Worm Breeder's Gazette 9(3): 94
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.
Seven recessive alleles of the gene glp-1 have now been isolated
using EMS mutagenesis, by both direct and complementation screens.
Five of these seven alleles, including the two isolated by
complementation, have the same phenotype; Z2 and Z3 divide a few times
resulting in a small number of germ cells which then all enter meiosis
prematurely and form sperm. This phenotype is completely penetrant and
is the strongest that we have seen for mutations in this gene (in the
other two alleles, Z2 and Z3 go through a greater number of divisions
before entering meiosis). The strength of this phenotype and frequency
of mutations with this phenotype suggest that it may be the null
phenotype of glp-1We have begun a mosaic analysis to determine in
which cell(s) the gene product is required for wildtype germline
development. An unstable free duplication was created by gamma-ray
mutagenesis of mnDp37 provided by Bob Herman. The new duplication,
qDp3, has breakpoints between dpy-17 and ncl-l on the left and between
glp-1 and unc-69 on the right. Because qDp3 covers the gene ncl-l, we
have been able to make use of the cell autonomous nature of the
phenotype of ncl-l(e1865) as a marker for the presence or absence of
the duplication, as suggested by Ed Hedgecock (WBG 9(1),66-67.).
The strain we are using for this mosaic analysis is ncl-l(e1865)unc-
36(e251)glp-1(q46)III;qDp3. The unc-36 phenotype depends on the
presence or absence of the wildtype unc-36 gene product in the AB.p
lineage (Kenyon,C. Cell 46, 477-487.). Therefore, by screening for
animals which are nonUnc but which are Glp it is possible to isolate
mosaic animals in which the duplication has been lost in cells that
require the wildtype glp-1 gene product for normal germline
development. These nonUnc Glp mosaics are then screened, by Nomarski,
for the presence of the Ncl phenotype to determine where the
duplication has been lost. Because the distal tip cells are known to
play an important role in the development of the germ line, we thought
it most likely that the wildtype glp-1 gene product would be required
in either the distal tip cells or in the germ line. For this reason,
we have concentrated on looking for duplication losses in cells
descended from Pl.
Our preliminary results are shown below. For each nonUnc Glp animal,
the presence or absence of qDP3 in the MS, C and D lineages was
determined by scoring the Ncl phenotype of body wall muscle cells
derived from these embryonic precursors. In the case of MS the sheath
cells and distal tip cells of the somatic gonad were also scored when
possible (in some animals only one distal tip cell nucleus could be
clearly observed). Because of the nonUnc phenotype of these animals it
was assumed that the duplication was present in at least some of the
cells derived from AB. The nonUnc Glp animals examined so far fall
into six classes based on their pattern of duplication loss. It should
be pointed out that in one group of animals (class VI) no Ncl nuclei
were observed. Such animals could be the result of a recombination
event between the duplication and the chromosome or they may have lost
the duplication in a tissue which could not be scored. For example,
because the nonUnc Glp animals are sterile, it is not possible to test
directly for the presence of qDp3 in the germ line.
So far, the results suggest that glp-1 acts in the germ line. The
presence of qDp3 in MS in the animals in class II indicates that the
presence of the glp-1 wildtype gene product in the somatic gonad is
not sufficient for normal germline development. In each of these
animals sheath cells in both gonadal arms and at least one distal tip
cell were scored and all were Ncl+. In addition, a mosaic animal has
been found (class VII) in which the anterior distal tip cell is Ncl
but germline development appears to be normal in both gonadal arms.
Thus, the wildtype glp-1 gene product in the distal tip cell is not
necessary for normal germline development. A large number of mosaic
animals (classes I, II and III) were found in which qDp3 is absent in
D. Because the D lineage generates only body wall muscle cells, we
think it likely that, in these animals, qDP3 has been lost in P3 or
before and that it is the absence of the wildtype glp-l gene product
in the germ line which is causing the abnormal germline development.
In two animals (classes IV and V) there is no evidence of a
duplication loss in the divisions leading up to the formation of the
germ line. Because many of the animals we have observed appear to have
had more than one duplication loss our working hypothesis is that, in
these animals, an additional, undetectable, duplication loss has
occurred in the germ line. Alternatively, the glp-1 gene product may
be required in other tissues in addition to the germ line and we are
currently testing
this
possibility.{Figure 1}