Cells have developed different monitoring mechanisms to target messenger RNA (mRNA) with mutations that otherwise would cause errors in protein synthesis and furthermore eradicate other incorrectly processed cellular RNAs. These processes function in both cell nucleus and cytoplasm. One of the best-observed RNA monitoring pathways is the Nonsense-Mediated mRNA Decay (NMD) pathway. (Mechanism and regulation…). This is a crucial RNA quality control mechanism which is conserved in all Eukaryotes. It retains cellular homeostasis through the elimination of aberrant transcripts harbouring premature termination codon (PTC). This process works in the cytoplasm and is closely related to translation termination. PTC can arise from errors in the metabolism of nucleic acids such as defects in splicing or genetic mutations. (Control of gene expression…). Recent evidence suggests that this pathway is not specifically committed to the degradation of transcripts having PTC but it also plays a vital role in regulating the expression of naturally occurring transcripts. (Mechanism and regulation…).
How are transcripts recognized as targets for NMD?
There are two chief models:-
The first model for NMD posits that in mammals, NMD is closely linked to pre mRNA splicing and mRNA having a PTC 50 to 55 nucleotides upstream of the final exon-exon junction are effectively degraded through NMD pathway. This is indicated by the appearance of a multi-subunit protein complex, exon-junction complex (EJC), which is placed 20 to 24 nucleotides upstream of an exon-exon junction throughout pre-mRNA splicing. (Mechanism and regulation…).
The second model postulates that the unusual long 3′ untranslated region (UTR) that is downstream of a PTC works as a signal to promote PTC identification. (Control of gene expression…). NMD can recognize PTC because they lead to inefficient termination of translation with resultant ribosome stalling. The reason behind this imperfectly dissociable termination complex appears to be the lack of termination stimulating signal that needs the intercommunication between the poly (A) tail binding protein and ribosome-bound eukaryotic release factor 3 (eRF3). Persistent with ‘faux 3′ UTR’ model, the physical distance between the poly (A) tail and the terminating ribosome is identified as a significant determinant for PTC recognition. (SMG6 promotes endonucleolytic…). Deposition UPF1 and other regulatory elements in the UTR’ region mediated the assembly of the other elements of NMD mechanism.
NMD PATHWAY MECHANISM:-
The first model is the classical NMD pathway and most highly observed. The NMD process initiates when NMD identify the transcript target as mentioned above. The process begins with UPF1 phosphorylation which is carried out by SMG1c complex, which is composed of the SMG1, a protein kinase, phosphoinositide 3-kinase (PI3K) and two subunits SMG8 and SMG9. In the beginning, UPF1 links with SMG1 and works as a clamp, communicating directly with the eRF1 and eRF3 for the formation of so-called surveillance complex (SURF) in the region of the PTC. SMG1 is associated tightly with two subunits of SMG1c complex, SMG8 and the NTPase SMG9, which regulate its activity by induction of conformational changes, with SMG8 which binds to preformed SMG1-SMG9 complex and maintain an inactive state of the kinase (blocking phosphorylation by SMG1 of UPF1). Two RNA helicases RUVBL1 and RUVBL2, due to their close association to the SMG1 kinase, contribute to the SMG1-8-9 complex regulation by promoting the SMG1 kinase molecule activation. Eventually, SURF complex interacts with UPF2, UPF3b and an EJC downstream of the PTC for the formation of the decay-inducing complex (DECID) which triggers UPF1 phosphorylation and leads to dissociation of eRF1 and eRF3. (Mechanism and regulation). RUVBL1 and RUVBL2 also interact with SURF complex and assist in the transition to the DECID complex by hydrolysis of ATP to ADP. DHX34, a DEHX/D box protein is found to associate with SURF complex and assist the transition to the DECID complex. DHX34 probably impacts the remodelling of the SURF complex by assisting the dissociation of eRF3 in an ATP dependent manner from SURF complex in the vicinity of the PTC. As a result of the remodelling of the NMD complexes, active helicase confirmation is adapted by UPF1 because of the reorganization of inhibitory domains by association with UPF2. The active UPF1 helicase works as RNPase, translocates along the side mRNA, resolves secondary structure and clears the mRNA from proteins and allowing access to nucleases. The activated NMD complex consists of UPF1, UPF2 and UPF3b translocate from its position upstream of the EJC towards the 3′ of the EJC. Eventually, phosphorylated UPF1 interacts with the phospho-binding proteins named SMG5, SMG6 and SMG7 and mRNA degradation factors and additional reorganization of this complex leads to degradation of mRNA. In the vicinity of the PTC, SMG6 cleaves the NMD targets which lead to the initiation of NMD-mediated degradation of RNA. (Mechanism and regulation…). The phosphorylated UPF1, SMG5 and SMG7 bind as a heterodimer. POP2, the catalytic subunit of the deadenylase complex is directly recruited by SMG7. Furthermore, SMG7 initiates decapping and also XRN1-catalyzed 5′-3′ degradation. (Mechanism and regulation…). SMG6 leads to the generation of unstable decay intermediate that does not have either 5’cap or 3′ poly (A) and thus is prone to cellular exonucleases. This leads to immediate exonucleolytic deterioration of the cleavage products. Finally, SMG5 and SMG6 furthermore add protein phosphatase 2A (PP2A) for the dephosphorylation of UPF1, thus recycling it for future rounds of NMD. (Leveraging the rules…).
Thus, NMD pathway is one of the most vital pathways which if not operates properly would lead to the generation of truncated proteins with anticipated detrimental impact on the organism. NMD and its regulation greatly impact the human disease development. Genetic mutations in factors of NMD leads to downregulation or complete absence of NMD activity. Some neurodevelopment disorders are intimately linked NMD dysregulation. Mutations in one of the NMD factor UPF3b has been associated to cause syndromic and non-syndromic Intellectual Disability (ID). UPF3b mutations are further associated with a range of disorders including autism, attention-deficit hyperactivity disorder and schizophrenia.
Mutations in other NMD factors such as UPF2 and SMG6 are linked to diverse neurodevelopmental disorders. SMG6 is found to have been associated with CAD. (Large-scale association analysis identifies new risk for CAD…) Abnormal NMD moreover is linked to cancer and inflammation. (Control of gene expression…).