Fig. 2. Expression pattern of
vha-8 in C. elegans. (A) Northern blot analysis with 32P-labeled
vha-8 probe detected two bands, 1.3 kb and 1.0 kb at all developmental stages: E, embryos; L2-L4, larval stages; A, adults. (B) Western blotting with anti-VHA-8 antibodies recognized a 29 kDa protein in wild-typeN2 worm lysate. (C-E) Green fluorescent protein (GFP) reporter gene expression pattern of
vha-8. Expression of GFP is under the control of 50 upstream regulatory sequences and coding regions of
vha-8. Strong signals were detected in H-shaped excretory cell (C,E). (F-K) Immunofluorescence localization of VHA-8 in wild-type worms using anti-VHA-8 antibody. Localization of VHA-8 in excretory canal was observed in 1.5-fold stage embryo (F), larva (I), and adult (K). (D,G,J) Nomarski images of (C,F,I), respectively. (H) DAPI- stained embryo shown in (F). (E,K) Magnified images of canal structures of the excretory cell. (L,M) Immunogold electron micrograph showing the sub-cellular localization of VHA-8. Signals of gold particles are seen to cluster in the cytoplasm of excretory canal (L, arrowheads). VHA-8 signals were also observed in stacked sheets of the apical plasma membrane of syncytial hypodermal cells (M, arrowheads). Scale bars, 20 um (C-K) or 1 um (L,M).