%0 Journal Article
%T ALG-2-interacting Tubby-like protein superfamily member PLSCR3 is secreted by an exosomal pathway and taken up by recipient cultured cells
%A Tatsutoshi Inuzuka
%A Akira Inokawa
%A Cen Chen
%A Kumiko Kizu
%J Bioscience Reports
%D 2013
%I Portland Press, Biochemical Society
%R 10.1042/bsr20120123
%X PLSCRs (phospholipid scramblases) are palmitoylated membrane-associating proteins. Regardless of the given names, their physiological functions are not clear and thought to be unrelated to phospholipid scrambling activities observed in vitro. Using a previously established cell line of HEK-293 (human embryonic kidney-293) cells constitutively expressing human Scr3 (PLSCR3) that interacts with ALG-2 (apoptosis-linked gene 2) Ca2+-dependently, we found that Scr3 was secreted into the culture medium. Secretion of Scr3 was suppressed by 2-BP (2-bromopalmitate, a palmitoylation inhibitor) and by GW4869 (an inhibitor of ceramide synthesis). Secreted Scr3 was recovered in exosomal fractions by sucrose density gradient centrifugation. Palmitoylation sites and the N-terminal Pro-rich region were necessary for efficient secretion, but ABSs (ALG-2-binding sites) were dispensable. Overexpression of GFP (green fluorescent protein)-fused VPS4BE235Q, a dominant negative mutant of an AAA (ATPase associated with various cellular activities) ATPase with a defect in disassembling ESCRT (endosomal sorting complex required for transport)-III subunits, significantly reduced secretion of Scr3. Immunofluorescence microscopic analyses showed that Scr3 was largely localized to enlarged endosomes induced by overexpression of a GFP-fused constitutive active mutant of Rab5A (GFP¨CRab5AQ79L). Secreted Scr3 was taken up by HeLa cells, suggesting that Scr3 functions as a cell-to-cell transferable modulator carried by exosomes in a paracrine manner.
%K apoptosis-linked gene 2 (ALG-2)
%K endosome
%K exosome
%K palmitoylation
%K unconventional secretion
%K vesicle
%U http://www.bioscirep.org/bsr/033/e026/bsr033e026.htm