Fig. 5.

Shh-coated beads cluster Cdo but not JLP or Bnip-2. (A) Photomicrographs of Shh-coated beads attached to cells that express Cdo or coexpress nonfluorescent Cdo plus JLP-GFP or Bnip-2-GFP. (Scale bar, 10 μm.) (B) Percentage of Shh-coated beads that clustered the indicated fluorescent protein. Values are means ± SD, n > 100 beads. *, P < 0.001. (C) Western blot analysis of pp38 and total p38 in C2C12 cultures treated with recombinant ShhN (0.5 μg/mL) for the indicated amounts of time. As a positive control cells were treated with NaCl (0.7 M) for 20 min. (D) qRT-PCR analysis of Gli1 expression in C2C12 cells treated ± ShhN (0.5 μg/mL) for 24 h. (E) Model for Ncad-stimulated p38 activation in myoblasts and for Cdo as a multifunctional coreceptor. Cdo bound in cis to ligated Ncad exists in a state that permits stable interaction with JLP/p38 and Bnip-2/Cdc42. In contrast, Cdo bound to Shh does not interact stably with these factors, although it permits the Shh signal to be transmitted to Ptch1, activating canonical Hedgehog pathway signaling. Note that activated MKK6 rescues the defective differentiation program caused by loss of Cdo or Bnip-2, but the role of MKK3/6 or other p38 activators has not been established, so a question mark accompanies their position. Note also that, Shh binds both Cdo and Ptch1, but the ternary complex shown is hypothetical.