Table 1.
DBL-1 is a dose-dependent regulator of body length and annular width.
Figure 1.
GFP-tagged DBL-1 is bioactive.
(A) Schematic diagram of GFP-tagged DBL-1 expressed from the dbl-1 promoter (dbl-1p). The GFP-tag (green) is inserted downstream of the prodomain (blue) and upstream of the DBL-1 mature domain (red). The construct also contains the dbl-1 specific 3′ untranslated region (UTR). (B–E) Body lengths of wild-type (B), dbl-1(++) (C), dbl-1(nk3) (D), and dbl-1(nk3) mutant animals expressing the GFP-tagged DBL-1 transgene (E). Scale bar = 100 µm.
Figure 2.
DBL-1 regulates cuticular permeability.
(A-D) Sensitivity to levamisole (A), tricaine (B), IP2P (C), and sodium azide (D) was measured over time in animals with wild-type, increased (dbl-1(++)), and reduced (dbl-1(nk3)) DBL-1 pathway signaling. (E) Sensitivity to different levamisole doses by animals with wild-type, reduced (dbl-1(nk3)), and increased (dbl-1(++)) DBL-1 pathway signaling. (F) Sensitivity to levamisole in animals with different DBL-1 levels treated with C06C3.5 (pseudogene control) or dpy-13 RNAi was measured over time. (G–I) Hoechst 33342 staining in dbl-1(++) (G) or dbl-1(nk3) (H) animals. Faint intestinal autofluorescence is visible in (G). Scale bar = 10 µm. The fraction of animals that stain with Hoechst 33342 is shown in (I). Error bars indicate the mean ± SEM. P-values compare data to wild type (***P≤0.0001; **P≤0.001; *P≤0.05) using the unpaired t-test. Chemical structures were drawn using Jmol: an open-source Java viewer for chemical structures in 3D. http://www.jmol.org/. Molecule key: gray, carbon; blue, nitrogen; purple, sodium; green, chloride; yellow, sulfur; and red, oxygen.
Figure 3.
DBL-1 signaling affects surface adhesion.
(A) Six worm-stars show larval and adult dbl-1(nk3) hermaphrodites knot by their tails. Scale bar = 0.5 mm. (B, C) dbl-1(nk3) adult animals become tangled by their tails, forming moderate (B) or dense (C) worm-star aggregates in liquid. Scale bars = 0.5 mm.
Figure 4.
DBL-1 signaling affects specific cuticular surface properties.
(A–C) Rhodamine-conjugated wheat germ agglutinin (WGA) staining in wild-type (A), dbl-1(++) (B), and dbl-1(nk3) (C) populations. Scale bar = 100 µm. (D–F) WGA staining in him-5(e1490); srf-5(ct115) animals with C06C3.5(RNAi) (pseudogene control RNAi) (D), lon-2(RNAi) (E), and dbl-1(RNAi) (F). Scale bar = 100 µm. (G–H) Staining of annuli in wild-type (G), dbl-1(++) (H), and dbl-1(nk3) (I) animals. Bars mark the length of 10 annuli and indicate the average length of 10 annuli for each strain. (J–L) COL-19:GFP expression in otherwise wild-type animals with C06C3.5(RNAi) (pseudogene control RNAi) (J), lon-2(RNAi) (K), and dbl-1(RNAi) (L). Scale bar = 10 µm.
Figure 5.
The DBL-1 pathway regulates cuticular organization and composition.
Transmission electron microscopy (TEM) micrographs of wild-type (A), dbl-1(++) (B), and dbl-1(nk3) (C) animals. C indicates cortical layer; M indicates medial layer; B indicates basal layer; and the arrow marks the surface coat and epicuticular layer. Scale bars = 1 µm.
Table 2.
DBL-1 levels affect the dimension of cuticular components.
Figure 6.
Model of DBL-1 pathway-mediated cuticular phenotypes.
Model of how DBL-1 controls organization and composition of the cuticle, which affects body length, permeability barrier function, and worm-star formation in animals with normal (A), increased (B), and decreased (C) DBL-1 pathway signaling. Cuticle layers are indicated on the left.