Fig. 2

Mechanisms for the production of residual proteins in knockout cells. A CRISPR–Cas introduces indels and PTC in early exon. N-terminal truncated protein could be produced by translation reinitiation using another AUG downstream PTC. B cis-elements on pre-mRNA regulates splicing. Colored cylinders represent exons, and gray lines represent introns. GU 5′ splice site, A the branch site, ISE intronic splicing enhancer, PPT the polypyrimidine tract, ISS intronic splicing silencer, AG 3′ splice site, ESE exonic splicing enhancer, ESS exonic splicing silencer. C The indels induce alternative splicing by disrupting the original splicing motif and/or activating cryptic splicing motif. Green lines indicate the joining sites during the alternative splicing. New transcript may restore the reading frame and produce internal deleted/inserted protein. For simplicity reasons, only the case of indels and PTC in the same exon is shown here. D The putative ORF-dependent NAS model proposes an unidentified macromolecular machinery within the nucleus examining the pre-mRNA and inducing alternative splicing in the presence of PTC