A highly sensitive prenylation assay reveals in vivo effects of bisphosphonate drug on the Rab prenylome of macrophages outside the skeleton

Ali, Naveid, Jurczyluk, Julie, Shay, Gemma, Tnimov, Zakir, Alexandrov, Kirill, Munoz, Marcia A., Skinner, Oliver P., Pavlos, Nathan J. and Rogers, Michael J. (2015) A highly sensitive prenylation assay reveals in vivo effects of bisphosphonate drug on the Rab prenylome of macrophages outside the skeleton. Small GTPases, 6 4: 202-211. doi:10.1080/21541248.2015.1085485


Author Ali, Naveid
Jurczyluk, Julie
Shay, Gemma
Tnimov, Zakir
Alexandrov, Kirill
Munoz, Marcia A.
Skinner, Oliver P.
Pavlos, Nathan J.
Rogers, Michael J.
Title A highly sensitive prenylation assay reveals in vivo effects of bisphosphonate drug on the Rab prenylome of macrophages outside the skeleton
Journal name Small GTPases   Check publisher's open access policy
ISSN 2154-1248
2154-1256
Publication date 2015-09-23
Sub-type Article (original research)
DOI 10.1080/21541248.2015.1085485
Open Access Status DOI
Volume 6
Issue 4
Start page 202
End page 211
Total pages 10
Place of publication Philadelphia, PA, United States
Publisher Taylor & Francis
Language eng
Abstract Bisphosphonate drugs such as zoledronic acid (ZOL), used for the treatment of common bone disorders, target the skeleton and inhibit bone resorption by preventing the prenylation of small GTPases in bone-destroying osteoclasts. Increasing evidence indicates that bisphosphonates also have pleiotropic effects outside the skeleton, most likely via cells of the monocyte/macrophage lineage exposed to nanomolar circulating drug concentrations. However, no effects of such low concentrations of ZOL have been reported using existing approaches. We have optimized a highly sensitive in vitro prenylation assay utilizing recombinant geranylgeranyltransferases to enable the detection of subtle effects of ZOL on the prenylation of Rab- and Rho-family GTPases. Using this assay, we found for the first time that concentrations of ZOL as low as 10nM caused inhibition of Rab prenylation in J774 macrophages following prolonged cell culture. By combining the assay with quantitative mass spectrometry we identified an accumulation of 18 different unprenylated Rab proteins in J774 cells after nanomolar ZOL treatment, with a >7-fold increase in the unprenylated form of Rab proteins associated with the endophagosome pathway (Rab1, Rab5, Rab6, Rab7, Rab11, Rab14 and Rab21). Finally, we also detected a clear effect of subcutaneous ZOL administration in vivo on the prenylation of Rab1A, Rab5B, Rab7A and Rab14 in mouse peritoneal macrophages, confirming that systemic treatment with bisphosphonate drug can inhibit prenylation in myeloid cells in vivo outside the skeleton. These observations begin a new era in defining the precise pharmacological actions of bisphosphonate drugs on the prenylation of small GTPases in vivo.
Keyword Bisphosphonate
Geranylgeranyl transferase
Macrophage
Prenylation
Rab
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status UQ

 
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Created: Thu, 03 Dec 2015, 18:47:29 EST by Susan Allen on behalf of Institute for Molecular Bioscience