Programmed cell death is a common mechanism for cell suicide in all multicellular organisms that is needed during development and tissue homeostasis. Several components of the C. elegans cell death machinery are highly conserved throughout the animal kingdom. For example, there are mammalian homologues for the C. elegans genes
ced-3 and
ced-9, the cell death caspases and the
bcl-2 gene, respectively. In contrast, other components of the C. elegans cell death machinery, for instance the adaptor molecule CED-4, are not highly conserved. Recent studies in the mouse and in Drosophilaindicate that the
apaf-1 genes of these organisms have a function similar to
ced-4. However, the
apaf-1 genes in the mouse and in Drosophila have additional structural domains not present in
Cel-ced-4. Therefore, the extent of conservation of the detailed molecular mechanism of programmed cell death remains an open question. In the ventral epidermis of Pristionchus pacificus, seven of 12 epidermal precursor cells die during late embryogenesis. We have isolated and characterized several cell-death defective mutants in which these cell deaths do not occur. These mutants fall into two complementation groups. The first complementation group corresponds to the cell death caspase
Ppa-ced-3 and results in animals defective in cell death execution (Sommer et al., 1998). We are currently focusing on the second group, which we have designated
ipa-2 (Inhibitor of P-ectoblast apoptosis). One candidate for this mutant is the gene encoding for the adaptor molecule CED-4. To test this hypothesis, we wish to clone the
Ppa-ced-4 gene. Having failed to clone
Ppa-ced-4 using PCR and low stringency hybridisation, we are now using the yeast two hybrid system to clone
Ppa-ced-4. For that purpose we cloned
mac-1 from P. pacificus.
mac-1 encodes an AAA-ATPase whose C. eleganshomologue binds specifically to C. elegans CED-4 in the yeast two hybrid system (Wu et al., 1999). PCR cloning of
Ppa-mac-1 revealed that the gene is SL1 spliced, unlike
Cel-mac-1, which is SL2 spliced. The yeast two hybrid screen, using Ppa-MAC-1 as a bait, is in progress. References: Sommer, R. J. et al., Development 125, 3865-3873 (1998) Wu, D. et al., Development 126, 2021-2031 (1999).