Caenorhabditis elegans (C. elegans)
An elegant worm
Why study worms?
Sydney Brenner
“Thus we want a multicellular organism which has a short life cycle, can be
easily cultivated, and is small enough to be handled in large numbers, like a
micro-organism. It should have relatively few cells, so that exhaustive studies
of lineage and patterns can be made, and should be amenable to genetic
analysis.” --Excerpts from Proposal to the Medical Research Council, 1963
C. elegans: the chosen one!
Easily cultivated: can grow thousands
on a petri dish, feed on non-hazardous
bacteria, and cheap to maintain
Short generation time: 3 days
Small: 1 mm (about the size of a pinhead)
Few cells: The adult has 959
hermaphrodrodite (XX) or 1031 (XO) cells
Amenable to genetic analysis:
maintained as hermaphrodites, but
males exist for genetic studies,
The genome is small- 100 Mb
Transparency: allows for development
to be analyzed from a single cell and all
cells to be lineage
Photo credit: Ian D. Chin-Sang (Queen's University, Kingston, ON, Canada).
Life cycle of C. elegans
Photo credit: http://www.scq.ubc.ca/genetic-studies-of-aging-and-longevity-in-model-organisms/
Anatomy of C. elegans
Pharynx
head
anterior
Intestine (yellow)
Gonad (pink)
Vulva
~1 mm
Fig. 5.42
Anus
Rectum
Epidermis
tail
posterior
Hermaphrodites do it by themselves
Hermaphrodite (XX)
Males (X0)
Photo credit: http://homepages.ucalgary.ca/~dhansen/worms.gif
The C. elegans gonad: an extremely
efficient reproductive system
Fig. 5.42
Within this lineage is
the secret of embryonic development
John Sulston
All neural synapses have been mapped
An entire C. elegans hermaphrodite worm consists of exactly
959 cells EVERY SINGLE TIME,
allowing one to follow the cell lineage.
Learn to read a lineage diagram!
Branching =
cell division
Increasing
age of worm
embryo
1st stage
larva
Line ending =
differentiated cell
2nd stage
larva
= Cell death
P0 zygote
Cleavage
Events
Lineage
Fig. 5.42
2 cell stage
4 cell stage
8 cell stage
Most lineages consist of multiple tissue types
but the P4, E and D cells gives rise to a single tissue type
Fig. 5.42
Mutations can alter lineages in many ways
Question:
1.) How many cell divisions took place in the wildtype lineage? ____
2.) In wild-type, how many total descendants will cell A have? ____
3.) How many differentiated cells from the wild-type lineage will be
a part of the adult worm? ____
4.) What is the best description of the defect in mutant 1?
How are the invariant lineages
established?
ie. How do cells know who they are and what they are doing?
• Control of apoptosis
•Partitioning of cytoplasmic determinants
•Timing of developmental events
•Cell-Cell interactions
Even cell death is programmed into the lineage
C. elegans was used to identify
the machinery that regulates
programmed cell death in vertebrates
See also p. 96 for further discussion of cell death pathways
The Nobel Prize in Physiology
or Medicine 2002
"for their discoveries concerning ’
genetic regulation of organ development
and programmed cell death'"
Sidney Brenner
H. Robert Horvitz
John Sulston
Partitioning of cytoplasmic determinants
blue nuclei
P-granules (green) are
cytoplasmic determinants
that are formed from
ribonucleoprotein
complexes that specify the
germ cells
P granules are
asymmetrically
segregated into one cell, the
P4 cell, which will give rise
to the germline
Fig. 5.44
P0
P1
AB
P3
P4
green P-granules
PARtition mutants (PAR) disrupt the
asymmetric distribution of p-granlues
Photo credit: http://mbg.cornell.edu/cals/mbg/research/kemphues-lab/images/par_phenotypes.gif
Timing of developmental events
Lof= loss of function, gene function is disrupted
wildtype
lin-14 (lof)
lin-4 (lof)
Moss E. 2007. Current Biology, R425.
Lin-14 is required for the timing of cell division in the L1 stage.
Lin-4 regulates transition from L1 to L2 stage.
wildtype
lin-14 (lof)
lin-4 (lof)
Graph of LIN-14 and LIN-4 levels in a wildtype embryo
Levels
LIN-4
L1
L2
L3
L4
LIN-14
Adult Time
If you have a mutation that results in an INCREASED level of
LIN-14 (gain of function) which lineage would you expect
wildtype
lin-14 (lof)
lin-4 (lof)
Graph of LIN-14 and LIN-4 levels in a wildtype embryo
Levels
LIN-4
L1
L2
L3
L4
LIN-14
Adult Time
lin-4 does not encode a protein—
It encodes a microRNA !!
lin-4
lin-4
lin-14
lin-4
lin-14
Translation blocked!
Cell-Cell Interactions: the P2 impact!
GLP-1
APX-1
Notch receptor Delta-like ligand
MOM-2
Wnt homolog
MOM-5
Frizzled homolog
Signal from P2 cell required to induce EMS cell
to produce E cell which forms the gut (see p. 198-199)
How do cell interactions result in the
formation of an organ?
Vulva formation!
C. elegans Vulva development
Early larval stage
Anchor cell (AC)
Figure 3.34
Gonad
VPCs
AC
Basement membrane
Gonad
P3.p-P8.p are the Vulva
Precursor Cells (VPCs)
Later larval stage
1° and 2° VPCs make
the vulva
3° VPCs are non-vulval
3°
2° 1° 2°
3°
Inductive and lateral signals induce the vulva
gonad
Anchor cell
VPCs
VPCs after
induction
P8
3° 3° 2°
1°
2°
3°
The primary and
secondary cells
form the vulva
Cell ablation studies helped identify key
players in vulva formation
Lecture notes: experiment 1
If anchor cell signaling is disrupted, all
VPCs cells adopt a non-vulva fate
anchor
cell
gonad
3° cell 3° cell
3° cell
3° cell
no vulva
3° cell
3° cell
The VPCs have multipotential
Early stage
Anchor cell
gonad
VPCs
Later stage
Anchor cell
gonad
3°
3°
2° 1° 2°
3°
What is causing the VPCs to be different?
Let’s do an experiment: what happens when the P6.p cell is
ablated?
Anchor cell
gonad
VPCs
3°
3° 2°
3°
2°
3°
3° 2°
2°
3°
3°
3° 3°
3°
3°
3°
2° 1°
2°
1°
A
B
C
Lecture notes: experiment 2
3°
What genes specify the VPC cell fate?
Looked for mutants that disrupted vulva formation
1) No vulva: worms hatch inside (yuck!!)
1) Too many vulvas
Lecture notes: experiment 3
Inductive and lateral signals induce the vulva
gonad
Anchor cell
VPCs
VPCs after
induction
P8
3° 3° 2°
1°
2°
3°
The primary and
secondary cells
form the vulva
The vulvaless mutations helped define the
Ras pathway
Lin-3/Epidermal Growth Factor (EGF)
Let-23/EGF Receptor
Let-60/RAS
Sem-5/GRB2
Lin-45/RAF
P6.p becomes the primary cell!
The Ras pathway is abnormally activated in many human
tumors
eg: pancreatic cancer, colorectal cancer, lung adenocarcinoma,
gall bladder cancer, bile duct cancer and thyroid cancer
Another representation of the RAS pathway
(VPC cells)
LIN-3 signal
A signal from P6.p actives notch (lin-12) in P5.p and P7.p
Figure 3.34
The transmembrane receptor is the Lin-12 protein, a
receptor protein related to Notch
“ Primary cell”
“ Secondary cells”
Figure 3.33
Both membrane and receptor are membrane bound!
Generation of Different Cell Types From Equivalent Cells in C. elegans:
Initial specification of the Anchor Cell also requires Notch
The signal:
lag-2
(delta)
Figure 3.35
The receptor:
lin-12 (notch)
Does the Notch pathway remind you of anything
you learned earlier?
No notch=neural!
The story of epidermal vs. neuronal fate in
Drosophila
Some cells become neuroblasts
and signal their neighbors to
remain epidermis
If signal is missing...
all cells eventually ingress
and become neuroblasts
Nervous system
Epidermis
Extra nervous
system
No epidermis!