A single nucleotide substitution (or point mutation) in a gene sequence, such as that observed in the example of achondroplasia just described, can alter the code in a triplet of bases and cause the nonsynonymous replacement of one amino acid by another in the gene product (see the genetic code in Table 3-1 and the example in Fig. 4-4). Such mutations are called missense mutations because they alter the coding (or “sense”) strand of the gene to specify a different amino acid. Although not all missense mutations lead to an observable change in the function of the protein, the resulting protein may fail to work properly, may be unstable and rapidly degraded, or may fail to localize in its proper intracellular position. In many disorders, such as β-thalassemia (Case 44), most of the mutations detected in different patients are missense mutations (see Chapter 11).
Point mutations in a DNA sequence that cause the replacement of the normal codon for an amino acid by one of the three termination (or “stop”) codons are called nonsense mutations. Because translation of messenger RNA (mRNA) ceases when a termination codon is reached (see Chapter 3), a mutation that converts a coding exon into a termination codon causes translation to stop partway through the coding sequence of the mRNA. The consequences of premature termination mutations are twofold. First, the mRNA carrying a premature mutation is often targeted for rapid degradation (through a cellular process known as nonsense-mediated mRNA decay), and no translation is possible. And second, even if the mRNA is stable enough to be translated, the truncated protein is usually so unstable that it is rapidly degraded within the cell (see Chapter 12 for examples).
Whereas some point mutations create a premature termination codon, others may destroy the normal termination codon and thus permit translation to continue until another termination codon in the mRNA is reached further downstream. Such a mutation will lead to an abnormal protein product with additional amino acids at its carboxyl terminus, and may also disrupt regulatory functions normally provided by the 3′ untranslated region downstream from the normal stop codon.
Mutations Affecting RNA Transcription, Processing, and Translation
The normal mechanism by which initial RNA transcripts are made and then converted into mature mRNAs (or final versions of noncoding RNAs) requires a series of modifications, including transcription factor binding, 5′ capping, polyadenylation, and splicing (see Chapter 3). All of these steps in RNA maturation depend on specific sequences within the RNA. In the case of splicing, two general classes of splicing mutations have been described. For introns to be excised from unprocessed RNA and the exons spliced together to form a mature RNA requires particular nucleotide sequences located at or near the exon-intron (5′ donor site) or the intron-exon (3′ acceptor site) junctions. Mutations that affect these required bases at either the splice donor or acceptor site interfere with (and in some cases abolish) normal RNA splicing at that site. A second class of splicing mutations involves base substitutions that do not affect the donor or acceptor site sequences themselves but instead create alternative donor or acceptor sites that compete with the normal sites during RNA processing. Thus at least a proportion of the mature mRNA or noncoding RNA in such cases may contain improperly spliced intron sequences. Examples of both types of mutation are presented in Chapter 11.
For protein-coding genes, even if the mRNA is made and is stable, point mutations in the 5′ and 3′-untranslated regions can also contribute to disease by changing mRNA stability or translation efficiency, thereby reducing the amount of protein product that is made.