The head inducer Cerberus is a multifunctional antagonist of Nodal, BMP and Wnt signals - PubMed (original) (raw)

. 1999 Feb 25;397(6721):707-10.

doi: 10.1038/17820.

Affiliations

The head inducer Cerberus is a multifunctional antagonist of Nodal, BMP and Wnt signals

S Piccolo et al. Nature. 1999.

Abstract

Embryological and genetic evidence indicates that the vertebrate head is induced by a different set of signals from those that organize trunk-tail development. The gene cerberus encodes a secreted protein that is expressed in anterior endoderm and has the unique property of inducing ectopic heads in the absence of trunk structures. Here we show that the cerberus protein functions as a multivalent growth-factor antagonist in the extracellular space: it binds to Nodal, BMP and Wnt proteins via independent sites. The expression of cerberus during gastrulation is activated by earlier nodal-related signals in endoderm and by Spemann-organizer factors that repress signalling by BMP and Wnt. In order for the head territory to form, we propose that signals involved in trunk development, such as those involving BMP, Wnt and Nodal proteins, must be inhibited in rostral regions.

PubMed Disclaimer

Figures

Figure 1

Figure 1

Cerberus protein binds to Xnr-1, BMP-4 and Xwnt-8. a, Cer–Flag secreted by Xenopus animal cap (AC) and cultured 293T cells. b, The two Cer protein products. c, HA-tagged activin, Vg1 and Xnr-1 secreted by Xenopus oocytes. d, Xnr-1 is bound specifically by Cer-S–Flag (lanes 2−4) or Cer-L–Flag (not shown). e, Cer binding inactivates Xnr-1 signalling. Animal cap explants were treated with oocyte control medium, activin (15 pM), Vg1 (0.3 nM) or Xnr-1 (1 nM) proteins, either alone (lanes 2−5) or together with 5 nM Cer-S (lanes 6−8) or Cer-L (lanes 9−11). f, Cer inhibits Xnr-1 with high affinity; 2 nM Xnr-1 was incubated with increasing concentrations of Cer-S (closed circles) or Cer-L (open circles). g, Cer-L–Flag binds BMP-4 with a _K_D of 0.6 nM. h, Binding of Cer-L–Flag to BMP-4 is competed for by BMP-2 but not by TGF-β-1, EGF or PDGF; Cer-S does not bind BMP-4. i, Cer-L (10 nM), but not Cer-S (20 nM), is a direct neural inducer of animal cap cells sensitized by brief dissociation–re-aggregation (but not of intact caps). j, Cer inhibits Xwnt8 but not β-catenin mRNA; induction of Siamois, a target of Wnt signalling, was assayed at stage 10+. k, Lanes 1, 2, soluble Xwnt-8–HA protein is secreted by mRNA-injected oocytes. Lanes 3−5, binding of Xwnt-8-HA (5 nM) to Cer–Flag proteins (10 nM). l, m, The Xwnt-8–HA, Xnr-1–HA and BMP binding sites in Cer-L are independent. Cer-L–Flag (2 nM) was bound to Xnr-1 (1 nM) or Xwnt-8 (2 nM) and competed with BMP-4 (10 nM), BMP-2 (27 nM) or Xnr-1 (8 nM). n, Multiple ligand-binding sites on Cerberus.

Figure 2

Figure 2

Phenotypic effects of cer-S mRNA, an inhibitor of nodal-related signals. Embryos were injected radially at the 4-cell stage. a–d, In situ hybridization for Xbra (a,b) and Xwnt-8 (c,d) in wild type and in embryos injected with 120 pg of cer-S mRNA per blastomere. e, Anterior endoderm formation requires early nodal-related signals in injected embryos: endogenous cerberus transcripts are inhibited by cer-S mRNA (200 pg total) and induced by Xnr-1 mRNA (50 pg). f, Dual function for Xnr-1; Xnr-1 mRNA (6,12, 24 pg), but not pCS2–Xnr-1 DNA (20, 40, 80 pg), can induce cer expression in animal caps. g–n, Head induction by simultaneous repression of BMP and nodal signals in explants of ventral marginal zone (VMZ). g, k, Wild-type VMZs consisting histologically of endoderm (en) and blood (bl). h, cer-S mRNA (150 pg); i, tBR mRNA (600 pg); j, m, co-injection of tBR mRNA (600 pg) and cer-S mRNA (150 pg) generates head-like structures (76%, n = 17), similar to those produced by cerberus mRNA (l, 50%, n = 10), consisting only of anterior structures with cement gland (CG), brain (br) and eye (ey) containing a lens (le). n, Blocking BMP and Nodal signalling by tBR (600 pg) and tALK4 (1.5 ng) RNAs is sufficient to induce head-like structures.

Figure 3

Figure 3

Head induction requires the triple inhibition of nodal-related, Wnt and BMP signals. Embryos were injected into one ventral blastomere with the indicated mRNAs or DNAs at the 4-cell stage. a, Ectopic head induction by a combination of chd (50 pg) and cer-S (200 pg) mRNA (58%, n = 115). b, pCSKA–Xwnt-8 DNA (50 pg) antagonizes head induction by chd/cer-S (1% ectopic heads, n = 111). c, Ectopic heads formed by injection of 150 pg cer mRNA (57%, n = 66) are abolished in d and e by co-injection of 1 ng CA–ALK4 mRNA (0%, n = 36) or 400 pg of CA–BMPR mRNA (0%, n = 21). f, Dorsal injection of pCS2-Xnr-1 (32 pg) causes head reduction (53%, n = 60). g, Co-injection of chd (25 pg) and Frzb-1 (200 pg) mRNAs mediates formation of a complete secondary axis with cyclopic head (49%, n = 175). h, Co-injection of 16 pg of pCS2–Xnr-1 DNA inhibits head induction by chd/Frzb-1 (5%, n = 133). i–n, Simultaneous repression of BMP and Wnt signalling leads to ectopic activation of cer in endoderm and concomitant downregulation of Xbra in mesoderm. Embryos were cut sagittally in two halves after fixation and before whole-mount hybridization. Arrowhead indicates the dorsal blastopore lip. Frzb-1 mRNA (300 pg) synergizes with tBR mRNA (800 pg) in upregulating cer in the ventral endoderm (arrow). Note in n the inhibition of Xbra in the injected area, correlating with the activation of cer (an anti-Nodal agent) observed in l.

Figure 4

Figure 4

Model of the formation and function of anterior endoderm in Xenopus head development.

Similar articles

Cited by

References

    1. Bouwmeester T, Kim SH, Sasai Y, Lu B, De Robertis EM. Cerberus is a head-inducing secreted factor expressed in the anterior endoderm of Spemann's organizer. Nature. 1996;382:595–601. - PubMed
    1. Thomas P, Beddington R. Anterior primitive endoderm may be responsible for patterning the anterior neural plate in the mouse embryo. Curr. Biol. 1996;6:1487–1496. - PubMed
    1. Shawlot W, Behringer RR. Requirement for Lim1 in head-organizer function. Nature. 1995;374:425–430. - PubMed
    1. Glinka A, Wu W, Onichtchouk D, Blumenstock C, Niehrs C. Head induction by simultaneous repression of Bmp and Wnt signalling in Xenopus. Nature. 1997;389:517–519. - PubMed
    1. Glinka A, et al. Dickkopf-1 is a member of a new family of secreted proteins and functions in head induction. Nature. 1998;391:357–362. - PubMed

Publication types

MeSH terms

Substances

LinkOut - more resources