Cellular Trafficking of FX-Ad5fiber35 Complex Figure 8. Electron microscopy of CHO-K1 cells incubated with HAdV5wt at 10,000 vp/cell, (A) in the absence (w/o), or (B) presence of FX (8 mg/ml) for 2 h at 376C. (A), (i) and (ii): General views of cell sections showing (i) intravesicular and (ii) cytoplasmic vector particles. In (iii) and (iv), a vector particle (Vir) is seen within a clathrin-coated vesicle (CCV); (iv), enlargement of the CCV shown in (iii), with measurements of the space between the vector particle and the inner leaflet of the vesicular membrane. N, nucleus; NPC, nuclear pore complexes viewed in a tangential section. (B), (i): vector particle within an endocytic vesicle in the vicinity of a nuclear pore; (ii), viral core seen in the process of traverse of the nuclear pore. doi:10.1371/journal.pone.0018205.g008 transduction efficiency of (i) the FX-HAdV5F35 complex, compared to FX-HAdV5wt complex (refer to Fig. 5, panels a and b), and (ii) of the FX-HAdV5F35 complex, compared to the HAdV5F35 vector alone (refer to Fig. 5, panels b). Discussion It was recently shown that HAdV5 interaction with human blood coagulation FX, which results in the formation of FX- HAdV5 complexes, is the major parameter responsible for the massive liver uptake of HAdV5 vector particles administered systemically. FX binds to the hexon capsomeres via its Gla domain, and subsequently interacts with cell surface-exposed HSPG molecules which are present in abundance at the surface of liver Kupffer cells [25–34]. This prompted several laboratories to engineer HAdV5-based vectors with hexon modifications designed to abolish the FX binding and reduce their hepatotropism (reviewed in [24]). However, the species B member HAdV35 PLoS ONE | www.plosone.org 11 May 2011 | Volume 6 | Issue 5 | e18205