Fig. 7. Shh fails to cross-link and switch from heparin to lower sulfated soluble variants.

a In stark contrast to soluble heparin added to the wash buffer A, Shh does not transfer to selectively desulfated heparins with a less negative net charge. b Shh does not switch to low-sulfated chondroitin sulfate (CS, a related type of glycosaminoglycan consisting of repeating sulfated galactosamine-glucuronic/iduronic disaccharides instead of sulfated glucosamine-glucuronic/iduronic disaccharides for HS), even at high CS concentrations. c Model of direct repeated Hh switching. Two binding sites for HS facilitate Shh diffusion in an HS matrix through competition and direct repeated switching between neighboring HS chains (bent arrows). d Functional impairment of one binding site reduces the propensity for direct competition, and thus the rate of switching between HS chains, apparently slowing Hh/Shh spread down. This is expected to result in a shorter and steeper gradient. e High-affinity HS association restricts Hh/Shh movement to the cellular surfaces where suitably sulfated HS resides (the “diffusible zone”). Hh/Shh movement is effectively confined within the zone in two dimensions, with a gradient forming from posterior producing cells (right) to anterior receiving cells (left). Undersulfated HS are poor acceptors for Shh if already bound to higher sulfated HS, which explains that clones deficient in HSPG expression block Hh transport from HS-expressing cells in the Drosophila wing disc¹⁵. In the absence of properly sulfated (sfl) anterior HS, or without HS (ttv), Hh clusters remain confined to cell surfaces with appropriately sulfated HS (reversed arrows)—representing the constricted diffusible zone. Fully functional GPI-linked Glp-HS chains at the plasma membrane are shown in dark blue; non-functional variants lacking HS chains or linked to undersulfated HS in light blue. P posterior, A anterior.