TY - JOUR
T1 - Adeno-associated virus 2 infection requires endocytosis through the CLIC/GEEC pathway
AU - Nonnenmacher, Mathieu
AU - Weber, Thomas
N1 - Funding Information:
We gratefully acknowledge Alice Dautry (Institut Pasteur, Paris, France), Mark A. McNiven (Mayo Clinic, Rochester, MN), Jürgen Kleinschmidt (Deutsches Krebsforschungszentrum, Heidelberg, Germany), and Sergei Zolotukhin (University of Florida, Gainesville, FL) for providing us with Eps15, dynamin2, pDG, and pTRUF12 plasmids, respectively. We thank Harvey T. MacMahon (Medical Research Council, Cambridge, UK) for the GRAF1 antibody and Kleopatra Rapti and Lifan Liang (Mount Sinai School of Medicine, New York, NY) for Ad5 vectors. We are grateful to Serine Avagyan, Lorenzo Battini, and Raphael Rozenfeld (Mount Sinai School of Medicine, New York, NY) for their help with flow cytometry and microscopy, and Christophe Lamaze (Institut Curie, Paris, France) for helpful discussions and advice. We thank Ronald Gordon and Heather Bell from the Pathology Core Electron Microscopy Facility of the Mount Sinai School of Medicine (New York, NY) for their help. We gratefully acknowledge the continuous support by Roger J. Hajjar (Mount Sinai School of Medicine, New York, NY; R.J.H.). This work was supported by U.S. National Institutes of Health Grants HL077322 (to T.W.), and HL100396 and HL088434 (to R.J.H.).
PY - 2011/12/15
Y1 - 2011/12/15
N2 - Adeno-associated viruses (AAVs) are nonpathogenic, nonenveloped, single-stranded DNA viruses in development as gene therapy vectors. AAV internalization was postulated to proceed via a dynamin-dependent endocytic mechanism. Revisiting this, we find that infectious endocytosis of the prototypical AAV, AAV2, is independent of clathrin, caveolin, and dynamin. AAV2 infection is sensitive to EIPA, a fluid-phase uptake inhibitor, but is unaffected by Rac1 mutants or other macropinocytosis inhibitors. In contrast, AAV2 infection requires actin cytoskeleton remodeling and membrane cholesterol and is sensitive to inhibition of Cdc42, Arf1, and GRAF1, factors known to be involved in the formation of clathrin-independent carriers (CLIC). AAV2 virions are internalized in the detergent-resistant GPI-anchored-protein-enriched endosomal compartment (GEEC) and translocated to the Golgi apparatus, similarly to the CLIC/GEEC marker cholera toxin B. Our results indicate that - unlike the viral entry mechanisms described so far - AAV2 uses the pleiomorphic CLIC/GEEC pathway as its major endocytic infection route.
AB - Adeno-associated viruses (AAVs) are nonpathogenic, nonenveloped, single-stranded DNA viruses in development as gene therapy vectors. AAV internalization was postulated to proceed via a dynamin-dependent endocytic mechanism. Revisiting this, we find that infectious endocytosis of the prototypical AAV, AAV2, is independent of clathrin, caveolin, and dynamin. AAV2 infection is sensitive to EIPA, a fluid-phase uptake inhibitor, but is unaffected by Rac1 mutants or other macropinocytosis inhibitors. In contrast, AAV2 infection requires actin cytoskeleton remodeling and membrane cholesterol and is sensitive to inhibition of Cdc42, Arf1, and GRAF1, factors known to be involved in the formation of clathrin-independent carriers (CLIC). AAV2 virions are internalized in the detergent-resistant GPI-anchored-protein-enriched endosomal compartment (GEEC) and translocated to the Golgi apparatus, similarly to the CLIC/GEEC marker cholera toxin B. Our results indicate that - unlike the viral entry mechanisms described so far - AAV2 uses the pleiomorphic CLIC/GEEC pathway as its major endocytic infection route.
UR - https://www.scopus.com/pages/publications/83655164541
U2 - 10.1016/j.chom.2011.10.014
DO - 10.1016/j.chom.2011.10.014
M3 - Article
C2 - 22177561
AN - SCOPUS:83655164541
SN - 1931-3128
VL - 10
SP - 563
EP - 576
JO - Cell Host and Microbe
JF - Cell Host and Microbe
IS - 6
ER -