Wnt proteins are lipid-modified and can act as stem cell growth factors

Abstract

Wnt signalling is involved in numerous events in animal development, including the proliferation of stem cells and the specification of the neural crest. Wnt proteins are potentially important reagents in expanding specific cell types, but in contrast to other developmental signalling molecules such as hedgehog proteins and the bone morphogenetic proteins, Wnt proteins have never been isolated in an active form. Although Wnt proteins are secreted from cells, secretion is usually inefficient and previous attempts to characterize Wnt proteins have been hampered by their high degree of insolubility. Here we have isolated active Wnt molecules, including the product of the mouse Wnt3a gene. By mass spectrometry, we found the proteins to be palmitoylated on a conserved cysteine. Enzymatic removal of the palmitate or site-directed and natural mutations of the modified cysteine result in loss of activity, and indicate that the lipid is important for signalling. The purified Wnt3a protein induces self-renewal of haematopoietic stem cells, signifying its potential use in tissue engineering.

P53 levels determine outcome during β-catenin tumor initiation and metastasis in the mammary gland and male germ cells

“…In the hematopoietic system, Wnt/ b-catenin signaling maintains cells in a pluripotent proliferative state (Willert et al, 2003). Similar findings were found in human embryonic stem cells where inhibition of GSK3b and subsequent b-catenin stabilization maintained the pluripotency of the cells in culture (Sato et al, 2004).…”

ERAD‐dependent control of the Wnt secretory factor Evi

“…Wnt proteins are palmitoylated at a conserved serine residue by the ER‐resident O‐acyltransferase Porcn (Kadowaki et al , 1996; Willert et al , 2003; Takada et al , 2006), which is required for the Wnt proteins to interact with Evi (Herr & Basler, 2012) (Fig 2A). To determine whether Wnt palmitoylation and consequently Evi‐Wnt interactions are required for the Wnt‐mediated increase in Evi protein, we treated wild‐type, Wnt3, or Wnt5B stable transfected HEK293T cells with the Porcn inhibitor LGK974 to block Wnt palmitoylation.…”

Wnt Signaling Inhibition Deprives Small Intestinal Stem Cells of Clonogenic Capacity

“…For Cre‐mediated recombination, cells were cultured for three days in 4‐OHT (final concentration 2 µM; Sigma); to prepare 1 mM stock solution, 4‐OHT was dissolved in ethanol. Mouse Wnt3a ligand was isolated from the culture medium of Wnt3a‐producing L cells according to the detailed protocol of Willert and colleagues (Willert et al ., 2003). …”