CRISPR-Based Screen Links an Inhibitor of Nonsense-Mediated Decay to eIF4A3 Target Engagement

Eukaryotic
initiation factor (eIF) 4A3 is a DEAD-box RNA helicase
and a core component of the exon-junction complex (EJC). The EJC marks
the location of exon:exon junctions following the removal of introns
by splicing and plays a critical role in an mRNA surveillance program
known as nonsense-mediated decay (NMD). NMD is often triggered by
the presence of a premature termination codon (PTC) upstream of the
EJC, leading to degradation of the variant mRNA which prevents synthesis
of a potentially harmful, truncated polypeptide. One approach by which
to treat rare diseases where the underlying cause is a PTC is thus
to prevent NMD, while stimulating readthrough of the PTC. Hence, there
is much interest in inhibiting NMD, and recently a set of small molecules,
1,4-diacylpiperazine derivatives, targeting eIF4A3 has been developed
and shown to harbor such activity. Herein, we undertake a CRISPR/Cas9-based
variomics screen to identify eIF4A3 alleles resistant to said compounds.
Our results provide genetic evidence linking compound bioactivity
to eIF4A3 engagement.