After a cell has died by programmed cell death, its corpse is quickly engulfed and broken down by one of its neighbors. Mutations in the genes
ced-1 and
ced-2 slow this process of phagocytosis, so that the corpses of dead cells accumulate during development and are easy to see. Although
ced-1 is a strictly zygotic gene, Hilary Ellis discovered several years ago that
ced-2(
e1752) mutants can be rescued if their mother provides wild-type
ced-2 product.
ced-2/+ animals born of a
ced-2 mother are also wild-type, so
ced-2 function can be provided by either the mother or the zygote. Hilary's observation explains why many
ced-1 alleles, but only one
ced-2 allele, have been found in screens of mutant F2 animals, and suggested that more genes like
ced-2 might exist. Recently, new mutations that cause cell corpses to persist have been isolated in our laboratory. Chand Desai found
n1812 as a spontaneous mutation in one of his stocks, and Michael Stern isolated
n1813 as a gamma-ray induced mutation. We tested these recessive alleles for complementation with
ced-1 and
ced-2 and with each other, and found that each mutation defines a new complementation group. Genetic mapping confirms that these are new ced genes:
n1812 is located on chromosome IV very near
mec-3, and
n1813 is on chromosome III between
lon-1 and the left end of nDf16. We have chosen the names
ced-5(
n1812) and
ced-6(
n1813) for these genes. Both
ced-5 and
ced-6 mutants show maternal rescue, just like
ced-2 mutants. However, careful examination of
ced-6 animals from a
ced-6/+ mother shows that though embryonic cell deaths are degraded normally, cell corpses formed later, during larval development, are not phagocytosed efficiently. This suggests that wild-type
ced-6 product put in the egg by the mother is used up or diluted out during larval development. We are now testing
ced-2 and
ced-5 for this phenotype. The gene
ced-5 is the only one of this group uncovered by a deficiency sDf2 fails to complement
ced-5(
n1812) for the phagocytosis of dead cells, and sDf2/ced-5 animals appear otherwise healthy, just as
ced-5 homozygotes do. When we compare newly hatched
ced-5(
n1812) 282) 91)/sDf2 animals with
ced-5(
n1812) 282) worms, we see twice as many cell corpses still present in the pharynx in the
ced-5/sDf2 animals. This suggests that
n1812 lowers but does not eliminate
ced-5 gene function. To identify more genes involved in the process of phagocytosing dead cells, we have developed a mutant screen that allows us to find genes of both the
ced-1 and
ced-2 varieties. We mutagenize
sem-4(
n1378) animals, which form bags-of-worms because they lack sex muscles ( Michael Stern, personal communication), and then screen F2 bags-of- worms using Nomarski microscopy to look for persistent cell corpses in the F3 embryos and larvae. Mutations that show maternal rescue like
ced-2 should appear as bags full of Ced worms. We are now analyzing several new ced mutations isolated from our first test of this procedure. Why is
ced-1 a zygotic gene while all of the other loci involved in the phagocytosis of dead cells show maternal rescue? One possibility is that the process of breaking down a dead cell involves two sets of genes: some are involved in specifically marking the cell corpse as an object that should be phagocytosed, and others encode products that are part of the cellular machinery necessary for phagocytosis. In this model it seems possible that genes in the first class could act only in dying cells, whereas genes in the second class could make products found in all cells, products that the mother also provides to the egg.