The field of developmental genetics investigates the genetic basis of the
changes in form that an organism passes through during its life cycle. One
cellular process that is common throughout these changes in form is cell
division. The two cell division events that need to be controlled are the
entry into the S-phase when DNA is replicated, and the entry into the M-phase
when mitosis occurs. In this regard, two timing events need to be monitored
by the cell. These are:
when to initiate replication (S-phase entry)
when to begin chromosomal condensations (M-phase entry)
Related to these events are four factors that appear to control the entry
into the M-phase.
The accumulation of a specific cellular mass is a factor for somatic cells.
This is called the mass factor.
Some cells need to obtain a specific growth rate for mitosis to begin. This
is called the growth rate factor.
The time between successive M-phases appears to be controlled by timer or
oscillator genes. This is the time factor and appears to be a factor
in embryo cells.
The entry into the M-phase also requires completion of the S-phase. This
insures that daughter cells receive complete DNA complements and is called
the completion of chromosomal replication factor.
For the cell to coordinate these different events, it must be able to monitor
the cell cycle. An important biological question that needs to be resolved
is how does the cell know where it is in the cell cycle. As you would expect,
genetics and biochemical characterization have provided an extensive, but
incomplete description of the process.
Cell cycle research has primarily been performed on mutant strains of the
fission yeast (Schizosaccharomyces pombe) and the budding yeast
(Saccharomyces cerevisae) that have genetic lesions in some phase
of the cell cycle. The cell division cycle (cdc) mutant strains have
been quite useful in elucidating important steps. The cell cycle in yeast
has two points where it is committed to proceed to the next stage in the
cycle. The first point called start occurs near the end of the G1,
and the cell becomes committed to DNA synthesis in the S phase of the cycle.
The second commitment point is at the beginning of the M phase when
the cell becomes committed to chromosomal condensation and the subsequent
The following diagrams illustrate the genetic and biochemical information
known about the entry into the M-phase of the cell cycle. As you can see
from the biochemical diagram a protein complex is formed at the two committal
points. Each complex consists of a protein called cyclin, and a protein
kinase called p34. The existence of such a complex was described
biochemically when a factor called maturation promoting factor (MPF)
was isolated that could initiate mitosis in certain mutant yeast strains
whose cell cycle was arrested at this stage. It was the coupling of this
type of biochemical research with genetics that defined and elucidated many
of the steps in the cycle.
To demonstrate the molecular events associated with the cell cycle in more
detail, a discussion that links the genetic research with the biochemical
research will be presented. In particular, the discussion will concentrate
only on the entry into the M-phase. The following table presents the mutants,
the product of these genes and their role in M-phase entry.
a 45,000-47,000 dalton protein that complexes with the protein kinase
p34cdc2 to form the MPF; its sequence is 30% conserved
over a 200 amino acid stretch in a wide range of species; M-phase entry can
be stimulated by adding this protein from clams to frog cells; its degradation
appears to be associated with the inactivation of
a serine-threonine protein kinase of 34,000 daltons that complexes with cyclin
to form the MPF; the inactive form of the protein is phosphorylated at threonine
(T) and tyrosine (Y) residues; the phosphorylation appears to performed by
p60src in humans; the active form of the protein is
dephosphorylated and it functions by phosphorylating a number of proteins;
this phosophorylation activity is coupled to the entry into the M-phase;
the protein must be associated with a normal cyclin protein for the M-phase
to be completed normally; association with deletion mutants of cyclin halts
the M-phase before it is completed
a protein of 80,000 daltons that assists with the dephosphorylation of
p34cdc2 by either inhibiting its phosphorylation or promoting
its dephosphorylation; its concentration increases as the cell approaches
the M-phase suggesting the accumulation of this protein to a specific
concentration is required to activate p34cdc2 ; its increase in concentration
appears to be coupled with the completion of the S-phase
a protein of 13,000 daltons which may be involved in the inactivation of
p34cdc2 late in mitosis by inhibiting its kinase activity or promoting its
It is clear from the genetic and biochemical studies that the appearance
of an active MPF occurs at the M-phase committal point. The following cellular
events have been associated with the onset of the protein kinase activity
of the cdc2 product.
nuclear envelope breakdown
cell shape changes
Each of these events is clearly required for cell division to occur. Furthermore,
the substrates of the p34cdc2 protein kinase are proteins
involved in the maintenance of the cell in the G2-phase. The phosphorylation
of these proteins may change their functions and permit the cell to enter
the M-phase. The key substrates of p34cdc2 protein kinase
Histone H1 - the phosphorylation of this protein may be important
for chromosomal condensation to occur
Centrosomal protein - these proteins are associated with centrioles,
the organizing center of the cell for microtubules associated with the
Lamin - this is a protein associated with the nuclear envelope
p60src - phosphorylation of the mitotic-specific
sites of this protein may influence the cytoskeleton and lead to changes
in the cell shape
other DNA binding proteins that need to be released for chromosomal
condensation to occur
The studies on yeast and other organisms has lead to the conclusion that
a universal control mechanism regulating entry into the M-phase is common
to all eukaryotic cells. The key features of the process are as follows.
Copyright © 1997. Phillip McClean
The protein kinase activity of p34cdc2 is central to the
model. This protein is thought to phosphorylate key proteins that lead to
the major events in the M-phase. High levels of this protein maintain the
cell in the M-phase, and its inactivation is required for exit from the phase.
The second key protein is cyclin that complexes with p34cdc2
to form the MPF. Cyclin is required for
p34cdc2 activation. Cyclin degradation is required for
the cell to exit the M-phase and probably the inactivation of
The activation of p34cdc2 is associated with the
dephosphorylation of the phosphorylated tyrosine and threonine residues of
the protein. Its kinase activity appears to be associated with the tyrosine
residue, so dephosphorylating this site appears essential.
Timing of the M-phase entry is associated with two other protein kinases
and the accumulation of p80cdc25. This timing event is
associated with the dephosphorylation of p34cdc2.
p13suc1 interacts with p34cdc2 and may
be involved in its rephosphorylation at the end of the M-phase.