Vacuolar-type ATPase in the accessory boring organ of Nucella lamellosa (Gmelin) (Mollusca : Gastropoda): role in shell penetration

Abstract

The structure and function of the accessory boring organ (ABO) of muricid gastropods has been described in numerous studies, and the ABO of Nucella lamellosa was found to be similar to those of other muricid species. The active cap region of the ABO is composed of tall, mitochondria-rich cells with distinct brush borders at their apicies, surrounding a hemolymph-containing central sinus. Using antibodies specific for vacuolar-type ATPase (V-ATPase), enzyme immunoreactivity was found to be limited to the brush border of the epithelial cells. Electron immunohistochemistry revealed that V-ATPase immunoreactivity resides in the plasma membranes of the microvilli. Immunodot blotting using yeast V-ATPase as a positive control confirmed the specificity of the reactions. SDS-PAGE of membrane suspensions from the ABO revealed protein bands of the requisite molecular weight for V-ATPase subunits. Western blots suggest that antibodies raised against mammalian V-ATPase subunits recognize subunits of the molluscan V-ATPase. The molecular weights of these identified subunits are similar to those in mammals. The V-ATPase-specific inhibitor bafilomycin A1 inhibited ATPase activity in samples of ABO homogenate by about 10% relative to control, providing further evidence for the presence of V-ATPase. Specific V-ATPase activity was about 67 picomoles of inorganic phosphate per microgram of protein per minute in the homogenate. Collectively this evidence strongly suggests that a vacuolar-type proton transporting ATPase is present in the brush border of the accessory boring organ of Nucella lamellosa, and is responsible for acidifying secretions from this gland. Similarities between the ABO, osteoclasts, and the mantle of freshwater bivalves also suggest that the mechanism for decalcification of calcareous substrates is conserved.

How to get into bones: proton pump and carbonic anhydrase inOsedaxboneworms

“…The dual 'drilling and feeding' function does not occur in the boring invertebrate animals mentioned earlier, but it is analogous to the mechanism for bone resorption in osteoclast cells of vertebrate animals [18]. The potential similarities between marine boring organisms and osteoclasts have previously been recognized for sponges [8] and snails [10], and also Osedax [16].…”

Novel and potential physiological roles of vacuolar-type H+-ATPase in marine organisms

“…For example, VHA is highly abundant in the brush border of cells in the accessory boring organ from the snail Nucella lamellosa (Clelland and Saleuddin, 2000), which drills through the calcium carbonate shells of its prey. And, although VHA has not yet been described in boring sponges, the etching cells (see Glossary) of these organisms have abundant vacuoles and CA activity (Pomponi, 1980), consistent with the presence of VHA.…”

Estimating the rotation rate in the vacuolar proton-ATPase in native yeast vacuolar membranes

“…Our activity assay and the use of inhibitors was based on (Serrano 1978;Clelland and Saleuddin 2000;Lunde and Kubo 2000;Padilla-Lopez and Pearce 2006), with modifications, as follows. The assay mixture contained the activity buffer (50 mM MES/TRIS, 5 mM MgCl 2 , pH 7.0), 5 mM sodium azide (to inhibit mitochondrial ATPase), 0.2 mM ammonium molybdate (to inhibit acid phosphatases), 100 μM sodium orthovanadate (to inhibit plasma membrane proton-ATPase).…”

“…Since the substrate of the reaction is MgATP and MgCl 2 is in excess to Na 2 ATP, the substrate concentration was close to 2 mM (at least in the beginning of the reaction). Several inhibitors that are known to inhibit other ATPases (Serrano 1978;Clelland and Saleuddin 2000;Lunde and Kubo 2000;Padilla-Lopez and Pearce 2006) were tested. These 'other' inhibitors removed only about 20% of the total ATPase activity, even at relatively high inhibitor concentration.…”