Mii Y and Takada S (2020), Heparan Sulfate Proteoglycan Clustering in Wnt ...

XB-ART-57251

Front Cell Dev Biol 2020 Jan 01;8:631. doi: 10.3389/fcell.2020.00631.

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Heparan Sulfate Proteoglycan Clustering in Wnt Signaling and Dispersal.

Mii Y , Takada S .

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Wnt, a family of secreted signal proteins, serves diverse functions in animal development, stem cell systems, and carcinogenesis. Although Wnt is generally considered a morphogen, the mechanism by which Wnt ligands disperse is still debated. Heparan sulfate proteoglycans (HSPGs) are extracellular regulators involved in Wnt ligand dispersal. Drosophila genetics have revealed that HSPGs participate in accumulation and transport of Wnt ligands. Based on these findings, a "restricted diffusion" model, in which Wnt ligands are gradually transferred by repetitive binding and dissociation to HSPGs, has been proposed. Nonetheless, we recently found that HSPGs are not uniformly distributed, but are locally clustered on cell surfaces in Xenopus embryos. HSPGs with N-sulfo-rich HS chains and those with N-acetyl-rich unmodified HS chains form different clusters. Furthermore, endogenous Wnt8 ligands are discretely accumulated in a punctate fashion, colocalized with the N-sulfo-rich clusters. Based on these lines of evidence, here we reconsider the classical view of morphogen spreading controlled by HSPGs.

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Species referenced: Xenopus
Genes referenced: frzb gpc1 gpc2 gpc4 gpc6 sdc2 wnt3a wnt8a

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FIGURE 1. Models explaining the diversity of HS chain modification of HSPGs. (A) N-sulfation of GlcA-GlcNAc units of HS chains by NDST. HS chains are synthesized by sequential actions of glycosyl transferases and modification enzymes. After polymerization of disaccharide units, elongated HS chains are extensively modified by sulfotransferases, including GlcNAc N-deacetylase/N-sulfotransferase (NDST), which catalyzes N-sulfation of GlcA-GlcNAc units of HS chains. (B) NA and NS domain model. Analysis of the oligosaccharides of HS chains obtained by digestion under conditions in which N-sulfated GlcA-GlcNAc units are selectively attacked, showed that various heparan sulfate samples all contained regions of consecutive N-sulfated GlcA-GlcNAc units, as well as contiguous N-acetylated ones (Gallagher and Walker, 1985; Maccarana et al., 1996; Bernfield et al., 1999). These findings suggest that modifications occur in clusters of variable length (N-sulfated or NS domains), which are interspersed among unmodified domains (N-acetylated or NA domains). It has been proposed that these two domains coexist on single HS chains. (C) N-acetyl-rich and N-sulfo-rich HS cluster model. Recently, Mii et al. found that N-sulfo-rich and N-acetyl-rich HSPGs are clustered independently on cell membranes of Xenopus embryos and on cultured cells (Mii et al., 2017). This new finding strongly suggests that NS and NA domains do not exist randomly on individual HS chains. Rather, the extent of N-sulfation appears to vary between HSPG clusters. Although N-sulfo-rich and N-acetyl-rich clusters rarely overlap on the cell surface, it cannot be excluded that HS chains in N-sulfo-rich and N-acetyl-rich HS clusters may contain some NA and NS domains, respectively. (D) N-acetyl-rich and N-sulfo-rich HS clusters in a Xenopus embryo. Double color immunostaining with direct-labeled NAH46 (anti-N-acetyl subunits) and HepSS-1 (anti-N-sulfo subunits) antibodies shows clustered distributions of HS chains recognized by these antibodies (Mii et al., 2017). Notably, NAH46 and HepSS-1 staining do not largely overlap, but rather show distinct distributions. (E) Endogenous Wnt8 colocalized with N-sulfo rich HS clusters. Double color immunostaining with anti-Wnt8 and HepSS-1 antibodies shows that Wnt8 staining mostly overlaps with HepSS-1 staining. Colocalization is indicated with closed (cell boundary) and open (inside cells) arrowheads. Scale bars, 20μm.

FIGURE 2. Models to control Wnt signaling and dispersal by HSPGs. (A) Restricted diffusion model. Based on genetic studies in Drosophila, it has been proposed that HSPGs mediate Wnt delivery by a restricted-diffusion mechanism, in which Wnt ligands are transported in a “bucket brigade” manner by repeated association and dissociation with HSPGs on cell membranes (Yan and Lin, 2009). (B) Model to explain Wnt signaling and delivery by clustering of HSPGs. Wnt8 preferentially binds to N-sulfo-rich HS clusters and Frzb binds to N-acetyl-rich clusters (upper; Mii et al., 2017). Accumulation of Wnt8 on N-sulfo-rich HS clusters leads to signalosome formation and internalization of Wnt8, which may contribute to degradation of Wnt8. When Frzb is abundant (lower), Wnt8-Frzb complexes bind to N-acetyl-rich HS clusters, which may reduce degradation of Wnt8 (Mii et al., 2017). Given that these two clusters are not distributed uniformly on the cell surface, it seems unlikely that Wnt ligands are transported in a “bucket brigade” manner between these clusters. Since N-sulfo-rich HS clusters are frequently internalized, this cluster appears to shorten the distribution range of Wnt8 (Mii and Taira, 2009). On the other hand, N-acetyl-rich HS clusters tend to remain on the cell surface, resulting in long-range distributions of Frzb as well as Wnt8-Frzb complexes (Mii and Taira, 2009). One possible model is that the balance of Wnt interactions between N-sulfo-rich HS clusters and N-acetyl-rich HS clusters may regulate Wnt signaling range in tissues.

References [+] :

Baeg, The Wingless morphogen gradient is established by the cooperative action of Frizzled and Heparan Sulfate Proteoglycan receptors. 2004, Pubmed