Exosome secretion is required for sonic hedgehog dispersal and signal gradient formation in the embryonic limb mesenchyme

Carrier-assisted diffusion and cytoneme transport have been postulated to disperse Hedgehog across diverse embryonic tissues, yet their relative contributions to patterning mesenchymal tissues remains poorly understood. Here, we combine novel signaling assays with quantitative microscopy to establish exosome secretion as a predominant and adaptable mechanism for Sonic Hedgehog (Shh) dispersal. Introducing a novel synchronous release system to visualize Shh trafficking in embryonic tissues, we demonstrate that Shh traffics through the exosome biogenesis pathway in the limb bud mesenchyme. Shh-bound exosomes diffuse through extracellular space, and can also bind and travel along cytonemes, providing a potential mechanism for directed and/or long-range transport. Using a synthetic patterning assay, we show that exosome secretion is essential to establish short-range Shh gradients in vitro. We propose that exosome-based Shh secretion, combined with different modes of extracellular transport, provides a tunable mechanism to sculpt Shh gradients on different length and time scales, across different embryonic tissues. HIGHLIGHTSO_LIA novel synchronous release system reveals trafficking dynamics in embryonic cells C_LIO_LISonic hedgehog is packaged and secreted on exosomes for extracellular dispersal C_LIO_LIExosome secretion is required to establish short-range Hedgehog gradients C_LIO_LIDiffusion and cytoneme transport provide tunable exosome dispersal strategies C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/692671v1_ufig1.gif" ALT="Figure 1"> View larger version (89K): org.highwire.dtl.DTLVardef@76b266org.highwire.dtl.DTLVardef@ba5b4borg.highwire.dtl.DTLVardef@bf9d67org.highwire.dtl.DTLVardef@1f6d699_HPS_FORMAT_FIGEXP M_FIG C_FIG