Cleavage and Breaking Symmetry
E. G. Conklin, 1905
Goals of Cleavage:
1) Make lots of cells
2) Begin making cells different from one another
How to give daughter cells DIFFERENT Identities
Extrinsic Mechanisms
Intrinsic Mechanisms
Autonomous specification
Cell fate determined by intrinsic mechanism
(e.g. cytoplasmic determinant)
Mosaic Development
Cells of embryo retain discrete fates when separated
(e.g. Ascidian embryo)
The “European Plan”: You develop according to where you were born and
lineage is important. (after S. Brenner)
As opposed toConditional specification
Cell fate determined by extrinsic mechanism
(e.g. signalling)
Regulative Development
Cells of embryo can modify cell fate to form new structures when isolated
(e.g. human identical twins)
The “American Plan”: Cells start off undetermined, albeit with some biases.
They are determined by their environment, sometimes after migrating long
distances, and tend to behave similarly to their neighbors. (after S. Brenner)
Cytoplasmic Rearrangement
in Ascidian Zygote
A) Before fertilization
Uniform cortical yellow cytoplasm
Gray, yolky cytoplasm inside
B) After sperm entry
Yellow cytoplasm streams towards
sperm nucleus
C and D)
Yellow cytoplasm moves with
migrating sperm nucleus
Yellow crescent is formed
Fate Mapping the Ascidian Embryo
Yellow Crescent
E. G. Conklin, 1905
Blastomere Isolation Experiments
Muscle
No Muscle
Whittaker, 1977
-Blastomere pair containing yellow crescent can form muscle when cultured
in isolation
-Muscle markers come on at appropriate time relative to control embryos
-Muscle markers come on even when cell division is blocked
-Remainder of embryo does not form muscle
The Ultimate in Blastomere Isolation:
Nishida (Roux’s Archives, 1992)
-Separate blastomeres after each division until 110 cell stage
-Allow blastomeres to develop separately
-Get many individual cell fates: muscle, gut, epidermis
BUT NOT: notochord or nervous system
Thus: muscle is AUTONOMOUSLY SPECIFIED
notochord is CONDITIONALLY SPECIFIED
Asymmetric cell division segregates cytoplasmic determinants
Need to:
1)
2)
3)
4)
Break Symmetry
Establish cell polarity
Localize cytoplasmic determinants
Control mitotic spindle
Asymmetric segregation of P granules
to P cell lineage
Susan Strome
Asymmetric Cell Division in C. elegans
C. elegans first cleavage asymmetries:
1)
2)
3)
4)
Cytoplasmic flow toward posterior
P granules/PIE-1 at posterior
Asymmetric cleavage (post. cell smaller)
Different cell fates
P Granules (Stome Lab)
PIE-1-GFP (Seydoux Lab)
Goldstein Lab
Asymmetric Cell Division
1) Break Symmetry
2) Establish Cell Polarity
3) Localize Determinants
4) Control Mitotic Spindle
Gonczy Nature Reviews, 2008
The C. elegans Hermaphrodite Gonad
Sperm Entry Point Determines Posterior in
C. elegans (Goldstein and Hird)
Sperm Entry Point Determines the D/V Axis in Xenopus
Sadler and Shakes
The paternal pronucleus is not required
for A/P patterning in C. elegans
wt
No sperm nucleus
Laser ablation of centrosomes
disrupts polarity
Time of ablation
(relative to establishment of polarity)
after
before
during
GFP-PAR2
Breaking Symmetry: It all starts at the cortex
1) Sperm contributes centrosomes and
Rho Gap
2) Creates asymmetry in cortical domains
3) Actin/myosin contractility more active at
anterior than posterior
4) Helps move “anterior” PARs to anterior
and allow “posterior” PARs to localize
Nance, 2005
Goldstein Lab
Asymmetric Cell Division
1) Break Symmetry
2) Establish Cell Polarity
3) Localize Determinants
4) Control Mitotic Spindle
Gonczy Nature Reviews, 2008
Cell Polarity: Specifying Sub-Cellular Identity
PAR6-GFP
PAR2-GFP
A conserved machinerly for cell polarity
Polarized
epithelium
Polarized
zygote
Anterior Complex
PAR3 PDZ Domain (prot-prot)
PAR6 PDZ Domain
PKC3 Atypical Prot. Kinase C
Who needs who for localization:
Anterior complex required for initial
localization of PAR2 (PKC3 inhibits
phosphorylates and blocks PAR2)
Posterior Complex
PAR2 Ring Finger
PAR1 Ser/Thr Kinase
PAR2 NOT required for initial localization
of anterior complex
Not Localized
PAR4 Ser/Thr Kinase
PAR5 14-3-3
PAR2 required for maintenance of anterior
complex (phos. of PAR3 by PAR1?)
PAR1 is most downstream: dependent on
all others for localization
Asymmetric Cell Division
1) Break Symmetry
2) Establish Cell Polarity
3) Localize Determinants
4) Control Mitotic Spindle
Gonczy Nature Reviews, 2008
Segregation of Cell Fate Determinants
Pie-1-GFP
Priess Lab
P Granules
Strome Lab
Seydoux Lab
Centrosomes
Actin/myosin
ZIF-1 Dependent
Degradation
Asymmetric Cell Division
1) Break Symmetry
2) Establish Cell Polarity
3) Localize Determinants
4) Control Mitotic Spindle
Gonczy Nature Reviews, 2008
Control of the Mitotic Spindle
Linking Cell Polarity to Spindle Orientation: A Conserved Model
Cell Polarity
MT Motor
/Par-3
Connect Cell Polarity to Motor
NuMA-type Dynein Binder
“GoLoco/TPR” Protein
Non-receptor Heterotrimeric G Protein
-Par complex localizes GPR1/2
-G-alpha activates GPR1/2
-GPR1/2 bind dynein complex
-dynein complex orients spindle
Siller and Doe, Nature Cell Biol. 2009
Worm
Fly
Mouse
Spindle Dynamics in Early C. elegans Zygote
GPR-1/2 at Anterior
GPR-1/2 at Posterior
Siller and Doe, Nature Cell Biol. 2009
The posterior “pulls harder” on the spindle to
generate different size daughter cells
Asymmetric Cell Division
1) Break Symmetry
2) Establish Cell Polarity
3) Localize Determinants
4) Control Mitotic Spindle
Anterior-Posterior Polarity in the Drosophila Oocyte
PARs and Oocyte Patterning in Drosophila
PAR-1
PAR-1 PAR-3-GFP
wt PAR-3-GFP
S-A PAR-3-GFP
STAU PAR-3-GFP
PAR-3 PAR-1
St. Johnston Lab
Asymmetric Cell Division in Neuroblasts
Apical Complex
e.g. Baz/Par3
Cell Fate Determinants
e.g. Miranda, Prospero
Baz = Par3
Wodarz, 2005
Bellaiche and Gotta, 2005