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.