478
Cellular Interactions
for active PKC to be downregulated is to
cleave its membrane and calcium binding
domains from the catalytic subunit. Once
cleaved, the catalytic subunit can diffuse
away from the plasma membrane while
the catalytic subunit remains as an ac-
tive kinase that is no longer calcium- or
membrane-dependent. In this state, it has
historically been referred to as PKM. Now
that PKM is not membrane tethered, it
can phosphorylate elements in the inte-
rior of the mammalian egg. In support of
this model, Gallicano et al. have demon-
strated that after activation of PKC, PKM
is generated, and that it phosphorylates
components of the cytoskeletal sheets and
other substrates in the interior of the egg.
PKC has both temporal and spatial pre-
cision because PKM can only form after
PKC is activated. Thus, as a result of the
fertilization-induced elevation in the level
of [Ca
2
+
]
i
, PKC initially modiFes the cell
periphery and then components in the cell
interior are later modiFed by PKM. Such a
model Fts well with the restructuring of an
egg into a zygote because, by modifying the
cell periphery (i.e. cortical granule exocyto-
sis) immediately, polyspermy is prevented
and then a more gradual modiFcation of
the interior can follow.
By using pharmacologic agents that
activate PKC, Moses and Kline have
investigated roles for PKC in mouse
eggs and have found that when such
agents are applied, metaphase II eggs are
driven into interphase, a characteristic of
egg activation. However, transition into
interphase had a different mechanism
than an egg was artiFcially activated by
calcium ionophore or inseminated by
sperm. Moses and Kline report that the
PKC agonist, PMA, caused abnormal and
greatly slowed metaphase to anaphase
transition. In part, this parallels Gallicano
and coworker’s results that show that eggs
exhibit an abnormal transit to anaphase
when PKC is activated by agonists and
the level of calcium is clamped low. In
agreement with this result, Moses and
Kline also show that application of PMA
to eggs at meiotic metaphase II does
not result in an elevation in the level
of [Ca
2
+
]
i
. These results suggest that the
metaphase to anaphase transition requires
a component of the cell other than PKC
that requires an elevation of the level of
[Ca
2
+
]
i
.
Whether activation of PKC with agonists
induces second polar body formation
wasa
lsoadd
ressedbyMosesandK
l
ine
.
The
investigators
conclude
that
even
though the cell is ultimately driven into
interphase, second polar body formation
is not induced by activation of PKC.
However, two important considerations
concerning polar body formation were
not taken into account. ±irst,
unlike
cytokinesis, polar body formation is a
two-step process in mammals, with an
initial extrusion of cytoplasm (bounded
by the plasma membrane) followed by
formation of a contractile ring at the
base of the extruded cytoplasm. Second,
when PMA is applied to mouse eggs,
the second polar body initially appears
(initial extrusion stage), but frequently,
because the contractile ring does not close,
the polar body is absorbed into the egg.
By the time the cell reaches telophase,
the extruded second polar body appears.
The cytoplasmic extrusion of the initiated
second polar body can be clearly seen in
Fgure 2b of the 1995 publication by Moses
and Kline, which shows a telophase stage
mouse egg. This inability of the egg to
completely close the contractile ring of
the second polar body suggests that some
other pathway not activated by PMA is
responsible for contractile ring closure.
As was discussed in the previous section,
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