Signalling: A new trick for an old lipid

  1. Hayley Sharpe  Is a corresponding author
  1. University of Cambridge, United Kingdom

Cholesterol is a lipid molecule that is a vital component of all animal cell membranes. It provides structural integrity, which is needed for the membrane to be an effective barrier, and is also required for the production of hormones and vitamin D. These roles mean the production and transport of cholesterol in cells is strictly regulated. This, combined with its poor solubility, has hindered efforts to study its specific molecular roles. Despite this, cholesterol has long been connected to the Hedgehog signalling pathway, which helps to regulate how tissues form in animals and is mutated in several types of cancer.

Now, in eLife, Rajat Rohatgi from Stanford University, Christian Siebold from the University of Oxford and colleagues – including Giovanni Luchetti and Ria Sircar as joint first authors – report a new role for cholesterol in activating the Hedgehog pathway through the receptor protein Smoothened (Luchetti et al., 2016). Similar results have also been recently reported by Adrian Salic and colleagues (Huang et al., 2016).

There are three main components in the Hedgehog pathway that allow cells to send and receive signals: the signalling protein Hedgehog, a transmembrane protein called Patched, and a transmembrane receptor protein called Smoothened. In the absence of Hedgehog, Patched inhibits Smoothened. However, when Hedgehog binds to Patched, this inhibition is blocked and Smoothened is able to activate other Hedgehog pathway components inside the cell. It is thought that Patched and Smoothened communicate using a small molecule rather than by direct contact (Taipale et al., 2002), but it is not clear exactly how this works.

Smoothened possesses two sites at which small molecules are able to bind: one is in its transmembrane domain region and the other is in its cysteine-rich domain on the external surface of the cell. A similar cysteine-rich domain is found in several other proteins, where it is known to be able to bind to lipids (Bazan et al., 2009). Earlier this year, Rohatgi, Siebold and colleagues presented the first complete crystal structure of the transmembrane domain region and cysteine-rich domain of Smoothened (Byrne et al., 2016). Unexpectedly, they found a cholesterol molecule occupied a hydrophobic (water-fearing) pocket in the cysteine-rich domain. Since disrupting cholesterol production in humans and mice affects Smoothened activity (Blassberg et al., 2016; Cooper et al., 2003), this raised the possibility that cholesterol might directly bind to and regulate Smoothened.

Cholesterol is a challenging molecule to work with because it is hydrophobic and can randomly integrate into membranes and modify the activities of many proteins. To overcome this problem both Luchetti et al. and Huang et al. used a chemical called methyl-β-cyclodextrin to deliver cholesterol to cells and show that it directly activates Smoothened through its cysteine-rich domain.

There are many common findings between the two studies. Firstly, both teams demonstrate that cholesterol stimulates Hedgehog signalling via Smoothened with a high degree of specificity. For example, cholestenol and other molecules that are similar to cholesterol were unable to do the same. Both teams were able to rule out the transmembrane domain region as the site of cholesterol binding by showing that cholesterol could activate Smoothened even in the presence of mutations that block the binding of small molecules to this region. By contrast, mutating or completely removing the cysteine-rich domain of Smoothened blocked both the cholesterol and Hedgehog responses. Furthermore, the presence of cholesterol and Hedgehog protein together led to higher levels of Hedgehog signalling activity than the presence of just Hedgehog protein, indicating a possible role for Patched in the regulation of Smoothened by cholesterol (Figure 1).

Model for how cholesterol may regulate the Hedgehog signalling pathway. 

Left: In the absence of Hedgehog protein, the transmembrane protein Patched (purple) inhibits the transmembrane receptor protein Smoothened (blue and grey) via an unknown mechanism. The findings of Luchetti et al. and Huang et al. suggest that Patched and Smoothened may communicate using the lipid molecule cholesterol (green), which is a core component of animal cell membranes (orange lines). Patched is similar to other proteins that transport small molecules across membranes, and might act to limit cholesterol access to Smoothened. In the absence of available cholesterol, Smoothened receptors on the cell surface are inactive. Right: Hedgehog protein binds to and inactivates Patched, potentially increasing cholesterol levels outside the cell. Cholesterol binds to a hydrophobic pocket in the Smoothened cysteine-rich domain (blue). Smoothened can now activate the Hedgehog signalling pathway, although many details of this process are not fully understood (see main article). In particular, it is not clear how cholesterol gains access to the Smoothened cysteine-rich domain. Cholesterol can be released from the membrane into the extracellular space by a process called desorption, but its insolubility makes this energetically unfavourable. Alternatively, a sterol-binding protein outside cells could deliver cholesterol to Smoothened.

How does cholesterol binding outside the cell translate to signalling within the cell? Luchetti et al. predict, based on previous structures (Byrne et al., 2016), that cholesterol binding to the cysteine-rich domain of Smoothened induces a clockwise rotation with respect to the transmembrane domain region. This change in shape could be sufficient to promote signalling inside the cell.

Together the findings of Luchetti et al. and Huang et al. strongly support a role for cholesterol in activating Smoothened in cells. However, it is worth noting that recent findings from other research groups favour an inhibitory role for sterol molecules instead (Roberts et al., 2016; Sever et al., 2016). Therefore, several critical questions remain. Does cholesterol binding itself alter Smoothened activity, or is cholesterol merely a cofactor that is needed for Smoothened to be activated by another molecule? Does Hedgehog protein affect cholesterol levels and is this mediated through the activity of Patched (Figure 1)? Since most cholesterol is trapped within the cell membrane, it will also be important to understand how cholesterol is able to access the cysteine-rich domain of Smoothened.

Nonetheless, this work reveals a new signalling role for cholesterol in controlling the Smoothened receptor and reiterates the possibility that Hedgehog signalling may have evolved from an ancient lipid-sensing pathway (Hausmann et al., 2009).


Article and author information

Author details

  1. Hayley Sharpe

    Cambridge Institute for Medical Research, University of Cambridge, Cambridge, United Kingdom
    For correspondence
    Competing interests
    The author declares that no competing interests exist.

Publication history

  1. Version of Record published: November 25, 2016 (version 1)


© 2016, Sharpe

This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.


  • 1,322
    Page views
  • 174
  • 0

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Open citations (links to open the citations from this article in various online reference manager services)

Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)

  1. Hayley Sharpe
Signalling: A new trick for an old lipid
eLife 5:e22492.

Further reading

    1. Biochemistry and Chemical Biology
    2. Cell Biology
    Haoyan Huang, Meng Qian ... Zongjin Li
    Research Article Updated

    Nitric oxide (NO), as a gaseous therapeutic agent, shows great potential for the treatment of many kinds of diseases. Although various NO delivery systems have emerged, the immunogenicity and long-term toxicity of artificial carriers hinder the potential clinical translation of these gas therapeutics. Mesenchymal stem cells (MSCs), with the capacities of self-renewal, differentiation, and low immunogenicity, have been used as living carriers. However, MSCs as gaseous signaling molecule (GSM) carriers have not been reported. In this study, human MSCs were genetically modified to produce mutant β-galactosidase (β-GALH363A). Furthermore, a new NO prodrug, 6-methyl-galactose-benzyl-oxy NONOate (MGP), was designed. MGP can enter cells and selectively trigger NO release from genetically engineered MSCs (eMSCs) in the presence of β-GALH363A. Moreover, our results revealed that eMSCs can release NO when MGP is systemically administered in a mouse model of acute kidney injury (AKI), which can achieve NO release in a precise spatiotemporal manner and augment the therapeutic efficiency of MSCs. This eMSC and NO prodrug system provides a unique and tunable platform for GSM delivery and holds promise for regenerative therapy by enhancing the therapeutic efficiency of stem cells.

    1. Biochemistry and Chemical Biology
    2. Cell Biology
    Maria Körner, Susanne R Meyer ... Alexander Buchberger
    Research Article Updated

    The ATPase p97 (also known as VCP, Cdc48) has crucial functions in a variety of important cellular processes such as protein quality control, organellar homeostasis, and DNA damage repair, and its de-regulation is linked to neuromuscular diseases and cancer. p97 is tightly controlled by numerous regulatory cofactors, but the full range and function of the p97–cofactor network is unknown. Here, we identify the hitherto uncharacterized FAM104 proteins as a conserved family of p97 interactors. The two human family members VCP nuclear cofactor family member 1 and 2 (VCF1/2) bind p97 directly via a novel, alpha-helical motif and associate with p97-UFD1-NPL4 and p97-UBXN2B complexes in cells. VCF1/2 localize to the nucleus and promote the nuclear import of p97. Loss of VCF1/2 results in reduced nuclear p97 levels, slow growth, and hypersensitivity to chemical inhibition of p97 in the absence and presence of DNA damage, suggesting that FAM104 proteins are critical regulators of nuclear p97 functions.