High-throughput interrogation of programmed ribosomal frameshifting in human cells

Martin Mikl; Yitzhak Pilpel; Eran Segal

By controlled sequence and structure manipulations in multiple contexts we aimed to dissect and 541 directly compare the regulatory architecture of 15 frameshifting events, revealing a great diversity in 542 regulatory strategies involving upstream and downstream sequence and structural elements and 543 amino acid properties. We demonstrate a high degree of optimization and specialization of regulatory 544 mechanisms. For example, no universally optimal slippery site exists, but in each of the PRF events 545 tested the native slippery site was found to be optimal given the sequence context. In addition, our analyses highlighted amino acid properties around the frameshifting sites as another 570 factor influencing PRF efficiency. Specifically, a charged amino acid at the last position before the 571 slippery site was associated with reduced frameshifting rate. Together with reports of a frameshift-572 promoting effect of specific amino-acid sequences upstream of the frameshifting site in the case of 573 +1 PRF in fungal OAZ genes (Yordanova et al., 2015) and the copA gene of Escherichia coli (Meydan 574 et al., 2017) , this observation provides additional evidence for a role of the nascent peptide in 575 regulating ribosomal frameshifting. 576