Worm Breeder's Gazette 11(5): 79
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 are interested in understanding the positional information in C. elegans which we believe directs the region-specific expression of mab- 5. To approach this question, we are using a strain with a chromosomally integrated mab-5 promoter: -gal fusion (Salser and Kenyon, this issue). We have found that in this strain (and also in several transformants carrying the same fusion extra-chromosomally), - gal is first expressed in the embryo, at about 260 minutes after the first cleavage. At this point elongation has not yet begun. Most of the cells that will later be affected by mab-5 mutations have been born and are in the posterior region of the embryo. We haven't yet determined exactly which cells stain for -gal, but -gal staining is in the posterior, consistent with the position of the cells that are known to be affected by mab-5 mutations. This embryonic staining was surprising, since the known phenotype of mab-5 mutations is almost exclusively postembryonic. Nevertheless, it suggests that positional information that directs the expression of mab-5 exists in the embryo. The mab-5 promoter: -gal fusion thus serves as a marker to allow us to study anterior-posterior pattern formation in C. elegans embryos. We have begun a series of experiments to try to learn how this positional information might be set up in the embryo. One can imagine a number of possibilities. 'Posterior' cytoplasmic determinants might be segregated through the first several cell divisions, like P granules and intestinal determinants, to a blastomere which gives rise to a cell or cells which are later positioned in the posterior. This cell or cells would then induce the expression of mab-5 in the correct region. If this were true, one might expect the pattern of mab-5 expression to be perturbed in par mutants, since par mutations affect the known segregation events in the first several cell divisions. In addition, it should be possible to identify, by embryonic ablations, the blastomeres to which the 'posterior determinants' were segregated. On the other hand, the posterior-specific expression of mab-5 could be directed by a par-independent mechanism. One possibility for such a mechanism would be a gradient of a molecule such as retinoic acid, which would be independent of cell boundaries. So far, we have tested the effect of one par mutation, par-2(e2030ts) , on the expression of the mab-5: -gal fusion. 1- to 4-cell par-2( e2030ts) embryos carrying the mab-5: -gal fusion were dissected from mothers that had been at 25 C (the non-permissive temperature) overnight, and incubated 4-5 hours longer at 25 C. This was long enough for wild-type embryos to develop to 1 1/2 fold, and to stain for -gal. 50% of the e2030 embryos had abnormal morphogenesis and no gut granules (the other 50% were less severely affected.) In these embryos, nevertheless, -gal staining was still localized to the posterior. It is not yet clear how to interpret this, since e2030 is a weak allele in which segregation of P granules is relatively normal in the first several cleavages, and only fails in later cell divisions (Kemphues et al., Cell 52, 311.). We are in the midst of constructing strains with other par alleles. We have also asked whether glp-1 affects mab-5 expression. glp-1 is known to be involved in signalling pathways, and in the embryo is required in inductive interactions that specify the normal development of at least some AB-derived cells (Priess et al. Cell 51, 601). glp- 1(q231ts) embryos carrying the fusion were shifted from 15 C to 25 C at the 1-cell or 2-cell stage, and incubated long enough for wild-type embryos to express -gal. Despite the fact that q231ts embryos shifted to 25 C at the 2-cell stage develop abnormally and do not hatch (see Austin and Kimble, Cell 51, 589), the pattern of -gal staining in these embryos was similar to wild-type. Thus, although glp-1 could affect mab-5 expression in some cells, it must not play a major part in setting up the pattern of mab-5 expression. We think it is unlikely that mab-5 expression is directed solely by lineage rather than by positional information (the other kind of hypothetically possible model) since the cells affected by mab-5 mutations are not related by lineage. However, looking at the pattern of -gal expression in cleavage-blocked embryos carrying the mab-5: - gal fusion should help to distinguish between this type of model, and a positional information model. At the four cell stage all four blastomeres give rise to cells which will later express the mab-5: - gal fusion (unpublished observations.) If this were due to lineage- specific segregation of mab-5 determinants, one might expect all four blastomeres to express -gal in a cleavage blocked embryo. Instead, in three out of six embryos which were blocked from further cleavage at the four cell stage using cytochalasin D, P2 alone expressed -gal. ( Two out of six had no -gal expression, and the sixth stained in a position that was most consistent with P2, but the orientation of the embryo was such that it was impossible to be sure.) Thus, there appears to be some mechanism by which only this one blastomere has the potential to express the mab-5: -gal fusion. This result is more consistent with a positional information model than with lineage- specific segregation of mab-5 determinants.