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Key Event Title
Antagonism, Smoothened receptor
|Level of Biological Organization|
Key Event Components
|regulation of receptor activity||smoothened||decreased|
Key Event Overview
AOPs Including This Key Event
|AOP Name||Role of event in AOP||Point of Contact||Author Status||OECD Status|
|Anatagonsim SMO leads to OFC||MolecularInitiatingEvent||Jacob Reynolds (send email)||Under development: Not open for comment. Do not cite|
|All life stages||High|
Key Event Description
The Smoothened (SMO) receptor is Class F G protein coupled receptor involved in signal transduction of the Sonic Hedgehog (SHH) pathway. It includes distinct functional groups including ligand binding pockets, cysteine rich domain (CRD), transmembrane helix (TM), extracellular loop (ECL), intracellular loop (ICL), and a carboxyl-terminal tail (C-term tail) (Arensdorf, Marada et al. 2016). SMO signaling is dependent upon its relocation to a subcellular location. This occurs in the plasma membrane for flies (Denef, Neubüser et al. 2000) and the primary cilium (PC) in vertebrates (Huangfu and Anderson 2005).
In the absence of Hedgehog (HH) ligand, the Patched (PTCH) receptor suppresses the activation of SMO. When HH ligand binds to PTCH, suppression on SMO is released and SMO is able to relocate, accumulate, and signal to intracellular effectors (Denef, Neubüser et al. 2000). This signaling to effectors results in the activation of the GLI transcription factors and the subsequent induction of HH target gene expression(Alexandre, Jacinto et al. 1996, Von Ohlen and Hooper 1997). The exact mechanism through which PTCH and SMO interact is not known.
An endogenous ligand for SMO has not been discovered although evidence for one exists and that PTCH controls SMO by controlling its’ availability or accessibility. To support this, it has been shown that PTCH and SMO do not physically interact (Chen and Struhl 1998). PTCH acts catalytically with SMO with one PTCH receptor capable of controlling many (~50) SMO receptors (Taipale, Cooper et al. 2002). Since PTCH includes a sterol sensing domain and shares characteristics of ancient bacterial transporters, a model of PTCH functioning by pumping a sterol-like MSO regulator has been proposed (Mukhopadhyay and Rohatgi 2014). SMO is constitutively active in the absence of PTCH suggesting that the elusive molecule is an agonist (Rohatgi and Scott 2007). Conversely, the discovery that oxysterols bind to the CRD binding domain acting as positive modulators suggest that the molecule could be an agonist with PTCH functioning to sequester away or limit cellular concentration (Corcoran and Scott 2006, Nachtergaele, Mydock et al. 2012)
The activity of SMO is controlled by ligand binding (Kobilka 2007). Two separate binding pockets, one in the groove of the extracellular CRD and the other in the helices of the TMD have been identified (Nachtergaele, Mydock et al. 2012, Rana, Carroll et al. 2013, Wang, Wu et al. 2013, Byrne, Sircar et al. 2016, Huang, Zheng et al. 2018). These two binding pockets have been shown to interact in an allosteric manner (Nachtergaele, Mydock et al. 2012). The binding pocket in the helices of the TMD binds several SMO agonists including SAG as well as antagonists Vismodegib and Sonidegib. The CRD binding pocket binds cholesterol and its’ oxidized derivates (Byrne, Luchetti et al. 2018). The antagonist cyclopamine binds to the TMD binding pocket and inhibits SHH signal transduction. However, in mSMO carrying the mutations D477G/E552K that disable the TMD binding pocket, cyclopamine binds to the CRD pocket and activates the pathway (Huang, Nedelcu et al. 2016). To date several oxysterols including 20(S)-hydroxylcholesterol, 22(S)-hydroxylcholesterol, 7-keto-25-hydroxylcholesterol and 7-keto-27-hydroxylcholesterol have been identified as activators of SMO (Dwyer, Sever et al. 2007, Nachtergaele, Mydock et al. 2012, Myers, Sever et al. 2013). A binding site for 24(S),25-epoxycholesterol has been identified in the TMD pocket using cryo-EM of SMO in complex with 24(S),25-epoxycholesterol (Qi, Liu et al. 2019).
How It Is Measured or Detected
Verification of binding and affinity for SMO can be measured using fluorescence binding assays and photoaffinity labeling respectively (Chen, Taipale et al. 2002). qRT-PCR can be used to determine the expression level of SMO (Lou, Li et al. 2020).
Domain of Applicability
- Sex- SMO is present in both male and females and differences in activation or antagonism between sex have not been demonstrated.
- Life stages- The Hedgehog pathway is a major pathway in embryonic development. Aberrant activation of HH signalling is known to cause cancer (Dahmane, Lee et al. 1997, Kimura, Stephen et al. 2005). For these reasons all stages of life are of relevance.
- Taxonomic- SMO is conserved in both vertebrates and invertebrates. SMO signaling is dependent upon its relocation to a subcellular location. This occurs in the plasma membrane for flies (Denef, Neubüser et al. 2000) and the primary cilium (PC) in vertebrates (Huangfu and Anderson 2005).
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