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doi: 10.1242/10.1242/dev.00284

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Polarization of the C. elegans zygote proceeds via distinct establishment and maintenance phases

¹ Department of Molecular Biology and Genetics, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205, USA
² Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, USA

View larger version (147K): [in a new window]   Fig. 1. GFP:PAR-2 and GFP:PAR-6 dynamics in wild-type and mutant embryos (Movies 1-2, 10-11, 20-21). (A) Pronuclear formation. (B) Cessation of ruffling in posterior. (C) Pseudocleavage. (D) Pronuclear meeting. (E) Spindle rotation. (F) Mitosis. (G) two-cell stage. Quicktime movies of this and subsequent figures are accessible at ftp://www.wormbase.org/pub/wormbase/datasets/seydoux_2003. C. elegans zygotes are approximately 50 µm in length.

View larger version (65K): [in a new window]   Fig. 2. GFP:PAR-2 and GFP:tubulin dynamics in wild-type embryos. (A-D) An embryo expressing both GFP:PAR-2 and GFP:tubulin (Movie 3). (E-H) An embryo expressing GFP:PAR-2 (Movie 4).

View larger version (95K): [in a new window]   Fig. 3. GFP:MEX-5 and PIE-1:GFP dynamics in wild-type embryos (Movies 5 and 6). (A) Pronuclear formation. (B-D) Pronuclear migration. (D) Pseudocleavage. (F) Pronuclear meeting. (G) Mitosis. (H) Two-cell stage. Note that PIE-1:GFP accumulates in the male pronucleus during pronuclear migration, on the posterior centrosome during mitosis, and in the P1 nucleus in the two-cell stage. PIE-1:GFP and GFP:MEX-5 also accumulate on the cytoplasmic P granules (best seen in the movies).

View larger version (116K): [in a new window]   Fig. 4. GFP:PAR-2 and GFP:PAR-6 dynamics in nmy-2(RNAi) embryos. A,D,G,J are embryos during pronuclear migration; B,E,H,K are at pronuclear meeting; C,F,I are in late mitosis; L is a 2-cell embryo. GFP:PAR-2 remains off the cortex in the most severely affected embryos (A-C; Movie 15), unless par-6 is also removed (D-F, Movie 16). In more weakly affected embryos [nmy-2(RNAiClassII)], GFP:PAR-2 appears on the cortex around pronuclear meeting (H, Movie 18). Note PAR-2 accumulation in foci around pronuclei and/or spindles (C,H,I). GFP:PAR-6 remains uniformly distributed throughout the cortex even in the most weakly affected embryos (J-L, Movie 17).

View larger version (123K): [in a new window]   Fig. 5. PAR-1 and MEX-5/6 function in a feedback loop to regulate expansion of the posterior domain. (A-F) par-1(RNAi) zygote expressing GFP:PAR-2 (Movie 22). (A) Pronuclear formation. (B) Pronuclear migration. (C) Pronuclear meeting. (D-E) Mitosis. (F) Two-cell stage. The GFP:PAR-2 domain is expanded at pseudocleavage compared to wild type (see Fig. 1). (G-L) mex-5(RNAi)mex-6(RNAi) zygote expressing GFP:PAR-2 (Movie 28). Stages same as in A-F. GFP:PAR-2 domain expansion is slowed down compared to wild type (see Fig. 1). (M) GFP:PAR-2 in par-1(RNAi) pie-1(RNAi) (Movie 27). Expansion of the PAR-2 domain is not suppressed (compare with B). (N) GFP:PAR-2 in par-1(RNAi) mex-5(RNAi)mex-6(RNAi) (Movie 26). Expansion of the PAR-2 domain is suppressed. Identical results were obtained in par-1(it51) mex-5(RNAi)mex-6(RNAi) embryos stained for PAR-2 (not shown). (O,P) Wild-type (O) and mex-5(zu199)mex-6(pk440) (P) zygotes stained with PAR-1 antibody. Arrows point to the ends of the PAR-1 domain in the wild-type embryo. PAR-1 is not visible on the cortex of this mex-5(zu199)mex-6(pk440) zygote. (Q,R) Wild-type (Q) and mex-5(zu199)mex-6(pk440) (R) zygotes stained with PAR-3 antibody. PAR-3 extends further posterior.

View larger version (111K): [in a new window]   Fig. 6. Regulation of PIE-1's nuclear:cytoplasmic ratio by MEX-5/6 is under PAR-1 control. GFP:MEX-5 is primarily cytoplasmic in all genotypes. In contrast, the nuclear:cytoplasmic ratio of PIE-1:GFP varies depending on the genotype as described in text.

View larger version (39K): [in a new window]   Fig. 7. Model for polarization of the C. elegans zygote. (A) Graphic depiction of the establishment and maintenance phases during polarization of the zygote. Proteins that localize to the anterior are in blue (PAR-3, PAR-6, PKC-3 on the cortex, MEX-5 and MEX-6 in the cytoplasm). Proteins that localize to the posterior are in pink (PAR-2 and PAR-1 on the cortex, PIE-1 in the cytoplasm). During meiosis, all are uniformly distributed throughout the zygote (purple color). Circles: pronuclei, Black lines: microtubules. (B) Sequential repression model [modified from Kemphues (Kemphues, 2000)]. Lines with bars depict antagonistic interactions, whereas lines with arrows depict positive interactions. In this model, we show that PAR-1 is restricted to the posterior by the anterior PARs. This hypothesis is supported by the observation that, in late one-cell embryos, PAR-1 is present throughout the cortex in par-3 mutants and par-3;par-2 double mutants (Etemad-Moghadam et al., 1995; Boyd et al., 1996). Further support for this model awaits analysis of PAR-1 dynamics in live embryos.

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