Essential Cell Biology 5th edition

HOW WE KNOW
DISCOVERY OF CYCLINS AND Cdks
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For many years, cell biologists watched the “puppet
show” of DNA synthesis, mitosis, and cytokinesis but
had no idea what was behind the curtain, controlling
these events. The cell-cycle control system was
simply a “black box” inside the cell. It was not even
clear whether there was a separate control system, or
whether the cell-cycle machinery somehow controlled
itself. A breakthrough came with the identification of
the key proteins of the control system and the realization
that they are distinct from the components of the
cell-cycle machinery—the enzymes and other proteins
that perform the essential processes of DNA replication,
chromosome segregation, and so on.
The first components of the cell-cycle control system
to be discovered were the cyclins and cyclin-dependent
protein kinases (Cdks) that drive cells into M phase.
They were found in studies of cell division conducted
on animal eggs.
Back to the egg
The fertilized eggs of many animals are especially suitable
for biochemical studies of the cell cycle because
they are exceptionally large and divide rapidly. An egg
of the frog Xenopus, for example, is just over 1 mm in
diameter (Figure 18−6). After fertilization, it divides rapidly
to partition the egg into many smaller cells. These
rapid cell cycles consist mainly of repeated S and M
phases, with very short or no G 1 or G 2 phases between
them. There is no new gene transcription: all of the
mRNAs and most of the proteins required for this early
stage of embryonic development are already packed
into the very large egg during its development as an
oocyte in the ovary of the mother. In these early division
cycles (cleavage divisions), no cell growth occurs, and all
the cells of the embryo divide synchronously, growing
smaller and smaller with each division (Movie 18.2).
Because of the synchrony, it is possible to prepare an
extract from frog eggs that is representative of the cellcycle
stage at which the extract is made. The biological
activity of such an extract can then be tested by injecting
it into a Xenopus oocyte (the immature precursor of
the unfertilized egg) and observing, microscopically, its
effects on cell-cycle behavior. The Xenopus oocyte is an
especially convenient test system for detecting an activity
that drives cells into M phase, because of its large
size, and because it has completed DNA replication and
is suspended at a stage in the meiotic cell cycle (discussed
in Chapter 19) that is equivalent to the G 2 phase
of a mitotic cell cycle.
Give us an M
In such experiments, Kazuo Matsui and colleagues
found that an extract from an M-phase egg instantly
drives the oocyte into M phase, whereas cytoplasm
from a cleaving egg at other phases of the cycle does
not. When they first made this discovery, they did not
know the molecules or the mechanism responsible, so
they referred to the unidentified agent as maturation
promoting factor, or MPF (Figure 18−7). By testing cytoplasm
from different stages of the cell cycle, Matsui and
colleagues found that MPF activity oscillates dramatically
during the course of each cell cycle: it increased
rapidly just before the start of mitosis and fell rapidly to
zero toward the end of mitosis (see Figure 18−5). This
oscillation made MPF a strong candidate for a component
involved in cell-cycle control.
When MPF was finally purified, it was found to contain a
protein kinase that was required for its activity. But the
kinase portion of MPF did not act alone. It had to have a
specific protein (now known to be M cyclin) bound to it
in order to function. M cyclin was discovered in a different
type of experiment, involving clam eggs.
0.5 mm
Figure 18−6 A mature Xenopus egg
provides a convenient system for
studying the cell cycle. (Courtesy of
Tony Mills.)