A colipase fold in the carboxy-terminal domain of the Wnt antagonists – the Dickkopfs
1998; Elsevier BV; Volume: 8; Issue: 14 Linguagem: Inglês
10.1016/s0960-9822(98)70309-4
ISSN1879-0445
Autores Tópico(s)Polyamine Metabolism and Applications
ResumoIt has recently been shown that a class of secreted proteins — the Dickkopfs — are released by the Spemann organizer of Xenopus and are potent antagonists of Wnt signaling [[1]Glinka A Wu W Delius H Monaghan AP Blumenstock C Niehrs C Dickkopf-1 is a member of a new family of secreted proteins and functions in head induction.Nature. 1998; 391: 357-363Crossref PubMed Scopus (1349) Google Scholar]. Although it has been reported that the Dickkopf (Dkk) proteins contain two cysteine-rich domains, no direct functional or structural conclusions have been drawn from their sequences. The role of Wnt signaling in animal development has recently become clearer, with the discovery of new components of this pathway [[2]Zorn AM Cell–cell signalling: Frog frizbees.Curr Biol. 1997; 7: R501-R504Abstract Full Text Full Text PDF PubMed Google Scholar], and is particularly well illustrated by experiments involving Xenopus head formation by the Spemann organizer [[2]Zorn AM Cell–cell signalling: Frog frizbees.Curr Biol. 1997; 7: R501-R504Abstract Full Text Full Text PDF PubMed Google Scholar] and wing development in Drosophila[[3]Tomlinson A Strapps WR Heemskerk J Linking Frizzled and Wnt signaling in Drosophila development.Development. 1997; 124 (98072327): 4515-4521Crossref PubMed Google Scholar]. These studies led to the characterization of the complete signaling pathway that is initiated extracellularly by the binding of Wnt to the Frizzled-like seven-transmembrane receptors [[2]Zorn AM Cell–cell signalling: Frog frizbees.Curr Biol. 1997; 7: R501-R504Abstract Full Text Full Text PDF PubMed Google Scholar], and proceeds intracellularly through a G-protein-coupled cascade [[4]Slusarski DC Corces VG Moon RT Interaction of Wnt and a Frizzled homologue triggers G-protein-linked phosphatidylinositol signalling.Nature. 1997; 390 (98049350): 410-413Crossref PubMed Scopus (548) Google Scholar] and the Armadillo and Lef proteins, which function in the nucleus by regulating transcription [[5]Behrens J von Kries JP Kuhl M Bruhn L Wedlich D Grosschedl R Birchmeier W Functional interaction of β-catenin with the transcription factor LEF-1.Nature. 1997; 390: 638-642Google Scholar]. This signaling cascade is necessary in vertebrate development to initiate posterior structures and is antagonized by molecules such as Dkk and Frzb, which are secreted by the Spemann organizer and are required for head induction [[6]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 (97472419): 517-519Crossref PubMed Scopus (300) Google Scholar]. To gain a better understanding of the role of Dkks in Wnt signaling, we carried out a detailed analysis of Dkk sequences and show here that they have a colipase fold. This observation may have direct implications for the function of the Dkks in signaling. It has been noted that the Dkks have two cysteine-rich domains ([[6]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 (97472419): 517-519Crossref PubMed Scopus (300) Google Scholar]; see also below), and we found additionally that the carboxy-terminal domain has the pattern of cysteines typical of the colipase domain (Figure 1). An iterative database search of the non-redundant protein database at the National Center for Biotechnology Information (NCBI) using the PSI-BLAST program [[7]Altschul SF Madden TL Schäffer AA Zhang J Zhang Z Miller W Lipman DJ Gapped BLAST and PSI-BLAST: a new generation of protein database search programs.Nucleic Acids Res. 1997; 25: 3389-3402Crossref PubMed Scopus (59810) Google Scholar], with the Dkk sequences as queries, recovered the colipases from different mammals at a statistically significant expectation (e) value of 10-4 in the second iteration. This was confirmed by reverse searches using different colipases, which retrieved the Dkks at similar e values within three iterations. Multiple alignment based secondary structure prediction using the PHD program [[8]Rost B Sander C Combining evolutionary information and neural networks to predict protein secondary structure.Proteins. 1994; 19 (94344959): 55-72Crossref PubMed Scopus (1337) Google Scholar] and subsequent threading [[9]Rost B Schneider R Sander C Protein fold recognition by prediction-based threading.J Mol Biol. 1997; 270 (97392440): 471-480Crossref PubMed Scopus (222) Google Scholar] through the protein structural database (PDB) suggested the existence of a similar fold in the carboxy-terminal domain of the Dkks and the colipases. On the basis of these observations, we predict that the disulfide-bonding pattern in the Dkks is the same as in the colipases (Figure 1). The colipase fold (Figure 2a) is typical of a wide range of small proteins, which are involved in a number of specific protein–protein interactions and include, in addition to the colipases, scorpion and snake toxins and protease inhibitors [[10]Hubbard TJP Murzin AG Brenner SE Chothia C SCOP: a structural classification of proteins database.Nucleic Acids Res. 1997; 25 (97169401): 236-239Crossref PubMed Scopus (222) Google Scholar]. These proteins have a series of short β strands with large connecting loops, which are held together by disulfide bonds. This results in a flat fold with finger-like protuberances, which act as interactive surfaces [[11]van Tilbeurgh H Sarda L Verger R Cambillau C Structure of the pancreatic lipase-procolipase complex.Nature. 1992; 359 (92396238): 159-162Crossref PubMed Scopus (321) Google Scholar]. The colipases bind to the pancreatic lipases and act as cofactors for them in lipid hydrolysis [[11]van Tilbeurgh H Sarda L Verger R Cambillau C Structure of the pancreatic lipase-procolipase complex.Nature. 1992; 359 (92396238): 159-162Crossref PubMed Scopus (321) Google Scholar]. It has been shown that the colipases are necessary for the interactions of the lipases with hydrophobic lipid micelles during the process of lipid digestion, and the exposed hydrophobic residues in the finger-like elements are involved in this lipophilic interaction [12van Tilbeurgh H Egloff MP Martinez C Rugani N Verger R Cambillau C Interfacial activation of the lipase-procolipase complex by mixed micelles revealed by X-ray crystallography.Nature. 1993; 362 (93241293): 814-820Crossref PubMed Scopus (637) Google Scholar, 13Hermoso J Pignol D Penel S Roth M Chapus C Fontecilla-Camps JC Neutron crystallographic evidence of lipase-colipase complex activation by a micelle.EMBO J. 1997; 16: 5531-5536Crossref PubMed Scopus (93) Google Scholar]. Several of these hydrophobic positions are conserved between the carboxy-terminal domain of Dkk and the colipases (Figure 1, Figure 2b), suggesting that Dkk — like the colipases — interacts with lipids. One direct functional implication of this observation is that the colipase-like domain of Dkk may be necessary for the membrane association of this protein, which in turn may be required for the inhibition of Wnt secretion or Wnt– receptor interaction. We thus propose a testable hypothesis that the inhibition of the Wnt function by Dkk is closely associated with the cell membrane and that the carboxy-terminal, colipase-like domain of Dkk mediates this association. The amino-terminal domain of Dkks may directly interact with Wnt or its receptor, Frizzled. The Wnt inhibition by Dkk thus appears to be distinct from the inhibition mediated by Frzb, which is a dominant-negative form of Frizzled ([[2]Zorn AM Cell–cell signalling: Frog frizbees.Curr Biol. 1997; 7: R501-R504Abstract Full Text Full Text PDF PubMed Google Scholar] and references therein). L Aravind, Department of Biology, Texas A&M University, College Station, Texas 77843, National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894, USA. e-mail: [email protected] EV Koonin, National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894, USA.
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