Figure 3

Western blotting analysis of vector transduced NSCs without (-) and with (+) 4070A infection. A. Equivalent whole cell extracts of control C17.2 NSCs and those expressing CasE, CasES or hrGFP, with and without 4070A infection, were assessed for CasBrE Env expression (αCasBrE Env; top) and Gag (αGag; bottom) after separation on 9% SDS-PAGE gels. CasES-NSCs produce a 60 kD Env isoform in contrast to the gp70 and precursor (pr85) isoforms made in CasE-NSCs [11]. The 60 kD isoform likely represents minimally glycosylated Env protein [40]. Note that CasBrE Env levels were elevated with 4070A infection, and p30gag levels are elevated in 4070A infected NSCs possessing CasBrE Env vectors. B. Examination of CasBrE Env released into the culture medium ± 4070A infection. Note that CasBrE gp70 release is reduced in the presence of 4070A infection for CasE-NSCs but not CasES NSCs. C. NSC culture supernatants were subjected to ultracentrifugation and virion pellets were assessed for CasBrE Env (top), 4070A Env (middle) and Gag (bottom). All blots were run in triplicate using equivalent protein loads that were confirmed by coomassie blue staining of equivalent gels run in parallel. Representative examples are shown.