Opinion
A conserved mechanism of Hedgehog gradient formation by lipid modifications

https://doi.org/10.1016/j.tcb.2006.11.002Get rights and content

Members of the Hedgehog (Hh) family of proteins are conserved morphogens that modulate cell fates in target tissues in different developmental systems. Dysregulation of Hh signaling results in a wide range of human diseases. The mature Hh is modified by lipids in two places, with palmitate at the N-terminus and cholesterol at the C-terminus. The lipid modifications are essential to the proper secretion and spreading of the morphogen throughout the extracellular matrix, interacting with heparan sulfate proteoglycans. However, the role of lipid modifications in regulating the range and activity of Hh proteins remains controversial. Here, we aim to resolve this issue by providing a model that is consistent with current and past reports. We propose that the cholesterol moiety functions to restrict the dilution and deregulated spread of the morphogen in the extracellular space.

Introduction

A key issue in developmental biology is how cells in a developing field acquire the positional information that will determine their fate. Secreted members of the Hedgehog (Hh) family are essential signaling molecules controlling growth and patterning in both vertebrates and invertebrates [1]. Hh proteins are considered to act as morphogens that can spread from localized sites of production to specify a diverse array of cell fates, ranging from segmental patterns in the cuticle of the Drosophila larva to neurons in the vertebrate neural tube, in a concentration-dependent manner 1, 2. The mature Hh is synthesized as a precursor protein that undergoes a series of posttranslational modifications, leading to covalent attachment of a cholesterol moiety at its C-terminus and palmitic acid at its N-terminus (Box 1, Figure 1). The cholesterol moiety of Hh has been shown to associate tightly with the cell membrane [3] and a specific cellular mechanism is needed for the release of the highly lipidated Hh from its source. The 12-pass transmembrane protein Dispatched (Disp) appears to fulfill this requirement [4], as it is required only for the release of cholesterol-modified Hh (Box 2). Thus, the cholesterol moiety of Hh is necessarily coupled to Disp function for the regulated release of Hh.

The extracellular spreading of Hh is a highly regulated process and is a crucial determinant of the morphogen gradient. The hydrophobic nature of Hh lipid modifications would be predicted to have a significant effect on the shape and range of activity gradients. Indeed, expression of different forms of Hh that lack either the cholesterol moiety (Hh-N or Shh-N) or palmitic acid (Hh-C85S or Shh-C25S; Figure 1b) in several animal models leads to profound alterations in diffusion and signaling properties of Hh. The loss of palmitoylation results in strong developmental defects, indicating that palmitate modification is required for Hh activity 5, 6, 7, 8, 9. The loss of activity in HhC85S might be associated with the observation that it is not internalized by its receptor Patched in Drosophila imaginal disc epithelia [10], although in vitro experiments indicate that purified ShhC24S can bind to Patched as efficiently as lipidated Hh [11]. By contrast, the cholesterol moiety does not seem to be necessary for Hh activity [12]. However, its role in regulating Hh spread and signaling in Drosophila and vertebrates is controversial. The main issue that needs to be resolved is whether the cholesterol moiety promotes or restricts Hh spreading to generate a defined activity gradient. Here, we provide a unifying view of the functions of the cholesterol moiety that is consistent with current and previous publications.

Section snippets

Cholesterol modification in the regulation of the Drosophila Hh gradient

Although the Hh signal was first described as a morphogen in the segmental patterning of the Drosophila larval cuticle [13], the adult wing pattern offers a unique readout of Hh signaling. The wing disc consists of an epithelial sack with a thick columnar pseudostratified epithelium on one side and the squamous epithelium, called the peripodial membrane, on the other. The apical surfaces of these two epithelia are oriented towards the disc lumen. Two populations of cells with different cell

Cholesterol modification in regulating the Shh gradient

Analogous to the Drosophila wing imaginal disc, the vertebrate limb bud has become an ideal model system to elucidate morphogen gradient action. The anterior–posterior asymmetry of digit patterns is controlled by a group of specialized mesodermal cells located at the posterior margin of the limb bud referred to as the zone of polarizing activity (ZPA). Cells in the ZPA secrete Shh, which functions as a classic morphogen in establishing the anterior–posterior polarity of the limb [24]. It is

Concluding remarks and future directions

The data suggest that the mechanisms of Hh secretion and spreading are generally conserved throughout evolution from Drosophila to vertebrates, despite some functional divergences in the intracellular signaling that Hh elicits in the receiving cells. It is evident that cholesterol modification of Hh is essential to restrict dilution and deregulated spreading of the morphogen through the extracellular environment. In its absence, Hh spreads far from the source and can elicit ectopic low

Acknowledgements

I.G. thanks all members of her laboratory, past and present, for their contribution to the understanding the role of lipid modifications in gradient formation: A. Callejo, N. Gorfinkiel, C. Torroja and J. Sierra. C.C. thanks Y. Litingtung, Y. Li, H. Zhang and X. Huang for their work on the cholesterol modification of Shh. We also thank the support of the Spanish D.G.I.C.Y.T, grant BFU2005–04183 (I.G.), the Fundación Areces for an institutional grant (I.G.), National Institutes of Health (C.C.)

References (42)

  • L. Koziel

    Ext1-dependent heparan sulfate regulates the range of Ihh signaling during endochondral ossification

    Dev. Cell

    (2004)
  • R.B. Pepinsky

    Identification of a palmitic acid-modified form of human Sonic hedgehog

    J. Biol. Chem.

    (1998)
  • A. Gallet

    Cholesterol modification of hedgehog is required for trafficking and movement, revealing an asymmetric cellular response to hedgehog

    Dev. Cell

    (2003)
  • Y. Ma

    Hedgehog-mediated patterning of the mammalian embryo requires transporter-like function of dispatched

    Cell

    (2002)
  • T. Caspary

    Mouse Dispatched homolog1 is required for long-range, but not juxtacrine, Hh signaling

    Curr. Biol.

    (2002)
  • Y. Nakano

    Inactivation of dispatched 1 by the chameleon mutation disrupts Hedgehog signalling in the zebrafish embryo

    Dev. Biol.

    (2004)
  • P.E. Kuwabara et al.

    The sterol-sensing domain: multiple families, a unique role?

    Trends Genet.

    (2002)
  • G. Michaux

    CHE-14, a protein with a sterol-sensing domain, is required for apical sorting in C. elegans ectodermal epithelial cells

    Curr. Biol.

    (2000)
  • P.W. Ingham et al.

    Hedgehog signaling in animal development: paradigms and principles

    Genes Dev.

    (2001)
  • T.M. Jessell

    Neuronal specification in the spinal cord: inductive signals and transcriptional codes

    Nat. Rev. Genet.

    (2000)
  • C. Peters

    The cholesterol membrane anchor of the Hedgehog protein confers stable membrane association to lipid-modified proteins

    Proc. Natl. Acad. Sci. U. S. A.

    (2004)
  • Cited by (67)

    View all citing articles on Scopus
    View full text