ARTICLE
Auteur(s) : Harry
Rubin
Department of Molecular and Cell Biology, Life Sciences
Addition, University of California, Berkeley, CA 94720-3200,
USA
Physiology of growth stimulation
The study of cell proliferation received an enormous boost with the
advent of monolayer cell culture in the mid 1900s. The cell culture
technique made it possible to observe and count all the cells in a
culture, to easily change individual components of the medium, vary
the cell population density and radioactively label the cells. A
number of major features of growth proliferation and its regulation
quickly became apparent, mainly from studies of fibroblasts and
their derivatives, established cell lines in culture (see [1, 2]
for complete primary references). In addition to the essential
micronutrients of a defined medium, the addition of animal serum or
its purified protein components is required for cell proliferation.
Once cells have adapted to culture, maximum proliferation occurs at
relatively low population densities, slows down as the population
approaches confluence, and is restricted to cell turnover at the
postconfluent saturation density. The saturation density is itself
proportional to the concentration of animal serum, which is the
source of protein growth factors.
The rate of DNA synthesis of course varies coordinately with the
rate of cell proliferation. The initiation of DNA synthesis can be
brought to a very low rate by sharply lowering the serum
concentration overnight in confluent cultures. The restoration of
serum to its original high level in such inhibited cultures is
followed by a lag of several hours before DNA synthesis begins to
increase, and it rises to a maximum in 15-20 hr. Several other
physiological processes which are inhibited by confluence and
withdrawal of serum are very rapidly stimulated by the restoration
of serum. These processes include the uptake of glucose and its
utilization, the phosphorylation of uridine, and most importantly,
the synthesis of protein. The increase in glucose uptake is not
essential for a single round of accelerated DNA synthesis, and the
uptake and phosphorylation of uridine is totally gratuitous since
there is ordinarily no uridine in the medium. In contrast, the
onset of DNA synthesis is acutely dependent on the rate of
synthesis and accumulation of protein, particularly of ribosomal
protein. The combination of these effects is known as the
coordinate response [3].
The stimulation provided by added serum must be continuously
maintained for the entire lag period to initiate an increase in the
rate of DNA synthesis [4]. In fact, acute inhibition of protein
synthesis by cycloheximide during the rise of DNA synthesis stops
any further increase in its rate [5]. Inhibition of RNA synthesis
by actinomycin D has a delayed effect on DNA synthesis, probably by
exhausting the supply of messenger RNA. The increase in protein
synthesis which occurs promptly upon the addition of serum is
dependent on the rate of the initiation step rather than the rate
of chain elongation. Although serum or a combination of the
purified polypeptide growth factors present in serum is necessary
to maintain the continuous proliferation of cells, the early
responses to such physiological agents as well as the later onset
of DNA synthesis can be stimulated by non-specific treatments. For
example, confluent chicken embryo fibroblasts can be stimulated in
this manner by subtoxic concentrations of heavier metals such as
zinc, cadmium, manganese, mercury and lead. Trypsin and other
proteolytic enzymes, in amounts too small to cause retraction much
less detachment of cells, also stimulate chicken embryo
fibroblasts. Such treatments are not able to stimulate established
lines of mouse fibroblasts, but sodium pyrophosphate at a sharply
defined low concentration, just sufficient to form floccular
precipitates with calcium in the medium, is a potent stimulator of
the coordinate response [6, 7].
The accumulated evidence indicates that all the stimulatory
agents, whether physiological, and therefore specific, or
non-physiological and not specific, initiate the coordinate
response by direct action on the plasma membrane. The restriction
of action to the plasma membrane is made particularly evident in
the case of pyrophosphate because of the strict dependence of
stimulation on the formation of insoluble floccules, which appear
suddenly at a fixed concentration that is dependent on the
concentration of calcium and inorganic phosphate in the medium [7].
If the floccules are prevented from combining with the plasma
membrane by completely filling a closed culture dish with medium
and inverting it, the floccules sink to the bottom and there is
neither contact with the cells nor stimulation [6]. A further
indicator of a central role for membrane activity in regulating the
coordinate response is the requirement for growth for normal cells
to attach to and spread on a solid substratum. If the cells are
kept in a spherical shape in suspension, or, are squeezed together
in a crowded culture, they will not proliferate.
Role of Mg2+ in regulating the coordinate response
of cells to growth factors and non-specific stimulators
Several features of the cellular response to growth-stimulating
treatment combined to draw attention to Mg2+ as a
plausible intracellular mediator of growth regulation. One of these
was the variegated nature of the response which includes increased
uptake and phosphorylation of several components [3]. A second
feature was the capacity of non-specific membrane-active treatments
to initiate the coordinate response. A third was the requirement of
Mg2+ for a very wide spectrum of cellular activities, in
particular the requirement of MgATP2- for every known
phosphoryl exchange reaction. Many of those reactions occurred at
critical control steps in biochemical pathways, such as
phosphofructokinase and pyruvate kinase in glycolysis. Of
particular significance is the high requirement for Mg2+
in the initiation of protein synthesis which must be maintained at
a high level for the entire G1 period of the mitotic cycle in order
to achieve the maximum effect on the initiation of DNA synthesis
and cell division.
The experimental strategy in examining the role of
Mg2+ in growth regulation was to reduce its
concentration in the medium and observe the effect on various
components of the coordinate response. The initial results were
erratic, since a drastic reduction in Mg2+ concentration
was required to achieve inhibition. As it turned out, it was
difficult to get the cells to give up their Mg2+, not
surprisingly given the essential nature of Mg2+ for so
many cellular activities. The use of EDTA to chelate
Mg2+ was no help since it has a much stronger preference
for Ca2+. However, pyrophosphate came to our aid because
it binds Mg2+ somewhat more than Ca2+.
Although very small amounts of pyrophosphate stimulate the
proliferation of Balb 3T3 fibroblasts in an excess of
Mg2+, when the pyrophosphate concentration exceeds that
of Mg2+ there is a reproducible reduction in synthesis
of protein, RNA and DNA.
It was later found that deprivation of Ca2+ in
confluent cultures of mouse fibroblasts inhibited DNA synthesis,
but the effect could be reversed with excess Mg2+[8]. In
contrast, inhibitory effects resulting from deprivation of
Mg2+ could not be reversed by raising the
Ca2+ concentration of the medium, indicating that
Mg2+ is the operational element. These experiments
showed that deprivation of Ca2+ concentration in the
medium facilitated the manipulation of the Mg2+
concentration in the cells over a wide range, as a function of the
external Mg2+ concentration. There is an optimal
concentration of Mg2+ for protein and DNA synthesis
beyond which there is a sharp decrease in both processes [9]. The
resulting bell-shaped curve for protein and consequently DNA
synthesis as a function of Mg2+ resembled the curve
produced for cell-free synthesis of protein by a purified
preparation of ribosomes [10]. Change in the rate of protein
synthesis is extremely sensitive to any change in the concentration
of Mg2+, up or down, indicating that the free
Mg2+ of the cell is normally set at a concentration that
would alter the rate of protein synthesis with any change in free
Mg2+, no matter how small. The Mg2+-dependent
change in protein synthesis precedes any change in DNA synthesis by
hours, consistent with the key regulatory roles of Mg2+
and protein synthesis in cell proliferation [11]. The cell responds
to a moderately inhibitory increase of intracellular
Mg2+ concentration by actively reducing its
concentration to normal levels over a period of hours, with a
consequent increase in protein synthesis and a delayed increase in
DNA synthesis, indicating how crucial Mg2+ is to the
cellular growth economy. It is notable that there is no direct
effect of Mg2+ concentration on the rate of DNA
synthesis. If the Mg2+ concentration is raised to
extremely high concentrations, the cell cannot lower its internal
Mg2+, and neither protein nor DNA synthesis recovers,
leading to death of the cells.
While protein synthesis is the crucial reaction to the
Mg2+ levels in cells that determine DNA synthesis and
proliferation, the effects are probably mediated through protein
kinase cascades. Insulin acts as a growth factor by combining with
a plasma membrane receptor and activating a tyrosine kinase on the
cytoplasmic side of the membrane [1, 2]. The tyrosine kinase in
turn activates the PI 3-K cascade consisting mainly of
serine/threonine kinases combined with inhibitory proteins which
lead to phosphorylation of the mTOR kinase. The mTOR kinase in turn
phosphorylates two proteins involved in the initiation of
translation. Increased synthesis of ribosomal proteins and
elongation factors are especially important in accumulating protein
and initiating DNA synthesis. While most of the protein kinases in
this pathway have a very low Km for MgATP2-, mTOR has a
high requirement that approximates the level of free
Mg2+ in the cell. The MMM model holds that any rise in
free Mg2+ increases MgATP2- and activates
mTOR, thereby accelerating the initiation of translation, and
speeding the passage of cells through the G1 period to DNA
synthesis and mitosis.
Direct evidence for the increase of free Mg2+ in
growth stimulation of cells
The key reaction in growth stimulation in the MMM model of cell
proliferation is an increase of total cellular Mg2+,
which raises the concentration of free Mg2+. That
increase in turn raises the concentration of MgATP2-
which is required for all the phosphoryl transfer reactions of the
cell. The earliest estimates of free Mg2+ were derived
from the equilibrium of Mg2+-dependent enzyme reactions
and indicated that free Mg2+ is only a small percentage
of the total Mg in cells [12]. Similar values for free
Mg2+ were obtained in computer modeling of energy
metabolism in the rat heart [13, 14]. In our own studies of
insulin- or serum-stimulated cells we had no way of measuring free
Mg2+ in cells, but we obtained very accurate
measurements of total cellular magnesium and of the amount bound to
the external surface of the cell [15, 16]. There was a 15 – 20%
increase in total Mg in the stimulated cells, which was maintained
through the G1 and S periods of the cell cycle. The increase in
total Mg indicates a significant increase in free Mg2+
since it has been reported that the latter rises disproportionately
with total magnesium [17].
Subsequently a Mg2+-sensitive indicator, mag-fura-2,
was developed [18, 19] which could be used to measure intracellular
free Mg2+ stimulated by growth factors. Epidermal growth
factor increased DNA synthesis 48-fold in a serum-starved line of
muscle cells [20]. Free Mg2+ increased after a
5 min lag period from an initial 0.32 mM to as high as 1.4 mM
at 20 min, and then leveled off. Swiss 3T3 cells stimulated by
insulin increased free Mg2+ in 30-60 min from a
basal level of 0.22 mM to a final value of 0.29-0.35 mM [21].
Estimates of the free Mg2+ level were unreliable beyond
1 hr because of mag-fura-2 leakage from the cells, but the results
were consistent with the long term increase in free Mg2+
as required to carry cells through the cell cycle. It is assumed
that a significant fraction of the free Mg2+ was
released from the internal surface of the perturbed membrane where
it is bound to fixed sites, but the increase in total
Mg2+ probably came from increased activity of Na/K
ATPase pumps of Mg2+ in the membrane [22].
The effect of Mg2+ and other cations on protein
synthesis could be determined in the oocytes of the frog Xenopus
laevis, which are large enough to allow direct injection of the
cations and measurement of protein synthesis in a single cell by
incorporation of 3H-leucine [23]. Initial results
indicated that injection of K+ into the oocytes
increased the rate of protein synthesis to a similar extent as did
exposure to gonadotropin, the physiological method for starting
embryonic development. There were several inconsistencies in the
results however that indicated that K+ played an
indirect role in the activation of protein synthesis [24]. The
injection of Mg2+, however, stimulated protein synthesis
at a much lower concentration than K+, giving a sharper
peak in a bell-shaped curve and exhibiting no inconsistencies. It
was concluded that K+ (or Na+) displaced
Mg2+ from bound sites and made it directly available for
protein synthesis. Ion-sensitive microelectrodes were used to
determine free K+ and Mg2+, and gave the
usual low values for the latter. How gonadotropin sets off these
changes remains to be determined but it may have to do with cation
exchanges initiated by perturbation of the cell membrane as
suggested for stimulation of somatic cells by growth factors [15,
16].
In contrast to the acceleration of protein synthesis as the key
response for entraining cell proliferation, the increased uptake of
uridine has no apparent physiological role in the process. That is
because uridine is not ordinarily added in the growth medium, and
its addition is devoid of effect on proliferation. However,
phosphorylation of uridine by uridine kinase is the limiting step
in its trapping by cells [25], a simple enzymatic reaction that
requires MgATP2- and can be studied in cells or in
cell-free extracts. Deprivation of Mg2+ in cultures
reduces the rate of uridine trapping by cells, in the same manner
as does the removal of serum, i.e., by reducing the Vmax for
Mg2+ of the reaction [26]. In contrast, the uptake of
thymidine is unaffected by changing the concentration of
Mg2+ in the medium. This difference can be explained by
the fact that the Km of uridine kinase for Mg2+ is
10-fold higher than the Km of thymidine kinase for Mg2+.
Hence, the phosphorylation of thymidine in cells has sufficient
cellular Mg2+ for maximum activity even without growth
factors or significant external Mg2+, while the
phosphorylation of uridine is sensitive to both treatments. Raising
the intracellular Mg2+ concentration high enough in the
presence of the divalent cation ionophore A21387 and high
extracellular Mg2+ increases uridine trapping to the
same extent as addition of serum to the cells. Mg2+ also
relieves feedback inhibition of uridine kinase by UTP and CTP [27].
The sum of these observations supports a regulatory role for
Mg2+ in the gratuitous uptake of uridine, and reinforces
its role in the other more essential responses to growth
factors.
Discussion
The advent of monolayer culture of cells brought with it
quantitative studies of the regulation of their proliferation by
growth factors and, in its wake, a number of hypotheses about the
underlying mechanism. These included the increased uptake of
nutrients following addition of serum to confluent,
contact-inhibited cultures that had been deprived of serum, and the
activation of any of the four major intracellular cations, or an
increase of intracellular pH. Of the essential nutrients, glucose
showed the largest increase of uptake, but a decrease in external
concentration to lower the amount taken up below that due to its
increase had no effect on the stimulation of DNA synthesis. Similar
experiments with Na+, K+ and Ca2+
likewise ruled out a central role for these cations in regulation
[28]. It was also realized that DNA synthesis itself was not the
direct target of growth factor stimulation since there was a lag of
5 to 12 h depending on cell type before there was an increase
in its synthesis. That shifted some of the emphasis to those
cellular responses that occurred within minutes after adding growth
factors. Strangely enough, one of the most informative of these
early reactions was the phosphorylation of uridine which is the
limiting factor in the rate of its uptake by the cell, but is
totally gratuitous to the growth and proliferation of the cells.
The rate of uptake of uridine proved to be sensitive to variations
in the intracellular concentrations of magnesium, as did the
activity of uridine kinase in cell-free extracts and feedback
inhibition by pyrimidine nucleotides. The rate of glucose uptake
was also dependent on intracellular Mg2+ content, which
supported a significant role for Mg2+ in coordinate
control.
The most significant early response of cells to growth factors
is an increased rate of translation which is governed by the
initiation step [29]. It has long been recognized that initiation
of protein synthesis has a higher Mg2+ requirement than
chain elongation [30, 31]. The rate of protein synthesis proved to
be directly proportional to change in the Mg2+ content
of cells [9, 11]. The changes in Mg2+ and protein
synthesis had to be maintained throughout the G1 period to produce
their full effect. At very high Mg2+ concentration there
was an inhibition of protein synthesis just as there is in
cell-free extracts. The sum of these observations lent strong
support to a central role of Mg2+ in coordinating
multiple biochemical responses of cells to growth factors. This
support was reinforced by measurements of free Mg2+
levels in cells after stimulation. A particularly significant
contribution to understanding the role of Mg2+ in
activating protein synthesis came from the separate injection and
measurement of the four major cellular cations into frog oocytes
with observation of their effects on the initiation of protein
synthesis, which left no doubt that increase in free
Mg2+ is the direct effector of the process which begins
growth and development of the frog set off physiologically by
gonadotropin [24].
Given the cumulative evidence for a central role of
Mg2+ in the coordinate response of cells to growth
stimuli, it is instructive to consider why there was so much
resistance to accepting the hypothesis. Possibly the most widely
read review of early signals in the mitogenic response gives
detailed attention to Na+, K+,
Ca2+ and H+ without mention of
Mg2+[32]. Nor does the review acknowledge the early and
persistent increase in protein synthesis and its essential relation
to the onset of DNA synthesis [5, 29, 33-35]. One might think that
the evidence for the essential role of increased protein synthesis
would call attention to its sensitivity to small changes in
Mg2+ as demonstrated in cell-free extracts [31, 36, 37].
However, a common attitude seemed to be that intracellular
Mg2+ is very well buffered and the cell goes to “some
pains to hold [it] steady” because it influences so many reactions
[38]. This attitude of course ignored the many reactions of the
coordinate response that had to be balanced in order to obtain
growth and proliferation, which makes Mg2+ the ideal
second messenger for the response. It also ignored the papers
showing there is a long term increase in Mg with growth factor
treatment, e.g. [15], and failed to predict the increase in free
Mg2+ demonstrated when specific fluorescent indicators
became available [20, 21, 39].
Another criticism was that growth-stimulated cells do not
exhibit a steep surge of Mg2+, as they do of
Ca2+, so they are “not set up to use Mg2+ as
a natural trigger” [40]. In this respect however, the relatively
small increase in Mg2+ sustained over many hours
parallels the increase in protein synthesis induced by growth
factors, while the brief trigger-like increase of Ca2+
is incompatible with those effects. A further criticism of
Mg2+ as regulator of the growth response was that
Mg2+ is an essential rather than a regulatory element of
the response [41]. This criticism is without foundation, however,
because the level of free Mg2+ is set at a point that
small increases or decreases are nowhere near the levels that
threaten cellular metabolism, but they do affect the rate of
coordinate response in a physiological manner. It is apparent that
there was, and possibly still is, a misconception of the nature of
the long term, finely graded nature of the coordinate response and
particularly the rate of protein synthesis that makes
Mg2+ the ideal candidate for the job.
It remained to be determined what specific biochemical process
was activated by Mg2+ to accelerate the initiation pf
protein synthesis. The precise mechanism, however, had to await the
elucidation of the molecular pathway leading from the receptor
binding of growth factors through intermediate steps to the
increased frequency of translation. Understanding of such a pathway
began to take shape in the late 1980s when the molecular era of
growth regulation commenced. One of the best understood pathways
for growth regulation is that instituted by the binding of insulin
to its surface receptor and the activation of its tyrosine kinase
function within the cell. This sets off the phosphatidyl inositol
3-kinase (PI-3K) pathway with its cascade of kinases that activates
mTOR, a PI-3K related kinase. mTOR then phosphorylates two proteins
that directly control the initiation of protein synthesis. All the
kinases in the PI-3K pathway except mTOR have a very low
requirement for ATP [42] and hence for free Mg2+. The
requirement of mTOR for ATP is almost two orders of magnitude
higher than that of the other kinases of the pathway. It was
originally proposed that mTOR activity is dependent on the level of
ATP in the cell, which was claimed to increase with cell
stimulation by insulin [42]. However, ATP actually decreases
slightly with cell stimulation [43]. Since the free Mg2+
level in the cell does increase [20, 21] and therefore increases
MgATP2-, which is the true substrate for kinases, it is
assumed that the free Mg2+ level is the regulatory
agency for protein synthesis. Thus the coordinate physiological
response and the molecular aspects of PI-3K pathway are joined
through the agency of Mg2+, consistent with the MMM
model of cell proliferation control [2].
Acknowledgements
I am grateful for the manuscript preparation and editing by Dorothy
M. Rubin. The effort for the paper was supported by the National
Institutes of Health grant G13LM07483-03.
References
1 Rubin H. Central roles of Mg2+ and of
MgATP2- in the regulation of protein synthesis and cell
proliferation: significance for neoplastic transformation. Adv
Cancer Res 2005; 93: 1-58.
2 Rubin H. Magnesium: The missing element in molecular
views of cell proliferation control. Bioessays 2005; 27:
311-20.
3 Rubin H. Central role for magnesium in coordinate control
of metabolism and growth in animal cells. Proc Natl Acad Sci USA
1975; 72: 3551-5.
4 Rubin H, Steiner R. Reversible alteration in the
mitotic cycle of chick embryo cells in various states of growth
regulation. J Cell Physiol 1975; 85: 261-70.
5 Kim JH, Gelbard AS, Perez AG. Inhibition of DNA
synthesis by actinomycin D and cycloheximide in synchronized HeLa
cells. Exp Cell Res 1968; 53: 478-87.
6 Bowen-Pope DF, Rubin H. Growth stimulatory
precipitates of Ca2+ and pyrophosphate. J Cell Physiol
1983; 117: 51-61.
7 Rubin H, Sanui H. Complexes of inorganic
pyrophosphate, orthophosphate and calcium as stimulants of 3T3
cells multiplication. Proc Natl Acad Sci USA 1977; 74: 5025-30.
8 Rubin H, Terasaki M, Sanui H. Magnesium
reverses inhibitory effects of calcium deprivation on coordinate
response of 3T3 cells to serum. Proc Natl Acad Sci USA 1978; 75:
4379-83.
9 Rubin H, Terasaki M, Sanui H. Major
intracellular cations and growth control: Correspondence among
magnesium content, protein synthesis, and the onset of DNA
synthesis in Balb/c 3T3 cells. Proc Natl Acad Sci USA 1979; 76:
3917-21.
10 Schreier MH, Staehelin T. Initiation of mammalian
protein synthesis: the importance of ribosome and initiation factor
quality for the efficiency of in vitro systems. J Mol Biol 1973;
73: 329-49.
11 Terasaki M, Rubin H. Evidence that intracellular
magnesium is present in cells at a regulatory concentration for
protein synthesis. Proc Natl Acad Sci USA 1985; 82: 7324-6.
12 Rose IA. The state of magnesium in cells as estimated
from the adenylate kinase equilibrium. Proc Natl Acad Sci USA 1968;
61: 1079-86.
13 Achs MJ, Garfinkel D. Computer simulation of energy
metabolism in anoxic perfused rat heart. Am J Physiol 1982; 232:
R164-R174.
14 Achs MJ, Kohn MC, Garfinkel D. Computer
simulation of metabolism in pyruvate-perfused rat heart. IV. Model
behavior. Am J Physiol 1979; 237: R174-R180.
15 Sanui H, Rubin H. Membrane bound and cellular
cationic changes associated with insulin stimulation of cultured
cells. J Cell Physiol 1978; 96: 265-78.
16 Sanui H, Rubin H. Changes of intracellular and
externally bound cations accompanying serum stimulation of mouse
Balb/c3T3 cells. Exp Cell Res 1982; 139: 15-25.
17 Corkey BE, Duszynski J, Rich TL,
Matschinsky B, Williamson JR. Regulation of free and
bound magnesium in rat hepatocytes and isolated mitochondria. J
Biol Chem 1986; 261: 2567-74.
18 Murphy E, Freudenrich CC, Levy LA,
London RE. Monitoring cytosolic free magnesium in cultured
chicken heart cells by use of the fluorescent indicator Furaptra.
Proc Natl Acad Sci USA 1989; 86: 2981-4.
19 Raju B, Murphy E, Levy LA, Hall RD,
London RE. A fluorescent indicator for measuring cytosolic
free magnesium. Am J Physiol 1989; 256: C540-C548.
20 Grubbs RD. Effect of epidermal growth factor on
Mg2+ homeostasis in BC3H-1 myocytes. Am J Physiol 1991;
260: C1158-C1164.
21 Ishijima S, Sonoda T, Tatibana M.
Mitogen-induced early increase in cytosolic free Mg2+
concentration in single Swiss 3T3 fibroblasts. Am J Physiol 1991;
261: C1074-C1080.
22 Lostroh AJ, Krahl ME. Insulin action: Accumulation
in vitro of Mg2+ and K+ in rat uterus: Ion
pump activity. Biochim Biophys Acta 1973; 291: 260-8.
23 Lau YT, Yassin RR, Horowitz SB. Potassium salt
microinjection into Xenopus oocytes mimics gonadotropin treatment.
Science 1988; 240: 1231-323.
24 Horowitz SB, Tluczek LJM. Gonadotropin stimulates
oocyte translation by increasing magnesium activity through
intracellular potassium-magnesium exchange. Proc Natl Acad Sci USA
1989; 86: 9652-6.
25 Rozengurt E, Stein WD, Wigglesworth N. Uptake
of nucleosides in density-inhibited cultures of 3T3 cells. Nature
1977; 267: 442-4.
26 Vidair C, Rubin H. Evaluation of Mg2+ as
an intracellular regulator of uridine uptake. J Cell Physiol 1981;
108: 317-25.
27 Vidair C, Rubin H. Mg2+ as activator of
uridine phosphorylation and other cellular responses to growth
factors. Proc Natl Acad Sci USA 2005; 102: 662-6.
28 Moscatelli D, Sanui H, Rubin H. Effects of
depletion of K+, Na+, or Ca2+, on
DNA synthesis and cell cation content in chick embryo fibroblasts.
J Cell Physiol 1979; 101: 117-28.
29 Stanners CP, Becker H. Control of macromolecular
synthesis in proliferating and resting Syrian hamster cells in
monolayer culture. J Cell Physiol 1971; 77: 31-42.
30 Revel M, Hiatt HH. Magnesium requirement for the
formation of an active messenger RNA-ribosome-S-RNA complex. J Mol
Biol 1965; 11: 467-75.
31 Schreier MH, Noll H. Conformational changes in
ribosomes during protein synthesis. Proc Natl Acad Sci USA 1971;
68: 805-9.
32 Rozengurt E. Early signals in the mitogenic response.
Science 1986; 234: 161-6.
33 Brooks RF. Continuous protein synthesis is required to
maintain the probability of entry into S phase. Cell 1977; 12:
311-7.
34 Castor LN. Responses of protein synthesis and
degradation in growth control of WI-38 cells. J Cell Physiol 1977;
92: 457-67.
35 Levine EM, Becker Y, Boone CW, Eagle H.
Contact inhibition, macromolecular synthesis, and polyribosomes in
cultured human diploid fibroblasts. Proc Natl Acad Sci USA 1965;
53: 350-6.
36 Brendler T, Godefroy-Colburn T, Yu S,
Thach RE. The role of mRNA competition in regulating
translation. J Biol Chem 1981; 256: 11755-61.
37 Ilan J. (ranslation of maternal messenger
ribonucleoprotein particles from sea urchin in a cell-free system
from unfertilized eggs and product analysis. Dev Biol 1978; 66:
375-85.
38 Rink TJ, Tsien RY, Pozzan T. Cytoplasmic pH
and free Mg2+ in lymphocytes. J Cell Biol 1982; 95:
189-96.
39 Ishijima S, Tatibana M. Rapid mobilization of
intracellular Mg2+ by bombesin in Swiss 3T3 cells:
mobilization through external Ca2+ and tyrosine
kinase-dependent mechanisms. J Biochem (Tokyo) 1994; 115:
730-7.
40 Whitfield JF. The roles of calcium and magnesium in cell
proliferation: An overview. In: Boynton A, McKeehan WL,
Whitfield JF, eds. Ions, Proliferation and Cancer. New York:
Acad. Press, 1982: 283-94.
41 Baserga R. In: The Biology of Cell Reproduction. Boston:
Harvard Univ. Press, 1985: 138-40.
42 Dennis PB, Jaeschke A, Saitoh M,
Fowler B, Kozma SC, Thomas G. Mammalian TOR: a
homeostatic ATP sensor. Science 2001; 294: 1102-5.
43 Fodge DW, Rubin H. Activation of
phosphofructokinase by stimulants of cell multiplication. Nature
New Biol 1973; 246: 181-3.
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