Glycosylation and palmitoylation of Wnt-3a are coupled to produce an active form of Wnt-3a.
Wnt-3a is a representative ligand that activates the beta-catenin-dependent pathway in Wnt signaling and is modified with glycans and palmitate. In this study, we analyzed the relationship between glycosylation and lipidation of Wnt-3a. Secretion of a Wnt-3a mutant that lacks glycosylation (Wnt-3a NQ) was impaired. Wnt-3a C77A, which lacks palmitoylation at Cys77, was secreted with similar efficiency to wild-type Wnt-3a (Wnt-3a WT), but did not induce the internalization of low-density lipoprotein receptor-related protein 6 (LRP6). Furthermore, removal of palmitate from Wnt-3a suppressed the ability to bind to its receptors Frizzled8 and LRP6. Wnt-3a C77A was glycosylated to an extent similar to Wnt-3a WT, while Wnt-3a NQ was not modified with palmitate. Expression of porcupine, which is a putative acyltransferase, enhanced palmitoylation of Wnt-3a WT greatly, but that of Wnt-3a NQ slightly. While Wnt-3a WT was present in both the endoplasmic reticulum (ER) and Golgi, Wnt-3a NQ was located to the ER only. Furthermore, Wnt-3a was not palmitoylated but was glycosylated in the cells treated with Brefeldin A, which inhibits transport of vesicles from the ER to the Golgi. These results indicate that glycosylation of Wnt-3a precedes palmitoylation and that both modifications are necessary for secretion of an active Wnt-3a.
Komekado
Hideyuki
H
Department of Biochemistry, Graduate School of Biomedical Sciences, Hiroshima University, 1-2-3 Kasumi, Minami-ku, Hiroshima 734-8551, Japan.
Yamamoto
Hideki
H
Chiba
Tsutomu
T
Kikuchi
Akira
A
D000595:Amino Acid Sequence; D000818:Animals; D001216:Asparagine / Q000378:metabolism; D002460:Cell Line; D051157:Frizzled Receptors / Q000235:genetics / Q000378:metabolism; D006031:Glycosylation; D006801:Humans; D026502:LDL-Receptor Related Proteins / Q000235:genetics / Q000378:metabolism; D060488:Low Density Lipoprotein Receptor-Related Protein-6; D051379:Mice; D008969:Molecular Sequence Data; D019308:Palmitic Acid / Q000378:metabolism*; D015398:Signal Transduction / Q000502:physiology*; D051153:Wnt Proteins / Q000235:genetics / Q000378:metabolism*; D060508:Wnt3 Protein; D060509:Wnt3A Protein
D051157:Frizzled Receptors; D026502:LDL-Receptor Related Proteins; C555475:LRP6 protein, human; D060488:Low Density Lipoprotein Receptor-Related Protein-6; C555476:Lrp6 protein, mouse; C555510:WNT3A protein, human; D051153:Wnt Proteins; D060508:Wnt3 Protein; D060509:Wnt3A Protein; C555511:Wnt3a protein, mouse; D019308:Palmitic Acid; D001216:Asparagine