SARS-CoV-2 uses −1 programmed ribosomal frameshifting (−1 PRF) to control expression of key viral proteins. Because modulating −1 PRF can attenuate the virus, ligands binding to the RNA pseudoknot that stimulates −1 PRF may have therapeutic potential. Mutations in the pseudoknot have occurred during the pandemic, but how they affect −1 PRF efficiency and ligand activity is unknown. Studying a panel of six mutations in key regions of the pseudoknot, we found that most did not change −1 PRF levels, even when base-pairing was disrupted, but one led to a striking 3-fold decrease, suggesting SARS-CoV-2 may be less sensitive to −1 PRF modulation than expected. Examining the effects of a small-molecule −1 PRF inhibitor active against SARS-CoV-2, it had a similar effect on all mutants tested, regardless of basal −1 PRF efficiency, indicating that anti-frameshifting activity can be resistant to natural pseudoknot mutations. These results have important implications for therapeutic strategies targeting SARS-CoV-2 through modulation of −1 PRF.

SARS-CoV-2使用-1编程的核糖体移码（-1 PRF）来控制关键病毒蛋白的表达。因为调节-1 PRF可以减弱病毒，所以与刺激-1 PRF的RNA假结结合的配体可能具有治疗潜力。大流行期间发生了假结突变，但是它们如何影响-1 PRF效率和配体活性尚不清楚。通过研究假结关键区域的六个突变，我们发现即使碱基配对被破坏，大多数突变也不会改变-1 PRF水平，但是一个突变导致三倍的下降，这表明SARS-CoV-2对-1 PRF调制的敏感性可能比预期的要低。研究了对SARS-CoV-2有活性的小分子-1 PRF抑制剂的作用，无论基础-1 PRF效率如何，它对所有测试的突变体都有相似的作用，表明抗移码活性可以抵抗天然假结突变。这些结果对于通过调节-1 PRF靶向SARS-CoV-2的治疗策略具有重要意义。