A direct screen for secreted proteins in Xenopus embryos identifies distinct activities for the Wnt antagonists Crescent and Frzb-1

Edgar M. Pera and E. M. De Robertis

Mech. Dev. 96, 183-195 (2000)

To determine the spectrum of secreted proteins that are present in the extracellular space of early Xenopus embryos, a direct secretion screen was performed. Surprisingly, 24% of previously identified bona fide secretory proteins corresponded to four secreted Wnt antagonists of the same family: frzb-1, sizzled, sfrp-2, and crescent. sfrp-2 and crescent are novel components of the growing cocktail of growth factor antagonists secreted by Spemann organizer cells in Xenopus. Crescent is first expressed at blastula, defining a deep endodermal region that may be homologous to the avian hypoblast. Unlike other members of this family of inhibitors, microinjection of crescent mRNA causes the development of cyclopic embryos, even though the amount of anterior neural tissue is normal. In crescent-injected embryos, studies with specific markers indicate that morphogenetic movements of the anterior midline are abnormal, resulting in a more posterior location of prechordal plate and ventral forebrain markers with respect to the developing eye field. The results are discussed in light of recent findings in zebrafish and Xenopus that suggest that Wnt signalling through non-canonical (non-catenin dependent) pathways plays a pivotal role in the regulation of morphogenetic movements.

	Fig. 1. Secretion cloning
	Fig. 2. Primary protein structure of Crescent
	Fig. 3. Expression of crescent, frzb-1, sfrp-2 and sizzled in Xenopus embryos by whole-mount in situ hybridization
	Fig. 4. Dynamic expression of crescent during early Xenopus development
	Fig. 5. Overexpression of crescent, but not of frzb-1, results in cyclopia
	Fig. 6. Microinjection of crescent mRNA does not inhibit development of anterior neural structures
	Fig. 7. Overexpression of crescent, but not of frzb-1 mRNA, perturbs the anterior extension of midline markers

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Crescent, a novel chick gene encoding a Frizzled-like cysteine-rich domain, is expressed in anterior regions during early embryogenesis

Peter Pfeffer, E. M. De Robertis and Izpisua-Belmonte, J.C.

Int. J. Dev. Biol. 41, 449-458 (1997)

We describe the isolation of a novel chicken gene that we have termed crescent, based on the most distinctive stage of its highly dynamic expression pattern during early embryogenesis. Crescent encodes a protein that in its N-terminal half shows the characteristic invariant 9 cysteine residues of the cysteine-rich domain (CRD) found in the Frizzled family of proteins, in Smoothened and in Collagen XVIII. The CRD of several Frizzled proteins have recently been shown to bind to Wg. Unlike Frizzled proteins, crescent does not contain a transmembrane domain and thus can not function as a receptor. Crescent expression is first found at stage XII (E-G&K) in the center of the area pellucida. On primitive streak formation, expression is detected in the entire anterior half of the area pellucida in the hypoblast layer. At maximal streak extension, crescent transcripts are localized primarily to the germinal crescent, where the primordial germ cells reside. During head process and head fold stages, crescent labels the anteriormost endodermal cells which will give rise to prospective foregut. With the commencement of somitogenesis, crescent expression rapidly wanes.

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