**1.2. PTCs and NMD mechanism**

Premature termination codons (PTCs) lead to premature translational termination resulting in absent functional proteins. Different inherited and acquired diseases can be attributable to PTCs, such as cystic fibrosis, Duchenne muscular dystrophy (DMD), mucopolysaccharidosis, spinal muscular atrophy (SMA) type I and type II, X-linked nephrogenic diabetes insipidus, rhinitis pigmentosa X-linked, nephropathic cystinosis, and β-thalassemia [5].

PTCs are responsible for about 10% of CF cases worldwide, but are particularly common in Ashkenazi Jews (40% of Israeli CF patients) [6]. The translation of PCT-containing mRNAs leads to the formation of truncated CFTR, in most cases lacking in its function [7].

During molecular evolution, a mechanism to eliminate these aberrant mRNAs containing PTCs has been developed, known as nonsense-mediated mRNA decay (NMD). This process of quality control is observed in all eukaryotic organisms and has an important role in the regulation of many cellular functions. Its physiological substrates act in a wide range of processes such as transcription, DNA repair, cell growth, intracellular transport, and NMD itself. Through this process, the mRNA levels containing PTCs are reduced but not completely eliminated, inducing a reduced level of proteins, which are synthesized in a truncated form [5].

In mammals the NMD process operates by the recognition of a premature stop codon. It is observed that the ability of PTCs in inducing NMD depends on its location with respect to the sequence downstream and associated proteins [8]. The NMD process often occurs in mam‐ malian cells after the splicing of pre-mRNA and, in most cases, is controlled by the exon junction complex (EJC). This complex consists of at least 10 proteins binding to regions consisting of 20–24 nucleotides upstream of the exon-exon junction and including UPF2 and UPF3 proteins.

According to the currently proposed models of the NMD mechanism, for the majority of mRNAs, the stop codons that are localized more than 50–55 nucleotides upstream of an exonexon junction are recognized as premature, since the EJC is found downstream to the stop codon. The EJC is recognized by UPF2 and UPF3 and ribosomes start to translate the mRNA until reaching the PTC. The termination of the translation is triggered by the recognition of PTCs by eRF1 and eRF3, release factors recruited by ribosomes. These proteins then recruit the NMD key factor UPF1, which in turn binds to the SMG1 kinase. These four proteins (eRF1, eRF3, UPF1, SMG1) constitute the SURF complex, responsible for ribosome blocking. When the PTC is upstream of this EJC, UPF1 at the termination site may interact with UPF2 (associ‐ ated with EJC) causing the phosphorylation of UPF1 by SMG1 and the dissociation of release factors (eRF1 and eRF3). The phosphorylated UPF1 recruits additional factors (SMG5, SMG6, and SMG7) triggering NMD. Once the NMD is stimulated, target transcripts are degraded by 5' and 3' exonucleases [5].
