The many paths to frameshifting: kinetic modelling and analysis of the effects of different elongation steps on programmed –1 ribosomal frameshifting

Liao, Pei-Yu; Choi, Yong Seok; Dinman, Jonathan D.; Lee, Kelvin H.

The kinetic model suggests that incomplete translocation of the pre-translocational ribosome aligning with deX XXY produces FS m . Previous studies suggested that FS m may result from slippage of a single P-site tRNA (1, 25, 26) . However, it is not clear when and/or how this could occur. In addition, the single slippage model does not explain experimental evidence regarding the influence of translocation on À1 PRF (22, 23) . Our model suggests that in both mechanisms, incomplete translocation and slippage of P-and A-site tRNAs participate in synthesizing frameshift proteins to varying extents for different À1 PRF signals. Frameshifting at the HIV-1 sequence was reported to generate $70% FS z and 30% FS m (1, 25) , indicating that Pathways II and/or III exert stronger influence on FS% than Pathway I. Notably, our protein analysis showed $80% FS z and 20% FS m for the frameshifting signal in HIV-1 group M subtype B. This small discrepancy may due to the use of different reporter systems, or differences in the quantitative methods employed for the assay. For the HTLV-1 pro-pol frameshift sequence, this study observed 39.4% FS m in total frameshift protein. This is the first study to demonstrate another frameshift sequence that generates a significant amount of FS m in addition to HIV-1. Interestingly, a small but observable lysine peak appeared at the corresponding position (-1 frame A-site codon in the recoding site) when HTLV-1 pro-pol frameshift proteins were sequenced in the previous study (31) , supporting the production of FS m . The stimulatory RNA of HTLV-1 pro-pol has been suggested to be a pseudoknot (31) , although no direct evidence for the RNA structure was shown in the same study. Interestingly, the mouse Edr frameshift sequence, which shares high similarity with the HTLV-1 pro-pol sequence, was suggested to involve a pseudoknot (38) . Because the length of HTLV-1 pro-pol stimulatory signals is not well-defined, the frameshift sequence incorporated in the TLV strain may not represent the whole stimulatory signal. However, the possible absence of a portion of the stimulatory signal did not prevent us from observing a significant amount of frameshift efficiency and the ratio of FS m to the total frameshift protein. Frameshift products from other À1 PRF signals were analysed previously (39, 40) . For SARS-CoV frameshifting, FS m was not found (39) . However, for the Alphavirus coding sequence 6k, both FS z and FS m were identified in the frameshift products, although the exact ratio was not determined (40) .