Microvesicle and tunneling nanotube mediated intercellular transfer of g-protein coupled receptors in cell cultures

• Published in: Experimental cell research 2012-03, Vol.318 (5), p.603-613
• Main Authors: Guescini, M., Leo, G., Genedani, S., Carone, C., Pederzoli, F., Ciruela, F., Guidolin, D., Stocchi, V., Mantuano, M., Borroto-Escuela, D.O., Fuxe, K., Agnati, L.F.
• Format: Article
• Language: English
• Subjects: 60 APPLIED LIFE SCIENCES; ADENOSINE; AMP; Animals; Biological Transport; Cell Communication; Cell culture; CELL CULTURES; Cells, Cultured; Cellular biology; Cercopithecus aethiops; Coculture Techniques; COS Cells; Exosomes; FLUORESCENCE; Fluorescence Resonance Energy Transfer; GPCRs; Green Fluorescent Proteins - metabolism; GTP-ASES; HEK293 Cells; Humans; Inter-cellular communication; Medicin och hälsovetenskap; Microscopy, Confocal; Microvesicles; NANOTUBES; Proteins; Receptor, Adenosine A2A - metabolism; RECEPTORS; Receptors, Dopamine D2 - metabolism; Recombinant Fusion Proteins - metabolism; Synaptic modulation; TRANSMISSION ELECTRON MICROSCOPY; Transport Vesicles - metabolism
• ISSN: 0014-4827; 1090-2422; 1090-2422
• DOI: 10.1016/j.yexcr.2012.01.005

Recent evidence shows that cells exchange collections of signals via microvesicles (MVs) and tunneling nano-tubes (TNTs). In this paper we have investigated whether in cell cultures GPCRs can be transferred by means of MVs and TNTs from a source cell to target cells. Western blot, transmission electron microscopy and gene expression analyses demonstrate that A2A and D2 receptors are present in released MVs. In order to further demonstrate the involvement of MVs in cell-to-cell communication we created two populations of cells (HEK293T and COS-7) transiently transfected with D2R-CFP or A2AR-YFP. These two types of cells were co-cultured, and FRET analysis demonstrated simultaneously positive cells to the D2R-CFP and A2AR-YFP. Fluorescence microscopy analysis also showed that GPCRs can move from one cell to another also by means of TNTs. Finally, recipient cells pre-incubated for 24h with A2AR positive MVs were treated with the adenosine A2A receptor agonist CGS-21680. The significant increase in cAMP accumulation clearly demonstrated that A2ARs were functionally competent in target cells. These findings demonstrate that A2A receptors capable of recognizing and decoding extracellular signals can be safely transferred via MVs from source to target cells.