TY - JOUR
T1 - Membrane-bound Gaussia luciferase as a tool to track shedding of membrane proteins from the surface of extracellular vesicles
AU - Zaborowski, Mikołaj Piotr
AU - Cheah, Pike See
AU - Zhang, Xuan
AU - Bushko, Isabella
AU - Lee, Kyungheon
AU - Sammarco, Alessandro
AU - Zappulli, Valentina
AU - Maas, Sybren Lein Nikola
AU - Allen, Ryan M.
AU - Rumde, Purva
AU - György, Bence
AU - Aufiero, Massimo
AU - Schweiger, Markus W.
AU - Lai, Charles Pin Kuang
AU - Weissleder, Ralph
AU - Lee, Hakho
AU - Vickers, Kasey C.
AU - Tannous, Bakhos A.
AU - Breakefield, Xandra O.
PY - 2019/11/22
Y1 - 2019/11/22
N2 - Extracellular vesicles (EVs) released by cells play a role in intercellular communication. Reporter and targeting proteins can be modified and exposed on the surface of EVs to investigate their half-life and biodistribution. A characterization of membrane-bound Gaussia luciferase (mbGluc) revealed that its signal was detected also in a form smaller than common EVs (<70 nm). We demonstrated that mbGluc initially exposed on the surface of EVs, likely undergoes proteolytic cleavage and processed fragments of the protein are released into the extracellular space in active form. Based on this observation, we developed a new assay to quantitatively track shedding of membrane proteins from the surface of EVs. We used this assay to show that ectodomain shedding in EVs is continuous and is mediated by specific proteases, e.g. metalloproteinases. Here, we present a novel tool to study membrane protein cleavage and release using both in vitro and in vivo models.
AB - Extracellular vesicles (EVs) released by cells play a role in intercellular communication. Reporter and targeting proteins can be modified and exposed on the surface of EVs to investigate their half-life and biodistribution. A characterization of membrane-bound Gaussia luciferase (mbGluc) revealed that its signal was detected also in a form smaller than common EVs (<70 nm). We demonstrated that mbGluc initially exposed on the surface of EVs, likely undergoes proteolytic cleavage and processed fragments of the protein are released into the extracellular space in active form. Based on this observation, we developed a new assay to quantitatively track shedding of membrane proteins from the surface of EVs. We used this assay to show that ectodomain shedding in EVs is continuous and is mediated by specific proteases, e.g. metalloproteinases. Here, we present a novel tool to study membrane protein cleavage and release using both in vitro and in vivo models.
UR - http://www.scopus.com/inward/record.url?scp=85075513481&partnerID=8YFLogxK
U2 - 10.1038/s41598-019-53554-y
DO - 10.1038/s41598-019-53554-y
M3 - Article
C2 - 31758005
AN - SCOPUS:85075513481
SN - 2045-2322
VL - 9
SP - 1
EP - 16
JO - Scientific Reports
JF - Scientific Reports
IS - 1
M1 - 17387
ER -