Suppression of Nonsense Mutations by New Emerging Technologies

Abstract

Nonsense mutations often result from single nucleotide substitutions that change a sense codon (coding for an amino acid) to a nonsense or premature termination codon (PTC) within the coding region of a gene. The impact of nonsense mutations is two-fold: (1) the PTC-containing mRNA is degraded by a surveillance pathway called nonsense-mediated mRNA decay (NMD) and (2) protein translation stops prematurely at the PTC codon, and thus no functional full-length protein is produced. As such, nonsense mutations result in a large number of human diseases. Nonsense suppression is a strategy that aims to correct the defects of hundreds of genetic disorders and reverse disease phenotypes and conditions. While most clinical trials have been performed with small molecules, there is an increasing need for sequence-specific repair approaches that are safer and adaptable to personalized medicine. Here, we discuss recent advances in both conventional strategies as well as new technologies. Several of these will soon be tested in clinical trials as nonsense therapies, even if they still have some limitations and challenges to overcome.

Keywords: ADAR-catalyzed editing; CRISPR technology; NMD; antisense oligo; box H/ACA RNA-guided modification; small molecules; stop codon read-through; suppressor tRNAs.

Figure 1

Nonsense mutation and its consequences. A point mutation (indicated) occurs in a gene, creating a premature termination codon (PTC) at the site >50–55 nts upstream of the exon–intron junction (indicated). After transcription, the nonsense mutation is passed onto pre-mRNA. Upon splicing, an exon–exon junction complex (EJC) forms (indicated). When the ribosome encounters the PTC, the eRF1–eRF3 complex enters the ribosome and recruits a number of factors to form the SMG1/UPF1/eRF1/eRF3 (SURF) complex. Together, EJC–SURF combination is called the decay-inducing (DECID) complex. Consequently, nonsense-mediated mRNA decay (NMD) is activated, and the mRNA is degraded. A small fraction of mRNA that escapes degradation is translated, but translation terminates at the PTC, generating a truncated protein (indicated). The normal stop codon is also indicated, and if the PTC is absent, translation will terminate at the normal stop codon, generating a full-length protein (also shown). DNA, pre-mRNA, mRNA, the exons and intron, and the amino acid protein chains are indicated.

Figure 2

Nonsense suppression by various approaches. A number of approaches (indicated) are developed to suppress nonsense mutations. Some can suppress either NMD or translation termination, while others can suppress both (indicated).