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First published online 13 June 2007
doi: 10.1242/dev.008268

Development 134, 2685-2695 (2007)
Published by The Company of Biologists 2007
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Binary cell fate specification during C. elegans embryogenesis driven by reiterated reciprocal asymmetry of TCF POP-1 and its coactivator ß-catenin SYS-1

Shuyi Huang, Premnath Shetty, Scott M. Robertson and Rueyling Lin*

Department of Molecular Biology, University of Texas Southwestern Medical Center, 6000 Harry Hines Boulevard, Dallas, TX 75390, USA.


Figure 1
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  		Fig. 1. SYS-1 is required for the activation of Wnt target genes in E and gut formation. (A-D) Micrographs of C. elegans embryos carrying the Psdz-23::gfp::H2B transgene, in wild-type, sys-1(RNAi) or pop-1(zu189) backgrounds. Embryos are shown at the two MS, two E stage, as DIC (C,D) and corresponding nuclear GFP::H2B fluorescence (A,B) images. Wild-type (left of the dashed line) and RNAi/mutant embryos (right of the dashed line) were imaged together. Note reduced GFP fluorescence in sys-1(RNAi) and pop-1(zu189) embryos as compared with wild-type embryos. Four GFP-positive cells were observed in pop-1(zu189) embryos owing to derepression in the MS lineage. (E-G) Representative wild-type, sys-1(RNAi) and pop-1(zu189) embryos from A-D showing reporter GFP expression. (H-K) Terminally differentiated sys-1(RNAi) (H,I) or teIs3(Pmed-1gfp::pop-1); sys-1(RNAi) (J,K) embryos shown under DIC (H,J) or birefringence optics revealing intestinal-specific gut granules (white speckles in I,K). Embryos lacking gut granules are outlined in white. Scale bars: 20 µm.

 

Figure 2
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  		Fig. 2. Reciprocal nuclear asymmetry of GFP::SYS-1 and POP-1 between A-P sisters. Immunofluorescence micrographs of teEx321(Pmed-1gfp::sys-1) C. elegans embryos. (A-D) Embryos at 1MS/1E stage stained with (A) anti-GFP antibody, (B) anti-POP-1 mABRL2 and (D) DAPI to visualize nuclei. (C) Merged image. (E-L) Anti-GFP (left) and corresponding DAPI staining (right) at 2E stage (E-H), 4E stage (I,J) and 8E stage (K,L) of embryogenesis. White lines connect nuclei of sister cells born from A-P divisions; black lines connect nuclei born from left-right divisions. The cortical GFP signal often appeared asymmetric, being observed at the anterior cortex of anterior cells (white arrowheads) but not at the posterior cortex of posterior sisters (open arrowheads). Scale bar: 10 µm.

 

Figure 3
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  		Fig. 3. GFP::SYS-1 nuclear asymmetry between A-P sisters is Wnt signal- and proteasome-dependent. Embryos are shown at the 2MS/2E stage, with lines connecting the MSa/MSp and Ea/Ep sisters. (A) Immunofluorescence micrographs of GFP::SYS-1 in various mutant backgrounds (as indicated to the left). Left column, anti-GFP; right column, DAPI. (B) Independent regulation of GFP::SYS-1 (a,d,g) and POP-1 (b,e,h) asymmetry. (a-c) Wild type; (d-f) mom-5(or57) showing symmetric GFP::SYS-1 and asymmetric POP-1; (g-i) wrm-1(RNAi) showing asymmetric GFP::SYS-1 and symmetric POP-1. (C) Wild-type (a-c) and rpn-8(RNAi) (d-f) embryos double stained with antibodies to GFP and PIE-1. Arrow, germline precursor; arrowheads, somatic nuclei that stain weakly with the PIE-1 antibody. Scale bar: 10 µm.

 

Figure 4
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  		Fig. 4. Increased level of SYS-1 causes extra gut at the expense of pharynx. Immunofluorescence of teEx321(Pmed-1gfp::sys-1) (A,C) and wild-type (B,D) C. elegans embryos stained with 3NB12 for pharynx (A,B) or ICB4 for gut (C,D). The anterior and posterior tips of the pharynx are indicated by open and closed arrowheads, respectively. Scale bar: 10 µm.

 

Figure 5
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  		Fig. 5. Altering the SYS-1-to-POP-1 ratio also affects cell fate decisions at other A-P divisions. (A) Schematic of the C. elegans pharynx, including the location of the eight pharyngeal muscle types (labeled 1 through 8), with 3NB12 staining indicated (dark gray, no staining). (B) Embryos derived from wild type, teEx321(Pmed-1gfp::sys-1) and sys-1(q544/+); teIs3(Pmed-1gfp::pop-1) stained with 3NB12. Schematics to the right indicate the three pm6 and three pm7 muscle cells in the wild type and predicted cell fate changes following an anterior-to-posterior (middle) or a posterior-to-anterior (bottom) fate transformation at the division giving rise to MSa and MSp. Triangles and squares denote cells derived from the MSa and MSp lineages, respectively. (C) The E lineage. Horizontal lines indicate divisions, whereas vertical lines indicate developmental time. All divisions are anterior (left) to posterior (right), except those labeled l/r, which indicate left/right divisions. Circles, the four cells that comprise intestinal ring 1; triangles, the two cells comprising ring 9. (D) PHO-1::GFP expression in wild-type and teEx321 larvae. Arrow, posterior end of the pharynx; brackets, cells in the first two intestinal rings in wild type and the corresponding region in teEx321. Scale bar: 10 µm in B; 30 µm in D.

 

Figure 6
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  		Fig. 6. Pathways regulating the reciprocal asymmetry of POP-1 and SYS-1 in C. elegans. Schematic of the two pathways regulating nuclear SYS-1 and POP-1 levels. SYS-1 levels are elevated by the MOM-2/MOM-5/APR-1 pathway, and nuclear POP-1 levels are decreased by the MOM-4/WRM-1/LIT-1 pathway with input from the MOM-2/MOM-5/APR-1 pathway. Each pathway contributes to an increase in the SYS-1-to-POP-1 ratio, which drives the posterior cell fate. The asterisk denotes that POP-1 activity could be regulated via post-translational modification.

 

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© The Company of Biologists Ltd 2007