Picture from Tcherepanova IY et al. (2000) J Biol Chem "Aspartic proteases from the nematode Caenorhabditis elegans. Structural ...."

Figure 7. Developmental expression of asp-1 mRNA. In A and B, C. elegans were fixed, permeabilized, and then hybridized with digoxigenin-labeled, ASP-1 antisense DNA as described under Experimental Procedures. RNA-DNA complexes were visualized by incubating the specimens serially with antidigoxigenin IgGs coupled to alkaline phosphatase and a chromogenic substrate. Alkaline phosphatase catalyzes the synthesis of an insoluble reaction product (shown as a black precipitate) in cells expressing ASP-1 mRNA. A, an L1 larva with ASP-1 mRNA located exclusively in the intestine. B, an L2 larva expressing ASP-1 mRNA in the intestine. The level of ASP-1 expression is significantly lower than that observed in younger nematodes. Arrows labeled ph and G indicate the posterior end of the pharynx and the developing gonad, respectively. C, total RNA was prepared from C. elegans embryos, L1 larvae, and adults and then subjected to Northern blot analysis. The amount of 32P-labeled ASP-1 probe bound was quantified by PhosphorImager analysis and then normalized to the level of myosin light chain mRNA as described previously (33). The data are means and standard deviations from three independent experiments.

Picture from Tcherepanova IY et al. (2000) J Biol Chem "Aspartic proteases from the nematode Caenorhabditis elegans. Structural ...."

Figure 6. Cell-specific expression of asp-1 promoter activity in C. elegans. Several independent lines of transgenic C. elegans carrying the asp-1/lacZ reporter transgene were created as described under Experimental Procedures. The histochemical stain for β-galactosidase produces an insoluble product in nuclei that are transcribing asp-1. A, reporter gene activity in the intestinal cells of developing embryos located in the uterus of a transgenic adult hermaphrodite. B, asp-1 promoter activity in the large nuclei of the intestinal cells of an L1 larva. The location of the int6 through int9 rings of intestinal cells is indicated. ph marks the location of the terminal bulb of the pharynx. C, high level asp-1 transcription in the posterior intestinal cells of an L3 larva. The nematodes were photographed using Nomarski optics as described previously (33).

Picture from Tcherepanova IY et al. (2000) J Biol Chem "Aspartic proteases from the nematode Caenorhabditis elegans. Structural ...."

Figure 8. Expression of ASP-1 in situ. The location of ASP-1 in intact C. elegans was determined by confocal immunofluorescence microscopy. A, the enrichment of ASP-1 along the intestinal lumen of an embryo at the 2- and 3-fold stages of development, 5 h prior to hatching (center) and 6 h prior to hatching (upper left). B, shows a higher magnification of a late embryo (3 h to hatch) in which aspartic protease-containing, punctate structures are observed adjacent to the intensely staining intestinal lumen. C, high level accumulation of ASP-1 along the intestinal lumen and in punctate structures of an L2 larva. The insert in each panel is a photograph using Nomarski optics of the identical field that is shown by immunofluorescence microscopy. The arrows indicate the location and orientation of the nematode pharynx, with the arrowhead directed toward the anterior of the pharynx.