Specificity of mRNA binding towards proteins within NMD machinery influenced in cancer

Monikaben Padariya^*

• University of Gdansk, Gdansk, Poland

The nonsense-mediated mRNA decay (NMD) process is recognized as the quality control of mRNAs, to maintain its integrity and production of functional proteins. Readthrough of aberrant mRNA containing premature termination codons (PTCs) can induce the production of truncated proteins with negative functionalities. To elucidate the structural and mechanistic basis of NMD components, we performed molecular dynamic simulations (MDS) to analyze their dynamic behavior across different stages of the process. We further investigated how cancer-associated mutations alter mRNA-binding protein (RBP) interactions within the NMD machinery. Over simulation time, the mRNA containing PTC underwent significant conformational rearrangements, ultimately forming stable interactions with the eukaryotic class-I release factor (eRF1). The efficiency of eRF1 for recognizing stop codons (UAG, UGA, or UAA) nitrogenous bases were identified, revealing a stronger preference towards UAA. Due to lower structural stability the AU-rich mRNA motifs showed a diminished eRF1 binding affinity, relative to other PTC containing transcripts. Among studied cancer variants, the D9Y, R10S, F56V, P89L, and I62M residues were found to either enhance or disrupt eRF1-mRNA interactions. Similarly, when evaluating EIF4A3 RBP from the exon junction complex (EJC), the P114L and G309A mutations significantly impaired protein-mRNA binding affinity. Surface residue mapping of SMG1 kinase revealed that it engages in a sequential binding order with SMG8, SMG9, and UPF1, displaying the highest affinity for SMG8. Overall, these findings contribute to the mechanistic understanding of molecular properties for different RBP from the NMD process, which can be the basis of developing new therapeutic strategies against genetic disease or cancer.