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@article{Ciepla:2014:10.1039/C4SC01600A,
author = {Ciepla, P and Konitsiotis, AD and Serwa, RA and Masumoto, N and Leong, WP and Dallman, MJ and Magee, AI and Tate, EW},
doi = {10.1039/C4SC01600A},
journal = {Chemical Science},
pages = {4249--4259},
title = {New chemical probes targeting cholesterylation of Sonic Hedgehog in human cells and zebrafish},
url = {http://dx.doi.org/10.1039/C4SC01600A},
volume = {5},
year = {2014}
}
TY - JOUR
AB - Sonic Hedgehog protein (Shh) is a morphogen molecule important in embryonic development and in theprogression of many cancer types in which it is aberrantly overexpressed. Fully mature Shh requiresattachment of cholesterol and palmitic acid to its C- and N-termini, respectively. The study of lipidatedShh has been challenging due to the limited array of tools available, and the roles of theseposttranslational modifications are poorly understood. Herein, we describe the development andvalidation of optimised alkynyl sterol probes that efficiently tag Shh cholesterylation and enable itsvisualisation and analysis through bioorthogonal ligation to reporters. An optimised probe was shown tobe an excellent cholesterol biomimetic in the context of Shh, enabling appropriate release of tagged Shhfrom signalling cells, formation of multimeric transport complexes and signalling. We have used thisprobe to determine the size of transport complexes of lipidated Shh in culture medium and expressionlevels of endogenous lipidated Shh in pancreatic ductal adenocarcinoma cell lines through quantitativechemical proteomics, as well as direct visualisation of the probe by fluorescence microscopy anddetection of cholesterylated Hedgehog protein in developing zebrafish embryos. These sterol probesprovide a set of novel and well-validated tools that can be used to investigate the role of lipidation onactivity of Shh, and potentially other members of the Hedgehog protein family
AU - Ciepla,P
AU - Konitsiotis,AD
AU - Serwa,RA
AU - Masumoto,N
AU - Leong,WP
AU - Dallman,MJ
AU - Magee,AI
AU - Tate,EW
DO - 10.1039/C4SC01600A
EP - 4259
PY - 2014///
SN - 2041-6520
SP - 4249
TI - New chemical probes targeting cholesterylation of Sonic Hedgehog in human cells and zebrafish
T2 - Chemical Science
UR - http://dx.doi.org/10.1039/C4SC01600A
UR - http://hdl.handle.net/10044/1/26917
VL - 5
ER -
Prof. Ed Tate
GSK Chair in Chemical Biology
Department of Chemistry
Molecular Sciences Research Hub, White City Campus,
82 Wood Lane, London, W12 0BZ
e.tate@imperial.ac.uk
Tel: +44 (0)20 759 + ext 43752 or 45821