C.elegans

Question 1

How can development of C.elegans be studied? Give 4 categories.

Answer 1

• Anatomy
  
  • EM-based constructions
• Embryology
  
  • Blastomere manipultion, lineage
• Transgenics
  
  • Expression patterns, cis-regulatory analysis, rescue)
• Genetics
  
  • Forward genetic screens, mapping, epistasis analysis

Question 2

Describe the story of embryonic cell fate specification: determinant.

Question 3

Classical developmental embryology has distinguished two major modes of cell fate specification. Define determinant and inductive.

Answer 3

• Determinant: intinsic, cell autonomous, coupled to lineal descent of cell, no regulation following cell loss
• Inductive: indeterminate development, non-cell autonomous, no obvious correlation between lineage and cell fate, regulation following cell loss

Question 4

Describe the story of LR asymmetry in C.elegans.

Answer 4

• The C.elegans offers a a unique oppurtunity to understand how asymetric anatomical structures confer functional laterality.
• The anatomy of the C.elegans has been visualised by sequetial EM slices. From this there are clear LR asymmetries
• All cells in the C.elegans are generated from invariant cell lineages.
• Most of the nervous system derives from the found AB cell
  
  • The two daughter cells of the AB undergo a division in LR axis instead of the AP axis like all the other deceendants.
  • It is this early divison that creates LR asymmetry
• The ASE neuronal pair are the main taste receptors of the C.elegans. Whilst cell postition, axon and dendritic morphology as well as other defining charcteristics are symmetric, the expression of the putative guanyly cyclase class are not. This asymmetric receptor expression confers increased sensory capabilites.
  
  • This neuronal pair can act as a paradigm for LR asymmetry
• The ASE asymmetry is sterotyped and not stochastic, i.e directional asymmetry. The asymmetry of ASE progesses from a symmetric state. What causes this asymmetric termination?
• Serial analysis of gene expression (SAGE) identified genes of the ASE gustatory class on the molecular level.
• In vivo cis-regultory analysis identified a a cis-regulatory motif ‘ASE motif’ required for the expression of many ASE genes.
• The zinc-finger ranscription factor che-1 binds to this motif and is the terminal selector of the ASE neurons - che-1 is continually required.
• What are the other factors that work together with Che-1?
• Genetic screens have shown mutants with defects in transcription factors to have symmetric ASE neruons.
  
  • These screens are done to identify the trans-acting factors
  • RNAi screens were done
  • E.g. Identified lys-22 and lys-5 mutants that display a double left phenotype and lys-2 double right
  • They used epistasis analysis to understand where they these proteins acted
  • These and other factors are asymmetrically expressed in the ASE neurons - how this is established remains unknown.
• Found the LR asymmetry to be regulated by a bi-stable feedback loop
• What biases this bistable feedback loop?
  
  • They removed signalling molecules such as Notch by genetics
  • They removed all possible signalling cells by ablation.
  • Found ASE asymmetry is dependent on ABa/ABp identify
    
    • Asymmetric divergence in the pair is due to the delta/notch signal pathway.

Question 5

What is SAGE analysis?

Answer 5

produces a snapshot of the mRNA present in a sample of interest in the form of small tags that correspond to the fragment of interest