Worm Breeder's Gazette 9(2): 70
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.
In an effort to understand the mechanism of determination in early C. elegans development, we have been using a screen devised by J. Priess et al. (C. elegans newsletter 8, #2, 5) to identify maternal- effect lethal (mel) mutations that affect the early embryonic cleavage pattern. This procedure requires first identifying the mel mutants, maintaining them in stock by selection, then observing early cleavages in embryos from mutant homozygotes of each strain. This approach has resulted in the identification of several mutants with aberrant cytoplasmic partitioning, defining three loci, designated par-1,2,3 ( Kemphues and Priess, Abstracts, 1985 C. elegans meeting, p15). However, because of the low frequency of par mutants among the total pool of maternal effect lethals, observing early development is not an efficient way to obtain large numbers of mutants with this phenotype. In addition, there may be mel mutants that affect determination without detectably affecting cytoplasmic partitioning. In an effort to overcome these problems we have begun to screen for mels that produce embryos arresting with large numbers of cells but which do not differentiate gut cells. This can be done very simply by scoring newly identified mel homozygotes (which are bags of dead embryos) under polarized light to visualize gut granules. Gut granules normally appear in cells of the gut lineage at about the 200 cell stage of embryogenesis. Mutants that produce embryos with large numbers of cells but no visible gut granules are potentially defective in some step in the determination or differentiation of the gut. In this screen, those mels that produce embryos with gut granules are discarded as are mels producing embryos with no gut granules and obviously small numbers of cells. We expect to find at least three interesting classes of mutations in such screens: 1) New alleles of par-1. Mutations in par-1 (b274, e2012) have been shown to affect partitioning of cytoplasm in the early cleavages and lead to arrested embryos with many differentiated cells, but no detectable gut granules. 2) Mutations in other loci that affect partitioning of cytoplasm. Because cytoplasmic localization appears to be important for gut determination, multiple loci controlling localization could mutate to a similar terminal phenotype. 3) Mutations that specifically affect the determination or differentiation of the gut. If there are maternally expressed molecules with a unique function in the determination or differentiation of the gut, mutations in genes encoding these molecules should surface in our screens. Our initial results are promising. We have screened roughly 3000 F1 clones from mutagenized adults and have identified four mel mutants whose progeny arrest in embryogenesis with large cell numbers but no gut granules. One of the mutants (it32) is a new allele of par -1 Two other mutations identify a new locus par-4 (it33, it47) V (near dpy-21). Progeny from par-4 mutants are defective in P granule localization and have synchronous early cell divisions, defects common to all par mutants. However, par-4 mutants differ from mutants at the other par loci in that the first division results in nearly normal size asymmetry. The fourth mutant has no obvious partitioning defect, but early divisions are slowed about two-fold compared to wild-type. Relative timing of divisions is normal, and the E lineage is normal at least to the 8 E-cell stage. The low frequency at which these mutants arise, coupled with the relative ease of screening should make it possible for us to satura te the genome for loci that can mutate to this phenotype.