Sea Urchin Embryogenesis, Cell Specification and Regulatory
Programme
As a model organism…
- Genome size: 800 Mb (~24000 genes)
- Echinoderm – deuterostomes closely related to chordata
- Large number of gametes are easily available
- Embryos develop synchronously (unless conditions are sub-optimal)
- Embryos are transparent and good for microscopy
- Easy gene transfer and reporter gene expression
- Easy to manipulate
- Genome fully sequenced
Used to study:
- Developmental biology (gene regulatory network)
- Genetics
- Cell cycle regulation (Tim Hunt, cyclins)
- Fertilisation studies: single female can produce 20x10 6 easily collectable eggs
- Biogerontology
Sea Urchin Lifecycle: Embryo (3 days) Larvae (6-8 weeks) Metamorphisis (up to
200 years)
Fertilisation is external
Egg sizes are 70-100µm (similar to human oocyte), maximum of
150 µm
Fertilisation membrane (= egg shell) rises too slowly to prevent
polyspermy but offers mechanical protection to the embryo
Elevation of the membrane due to -
Sea urchins live by sea shore, so tembryos can be dashed by
waves against rocks after fertilisation
Another possible function of the egg shell is in preventing bacterial infection
Sea Urchin Cleavage
, Exhibit radial holoblastic cleavage
First seven cleavage divisions are
“stereotypic” – same pattern is followed
in every individual of the same species
First and second cleavages are both
meridional and are perpendicular to each
other (cleavage furrows pass through
animal and vegetal poles)
Third cleavage is equatorial,
perpendicular to the first 2 cleavage
planes and separates animal and vegetal hemispheres from each other
Fourth cleavage = four cells of the animal tier divide meridionally into 8
blastomeres, each with same volume (= mesomeres); vegetal tier undergoes unequal
equatorial cleavage to produce four large cells (= macromeres) and four smaller cells
(= micromeres) at the vegetal pole
As 16-cell embryo cleaves: 8 mesomeres divide equatorially to produce 2 tiers (an 1
and an2), one staggered above the other
- Macromeres divide meridionally forming a tier of 8 cells below an 2
Micromeres divide unequally, producing a cluster of 4 small micromeres at the tip of
the vegetal pole, beneath a tier of 4 large micromeres
- Small micromeres divide once more then cease dividing until larval stage (thought to
give rise to germ cells and migrating to reside in coelomic pouches of larva)
- Large micromeres (reside above the small micromeres) become the skeletogenic
cells (primary mesenchyme cells – PMCs)
- 60-cell stage – embryo composed of 6 tiers of cells whose fates have been mapped
Sixth division : animal hemisphere cells divide meridionally while vegetal cells divide
equatorially, pattern reversed in the seventh division (= 120-cell blastula)
Blastula Formation
120-cell blastula in which cells form a hollow sphere surrounding a central cavity
(blastocoel) – pattern of divisions becomes less regular
Cells are the same size, micromeres slowed down their cell divisions
Every cell is in contact with proteinaceous fluid of the blastocoel on the inside and
with the hyaline layer on the outside
Tight junctions unities the loosely connected blastomeres into seamless epiterlial
sheet that encircles the blastocoel
Cells divide, blastula remains one cell layer thick, thining out as it expands –
accomplished by adhesion of the blastomeres to the hyaline layer and by an influx
of water that expands the blastocoel (Dan, 1960)
Rapid and invariant cell cleavages last through ninth and tenth division depending on
species (fates of cells have become specified)
, Each cell becomes ciliated on the region of the cell membrane farthest from the
blastocoel
Ciliated blastula begins to rotate within the fertilisation envelope
Differences also seen in the cells
- Cells at vegetal pole of the blastula begin to thicken, forming a vegetal plate
- Cells of animal hemisphere synthesise and secrete a hatching enzyme that digests
their fertilisation envelope (Lepage et al., 1992) – embryo now a free-swimming
hatched blastula
Morphogenetic Movements
PMC undergoes to Epithelial Mesenchyme Transition (EMT) (Lyons et al.. 2011)
After hatching of blastula, large micromeres (at the
vegetal plate) change their shape, lose adhesions to
their neighbouring cells and then break away from
apical layer to enter the blastocoel as skeletogenic
mesochyme cells (PMCs) (undergoing classic EMT)
Once they have ingressed, Skeletongenic mesenchyme
cells then begin extending and contracting long thin
processes (=filopoda)
- Eventually filopodia become localised within
prospective ventrolateral region of the blastocoel – after active making and braking
filopodial connections to the wall of the blastocoel
- At the ventolateral region – fuse into syncytial cables which will form the axis of
the calcium carbonate spicules of the larval skeletal rods
Ingression in sea urchins is a movement of individual cells or small clusters of cells
(unlike gastrulation in fly and frog, which is an involution – movement of sheets of
cells(?))
PMC ingression studies using TEM and SEM – apical surface of the epithelium is
covered with the hyaline layer (containing hyaline, laminin, collagens and
echinonectin
- Cilium of each PMC is lost prior to ingression, which frees the cells from the hyaline
layer, while the small micromere progeny retain their cilia
- As PMCs ingress – loosen connections to neighbours, change shape and breach basal
lamina to enter the blastocoel
- Apical side of the epithelium faces the external environment and adherens
junctions connect cells to one another – changes in cell adhesion accompany
ingression
- Cells that ingress they lose affinity to apical lamina and gain affinity to the basal
lamina and blascoelar matrix at time of ingression
Programme
As a model organism…
- Genome size: 800 Mb (~24000 genes)
- Echinoderm – deuterostomes closely related to chordata
- Large number of gametes are easily available
- Embryos develop synchronously (unless conditions are sub-optimal)
- Embryos are transparent and good for microscopy
- Easy gene transfer and reporter gene expression
- Easy to manipulate
- Genome fully sequenced
Used to study:
- Developmental biology (gene regulatory network)
- Genetics
- Cell cycle regulation (Tim Hunt, cyclins)
- Fertilisation studies: single female can produce 20x10 6 easily collectable eggs
- Biogerontology
Sea Urchin Lifecycle: Embryo (3 days) Larvae (6-8 weeks) Metamorphisis (up to
200 years)
Fertilisation is external
Egg sizes are 70-100µm (similar to human oocyte), maximum of
150 µm
Fertilisation membrane (= egg shell) rises too slowly to prevent
polyspermy but offers mechanical protection to the embryo
Elevation of the membrane due to -
Sea urchins live by sea shore, so tembryos can be dashed by
waves against rocks after fertilisation
Another possible function of the egg shell is in preventing bacterial infection
Sea Urchin Cleavage
, Exhibit radial holoblastic cleavage
First seven cleavage divisions are
“stereotypic” – same pattern is followed
in every individual of the same species
First and second cleavages are both
meridional and are perpendicular to each
other (cleavage furrows pass through
animal and vegetal poles)
Third cleavage is equatorial,
perpendicular to the first 2 cleavage
planes and separates animal and vegetal hemispheres from each other
Fourth cleavage = four cells of the animal tier divide meridionally into 8
blastomeres, each with same volume (= mesomeres); vegetal tier undergoes unequal
equatorial cleavage to produce four large cells (= macromeres) and four smaller cells
(= micromeres) at the vegetal pole
As 16-cell embryo cleaves: 8 mesomeres divide equatorially to produce 2 tiers (an 1
and an2), one staggered above the other
- Macromeres divide meridionally forming a tier of 8 cells below an 2
Micromeres divide unequally, producing a cluster of 4 small micromeres at the tip of
the vegetal pole, beneath a tier of 4 large micromeres
- Small micromeres divide once more then cease dividing until larval stage (thought to
give rise to germ cells and migrating to reside in coelomic pouches of larva)
- Large micromeres (reside above the small micromeres) become the skeletogenic
cells (primary mesenchyme cells – PMCs)
- 60-cell stage – embryo composed of 6 tiers of cells whose fates have been mapped
Sixth division : animal hemisphere cells divide meridionally while vegetal cells divide
equatorially, pattern reversed in the seventh division (= 120-cell blastula)
Blastula Formation
120-cell blastula in which cells form a hollow sphere surrounding a central cavity
(blastocoel) – pattern of divisions becomes less regular
Cells are the same size, micromeres slowed down their cell divisions
Every cell is in contact with proteinaceous fluid of the blastocoel on the inside and
with the hyaline layer on the outside
Tight junctions unities the loosely connected blastomeres into seamless epiterlial
sheet that encircles the blastocoel
Cells divide, blastula remains one cell layer thick, thining out as it expands –
accomplished by adhesion of the blastomeres to the hyaline layer and by an influx
of water that expands the blastocoel (Dan, 1960)
Rapid and invariant cell cleavages last through ninth and tenth division depending on
species (fates of cells have become specified)
, Each cell becomes ciliated on the region of the cell membrane farthest from the
blastocoel
Ciliated blastula begins to rotate within the fertilisation envelope
Differences also seen in the cells
- Cells at vegetal pole of the blastula begin to thicken, forming a vegetal plate
- Cells of animal hemisphere synthesise and secrete a hatching enzyme that digests
their fertilisation envelope (Lepage et al., 1992) – embryo now a free-swimming
hatched blastula
Morphogenetic Movements
PMC undergoes to Epithelial Mesenchyme Transition (EMT) (Lyons et al.. 2011)
After hatching of blastula, large micromeres (at the
vegetal plate) change their shape, lose adhesions to
their neighbouring cells and then break away from
apical layer to enter the blastocoel as skeletogenic
mesochyme cells (PMCs) (undergoing classic EMT)
Once they have ingressed, Skeletongenic mesenchyme
cells then begin extending and contracting long thin
processes (=filopoda)
- Eventually filopodia become localised within
prospective ventrolateral region of the blastocoel – after active making and braking
filopodial connections to the wall of the blastocoel
- At the ventolateral region – fuse into syncytial cables which will form the axis of
the calcium carbonate spicules of the larval skeletal rods
Ingression in sea urchins is a movement of individual cells or small clusters of cells
(unlike gastrulation in fly and frog, which is an involution – movement of sheets of
cells(?))
PMC ingression studies using TEM and SEM – apical surface of the epithelium is
covered with the hyaline layer (containing hyaline, laminin, collagens and
echinonectin
- Cilium of each PMC is lost prior to ingression, which frees the cells from the hyaline
layer, while the small micromere progeny retain their cilia
- As PMCs ingress – loosen connections to neighbours, change shape and breach basal
lamina to enter the blastocoel
- Apical side of the epithelium faces the external environment and adherens
junctions connect cells to one another – changes in cell adhesion accompany
ingression
- Cells that ingress they lose affinity to apical lamina and gain affinity to the basal
lamina and blascoelar matrix at time of ingression
