Single live cell TGF-β signalling imaging: breast cancer cell motility and migration is driven by sub-populations of cells with dynamic TGF-β-Smad3 activity.
BACKGROUND
Metastasis is a process where only a small subset of cells is capable of successfully migrating to and propagating at secondary sites. TGF-β signalling is widely known for its role in cancer metastasis and is associated with cell migration in whole cell populations.


RESULTS
We extend these findings by investigating the role of TGF-β signalling in promoting migration and motility by imaging the signalling activity in live, individual MDA-MB-231 cancer cells utilizing a novel Smad3 Td-Tomato reporter adenovirus. Here we find that not all MDA-MB-231 cancer cells have similar TGF-β mediated Smad3 transcription activity and display at least two distinct migratory populations. Importantly, Smad3 activity was significantly higher within migratory cells compared to non-migrated cells in wound healing and transwell assays. Furthermore, time-lapse experiments showed that MDA-MB-231 cells displaying Smad3 activity moved faster and a greater distance compared to cells not displaying Smad3 reporter activity. Interestingly, despite being more motile than cells with undetectable levels of Smad3 activity, high Smad3 activity was detrimental to cell motility compared to low and medium level of Smad3 activity.


CONCLUSIONS
We have developed a method enabling real-time visualization of TGF-β signalling in single live cells. Breast cancer cell motility and migration is driven by sub-populations of cells with dynamic TGF-β-Smad3 activity. Those sub-populations may be responsible for tumor invasion and metastasis.
Department of Surgery (RMH), The University of Melbourne, The Royal Melbourne Hospital, Parkville, VIC, 3050, Australia. rluwor@unimelb.edu.au.
Rodney B
RB
Luwor
Department of Surgery (RMH), The University of Melbourne, The Royal Melbourne Hospital, Parkville, VIC, 3050, Australia. d.hakmana@student.unimelb.edu.au.
Dulani
D
Hakmana
Department of Surgery (RMH), The University of Melbourne, The Royal Melbourne Hospital, Parkville, VIC, 3050, Australia. jiaria@unimelb.edu.au.
Josephine
J
Iaria
Department of Surgery (RMH), The University of Melbourne, The Royal Melbourne Hospital, Parkville, VIC, 3050, Australia. thao.nheu@gmail.com.
Thao V
TV
Nheu
Department of Biochemistry, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, VIC, 3086, Australia. Richard.Simpson@latrobe.edu.au.
Richard J
RJ
Simpson
Department of Surgery (RMH), The University of Melbourne, The Royal Melbourne Hospital, Parkville, VIC, 3050, Australia. hongjian@unimelb.edu.au.
Hong-Jian
HJ
Zhu
D001943:Breast Neoplasms / Q000235:genetics*; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics*; D005260:Female; D006801:Humans; D015398:Signal Transduction / Q000235:genetics*; D051900:Smad3 Protein / Q000235:genetics*; D014158:Transcription, Genetic / Q000235:genetics; D016212:Transforming Growth Factor beta / Q000235:genetics*
C497790:SMAD3 protein, human; D051900:Smad3 Protein; D016212:Transforming Growth Factor beta