GeneticsTier 1Medical Sciences concept

Splice site mutations - effect on protein length/frame

Core concept

Splice sites (donor -- GT at intron start; acceptor -- AG at intron end) are recognised by the spliceosome to remove introns and join exons -- mutations here disrupt normal splicing
Exon skipping: splice site mutation causes the adjacent exon to be left out entirely -- if the skipped exon length is not a multiple of 3, this shifts the reading frame downstream -> frameshift, usually a premature stop codon
Intron retention: splicing fails and an intron is retained in the mature mRNA -- introduces intronic sequence (usually containing an in-frame stop codon) -> premature truncation

Calculating the effect: if the affected exon's length is a multiple of 3, skipping it removes a whole number of codons -- an in-frame deletion (protein shorter but reading frame preserved downstream) -- generally a milder phenotype
If the exon length is not a multiple of 3, skipping shifts the frame for every downstream codon -> frameshift and usually premature termination -- generally a more severe phenotype (analogous to a frameshift indel)
This exact calculation (exon length mod 3) is the type of question she will be asked to work through given a stated exon size, not simply asked to define "splice site mutation"

Explains phenotype variability seen in conditions like Duchenne vs Becker muscular dystrophy: DMD (out-of-frame deletion, no functional protein, severe) vs BMD (in-frame deletion, partially functional truncated dystrophin, milder) -- the same underlying gene, frame effect determines severity
Splice site variants are classified as likely pathogenic with high confidence when they disrupt the canonical GT/AG dinucleotide, even without direct RNA/protein evidence, because the mechanism is well established
Therapeutic exon-skipping drugs (e.g. for DMD) deliberately induce additional exon skipping to restore the reading frame, converting an out-of-frame mutation into an in-frame (and thus milder) one -- exploits this exact frame-mod-3 principle therapeutically

Directly extends assessing variant pathogenicity - splice site disruption is one of the strongest single pieces of pathogenicity evidence given its predictable mechanism
Same frame-shift-vs-in-frame logic applies to insertion/deletion (indel) variants generally, not just splice-site-driven exon skipping
Underlies the mechanism behind exon-skipping antisense oligonucleotide therapy, a specific gene-therapy-adjacent strategy distinct from gene addition or editing

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