Specifics of viral entry in epidemic keratoconjunctivitis

Abstract

Cellular entry pathways for viruses dictate the types of immune responses elicited upon infection. Human adenovirus species D type 37 (HAdV‐D37) causes epidemic keratoconjunctivitis (EKC), associated with severe ocular surface inflammation. However, viral entry is known to be cell type and virus specific. To examine basic mechanisms of HAdV‐D37 pathogenesis in EKC, we studied entry of HAdV‐D37 in vivo in our mouse corneal adenovirus keratitis model, and in vitro in cultured human corneal cells. In the mouse model at 1 hour post infection, viruses appeared to be entering cells by macropinocytosis. By 8 hours post infection, viruses formed regular packed intracytoplasmic arrays adjacent to cell nuclei, with minimal capsid uncoating. Therefore, viral entry occurred, but transport of virus to the cell nucleus appeared abortive, possibly explaining the failure of HAdV‐D37 replication in the mouse cornea. In vitro, inhibition of PKCα reduced viral entry and phosphorylation of both Src and caveolin‐1 in lipid raft fractions, suggesting that PKCα activity occurs upstream of both molecules. Phosphorylated PKCα and Src were found in same endosomal fractions and may be physically associated. These results suggest a central role for PKC in HAdV‐D37 entry and regulation of downstream molecules in corneal cells, including caveolin‐1. Interestingly, the small GTPase dynamin II, did not appear necessary for HAdV‐D37 entry into corneal epithelial cells, but was critical for viral entry in corneal fibroblasts.