Roles of two types of heparan sulfate clusters in Wnt distribution and signaling in Xenopus

Abstract

Wnt proteins direct embryonic patterning, but the regulatory basis of their distribution and signal reception remain unclear. Here, we show that endogenous Wnt8 protein is distributed in a graded manner in Xenopus embryo and accumulated on the cell surface in a punctate manner in association with “N-sulfo-rich heparan sulfate (HS),” not with “N-acetyl-rich HS”. These two types of HS are differentially clustered by attaching to different glypicans as core proteins. N-sulfo-rich HS is frequently internalized and associated with the signaling vesicle, known as the Frizzled/Wnt/LRP6 signalosome, in the presence of Wnt8. Conversely, N-acetyl-rich HS is rarely internalized and accumulates Frzb, a secreted Wnt antagonist. Upon interaction with Frzb, Wnt8 associates with N-acetyl-rich HS, suggesting that N-acetyl-rich HS supports Frzb-mediated antagonism by sequestering Wnt8 from N-sulfo-rich HS. Thus, these two types of HS clusters may constitute a cellular platform for the distribution and signaling of Wnt8.

Direct visualization of a native Wnt in vivo reveals that a long-range Wnt gradient forms by extracellular dispersal

“…Due to their hydrophobic nature ( Willert et al, 2003 ), fully processed Wnts are not thought to be capable of freely moving over long distances in the extracellular space on their own ( Langton et al, 2016 ), and it remains to be determined where in the extracellular environment Wnt/EGL-20 spreads and if spreading involves association with cell surface or extracellular matrix proteins ( Han et al, 2005 ; Fuerer et al, 2010 ; Mii et al, 2017 ), structures such as lipoprotein particles ( Neumann et al, 2009 ; Panáková et al, 2005 ) or exosomes/microvesicles ( Korkut et al, 2009 ; Gross et al, 2012 ; Greco et al, 2001 ), and/or other interacting molecules to modulate Wnt solubility ( Mii and Taira, 2009 ; Mulligan et al, 2012 ; Chang and Sun, 2014 ). While we have shown that extracellular dispersal is required for shaping a long-range Wnt gradient, the endogenous factors that promote and/or hinder ligand spreading await further characterization.…”

Assembly of protein complexes restricts diffusion of Wnt3a proteins

“…We recently showed that two different types of heparan sulfate proteoglycans (HSPG), which are separately clustered on the cell membrane, are differently associated with Wnt and sFRP 35 . While one type of the HSPG, which is enriched with N-sulfation on the heparin sulfate chain, is preferentially associated with Wnt8 and involved in the signaling, the other type, which is enriched with N-acetylation, is preferentially associated with Frzb, Xenopus sFRP3, and Wnt8/Frzb complexes.…”

Functional link between plasma membrane spatiotemporal dynamics, cancer biology, and dietary membrane-altering agents

“…For example, in the presence of Wnt, large cluster patches containing LRP6 have been observed (> 200 nm). More recently, evidence from various studies suggests that select proteins, lipids, and even saccharides are involved in the formation of Wnt/β-catenin signaling-associated nanoclusters [14, 111–114]. The Wnt/β-catenin signaling pathway positive feedback regulator Ly6 family protein LY6/PLAUR domain-containing 6 (Lypd6) is required for LRP6 phosphorylation and activation in lipid rafts.…”

“…Similarly, other plasma membrane components such as heparin sulfate (HS), cholesterol, and phosphatidylinositol-4,5-bisphosphate (PIP 2 ) might also play a role in Wnt/β-catenin pathway-related nanoclustering. Two types of HS molecules, “N-sulfo-rich HS” (N-sulfo HS) and “N-acetyl-rich HS” (N-acetyl HS), have been shown to form distinct 200-nm clusters, as measured by STED microscopy [114]. Interestingly, N-sulfo HS, not N-acetyl-HS, preferably colocalize with the Wnt receptors Fz and LRP6 as well as LRP6’s active counterpart, phosphor-LRP6 [114].…”

“…Two types of HS molecules, “N-sulfo-rich HS” (N-sulfo HS) and “N-acetyl-rich HS” (N-acetyl HS), have been shown to form distinct 200-nm clusters, as measured by STED microscopy [114]. Interestingly, N-sulfo HS, not N-acetyl-HS, preferably colocalize with the Wnt receptors Fz and LRP6 as well as LRP6’s active counterpart, phosphor-LRP6 [114]. N-sulfo HS clusters have also been shown to colocalize with Wnt ligands inside cells and have been postulated to function as pre-existing cores for signalosome formation [122].…”