cell news 2/2013
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During early phases of epiboly (30-50% epiboly, 4.66-5.25
hpf), marginal EVL cells exhibit actin-rich flopodia protrusion
and have been therefore proposed to participate actively in the
spreading
(24)
. In contrast, flopodia are no longer detected once
the EVL margin reaches the equator of the sphere (50% epiboly)
(18, 25)
. At the same time (30-40% epiboly) a ring-like actomy-
osin structure forms within the YSL adjacent to the EVL margin.
It spans around the circumference of the yolk cell and becomes
increasingly pronounced
(17, 25)
. The marginal EVL cells are con-
nected to the YSL via tight junctions
(17, 26)
. Altering myosin
activity specifcally within the YSL via the mitogen-activated
protein (MAPK) pathway components traf2/nika and mapkapk2
suggest a critical role of the YSL actomyosin ring for EVL epiboly
(17, 27)
. To test if the YSL actomyosin ring generates the forces
that drive EVL movements, we utilized a UV-Laser ablation setup
to locally disrupt the actomyosin ring in close proximity of the
EVL cells
(15)
. We found considerable delays in EVL cell move-
ments adjacent to the ablation site, supporting the role of the
actomyosin ring as a driving force for the epithelial spreading.
However, the key question still remains: How does the actomy-
osin ring generate the necessary force to accomplish this task?
Actomyosin ring contraction
Contractile actomyosin rings function in many different mor-
phogenetic processes. On the cellular scale, actomyosin rings are
assembled at the equatorial plane of dividing cells to drive cyto-
kinesis
(28)
. They also participate in the apical constriction of in-
vaginating cells
(29)
. Supracellular actomyosin rings form upon
wounding of an epithelium, and they promote the closing of the
wound
(30)
. This is also how ‘dorsal closure’ proceeds in
Droso-
phila
. Here, a supracellular actomyosin cable contributes to the
closing of an epithelium on the dorsal side of the embryo
(31,
32)
. The common view of how actomyosin rings function in these
processes is through a purse-string mechanism. Contractile my-
osin motor protein activity on the circular actin structure causes
the ring to radially constrict. This triggers shape changes upon
the cell/tissue structures to which it is associated. One possible
mechanism for the function of the actomyosin ring in zebrafsh
epiboly therefore is that myosin motor function generates active
tension along the circumference of the ring (circumferential ac-
tive tension), which, due to the spherical geometry of the embryo
gives rise to a force pulling on the EVL margin once the ring has
passed the equator.
This hypothesis concerns cell and tissue mechanics underlying
epiboly. Testing it requires in vivo mechanical measurements of
the tension within the epibolyzing actomyosin ring. Here, corti-
cal laser ablation (COLA) has proven to be a very versatile tool
(9,
11, 14, 32)
. The basic principle of cortical laser ablation is that a
cortex under tension will rapidly recoil in response to a cut, with
a recoil velocity that is proportional to the mechanical tension
present before the cut. As such, COLA allows to compare relati-
ve tension along different orientations or at different times. We
systematically mapped the tension within the actomyosin ring
over the course of zebrafsh epiboly by use of a pulsed UV laser.
In agreement with the hypothesis that the YSL actomyosin ring
acts as a purse-string, we measured signifcant circumferential
tension within the ring (Fig. 2a), which depends on myosin-2
activity and increases during epiboly progression
(15)
. Due to
the spherical geometry of the yolk cell, circumferential active
tension results in a net force pulling on the EVL margin, which is
zero when the ring is at the equator and grows as the ring ap-
proaches the vegetal pole. Consequently, when the actomyosin
ring is near the equator of the yolk cell, tension within the ring in
the animal-vegetal (AV) direction is expected to be much smaller
than circumferential tension
(15)
.
To our surprise we found that the AV tension within the ring at
60-70% epiboly (Fig. 2a) is roughly half of the circumferenti-
al tension, and remains constant throughout epiboly (15). These
results are incompatible with the actomyosin ring functioning as
a simple purse-string only. To fnd out where the additional ten-
sion in the AV direction comes from, we turned towards analy-
zing actin and myosin dynamics. Interestingly, we observed that
the formation of the ring is accompanied by substantial fow of
actin and myosin from vegetal parts of the yolk cell towards the
margin of the EVL, and in a direction opposite to those of epiboly
movements (Fig. 2b). These retrograde cortical fows are initially
slow and long-ranged, but increase their speed at later stages of
epiboly
(15)
.
Cortical fows are a wide spread phenomena in animal cell mor-
phogenesis
(3)
. The emergence of cortical fows has been associ-
ated with gradients of myosin contractility and recent studies in
different model organisms have highlighted their pivotal role in
fundamental morphogenetic processes ranging from cellular po-
larization to cell intercalation or tissue invagination
(11, 13, 14,
33, 34).
To elucidate how retrograde cortical fows might con-
tribute to zebrafsh epiboly, we require a biophysical framework
that identifes the relevant mesoscale physical properties and
their relation with deformation and fow. To this end we develo-
ped a theoretical description of actomyosin network mechanics
in the YSL as well as in the EVL tissue. This description is concep-
tually similar to the one utilized for unraveling the physical basis
of cortical fow in the
C. elegans
zygote
(11).
Flow-friction, a crawling motor
The YSL cortex is a thin (1-2 µm) network on the surface of the
yolk cell that expands along the AV direction with a width of
30-80 µm and spans around the circumference of the spherical
embryo having a radius of roughly 350 µm. It consists of a high-
ly dynamic network of cross-linked actin flaments, upon which
myosin motor proteins can exert active forces through the con-
sumption of ATP
(8)
. The dynamics of this collective network are
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