Coordination of -1 Programmed Ribosomal Frameshifting by Transcript and Nascent Chain Features Revealed by Deep Mutational Scanning

Carmody, P. J.; Zimmer, M. H.; Kuntz, C. P.; Harrington, H. R.; Duckworth, K. E.; Penn, W. D.; Mukhopahyay, S.; Miller, T. F.; Schlebach, J. P.

doi:10.1101/2021.03.11.435011

bioRxiv · 10.1101/2021.03.11.435011

Coordination of -1 Programmed Ribosomal Frameshifting by Transcript and Nascent Chain Features Revealed by Deep Mutational Scanning

Carmody, P. J.·Zimmer, M. H.·Kuntz, C. P.·Harrington, H. R.·Duckworth, K. E.·Penn, W. D.·Mukhopahyay, S.·Miller, T. F.·Schlebach, J. P.

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Abstract

Programmed ribosomal frameshifting (PRF) is a translational recoding mechanism that enables the synthesis of multiple polypeptides from a single transcript. In the alphavirus structural polyprotein, -1PRF is coordinated by a "slippery" sequence in the transcript, an RNA stem-loop, and a conformational transition in the nascent polypeptide chain. To characterize each of these effectors, we measured the effects of 4,530 mutations on -1PRF by deep mutational scanning. While most mutations within the slip-site and stem-loop disrupt -1PRF, mutagenic effects upstream of the slip-site are far more variable. Molecular dynamics simulations of polyprotein biogenesis suggest many of these mutations alter stimulatory forces on the nascent chain through their effects on translocon-mediated cotranslational folding. Finally, we provide evidence suggesting the coupling between cotranslational folding and -1PRF depends on the translation kinetics upstream of the slip-site. These findings demonstrate how -1PRF is coordinated by features within both the transcript and nascent chain.

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Carmody, P. J., Zimmer, M. H., Kuntz, C. P., Harrington, H. R., Duckworth, K. E., Penn, W. D., Mukhopahyay, S., Miller, T. F., Schlebach, J. P.. 2021-03-11. Coordination of -1 Programmed Ribosomal Frameshifting by Transcript and Nascent Chain Features Revealed by Deep Mutational Scanning. https://doi.org/10.1101/2021.03.11.435011

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