The structure of SHH in complex with HHIP reveals a recognition role for the Shh pseudo active site in signaling

Abstract

Hedgehog (Hh) signaling is crucial for many aspects of embryonic development, whereas dysregulation of this pathway is associated with several types of cancer. Hedgehog-interacting protein (Hhip) is a surface receptor antagonist that is equipotent against all three mammalian Hh homologs. The crystal structures of human HHIP alone and bound to Sonic hedgehog (SHH) now reveal that HHIP is comprised of two EGF domains and a six-bladed beta-propeller domain. In the complex structure, a critical loop from HHIP binds the pseudo active site groove of SHH and directly coordinates its Zn2+ cation. Notably, sequence comparisons of this SHH binding loop with the Hh receptor Patched (Ptc1) ectodomains and HHIP- and PTC1-peptide binding studies suggest a 'patch for Patched' at the Shh pseudo active site; thus, we propose a role for Hhip as a structural decoy receptor for vertebrate Hh.

All Mammalian Hedgehog Proteins Interact with Cell Adhesion Molecule, Down-regulated by Oncogenes (CDO) and Brother of CDO (BOC) in a Conserved Manner

“…The specific crystal lattice contact in the original ShhN structure that was postulated as a possible Hh oligomerization interface involves Arg 73 (Lys 132 in Drosophila HhN) (23). This interface buries only 180 Å 2 of surface area and does not occur in any other HhN-containing crystal structures, either alone or complexed with Ihog, CDOFn3, BOCFn3, or Hip (22,32,36,37) and thus seems unlikely to represent a conserved self-interaction among HhN molecules.…”

“…A second category, R128N, T154I, and T130N, interrupts hydrogen-bonding networks formed between IhhN and Asp 872 of CDO or the equivalent residue, Asp 758 , of BOC (34,35). Superposition of IhhN on DhhN in the DhhN⅐Hip structure suggests that these mutations would also disrupt a similar hydrogen-bonding network involving Glu 380 of Hip (32,36) and thus be likely to disrupt interactions between HhN proteins and multiple binding partners.…”

“…The set of HhN structures analyzed includes both vertebrate and invertebrate HhNs; HhN in the presence or absence of calcium; and HhN in the presence or absence of the binding partners Ihog, CDO, BOC, and Hip (22,32,36,37) (supplemental Table 2). HhN-HhN contacts in each of 14 unique crystal lattices were analyzed, which yielded roughly 100 pairwise HhN interactions for subsequent analysis.…”

Dynamics of Sonic hedgehog signaling in the ventral spinal cord are controlled by intrinsic changes in source cells requiring Sulfatase 1

“…8D). Lack of 5E1 signal in the receiving field might reflect either low amounts of Shh, below the limit of detection, or, as the 5E1 epitope is masked by Shh binding to its receptors (Bosanac et al, 2009;Maun et al, 2010), inability of the antibody to access its epitope. In support of the latter interpretation, detection of 5E1 immunoreactivity at the surface of LFP cells is temporally correlated with downregulation of the receptor ptc in these cells (Touahri et al, 2012).…”

“…In these experiments, we used the 5E1 monoclonal antibody that recognizes the biologically active form of Shh (Ericson et al, 1996). This conformation-dependent antibody indeed specifically binds to the Shh zinc coordination site, also identified as the binding site for Shh receptors (Bishop et al, 2009;Bosanac et al, 2009;Maun et al, 2010). We first examined Shh distribution over the culture period, corresponding to the time window of active patterning rearrangement (Fig.…”