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Cell Rep. 2017 Oct 3;21(1):274-288. doi: 10.1016/j.celrep.2017.09.022.

A RhoA-FRET Biosensor Mouse for Intravital Imaging in Normal Tissue Homeostasis and Disease Contexts.

Author information

1
The Garvan Institute of Medical Research, St. Vincent's Clinical School, Faculty of Medicine, University of New South Wales, Sydney, NSW 2010, Australia.
2
Cancer Research UK Beatson Institute, Switchback Road, Bearsden, Glasgow G611BD, UK.
3
Neuromuscular and Regenerative Medicine Unit, University of New South Wales, Sydney, NSW 2010, Australia; Oncology Research Unit, School of Medical Sciences, University of New South Wales, Sydney, NSW 2010, Australia.
4
Neuromuscular and Regenerative Medicine Unit, University of New South Wales, Sydney, NSW 2010, Australia.
5
Oncology Research Unit, School of Medical Sciences, University of New South Wales, Sydney, NSW 2010, Australia.
6
Developmental and Stem Cell Biology Division, Victor Chang Cardiac Research Institute, Sydney, NSW 2010, Australia; St. Vincent's Clinical School of Biotechnology and Biomolecular Sciences, University of New South Wales, Sydney, NSW 2052, Australia.
7
Centre for Cancer Biology, SA Pathology and University of South Australia School of Medicine, University of Adelaide, Adelaide, SA 5000, Australia.
8
Signalling Programme, Babraham Institute, Cambridge CB223AT, UK.
9
Francis Crick Institute, London NW11AT, UK. Electronic address: kurt.anderson@crick.ac.uk.
10
The Garvan Institute of Medical Research, St. Vincent's Clinical School, Faculty of Medicine, University of New South Wales, Sydney, NSW 2010, Australia. Electronic address: p.timpson@garvan.org.au.

Abstract

The small GTPase RhoA is involved in a variety of fundamental processes in normal tissue. Spatiotemporal control of RhoA is thought to govern mechanosensing, growth, and motility of cells, while its deregulation is associated with disease development. Here, we describe the generation of a RhoA-fluorescence resonance energy transfer (FRET) biosensor mouse and its utility for monitoring real-time activity of RhoA in a variety of native tissues in vivo. We assess changes in RhoA activity during mechanosensing of osteocytes within the bone and during neutrophil migration. We also demonstrate spatiotemporal order of RhoA activity within crypt cells of the small intestine and during different stages of mammary gestation. Subsequently, we reveal co-option of RhoA activity in both invasive breast and pancreatic cancers, and we assess drug targeting in these disease settings, illustrating the potential for utilizing this mouse to study RhoA activity in vivo in real time.

KEYWORDS:

FLIM-FRET; actin; biosensors; breast cancer; cell biology; development; immunology; intravital imaging; pancreatic cancer; small GTPase RhoA

PMID:
28978480
DOI:
10.1016/j.celrep.2017.09.022
[Indexed for MEDLINE]
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