The PEG10 Gag domain generates virus-like particles.

(A) Western blots of HEK293T cells or extracellular vesicles recovered from the culture medium after transfection with VSV-G and increasing amounts of Gag-Pol cDNA. Results are representative of 2 independent experiments. (B) Western blots of extracellular vesicles in panel A after sucrose gradient density centrifugation. The upper blot shows cells transfected with HIV-1 Gag, whereas the lower blot shows cells transfected with PEG10-RF1. We speculate that the faster migrating PEG10-RF1 species is a processed form of the protein. Results are representative of 2 independent experiments. (C) Western blots of VLPs. Results are representative of 3 independent experiments. (D) Electron micrographs of negatively stained VLPs. Scale bar, 20 nm. Results are representative of 2 independent experiments. (E) Western blots of wild-type ESCs and TSCs. Results are representative of 2 independent experiments. (F) Western blots of HEK293T cells ectopically expressing wild-type (WT) mouse PEG10 or a PEG10 frameshift (FS) mutant that can only make PEG10-RF1/2. Results are representative of 2 independent experiments. (G) Western blots of extracellular vesicles shed from WT or PEG10-deficient (KO) TSCs and enriched by differential centrifugation. Results are representative of 5 independent experiments. (H) Western blots of extracellular vesicles shed from WT TSCs and analyzed by sucrose gradient density centrifugation. Results are representative of 2 independent experiments. (I) Electron micrographs of TSC extracellular vesicles after immuno-gold labeling for PEG10. Scale bar, 50 nm.