oxidized by specific
polyamine or diamine oxidases. Some of the oxidation products exhibit biological activity. Thus J-aminobutyric acid,
which plays an important role in neural function, can be
formed from putrescine.
Acetylation. Acetylation is another metabolic pathway that
leads to a decrease in active polyamine levels (6). This reaction
is catalyzed by specific acetyltransferases and causes the reduction in the net positive charge density of the polyamines.
Acetylpolyamines and diamines are not tightly bound to cellular
negatively charged molecules (such as nucleic acids, phosphoproteins, or phospholipids) and thus may destabilize nucleic
acids. Polyamine excretion is also enhanced by acetylation.
Despite the wide distribution of polyamines in nature, their
exact biological functions have not yet been elucidated. Many
biologists were puzzled by two questions: 1) is there a need for
three different naturally occurring polyamines and 2) does
each of them fulfill a specific function?
In this short review, we will try to explain some specific
functions of the different polyamines. It will be shown that
spermidine enhances the phosphorylation of threonine and
tyrosine residues in ERK1 and ERK2 and that both the diamine
putrescine and triamine spermidine stimulate the phosphorylation of proteins by tyrosine kinases. The activation of kinases
by polyamines triggers the expression of nuclear oncogenes.
Putrescine stimulates the transcription of c-fos and c-jun,
whereas spermidine enhances the synthesis of c-Myc.
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Polyamines and the expression of nuclear oncogenes
Growth-associated genes, such as c-fos and c-myc protooncogenes, are activated during cellular proliferative
processes. This activation is due to signals transmitted from the
cellular membrane to its nucleus. Following mitogenic stimulation, a simultaneous activation of polyamine biosynthesis
and the transcription of the c-fos protooncogene has been
observed. Similarly, malignant transformation leads to an
FIGURE 4. Effect of putrescine on signal transduction events. The diamine
putrescine, which is formed from ornithine (by ODC), stimulates tyrosine
phosphorylation by tyrosine kinases and the expression of the nuclear protooncogenes c-fos and c-jun.
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The Ras/MAPK cascade is the best-defined pathway
involved in cell proliferation. In this pathway, a central role is
played by proteins p42 and p44. Various recent studies suggested that polyamines are involved in the expression and activation of MAPKs. Thus ODC-overproducing transfectants
showed enhanced MAPK (7) and tyrosine kinase (2) activities.
In those studies, no differential effect of one of the polyamines
has been reported. We have recently demonstrated (4) that
spermidine preferentially stimulated the phosphorylation of
p42 and p44 (Fig. 3). The MAPK pathway is essential for
growth. PD-98059 [2-(2-amino-3c-methoxyphenyl)-oxanaphthalen-4-one] selectively inhibits the MAPK-activating
enzymes (MEKs). This inhibitor inhibits ERK phosphorylation
and ODC activity (7).
Polyamines and signal transduction
It can be seen in Fig. 1 that signals are transduced from the
cellular membrane to its nucleus by activating kinases, including tyrosine kinases. Subsequently ras, MEKs, and ERKs are
activated. During the last steps of the signal transduction
process, nuclear protooncogenes and transcription factors are
expressed. It is remarkable that many of these steps are regulated by polyamines and that DFMO, which inhibits
polyamine biosynthesis, prevents the transfer of information
from the membrane to the nucleus. All of this strongly suggests
that polyamines play a role in signal transduction.
An ODC-oncogene loop
Protooncogene products, expressed downstream of the
MAPK cascade, include the transcription factors activator protein 1 (AP-1) or c-Myc. AP-1 is either a homodimer of Jun or a
heterodimer of Jun and Fos, which bind to a common DNA
binding site located in introns 3, 5, and 11 of the ODC gene
(13). The Myc protein, on the other hand, is a transcription factor that regulates the expression of genes by binding to the specific DNA sequence CACGTG (5). Among the target genes for
Myc regulation is ODC (5). Both Myc and AP-1 are substrates
of phosphorylation by MAPK. If polyamines, indeed, regulate
the formation of Myc and AP-1, then a reciprocal effect of
these nuclear oncogenes and polyamine synthesis could be
conceived. Such a polyamine-oncogene loop has been proposed by Flamingi (7).
The expression of ODC can also be regulated by cAMP (3).
Spermidine may regulate the activity of protein kinase A (8).
The ODC gene has a cAMP binding site. Therefore, another
cAMP loop may be envisaged.
Conclusions
It now appears that the naturally occurring polyamines not
only stabilize cellular nucleic acids and/or membranes but also
play a pivotal role in activating protein kinases and transcription
factors. These findings may explain the role of polyamines in cellular proliferation and differentiation processes. Moreover, carcinogenesis could be related to the accumulation of polyamines
in cancer cells. The expression of ODC, which catalyzes the formation of putrescine from ornithine, can be regulated by factors
that are controlled by polyamines such as Myc, Jun, Fos, and
cAMP, thus forming several reciprocal loops. All of these findings stress the importance of naturally occurring polyamines in
controlling essential physiological events and may explain the
distribution of more than a single polyamine in nature.
The research from our laboratory was supported by the Joseph H. Sciaky
Memorial Foundation.
References
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increase in polyamine biosynthesis, deregulation of ODC, and
the amplification of protooncogenes. It has been demonstrated
that polyamines stimulate the transcription of c-myc and c-fos
(11), but the preferential role of each of the polyamines on protooncogene expression has not been explored. The involvement of polyamines in the expression of nuclear protooncogenes has previously been confirmed by the finding that
DFMO blocked the expression of c-fos and c-myc in cultured
cells (11). We have recently reported that spermidine at micromolar concentrations stimulated the transcription and translation of c-myc (Fig. 3) in cultured rat kidney epithelial cells (12).
On the other hand, putrescine was more active (Fig. 4) in preferentially stimulating the expression of c-fos and c-jun in those
cells (12). These findings suggest that each of the polyamines
has a preferential effect in controlling nuclear protooncogene
expression.