Worm Breeder's Gazette 14(5): 33 (February 1, 1997)

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.

Mosaic analysis of let-60 suggests a limited role for Ras in later development in C. elegans

John Yochem, Meera Sundaram, Min Han

Dept. of MCD Biology University of Colorado Boulder, CO 80309-0347

The requirements for LET-60 Ras during later stages of development were
determined by means of generating animals mosaic for let-60(sy101sy127),
a likely null allele [the strain used has the genotype: ncl-1(e1865)
unc-36(e251); let-60(sy101sy127); him-5(e1490);
kuEx72(ncl-1+unc-36+let-60+)]. As previously reported, an analysis of
hermaphrodites has revealed a requirement for let-60 activity in the sex
myoblasts for their terminal positioning (Sundaram et al., 1996). Mosaic
animals have also been examined for other properties, revealing several
expected and unexpected observations, including a very restricted lethal
focus for embryonic expression of the gene.

An expectation based on laser ablations and genetic analyses is a
requirement for let-60 in the vulval precursor cells. This expectation
has been completely met: seven of seven larvae observed at the L3 stage
as lacking the array in all six vulval presursor cells as a consequence
of two independent losses had a complete failure of induction. Based on
hermaphrodites in which the vulval precursor cells were themselves
mosaic, let-60 activity is absolutely required for induction of the
primary cell fate but not for the secondary fate, in complete agreement
with mosaic analyses of let-23 (Simske & Kim, 1995; Koga & Oshima,
1995). An absence of let-60 activity in the germ line results in a
failure of germ nuclei to exit pachytene during oogenesis, in agreement
with mosaic analysis of the gene encoding MAP kinase (Church et al.
1995). The morphology of the copulatory spicules is affected in males
lacking let-60 activity in parts of the cell lineage generating the
blast cell B. In contrast, males in which B but not F retains the array
have morphologically normal spicules, in agreement with a model proposed
by Chamberlin and Sternberg (1994). 

Animals homozygous for let-60(sy101sy127) die in the first larval stage
with a fluid-filled morphology. The cause of death has remained unknown,
but the mosaic analysis gave the first indication that it results from
an absence of an excretory duct cell, as follows. The lethal focus for
embryonic expression of let-60 is very restricted in the cell lineage
and exhibits lineal plasticity: the only losses not seen were AB and
ABp, but animals with losses in ABpl or ABpr were readily obtained. This
paradox was resolved by an examination of the excretory duct cell
(normally ABplpaaaapa) in 54 animals with losses in ABpl or ABplp. The
nucleolus of this cell was nonNcl in each of these animals, indicating
that it had inherited the array and could not have derived from ABpl, as
in normal development. A lack of let-60 activity in ABpl versus ABpr
appears to result in a switch of cell fates reminiscent of the switch
that occurs when ABplpaaaap, normally the mother of the duct cell, is
ablated (Sulston et al., 1983). Moreover, animals homozygous for
let-60(sy101sy127) lack a duct cell, whereas being homozygous for an
activating allele (n1046) of let-60 frequently results in the presence
of two duct cell nuclei in one animal. A discrepancy between the lethal
focus for let-60 and those (ABplp and part of ABal) for let-23(mn23)
(Koga & Oshima) might be explained by different degrees of
maternally-provided activities.

Chamberlin & Sternberg, Development 120: 2713 (1994); Church et al.,
Development 121: 2525 (1995); Koga & Oshima, Development 121: 2655
(1995); Simske & Kim, Nature 375: 142 (1995); Sulston et al., Dev. Biol.
100: 64 (1983); Sundaram et al., Development 122: 2823 (1996).