Minimal impact of ZAP on lentiviral vector production and transduction efficiency

• Published in: Molecular therapy. Methods & clinical development Vol. 23; pp. 147 - 157
• Main Authors: Sertkaya, Helin, Hidalgo, Laura, Ficarelli, Mattia, Kmiec, Dorota, Signell, Adrian W., Ali, Sadfer, Parker, Hannah, Wilson, Harry, Neil, Stuart J.D., Malim, Michael H., Vink, Conrad A., Swanson, Chad M.
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 10.12.2021; American Society of Gene & Cell Therapy; Elsevier
• Subjects: HIV-1; lentiviral vector; Original; retroviral vector; ZAP; ZC3HAV1; zinc-finger antiviral protein; HIV-1; ZAP; zinc-finger antiviral protein; ZC3HAV1; retroviral vector; lentiviral vector
• ISSN: 2329-0501; 2329-0501
• DOI: 10.1016/j.omtm.2021.08.008

The antiviral protein ZAP binds CpG dinucleotides in viral RNA to inhibit replication. This has likely led to the CpG suppression observed in many RNA viruses, including retroviruses. Sequences added to retroviral vector genomes, such as internal promoters, transgenes, or regulatory elements, substantially increase CpG abundance. Because these CpGs could allow retroviral vector RNA to be targeted by ZAP, we analyzed whether it restricts vector production, transduction efficiency, and transgene expression. Surprisingly, even though CpG-high HIV-1 was efficiently inhibited by ZAP in HEK293T cells, depleting ZAP did not substantially increase lentiviral vector titer using several packaging and genome plasmids. ZAP overexpression also did not inhibit lentiviral vector titer. In addition, decreasing CpG abundance in a lentiviral vector genome did not increase its titer, and a gammaretroviral vector derived from murine leukemia virus was not substantially restricted by ZAP. Overall, we show that the increased CpG abundance in retroviral vectors relative to the wild-type retroviruses they are derived from does not intrinsically sensitize them to ZAP. Further understanding of how ZAP specifically targets transcripts to inhibit their expression may allow the development of CpG sequence contexts that efficiently recruit or evade this antiviral system. [Display omitted] ZAP is an antiviral protein that targets CpG dinucleotides in viral RNA. Despite retroviral vectors containing a much higher CpG abundance than the viruses they are derived from, ZAP does not substantially restrict vector production or transduction efficiency. Therefore, ZAP does not appear to restrict the core retroviral vector platform.