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The expression of TGFß signal transducers in the hypodermis regulates body size in C. elegans

Department of Biology, Queens College, and The Graduate School and University Center, CUNY, Flushing, NY 11367, USA

View larger version (27K): [in a new window]   Fig. 1. Seam cell, pharynx and body length measurements of dbl-1, sma-3, sma-4 and sma-1 mutants as a proportion of wild-type (N2) size.

View larger version (99K): [in a new window]   Fig. 2. The sma-3::gfp(C) expression stage and pattern in wild type. (A,C,D,F,H) Direct fluorescence from GFP; (B,E,G,I) Nomarski images of the same samples to their left. (A,B) The expression begins at the late embryo stage, 2.5-fold stage. (C) The gene expression pattern in L3 stage worms. At L4 stage, sma-3 is expressed in pharynx, head region hypodermis (D,E), intestine (F,G) and body hypodermis (H,I). The arrows indicate the hypodermal nuclei. The arrowhead indicates an intestinal nucleus.

View larger version (80K): [in a new window]   Fig. 6. Expression of SMA-3::GFP C-terminal fusion protein in different genetic backgrounds. (A-F) sma-3::gfp C-terminal fusion was co-injected with sma-3 genomic fragment, which contains the whole sma-3 gene sequence. (A,B) sma-3(wk30) mutant background. (C,D) sma-4(e729) mutant background. (E,F) sma-6(wk7) mutant background. (G,H) sma-3::gfp C-terminal fusion was co-injected with sma-3 genomic lacking coding sequences. All of the photos are taken with the same exposure time. The prominent fluorescence in B is autofluorescence of the intestine. (I) Western blot of total protein extracts from strains carrying sma-3::gfp(C) constructs. The extra-chromosomal arrays were integrated into N2 (qcIs12[sma-3::gfp(C) + sma-3 + rol-6] and qcIs16[sma-3::gfp(C) + sma-3no-code + rol-6]) and crossed into different genetic backgrounds. Equal amounts of total protein were loaded in each lane and SMA-3::GFP fusion protein was detected by western blot with anti-GFP antibody.

View larger version (108K): [in a new window]   Fig. 3. The sma-3:gfp(N) expression and localization in different mutant backgrounds: (A,B) sma-3(wk30); (C,D) sma-4(e729); (E,F) sma-2(e502); and (H,I) sma-6(wk7). (A,C,E,H) show direct fluorescence from GFP; (B,D,F,I) are the Nomarski images of the same samples. The arrows indicate hypodermal nuclei (A,C,E). The arrowhead indicates an intestinal nucleus (A).

View larger version (90K): [in a new window]   Fig. 4. The sma-3 expression with tissue-specific promoters. Worms carrying each array are picked during L4 stage and photographed 24 hours later. The A1-H1 photos (first and third columns) show the overview of the expression pattern and the body size. The A2-H2 photos (second and fourth columns) focus on the region that has strong GFP expression. The tissue specific promoter used in each set is indicated in the A1-H1 photos. Scale bar: 0.5 mm in A1-H1 photos.

View larger version (117K): [in a new window]   Fig. 5. The distinct intensity of fluorescence in worms with different sma-3::gfp constructs in the sma-3(wk30) background. (A,B) sma-3::gfp(N); (C,D) sma-3::gfp(C); (E,F) co-injection of sma-3::gfp(N) and sma-3::gfp(C). (A,C,E) The head region; (B,D,F) the body hypodermis. All of the worms (L4 stage) are grown under the same conditions and the photos are taken with the same exposure time.