Abstract
The mechanism of intercellular transport of Wnt ligands is still a matter of debate. To better understand this issue, we examined the distribution and dynamics of Wnt8 in Xenopus embryos. While Venus-tagged Wnt8 was found on the surfaces of cells close to Wnt-producing cells, we also detected its dispersal over distances of 15 cell diameters. A combination of fluorescence correlation spectroscopy and quantitative imaging suggested that only a small proportion of Wnt8 ligands diffuses freely, whereas most Wnt8 molecules are bound to cell surfaces. Fluorescence decay after photoconversion showed that Wnt8 ligands bound on cell surfaces decrease exponentially, suggesting a dynamic exchange of bound forms of Wnt ligands. Mathematical modeling based on this exchange recapitulates a graded distribution of bound, but not free, Wnt ligands. Based on these results, we propose that Wnt distribution in tissues is controlled by a dynamic exchange of its abundant bound and rare free populations.
Highlights
The Wnt family of secreted signaling proteins has diverse roles in animal development, stem cell systems, and carcinogenesis (Clevers et al, 2014; Loh et al, 2016; Nusse and Clevers, 2017)
We recently showed that heparan sulfate proteoglycans (HSPGs) on cell surfaces are discretely distributed in a punctate manner, which varies with heparan sulfate (HS) modification, forming two different types of HS clusters, N-sulfo-rich and N-acetyl-rich forms (Mii et al, 2017)
As we have previously shown (Mii et al, 2017), Wnt8 and Frzb fused with monomeric Venus were visualized along cell boundaries when expressed in Xenopus embryos (Figure 1A)
Summary
The Wnt family of secreted signaling proteins has diverse roles in animal development, stem cell systems, and carcinogenesis (Clevers et al, 2014; Loh et al, 2016; Nusse and Clevers, 2017). We examined extracellular dynamics of Wnt and Frzb, both of which are involved in anteroposterior patterning of vertebrate embryos (Clevers and Nusse, 2012; Kiecker and Niehrs, 2001; MacDonald et al, 2009; Mii et al, 2017) We visualized their localization in Xenopus embryos by fusing them with fluorescent proteins and we examined their dispersion by capturing them in distant cells. We refined FDAPbased analysis by focusing on a limited area at the cell boundary, which enabled us to quantify dynamics comparable to those measured by FCS Based on these results and our previous findings, we propose a basic mathematical model to explain distribution and dispersion of secreted proteins
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