VL5 - c. elegans I Flashcards
- Caenorhabditis elegans
Basic Characteristics:
* Benign for humans, optically transparent, miniature in size.
* Well-established protocols for mutagenesis, transgenesis, and molecular biology.
* Easy to breed, small size, rapid life cycle.
* Large brood size and hermaphrodites,
* can be frozen and stored for over 10 years at -80°C.
Genome and Genetics:
* Six chromosomes (linkage groups): Five autosomes (I-V) and one sex chromosome (X)
* Contains 20,444 protein-coding genes
* Two genders: Hermaphrodites (self-fertilization, clones of the parent) and males.
Genome Sequencing and Significance:
* First multicellular eukaryotic organism with a fully sequenced genome in 1998
* 38% of C. elegans protein-coding genes have predicted orthologs in the human genome
* Invariant development with constant cell numbers (951 in hermaphrodites, 1031 in males).
Limitations:
* Small size makes dissection almost impossible.
* No cell culture lines exist.
* Not all metazoan genes are found in the C. elegans genome.
Reproiduction of c.elegans
hermaphrodites:
* Selfing/cloning: ca. 300 progenies
* Mating: ca. 1000 progenies (males sperms preferred)
What is special about the somatic muscle from c.elegans
- 95 diamond shaped body-wall muscle cells
- 4 quadrant run along side legth of animal
- lead to sinusoidal movement of the animals
Basically a tube within a tube
*Model for Duchenne´s Muscular Dystrophy and cardiomyoparthies
What is special about the c. elegany intestine?
Structure
* 20 large epithelial cells form a long tube around the lumen
* only fixed to pharynx and rectum
* only one associated muscle at the posterior end
* not directly innervated
Functions
* Digestion of food
* absorption of processed nutrients
* synthesis and storage of macromolecules
* initation an innate immune response to pathogens
* nutring of germ cells by producing yolk
Serves as a model
* Organogenesis
* Morphogenesis
* Cell fate (pharynx)
* infection and response to infection (intestine)
What is special abot the c. elegans secretory-excretory system?
Structure:
* constist of 3 tubular cells
–> likely needed for osmoregulation
- great model for tube formation
What is special about the c.elegany nevous system?
Structure
* Hermaphrodites 302 neurons / males 382 neurons
* mostly simple structure with 1 or 2 neurites exiting from the cell body
* passive nerve condunction: No sodium-dependet action potentials
* High membrane rsistance
* carious neurotransmitters and receptors involved
* Multi-functional neurons (polymodality) –> probably because of low number
–> entire neuronals circuitry has been mapped all 7000 synapses & gap junctions + connectome known!
model for:
* cell lineage studie
–> detail characaterization of the neuron connection is a major advantage for analyzing neuronal function and pathways
–> detail characterization of neurons is key for rare neurological diorders
- What is special about the c.elegans reproductive system?
- Hermaphrodiric Nature
Heraphrodities asre the predominant form. Male also exist (arise from X0 chromosomal constitution)
–> self fertilization is possible - defined number of cells: (entire development lineage is kown. Includes the number of cells in each organ and tissue)
Model
* Morphogenesis
* Sex-specific anatomy
* programmed cell death
The vulva development is a powerful tool to understand `signal transduction´
Vulva development in c.elegans
- defined set of diffrentiated cell types:
-1° (Central Vulva Cells)
-2° (Lateral Vulva cells)
-3° (Hypodermis) - interesting communication between diffrent cell types
- Vulva forms during L3 Larve stage from 6 VPC (Vulva precursor cells) p3.p -p8.p
- VPC can become a vulva cell or a hypodermis cell. Which cell becomes what depends from inductive signal from anchor cell
- Anchor cell signal –> LIN3 (Protein)
- P6.p is closest to anchor cell and recieves most LIN3 –> turns into VPC 1°
- p5.p and p7.p –> VPC 2°
- P8.p, P4.p and p3.p no LIN 3 signal at all –> VPC 3°
NOTE:
* P6.p turns into 1° VPC because of LIN3
* P6.p supress the inductive signal so p5.p and p7.p becomes 2° VPC (lateral Signal)
* P3.p, P4.p and P8.p develop into hyodermis cells becaus of lack of LIN3
Scenario 1:
Laser ablation of the anchor cell during L1 results in a vulvaless phenotype.
In a vulvaless mutant, the prescence of the anchor cell does not change the cell fate (loss-of-function,reduction-of-function)
Scenario 2:
Overexpression of LIN3 –> multivulva phenotype
Mutagenesis strategies in c.elegans
1. Genome-wide mutagenesis (foward genetics)
The whole genome is mutagenized to identify genes that are involved in a specific process, e.g developmental or physiological
* Chemical mutagens
* radiation mutagensis
* Transposon insertional mutagenesis
2. Target-selected mutagenesis (reverse genetics)
The whole genome is mutagenized, but only a specific target gene is screened for mutations
3. Gene-targeted mutagenesis (reverse genetics)
A target gene is specially mutagenized
Mutation in c.elegans occur spontaneously and are caused by repair erros, endogenous oxidative damage, exogenous environmental insult
Worms in space
Cell lineage study in c.elegans
John Sulston and Syndey Brenner started to collect knwoledge about the development of the nervous system of c.elegans
Method:
* DIC (differential interference contrast) optics based on Normarski
* Animal has to be monitored alive
Results
* Invariant cell division generated a fixed number of progency cells of rigidly determined fates
* Three main events occured during post-embryonic development
-Cell division
-Cell migration
-Cell death
John Sulston and Robert Horvitz publsihed the embryoinic cell linegae of c.elegans
–> Nobel prize to Sydney Brenner, Robert Hovritz and John Sulston in 2002
Apoptosis in C.elegans
–> John Sulston, Robert Horvitz and Sydney Brenner observed the first apoptosis event in c.elegans during the cell lineage studies
2 Types of programmed cell deaths occure in c. elegans
1. Developmental cell death
113 cells die during embryonic development
250-40 min post fertilization)
18 cells die during post-embryonic development
2. Germ cell death
(>50% of all germ cells die)
- c.elegans does not need apoptosis for proper development
- Genes involved in apoptosis might be essential for innate immune response
- Mutant taht effect the apotosis are viable and fertile but:
-grow slower
-smaller brood size
-fail to elimniate damage germ cells following gentoxic stress
-hyper-susceptible to infections
The decision for a cell to undergo apoptosis in C. elegans is influenced by developmental cues or external signals.
* Activation of the egl-1 gene by inhibiting ced-9 (which normally blocks apoptosis).
* This allows pro-apoptotic genes to initiate cell suicide
* The dead cell is then recognized and engulfed by neighboring cells (not macrophages) through specific genetic pathways and additional genes (ced-10) participate in the degradation of the dead cell.
- egl-1: identified in a mutagenesis screen for egg-laying defective mutants. The hermaphrodite specific neurons is missing in these mutants. The HSN dies by programmed cell death but not in egl-1/ced-3 double mutants.
-
ced-9: The protecting gene
(Anti-apoptotic Bcl-2-like protein)
How to kill a c.elegans cell?
–> upregulating egl-1
Cancer and apoptosis: cancer involves the unchecked growth and accumulation of cells, and a crucial aspect of this process is the imbalance between cell proliferation and programmed cell death, particularly the resistance of cancer cells to undergo apoptosis. Understanding these dynamics is essential for developing strategies to target and treat cancer.
