Ugrás a tartalomhoz

Molecular diagnostics

Dr. István Balogh, Dr. János Kappelmayer, Dr. József Tőzsér (2011)

University of Debrecen

Chapter 4. 4. Special mutation consequences

Chapter 4. 4. Special mutation consequences

Table of Contents

Three codons of the 64 in the genetic code result in stop signal. These are the TAA, TAG and TGA codons. Therefore, terminating mutations are not rare, in fact, they account for 1/6th of all point mutations. Truncated proteins pose a significant danger to the cell. Their potential deleterious effect does not consist only in the fact that the resources are used for the generation of a useless end-product,but they are also prone to form aggregates in some cases, which might be poisonous for the cell. The cell has different protection mechanisms for the elimination of the potentially harmful truncated proteins. One of these mechanisms is nonsense-mediated mRNA decay (NMD), which affects the mRNA, in order to avoid the construction of the truncated protein. The mature mRNA has special protein complexes at the exact sites of exon junctions, called exon junction complexes (EJCs). As the ribosome moves in the mRNA, the EJCs are dislocated from the mRNA. However, in the case of an early (premature) stop codon, not all EJCs will be dislocated during the completion of the pioneer round of translation and the EJC that remained bound to the mRNA will trigger the NMD mechanism resulting in the degradation of the mRNA molecule (Figure 4.1.). NMD is not perfectly efficient, if the stop codon is in the last exon or too close to the EJC.

Figure 4.1. Figure 4.1. Nonsense-mediated mRNA decay (NMD)

Figure 4.1. Nonsense-mediated mRNA decay (NMD)

As has been shown above, a detected molecular alteration can be tested to prove its harmful effect on different levels. If the mutation has been described in the literature as pathogenic alteration, this information is usually accepted without further investigation. The situation is different, however, in the case of novel, potentially pathogenic mutations. If a point mutation introduces a premature stop codon, or causes frameshift, no further experimental work is needed. The case of the most frequent mutations, the missense mutations, is far more complicated. In addition to the experimental systems mentioned above, the physico-chemical consequence of the mutation (amino acid polariry, side chain composition, size) can be tested. Cis segregation of the detected mutation with the disease through generations is also an important indirect proof. Therefore, in order for a mutation to be qualified as pathogenic, it must cause some fundamental change in the structure and/or amount of the protein. This dogma, however, has been questioned recently.

Silent or synonymous mutation is a genetic alteration which does not affect the amino acid sequence of the protein. In the case of such a mutation, no phenotypic consequence is expected. In some cases, however, when a mutation affects a functionally important element, splicing defects might occur. In Figure 4.2, another mechanism is shown, where silent mutation might lead to gain-of-function of a protein. In this case, the kinetics of the translation will be impaired, which results in the changes of the conformation of the newly expressed protein. A c.3435C>T mutation has been described in the 26th exon of the MDR1 gene coding P-glycoprotein (Figure 4.2.). The originally coded isoleucine at the amino acid position 1145 will not be replaced by any other amino acid but the activity profile of the mutant P-glycoprotein will be somewhat different compared to the wild type. This difference might be attributed to the different tRNA that is needed for the mutant codon. The new tRNA can be considered to be a rare codon, which is why its availability is limited during the translation. The limitation will lead to different translation kinetics. The example above further complicates the problematics of potentially pathogenic mutations involving a single nucleotide, as the harmless nature of a synonymous mutation had been rendered unquestionable before the this description was formulated. However, there is no information as yet about whether such an effect is responsible for disease in any other gene.

Figure 4.2. Figure 4.2. Pathogenic silent mutation

Figure 4.2. Pathogenic silent mutation

As it has been shown, the genetic code consists of three letters and there are four possibilities within the codon, which allows for 64 possible combinations altogether. This number is more than enough to encode 20 amino acids. The third nucleotide of the codon often wobbles, which means that irrespective of the third nucleotide, the first two are responsible for the encoded amino acid. In Figure 4.3., the red numbers in the picture show the frequency of the given codon. This depends primarily on the intracellular availability of the given tRNA. The use of rare codons might affect the efficiency of the translation, which is a phenomenon that has been described in prokaryotic organisms. As regards humans, such a mutation has been shown to be present in the MDR1 gene (see above). In this case, the synonymous mutation causes change in the translation kinetics resulting in gain-of-function of the expressed P-glycoprotein.

Figure 4.3. Figure 4.3. The use of the codons in humans

Figure 4.3. The use of the codons in humans