Worm Breeder's Gazette 7(2): 25
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 analyzing zgy-11 II (0.8 map units to the right of dpy-10; between tra-2 and vab-9 by deficiency mapping*). This locus was first identified by tsb2, a maternal effect zygotic lethal. Preliminary observations of tsb2 embryos suggested that zgy-11 plays a role in the establishment or maintenance of polarity in the early embryo. In eggs from homozygous tsb2 at 25 C, the first cleavage furrow is often improperly positioned, giving rise to even cleavages and cleavages with reversed asymmetry. Three new alleles of zgy-11 (tsb271, b272, tsb273) were isolated in complementation screens of 6000 heterozygous F1 progeny of tsb2 males and mutagenized dpy-10 hermaphrodites. Two other alleles, mn40 (from R. Hernan) and ct-1 (isolated by J. Laufer) were identified by complementation tests. mn40 is suppressible by sup-7 X (Newsletter, Vol.7, no.l, p.87). Phenotypic analysis thus far has concentrated primarily on tsb2 and mn40 and has utilized Nomarski microscopic observations of early development in live worms, tcrit analysis using short pulses of high temperature in individual embryos, and staining of fixed worms and embryos with DAPI and/or anti-tubulin antibody (as per Albertson et al. , Newsletter Vol.7, no.1, p.73). All alleles of zgy-11 show similar defects but the expressivity and penetrance differ. The first detectable abnormality occurs at metaphase II of meiosis. Embryos cut from tsb2 and mn40 homozygotes and stained with DAPI show a three fold increase, relative to N2, in the proportion of one-cell embryos with metaphase II figures. This observation is consistent with an increase in the proportion of metaphase II spindles observed in preps stained with anti-tubulin. Normal resolution of meiosis II seems to fail completely--with DAPI staining second polar bodies are seen with low frequency in tsb2 embryos and have not been observed in mn40 embryos. Furthermore, the time between the extrusion of the first polar body and the appearance of pronuclei is increased three to four-fold in tsb2 and mn40 respectively. During this extended period, large areas of cytoplasm form which exclude cytoplasmic granules, and extremely vigorous cytoplasmic movements occur. When (and if) pronuclei form, they often appear abnormal in position, number, size, and morphology. The effect of these early defects on the first cleavage is variable (even in mn40, a presumed null allele). Spindles can form with reversed or normal asymmetry or with near symmetry (as was first reported for tsb2). Often, however, spindles in one-cell embryos are quite abnormal (e.g. multipolar or disorganized). Finally, in most cases, multiple nuclei form at the end of first cleavage, irrespective of the polarity of cleavage. These nuclei often appear to contain fragmented chromosomes. We have been able to elicit the mutant phenotype in tsb2 embryos with a temperature pulse of 25 C for 15 minutes. Embryos are refractory to shifts up after pronuclear fusion and seem to be most sensitive around the time of meiosis. We do not yet know to what extent the tcrit extends into oogenesis. Based on our previous results with sup-7 suppression of mn40, zgy-11 appears to encode a protein which is translated prior to the time of zygotic gene expression. The more recent observations suggest that the time of action of the zgy-11 protein is near the time of meiosis and may be specific to a time between meiosis I and pronuclear formation. Expression of zgy-11 does not seem to be required at any time other than oogenesis. Spermatogenesis appears normal in mutant hermaphrodites, and in at least one non-conditional mutant, b272, male fertility is unaffected. Most alleles are not rescued by mating to wild type males. However, tsb273, the least severe zgy-11 mutant, can be rescued by mating with N2 males. Rescue is not dependent on the wild type allele and cannot be effected by tsb273 males raised at high temperatures. This suggests that the zyg-11 gene, while not essential for spermatogenesis, may be expressed and its products stored in sperm. * Deficiencies generously provided by Chris Sigurdson.