Entry of human parechovirus 1

Human parechovirus 1 (HPEV-1) is a prototype member of parechoviruses, a re cently established picornavirus genus. Although there is preliminary eviden ce that HPEV-1 recognizes alpha (V) integrins as cellular receptors, our un derstanding of early events during HPEV-1 infection is still very limited. The aim of this study was to clarify the entry mechanisms of HPEV-1, includ ing the attachment of the virus onto the host cell surface and subsequent i nternalization. In blocking experiments with monoclonal antibodies against different receptor candidates, antibodies against alpha (V) and beta (3) in tegrin subunits, in particular in combination, appeared to be the most effi cient ones in preventing the HPEV-1 infection. To find out whether HPEV-1 u ses clathrin-coated vesicles or other routes for the entry into the host ce ll, we carried out double-labeling experiments of virus-infected cells with anti-HPEV-1 antibodies and antibodies against known markers of the clathri n and the caveolin routes. At the early phase of infection (5 min postinfec tion [p.i.]) HPEV-1 colocalized with EEA1 (early endosomes), and later, aft er 30 min p.i., it colocalized with mannose-6-phosphate receptor (late endo somes), whereas no colocalization with caveolin-1 was observed. The data in dicate that HPEV 1 utilizes the clathrin-dependent endocytic pathway for en try into the host cells. Interestingly, endocytosed HPEV-1 capsid proteins were observed in the endoplasmic reticulum and cis-Golgi network 30 to 60 m in p.i. Depolymerization of microtubules with nocodazole inhibited transloc ation of the virus to the late endosomes but did not block HPEV-1 replicati on, suggesting that the RNA genome may be released early during the entry p rocess.