Secondary structure of the SARS-CoV-2 genome is predictive of nucleotide substitution frequency

Accurate estimation of the effects of mutations on SARS-CoV-2 viral fitness can inform public-health responses such as vaccine development and predicting the impact of a new variant; it can also illuminate biological mechanisms including those underlying the emergence of variants of concern¹. Recently, Lan et al reported a high-quality model of SARS-CoV-2 secondary structure and its underlying dimethyl sulfate (DMS) reactivity data². I investigated whether secondary structure can explain some variability in the frequency of observing different nucleotide substitutions across millions of patient sequences in the SARS-CoV-2 phylogenetic tree³. Nucleotide basepairing was compared to the estimated “mutational fitness” of substitutions, a measurement of the difference between a substitution’s observed and expected frequency that is correlated with other estimates of viral fitness⁴. This comparison revealed that secondary structure is often predictive of substitution frequency, with significant decreases in substitution frequencies at basepaired positions. Focusing on the mutational fitness of C→T, the most common type of substitution, I describe C→T substitutions at basepaired positions that characterize major SARS-CoV-2 variants; such mutations may have a greater impact on fitness than appreciated when considering substitution frequency alone.