Worm Breeder's Gazette 8(1): 18
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.
Between the time of fertilization and first cleavage of C. elegans zygotes, several directed movements of cellular components, which reflect the asymmetry of the zygote, occur. The zygote undergoes contractions of the anterior membrane and pseudocleavage. The egg and sperm pronuclei migrate from opposite poles of the zygote, meet in the posterior hemisphere, and then move to the center. The first mitotic spindle forms in the center of the zygote, but becomes asymmetric with respect to its position along the anterior-posterior axis and the behavior and morphology of its asters. The segregation of P granules to the posterior pole of the zygote, which occurs concurrently with pseudocleavage and pronuclear migration, provides one of the most striking manifestations of zygote asymmetry. In order to determine whether specific cytoskeletal fiber systems are involved in P-granule segregation and other early movements and whether these early events are interdependent, I have analyzed mutant embryos and wild-type embryos treated with microtubule (MT) and microfilament (MF) inhibitors. Analysis of zyg-9(b244) embryos (described in a previous newsletter) and embryos treated with low levels of MT inhibitors (which partially phenocopy the b244 defect) demonstrate that P-granule segregation is not mediated by the mitotic spindle; even though spindle orientation is altered in mutant and treated embryos, P granules nevertheless coalesce and become localized in the posterior pole. Treatment of zygotes with levels of MT inhibitors (colcemid, vinblastine, or griseofulvin) that prevent mitosis does not prevent proper P-granule segregation or pseudocleavage but does inhibit pronuclear migration. Conversely, treatment of embryos with MF inhibitors (cytochalasin D or B) inhibits P-granule segregation and pseudocleavage without preventing pronuclear migration. In addition, the resulting spindle does not become asymmetric in position or in aster behavior in cytochalasin treated zygotes. The results suggest that P-granule segregation does not require either the spindle or cytoplasmic MTs, but that this process as well as generation of other asymmetries do require cytoskeletal functions that depend on MFs. Although MT-mediated events (pronuclear migration) and MF-mediated events (pseudo-cleavage, P-granule segregation) are both normally initiated at fertilization, these two classes of processes can be uncoupled by drugs or mutation and therefore appear to be independent. Details of these experiments will appear in Cell 35, pp. 15-25. In immunoprecipitation experiments designed to identify the molecular components of P granules, one of the anti-P-granule monoclonal antibodies (K76) precipitates a 37,000-dalton polypeptide and another anti-P-granule antibody (L416) precipitates an 83,000- dalton polypeptide from [35S]-worm homogenates. In gel blot experiments both of these antibodies bind to a ~37,000-dalton polypeptide in worm and egg homogenates. However, the K76 antibody also binds a higher and lower molecular weight species as well. Further investigation of the polypeptides recognized by these antibodies, as well as two other anti-P-granule antibodies, is under way.