Vitor J. Barbosa, Frankie Kimm, Ruth Lehmann.
Developmental Genetics, NYU, Skirball Inst, New York, NY.
The establishment of dorsal-ventral polarity in the Drosophila
oocyte requires the activity of the germline Gurken/TGF
α (GRK) ligand to be correctly controlled both in time and in space.
We did a mutagenesis screen using the ventralization of the eggshell
(the “spindle” phenotype) as readout of defective GRK signaling in
order to understand how events such as recombinational double strand
break repair, microtubule-based transport and cytoskeleton polarity
influence oocyte organization. Ventralized eggs were derived from
germ line clones homozygous for the mutagenized 2R chromosome arm.
After screening 8179 lines we isolated 119 “spindle” mutants. These
have been divided, thus far, into 16 groups by complementation
analysis, which is still in progress. Preliminary analysis of the
phenotype in 6 of these groups showed a consistent correlation
between the ventralization of the eggshell and Gurken activity
either by lack of Gurken protein or by its mislocalization within
the oocyte. We also performed epistatic analysis with these groups
and mutants of both the meiotic checkpoint and the pathway leading
to the formation of double-strand DNA breaks prior to meiotic
recombination. These tests showed that the activity of these genes
is independent of the meiotic checkpoint or downstream of it.
Curiously, two of these mutations also affected the architecture of
the egg chamber: one leading to misplacement of the oocyte nucleus,
which was identified as a mutation in the kinesin heavy chain
(Khc) gene and the other to a misplaced oocyte within the egg
chamber. Two other complementation groups appear to be involved in
the process of oocyte specification. In parallel with the phenotypic
studies we are currently in the process of mapping and molecularly
identifying the genes isolated.