A cDNA encoding Tc-MUC-5, a mucin from Toxocara canis larvae identified by expression screening.

Toxocara canis is an ascarid nematode parasite of canids. Larvae infect a wide range of accidental hosts including humans, in whom they are the aetiologic agent of visceral and ocular Larva migrans. The labile surface coat of T. canis larvae consists of a family of mucin glycoproteins termed TES-120, for which the cDNAs have recently been cloned. In this paper, we describe the identification of a novel cDNA (Tc-muc-5) encoding an apomucin by expression screening of a cDNA library with antiserum raised to T. canis excretory/secretory products, and compare the predicted Tc-MUC-5 protein with those of other T. canis mucins (Tc-MUC-1-Tc-MUC-4) that include the TES-120 surface coat glycoproteins. Tc-MUC-5 has both a larger open reading frame and a more divergent sequence than the other T. canis mucins. It contains a putative signal peptide followed by two six-cysteine (SXC) domains, an extended threonine-rich central mucin core domain and two C-terminal SXC domains. Amino acid composition analysis of secreted TES-120 glycoproteins revealed a distinct lack of lysine residues; while this finding is in agreement with the primary sequences of Tc-MUC-1-Tc-MUC-4, Tc-MUC-5 is conspicuous by its relative abundance of lysines (6.7%), suggesting that this protein is not part of the TES-120 family of surface coat proteins.

Zebrafish E-cadherin: expression during early embryogenesis and regulation during brain development.

Zebrafish E-cadherin (cdh1) cell adhesion molecule cDNAs were cloned. We investigated spatial and temporal expression of cdh1 during early embryogenesis. Expression was observed in blastomeres, the anterior mesoderm during gastrulation, and developing epithelial structures. In the developing nervous system, cdh1 was detected at the pharyngula stage (24 hpf) in the midbrain-hindbrain boundary (MHB). Developmental regulation of MHB formation involves wnt1 and pax2.1. wnt1 expression preceded cdh1 expression during MHB formation, and cdh1 expression in the MHB was dependent on normal development of this structure.

Differential expression of cadherin-2 and cadherin-4 in the developing and adult zebrafish visual system.

Cadherins are homophilic cell adhesion molecules that control development of a variety of tissues and maintenance of adult structures. Although cadherins have been implicated in the development of the brain, including the visual system, in several vertebrate species, little is known of their role in zebrafish. In this study, we examined distribution of cadherin-2 (Cdh2, N-cadherin) in the visual system of developing and adult zebrafish using both immunocytochemical and in situ hybridization methods, and we compared Cdh2 distribution to that of the previously reported and closely related cadherin-4 (Cdh4, R-cadherin). As in other vertebrates, in zebrafish embryos Cdh2 was widely expressed in the early nervous system, but its expression became more restricted as development proceeded. Cdh4 was not detectable until later in development, at about the time when the first ganglion cells are generated. Cdh2 and Cdh4 were expressed in distinct regions of developing visual structures, including the lens. We hypothesize that the differential expression of these two cadherins in developing zebrafish visual structures reflects functionally different roles in the development of the vertebrate visual system.

Identification of a new C-type lectin, TES-70, secreted by infective larvae of Toxocara canis, which binds to host ligands.

Infective larvae of the dog roundworm Toxocara canis survive in the tissues of their hosts for extended periods in a state of developmental arrest, successfully evading immune destruction. This survival strategy is thought to be mediated by T. canis excretory/secretory (TES) products which downregulate or divert the immune response. We purified one of the major TES products, TES-70 and gained amino acid sequence from 4 tryptic peptides. These peptides were matched to a predicted protein from a cDNA that was isolated by expression screening a T. canis cDNA library with mouse anti-TES serum. The predicted protein (Tc-CTL-4) is similar to, but larger than, Tc-CTL-1, a 32-kDa C-type lectin secreted by T. canis larvae. Tc-CTL-4 has a signal peptide, 2 Cys-rich domains and a C-terminal calcium-dependent C-type lectin domain that shares sequence similarity with host immune cell receptors such as macrophage mannose receptor and CD23. The lectin domain was expressed in bacteria and antiserum to the purified recombinant protein was used to confirm that Tc-ctl-4 did encode the native TES-70 glycoprotein. TES-70 selectively bound to ligands on the surface of Madin-Darby Canine Kidney cells in vitro in a calcium-dependent manner, inhibitable by mammalian serum, indicating that a host glycan is the native ligand for this new parasite lectin.