ShAR1alpha and ShAR1beta: novel putative nicotinic acetylcholine receptor subunits from the platyhelminth blood fluke Schistosoma.

The cDNAs for two novel neuronal-type nicotinic acetylcholine receptor (nAChR) subunits have been cloned and characterised from the parasitic trematode blood fluke Schistosoma haematobium. One of these encodes a putative nAChR alpha-subunit named ShAR1alpha, whilst the second encodes a potential non-alpha subunit, ShAR1beta. These ShARs possess the key structural features common to all nAChRs, but they are unusual in that they have very large cytoplasmic domains spanning M3 and M4. Overall, the ShAR1alpha and ShAR1beta proteins share 37% identity and 53% similarity, but excluding the residues of the M3-M4 domain this rises to 52% identity and 71% similarity. Sequence comparisons with other nAChR polypeptides indicate that both ShARs are most similar to the invertebrate alpha7-like subunits identified in insects and nematodes, and to the vertebrate subunits alpha7 and alpha8. Outside of the M3-M4 domain, 45% and 40%, respectively, of the ShAR1alpha and ShAR1beta residues are conserved in the ACR-16 subunit from Caenorhabditis elegans. Phylogenetic analysis suggests that the ShARs share a common lineage with members of the ACR-16 group as well as alpha7 and alpha8. Immunolocalisation studies revealed distinct and non-overlapping patterns of distribution for ShAR1alpha and ShAR1beta within the parasite. ShAR1beta was localised within the musculature and on discrete cell bodies within the connective parenchyma. In contrast, ShAR1alpha was localised exclusively to the surface membranes, suggesting it may contribute to the regulatory nAChR we have characterised previously. In Xenopus oocyte expression studies, ShAR1alpha did not form functional channels on its own or in combination with ShAR1beta or the chick beta2 subunit. Furthermore, a chimera in which the M3-M4 domain of ShAR1alpha was replaced with that of chick alpha7 was also non-functional. We discuss our findings in the context of the proposed role for surface nAChRs in the regulation of glucose uptake in the parasite, and the potential exploitation of these receptors as targets for cholinergic schistosomicides.

Molecular characterization of an acetylcholinesterase implicated in the regulation of glucose scavenging by the parasite Schistosoma.

Acetylcholinesterase (AChE) present on the surface of the trematode blood fluke Schistosoma has been implicated in the regulation of glucose scavenging from the host blood. Determination of the molecular structure and functional characteristics of this molecule is a crucial first step in understanding the novel function for AChE and in evaluating the potential of schistosome AChE as a target of new parasite control methods. We have determined the primary structure of acetylcholinesterase from Schistosoma haematobium. Immunolocalization studies confirmed that the enzyme was present on the parasite surface as well as in the muscle. The derived amino acid sequence possesses features common to acetylcholinesterases: the catalytic triad, six cysteines that form three intramolecular disulphide bonds, and aromatic residues lining the catalytic gorge. An unusual feature is that the fully processed native enzyme exists as a glycoinositol phospholipid (GPI)-anchored dimer, but the sequence of the C?terminus does not conform to the current consensus for GPI modification. The enzyme expressed in Xenopus oocytes showed conventional substrate specificity and sensitivity to established inhibitors of AChE, although it is relatively insensitive to the peripheral site inhibitor propidium iodide. Distinctions between host and parasite AChEs will allow the rational design of schistosome-specific drugs and vaccines.