The toposome, essential for sea urchin cell adhesion and development, is a modified iron-less calcium-binding transferrin

Abstract

We describe the structure and function of the toposome, a modified calcium-binding, iron-less transferrin, the first member of a new class of cell adhesion proteins. In addition to the amino acid sequence of the precursor, we determined by Edman degradation the N-terminal amino acid sequences of the mature hexameric glycoprotein present in the egg as well as that of its derived proteolytically modified fragments necessary for development beyond the blastula stage. The approximate C-termini of the fragments were determined by a combination of mass spectrometry and migration in reducing gels before and after deglycosylation. This new member of the transferrin family shows special features which explain its evolutionary adaptation to development and adhesive function in sea urchin embryos: (i) a protease-inhibiting WAP domain, (ii) a 280 amino acid cysteine-less insertion in the C-terminal lobe, and (iii) a 240 residue C-terminal extension with a modified cystine knot motif found in multisubunit external cell surface glycoproteins. Proteolytic removal of the N-terminal WAP domain generates the mature toposome present in the oocyte. The modified cystine knot motif stabilizes cell-bound trimers upon Ca-dependent dissociation of hexamer-linked cells. We determined the positions of the developmentally regulated cuts in the cysteine-less insertion, which produce the fragments observed previously. These fragments remain bound to the hexameric 22S particle in vivo and are released only after treatment of the purified toposome with reducing agents. In addition, some soluble smaller fragments with possible signal function are produced. Sequence comparison of five sea urchin species reveals the location of the cell-cell contact site targeted by the species-specific embryo dissociating antibodies. The evolutionary tree of 2-, 1-, and 0-ferric transferrins implies their evolution from a basic cation-activated allosteric design modified to serve multiple functions.

Evolution of yolk protein genes in the Echinodermata

“…; Noll et al. ). The protein may also have an immune function, since it comprises more than half of the protein present in the coelomic fluid of echinoids (Cervello et al.…”

“…Within the phylum Echinodermata, the MYP of the Echinozoa (Echinoidea + Holothuroidea) is a metal binding glycoprotein from the transferrin (Tfn) superfamily. The gene encoding echinozoan MYP is therefore unrelated to ancestral vtg (Unuma et al 2001;Brooks and Wessel 2002;Yokota et al 2003;Noll et al 2007;Rojas-Cartagena et al 2007).…”

“…In the gonads, Tfn-like MYP is stored within granules in somatic cells (nutritive phagocytes) and is suggested to serve as a nutrient source for spermatogenesis, oogenesis, and embryonic development (Scott et al 1990;Unuma et al 1998Unuma et al , 2003. Sea urchin MYP apparently transports both iron and zinc to developing oocytes (Brooks and Wessel 2002;Unuma et al 2007), although the geometry of the iron binding site differs from other Tfns (Lambert et al 2005;Noll et al 2007). The protein may also have an immune function, since it comprises more than half of the protein present in the coelomic fluid of echinoids (Cervello et al 1994;Ramirez-Gomez et al 2009).…”

The sea urchin major yolk protein is synthesized mainly in the gut inner epithelium and the gonadal nutritive phagocytes before and during gametogenesis

“…The coelomic fluid-type MYP is a precursor of the egg-type MYP (Harrington and Easton, 1982) and was formerly called sea urchin vitellogenin (Shyu et al, 1986;Cervello et al, 1994;Unuma et al, 2001;Yokota and Sappington, 2002) after the yolk protein precursor vitellogenin found in the blood of oviparous vertebrates (Wallace, 1985). However, the sequencing of MYP cDNA from Pseudocentrotus depressus (Unuma et al, 2001) and other species (Brooks and Wessel, 2002;Yokota et al, 2003;Noll et al, 2007) has revealed that MYP is not homologous to vertebrate vitellogenins but is slightly homologous to transferrins, a family of iron-binding protein. Therefore, to avoid confusion we refer to the coelomic fluid-type MYP (180 kDa) as CFMYP and to the egg-type MYP (170 kDa) as EGMYP and we use the term MYP in a broad sense when the type is not specified (Unuma et al, 2007).…”

Exogastrulation and interference with the expression of major yolk protein by estrogens administered to sea urchins

“…The physiological roles of MYP as a nutrient storage and zinc carrier (Unuma et al 2007) are similar to those of vertebrate vitellogenin. MYP also has a role in cell adhesion during embryogenesis and has been called toposome by some researchers (Noll et al 1985(Noll et al , 2007Cervello and Matranga 1989;Hayley et al 2006). The expression of sea urchin MYP is not female-specific and not restricted to the breeding season (Shyu et al 1986;Unuma et al 1998Unuma et al , 2001Yokota and Sappington 2002).…”