Worm Breeder's Gazette 12(1): 48 (September 1, 1991)
These abstracts should not be cited in bibliographies. Material contained herein should be treated as personal communication and should be cited as such only with the consent of the author.
We have previously described the isolation and preliminary characterization of ced-9 ,a gene involved in the control of programmed cell death in C. elegans (Hengartner, Ellis, and Horvitz, WBG 11 (2), 102, 1990). The gain-of-function allele ced-9 ( n1950sd )prevents cell deaths, whereas loss-of-function ced-9 alleles result in partial sterility and maternal-effect (F2) lethality. Mutations in ced-3 and ced-4 ,which block essentially all programmed cell deaths, suppress these ced-9 (lf)phenotypes, suggesting that the defects associated with the loss of ced-9 function are caused by inappropriate activation of the pathway for programmed cell death.
We now have studied ced-9 (lf)progeny of ced-9 (lf)mothers, hoping to identify the defect(s) leading to the maternal-effect lethality. Embryos generated by mothers homozygous for the weak allele ced-9 ( n1950 n2161 )develop normally up to approximately the 200-cell stage (AB128), at which point many cell corpses gradually start to appear. Using the 4D-microscope, we determined the cell lineages in a single ced-9 ( n1950 n2161 )embryo and were able to identify 49 of the earlier appearing corpses (which represent less than half of the total number of corpses eventually present). These studies revealed: (1) Four of those 49 corpses were cells that also die in the wild type; the others were the result of ectopic cell death. (2) Most of the ectopic deaths occurred in the AB lineage, although some MS deaths and at least one C death were also observed. (3) The 45 ectopic cell deaths resulted in 78 cells not being generated. Most (50) of these cells were neurons or glial cells. However, mothers of hypodermal cells and muscle cells (body wall, pharyngeal, and defecation muscles) as well as a few postembryonic blast cells were also observed to die. (4) Although some of the ectopic cell deaths were of cells that in the wild type generate descendents that die, some were not, indicating that they were not simply a consequence of "premature activation" of the pathway for programmed cell death.
We were unable to discern any obvious pattern to the ectopic cell deaths. It is possible that we missed an underlying pattern because we defined only partially the set of cells that can die in ced-9 (lf)animals. Although we extensively studied only a single animal, we believe that there is variability in the cells that die, since the homologs of cells that undergo ectopic deaths often did not die. The time of appearance of the first corpses and the general extent of ectopic cell deaths seem to be fairly reproducible among animals.
We have also studied embryos from mothers homozygous for the strong allele ced-9 ( n1950 n2077 ).Surprisingly, the defects and terminal phenotype associated with this allele are quite different from what we observed for n1950 n2161 embryos. F2 n1950 n2077 embryos arrest much earlier in development, with different individuals having from a few dozen to a few hundred cells at most. The embryos invariably look sick; cell divisions are slow and asynchronous. In those animals that develop sufficiently far cell corpses start appearing at about the same point in development as in n1950 n2161 embryos. We followed the lineage of a single ced-9 ( n1950 n2077 )embryo with the 4D-microscope. This embryo arrested with 57 cells (AB(32), MS(4,) C(4)). Nothing resembling a programmed cell death was observed. However, defects in cell division and cytokinesis were apparent: two of the AB(8) cells divided one and two generations later than the other AB cells and gave rise to four and seven daughters, respectively. These results suggest that DNA replication and centriole duplication occurred in these cells, but that M phase was blocked. Cells damaged with a laser beam can behave in this way. A few examples of incomplete cytokinesis were also observed, in which cases binucleate cells were formed. The nature of these defects and the general sickness of these embryos raise the question as to whether they are direct effects of a lack of zygotic ced-9 function or just secondary consequences of events in the maternal germline. Either way, the fact that these defects are completely suppressed by mutations in ced-3 or ced-4 strongly suggests that they're a consequence, either direct or indirect, of ectopic activation of the pathway for programmed cell death.