Collapsing retroviruses for efficient delivery of viro-toxic cargoes

Abstract

Retroviruses are excellent tools for delivering and expressing transgenic cargoes with broad utility in research and therapy. However, many cargoes including virus restriction factors can dramatically limit virus production and/or infectivity. An extreme viro-toxic cargo is the DNA cytosine deaminase APOBEC3B, which potently restricts retrovirus infectivity by a direct cDNA deamination-dependent mechanism. To overcome viro-toxicity, an APOBEC3B minigene cargo is disrupted by a translation stop cassette flanked by a direct repeat of its own sequence. The integrity of the minigene is reconstituted naturally by retroviral recombination during transduction. Efficiency can be improved from 90% to nearly 100% by coupling the minigene to the translation of a downstream selectable marker. Collapsing retrovirus (CRV) technology enables the functional delivery of APOBEC3B to a target cell population and may have broad utility for delivering viro-toxic cargoes. MotivationMany different proteins are detrimental to retrovirus production and/or to virus-producing cells. This problem can be circumvented by interrupting the genetic cargo with a direct repeat-flanked translation stop cassette and allowing transgene integrity to be restored naturally by recombination during reverse transcription. As exemplified by the retrovirus restriction factor APOBEC3B, this process is very efficient. HighlightsO_LICollapsing retroviruses (CRVs) prevent retrovirus restriction by viro-toxic proteins C_LIO_LITruncated tandem repeats recombine during transduction and restore virus integrity C_LIO_LITranslation-coupled selection enables near 100% transduction efficiency C_LIO_LICRVs have broad utility for delivering viro-toxic cargoes such as APOBEC3B C_LI Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=186 SRC="FIGDIR/small/735095v1_ufig1.gif" ALT="Figure 1"> View larger version (33K): org.highwire.dtl.DTLVardef@e96092org.highwire.dtl.DTLVardef@1c343f9org.highwire.dtl.DTLVardef@a3b53dorg.highwire.dtl.DTLVardef@1cfeb69_HPS_FORMAT_FIGEXP M_FIG C_FIG