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}