Early steps of clathrin-mediated endocytosis involved in phagosomal escape of Fcgamma receptor-targeted adenovirus.
Adenovirus type 2 (Ad2) and Ad5 enter epithelial cells via the coxsackievirus B Ad receptor (CAR) and alpha(v) integrin coreceptors. In the absence of CAR, they can be directed to the Fcgamma receptor 1 of hematopoietic cells by an adaptor comprising the extracellular CAR domain and the Fc portion of a human immunoglobulin G (CARex-Fc). This gives rise to Ad aggregates and single particles which together enhance gene delivery up to 250-fold compared to adaptor-less viruses. A small interfering RNA knockdown of the clathrin heavy chain and quantitative electron microscopy of hematopoietic leukemia cells showed that the majority of Ads were phagocytosed as clusters of 1 to 3 microm in diameter and that about 10% of the particles entered cells by clathrin-mediated endocytosis. The clathrin knockdown did not affect phagocytosis but, surprisingly, inhibited viral escape from phagosomes. Similarly, blocking an early stage of clathrin-coated pit assembly inhibited phagosomal escape and infection but not aggregate uptake, unlike blocking of a late stage of clathrin-coated pit formation. We propose a cooperative interaction of clathrin-mediated endocytosis and phagocytosis triggering phagosomal lysis and infection.
Meier
Oliver
O
Zoologisches Institut, University of Zürich, 8057 Zürich, Switzerland.
Gastaldelli
Michele
M
Boucke
Karin
K
Hemmi
Silvio
S
Greber
Urs F
UF
D000260:Adenoviruses, Human / Q000502:physiology*; D002460:Cell Line; D002966:Clathrin / Q000502:physiology; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; D004705:Endocytosis / Q000502:physiology*; D006801:Humans; D010587:Phagocytosis / Q000502:physiology; D017452:Receptors, IgG / Q000502:physiology*; D011991:Receptors, Virus / Q000502:physiology*
C568771:CLMP protein, human; D002966:Clathrin; D062727:Coxsackie and Adenovirus Receptor-Like Membrane Protein; D017452:Receptors, IgG; D011991:Receptors, Virus