Dispatched mediates Hedgehog basolateral release to form the long-range morphogenetic gradient in the Drosophila wing disk epithelium

Abstract

Hedgehog (Hh) moves from the producing cells to regulate the growth and development of distant cells in a variety of tissues. Here, we have investigated the mechanism of Hh release from the producing cells to form a morphogenetic gradient in the Drosophila wing imaginal disk epithelium. We describe that Hh reaches both apical and basolateral plasma membranes, but the apical Hh is subsequently internalized in the producing cells and routed to the basolateral surface, where Hh is released to form a longrange gradient. Functional analysis of the 12-transmembrane protein Dispatched, the glypican Dally-like (Dlp) protein, and the Iglike and FNNIII domains of protein Interference Hh (Ihog) revealed that Dispatched could be involved in the regulation of vesicular trafficking necessary for basolateral release of Hh, Dlp, and Ihog. We also show that Dlp is needed in Hh-producing cells to allow for Hh release and that Ihog, which has been previously described as an Hh coreceptor, anchors Hh to the basolateral part of the disk epithelium.

Essential basal cytonemes take up Hedgehog in the Drosophila wing imaginal disc

“…This model posits that the soluble multimer, either alone or complexed with chaperones, disperses in the extracellular milieu of the Hhresponding cells. There is, however, strong evidence for a direct transfer mechanism based on cytonemes that extend from the Hhexpressing cells in the P compartment (Bischoff et al, 2013;Callejo et al, 2011).…”

“…The path that Hh takes prior to release at cytoneme tips is complex, initially involving placement in the apical membrane of Hh-producing cells, and dynamin-dependent endocytosis prior to transit to the basolateral compartment in a vesicular form (Callejo et al, 2011 (Matusek et al, 2014), work from the Guerrero group suggests that Hhcontaining vesicles that are associated with cytonemes may appear to be 'exosomes' if the cytonemes are not marked and simultaneously imaged (Gradilla et al, 2014).…”

“…Disc cells also have cytonemes that extend from the basolateral membrane that have been implicated in Hh signaling (Callejo et al, 2011). These basolateral cytonemes mostly orient toward the A/P compartment border (Callejo et al, 2011;Roy et al, 2011), and some extend across the A/P border and appear to transport Hh to A compartment cells in motile puncta (Bilioni et al, 2013;Bischoff et al, 2013;Callejo et al, 2011).…”

“…They protrude from many cell types, including the Dpp-receiving cells of the wing disc that have cytonemes that extend across the apical surface toward the Dpp-expressing cells at the A/P border (Hsiung et al, 2005;Ramirez-Weber and Kornberg, 1999;Roy et al, 2011). Disc cells also have cytonemes that extend from the basolateral membrane that have been implicated in Hh signaling (Callejo et al, 2011). These basolateral cytonemes mostly orient toward the A/P compartment border (Callejo et al, 2011;Roy et al, 2011), and some extend across the A/P border and appear to transport Hh to A compartment cells in motile puncta (Bilioni et al, 2013;Bischoff et al, 2013;Callejo et al, 2011).…”

“…The complex choreography that prepares Hh for delivery to A compartment cells was discovered in studies on Hh and Ihog, which were overexpressed as chimeras fused with either GFP or YFP (Bischoff et al, 2013;Callejo et al, 2011). These studies used the Gal4-UAS system, which elevates expression levels an estimated 10× (Yagi et al, 2010); the studies circumvented the cell lethality of Ihog overexpression with a time-limited pulse, and succeeded despite the apparent partial functionality of Hh:GFP expressed by this method (Torroja et al, 2004).…”