Synonymous Variants of Uncertain Silence

Abstract

Synonymous variants, traditionally regarded as silent mutations due to their lack of impact on protein sequence, structure and function, have been the subject of increasing scrutiny. This commentary explores the emerging evidence challenging the notion of synonymous variants as functionally inert. Analysis of the activity of 70 synonymous variants in the HIV Tat transcription factor revealed that 50% of the variants exhibited significant deviations from wild-type activity. Our analysis supports previous work and raises important questions about the broader impact of non-silent synonymous variants in human genes. Considering the potential functional implications, the authors propose classifying such variants as "synonymous variants of uncertain silence" (sVUS), highlighting the need for cautious interpretation and further investigations in clinical and genetic testing settings.

Keywords: sequence; silent variant; synonymous variant; variant interpretation.

Figure 1

Map of synonymous variants in Tat with their effect on transcriptional function. Each reference amino acid in Tat has from 0 to 5 synonymous substitutions in each codon due to redundancy in the genetic code. hEK-293T and Jurkat cells were analyzed, and a maximum of three different codons were randomly sampled for each position. The map shows the effect of the synonymous variants (codons 1-3) on Tat transcriptional activity at each position for each cell line. Key: loss of function = red; gain of function = green; wild-type activity = grey; no data = white; AA = amino acid, SS = secondary structure, PTM = post-translational modification. The map does not include 10 variants that produced different results among technical replicates or cell lines and are considered indeterminate (also colored white). Minimum, maximum, and average transcriptional activities for the 19 amino acid substitutions at each position are shown as a gradient heatmap, with red indicating WT activity and yellow indicating LOF activity. Matthew’s correlation coefficient (MCC) was used to indicate specificity of each amino acid position for physiochemical characteristics. A gradient shows scores ranging from −1 (blue) to 0 (white) to 1 (magenta). White indicates no specificity, magenta indicates high specificity for the physiochemical group, and blue indicates a high specificity for negative preference against the physiochemical group. Several protein-protein interactions are shown with separate coloring as previously defined [18,19].