Functional Mechanisms of Patched1, the Hedgehog Pathway Receptor

Abstract

The Hedgehog signaling pathway is a fundamental signaling pathway in metazoans that governs the development, growth, and maintenance of numerous tissues throughout a metazoan organism’s life cycle. How Hedgehog signaling is regulated in healthy and diseased states is poorly understood. Major regulators of the pathway are the Patched (Ptch) sub-family of cell-surface receptors, with Ptch1 being the main regulator of Hedgehog signaling in vertebrates. Ptch1 serves as the receptor for Hedgehog signaling molecules (Hh). In the absence of Hh, Ptch1 acts to inhibit the cell-surface protein, and obligate Hedgehog pathway activator, Smoothened (Smo). In the presence of Hh, Ptch1 is inhibited and Smo de-repressed. Here we investigate the mechanism of Smo inhibition by Ptch1 and the mechanism of Ptch1 inhibition by Hh. We find that Ptch1 inhibits Smo enzymatically, operating in a hypersensitive regime. We identify the molecular etiology of holoprosencephaly-associated mutants in human Ptch1 as being either (i) increased affinity for Smo, (ii) increased enzymatic activity toward Smo, or (iii) insensitivity to Hh. We also find that Ptch1 enzymatic activity is consistent with potassium-driven, cholesterol antiport. Finally, we find that Ptch1 inhibition by Hh is mediated by a palmitic acid moiety appended to the amino-terminus of Hh, where the fatty acid directly binds to specific sites within Ptch1.