Analysis of 4070A envelope levels in retroviral preparations and effect ontarget cell transduction efficiency

A number of stable producer cell lines for high-titer Mo-MuLV vectors have been constructed. Development has previously centered on increasing end-poi nt titers by producing maximal levels of Mo-MuLV Gag/Pol, envelope glycopro teins, and retroviral RNA genomes. We describe the production yields and tr ansduction efficiency characteristics of two Mo-MuLV packaging cell lines, FLYA13 and TEFLYA. Although they both produce 4070A-pseudotyped retroviral vectors reproducibly at >1 X 10(6) LFU ml(-1), the transduction efficiency of unconcentrated and concentrated virus from FLYA13 lines is poor compared with vector preparations from TEFLYA lines. A powerful inhibitor of retrov iral transduction is secreted by FLYA13 packaging cells. We show that the i nhibitory factor does not affect transduction of target cells by RD114-pseu dotyped vectors. This suggests that the inhibitory factor functions at the level of envelope-receptor interactions. Phosphate starvation of target cel ls shows a two-fold increase in Pit2 receptor mRNA and causes some improvem ent in FLYA13 virus transduction efficiency. Western blots show that FLYA13 viral samples contain an eight-fold higher ratio of 4070A envelope to p30( gag) than that of virus produced by TEFLYA producer cell lines. This study correlates overexpression of 4070A envelope glycoprotein in retroviral prep arations with a reduction of transduction efficiency at high multiplicities of infection. We suggest that TEFLYA packaging cells express preferable le vels of 4070A compared with FLYA13, which not only enables high-titer stock s to be generated, but also facilitates a high efficiency of transduction o f target cells.