Switch from Cap- to Factorless IRES-Dependent 0 and +1 Frame Translation during Cellular Stress and Dicistrovirus Infection

Wang, Qing S.; Jan, Eric

To increase coding capacity, some RNA viruses utilize frameshifting or termination/reinitiation strategies to translate overlapping ORFs [47] . Frameshift events are tightly regulated under different cellular conditions. For example, the extent of programmed -1 frameshifting in human immunodeficiency virus (HIV), which is responsible for generating the precursor of Gag-Pol enzymes, is critical for viral assembly and maturation. Alterations in frameshift efficiency can inhibit viral replication [56, 57] . As a result, the frameshift event has become a strategic antiviral target [58] . Alternatively, it is known that the dynamic levels of polyamines can in part regulate a +1 frameshifting event of the antizyme ORF to control polyamine biosynthesis in cells [59] . The +1 frame protein ORFx of IAPV is detected in virusinfected honey bees suggesting that it has a function during virus infection [25] . However, the function of ORFx has been elusive. Exploring the regulation of +1/0 frame translation may provide clues as to when ORFx is needed during infection. Since translation of both 0 and +1 frames are dependent on the integrity of IAPV IRES, it was important to determine whether the alternate frames are translated either in a fixed ratio or altered during cellular stress or virus infection. In this study, a 20% ratio of +1/0 frame translation was observed using reporter RNA transfections in insect cells, which is similar to the ratio in vitro ( Figure 4D ) [25] . We also find that IAPV IGR IRES +1 frame translation is stimulated to varying extents during cellular stress, similar to that observed monitoring 0 frame translation ( Figure 5 ). Both 0 and +1 frame translation mediated by the IAPV IGR IRES was stimulated to the same extent during CrPV infection in S2 cells, suggesting that translation in both frames are regulated similarly by the IRES (Figure 7) . However, these experiments were performed using CrPV infection in S2 cells and it remains to be determined whether the ratio of +1/0 frame translation is regulated under conditions better representing physiological environments such as in IAPV-infected honey bee cells.