Cancer-associated synonymous mutations reveal stress signal-dependent mRNA folding that selectively modulates protein function.

Recent technical advances have facilitated studies on changes in mRNA structures in response to signaling pathways. However, if mRNA structures can affect the function of the encoded protein remains poorly understood. In-cell RNA structural probing (SHAPE-MaP) demonstrates how two cancer-associated synonymous mutations (CASMs) at proline codon 34 (c.102 C>A and c.102 C>G) prevent DNA damage-induced TP53 mRNA folding, whereas the non-cancer-associated c.102C>U mutation does not. Transcript and chromatin immunoprecipitation (ChIP) analysis reveal that p53 expressed from CASM34 has reduced promoter binding and reduced induction of p53 downstream target genes PUMA and 14-3-3-{sigma}, but not p21CDKN1A. Transcriptome analysis reveals a CASM34-mediated global attenuation of DNA damage-responsive gene expression. Together, the results demonstrate that CASM34 interferes with signal-induced p53 mRNA folding during DNA damage, leading to selective modulation of protein activity. More broadly, our findings highlight a general concept by which cancer-associated synonymous mutations target signal-induced mRNA structures that influence the encoded protein. Significance statementRecent studies have revealed that synonymous mutations can target RNA metabolism and are associated with numerous diseases; however, how these mutations affect the function of the encoded protein remain unclear. Using in-cell RNA structural probing, we show that two cancer-associated synonymous mutations in TP53 disrupt DNA damage-induced folding of p53 mRNA. These mutations selectively impair p53 promoter binding and transcriptional activation of specific downstream target genes. Our results demonstrate that disruption of signal-induced mRNA folding by synonymous mutations can modulate the activity of the encoded protein. More broadly, this work identifies signal-responsive mRNA structures as functional targets of cancer-associated synonymous mutations.