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Article

Molecular Determinants and Specificity of mRNA with Alternatively-Spliced UPF1 Isoforms, Influenced by an Insertion in the ‘Regulatory Loop’

1
International Centre for Cancer Vaccine Science, University of Gdansk, ul. Kładki 24, 80-822 Gdansk, Poland
2
Inserm UMRS1131, Institut de Génétique Moléculaire, Université Paris 7, Hôpital St. Louis, F-75010 Paris, France
3
Department of Medical Biosciences, Umeå University, Building 6M, 90185 Umeå, Sweden
4
Regional Centre for Applied Molecular Oncology (RECAMO), Masaryk Memorial Cancer Institute, Zlutykopec 7, 65653 Brno, Czech Republic
5
Institute of Genetics and Cancer, University of Edinburgh, Edinburgh EH4 2XR, UK
*
Authors to whom correspondence should be addressed.
Academic Editors: Naoyuki Kataoka and Luc Rochette
Int. J. Mol. Sci. 2021, 22(23), 12744; https://doi.org/10.3390/ijms222312744
Received: 30 September 2021 / Revised: 19 November 2021 / Accepted: 22 November 2021 / Published: 25 November 2021
(This article belongs to the Special Issue Manipulating Immune Tolerance with Cellular Therapies)
The nonsense-mediated mRNA decay (NMD) pathway rapidly detects and degrades mRNA containing premature termination codons (PTCs). UP-frameshift 1 (UPF1), the master regulator of the NMD process, has two alternatively-spliced isoforms; one carries 353-GNEDLVIIWLR-363 insertion in the ‘regulatory loop (involved in mRNA binding)’. Such insertion can induce catalytic and/or ATPase activity, as determined experimentally; however, the kinetics and molecular level information are not fully understood. Herein, applying all-atom molecular dynamics, we probe the binding specificity of UPF1 with different GC- and AU-rich mRNA motifs and the influence of insertion to the viable control over UPF1 catalytic activity. Our results indicate two distinct conformations between 1B and RecA2 domains of UPF1: ‘open (isoform_2; without insertion)’ and ‘closed (isoform_1; with insertion)’. These structural movements correspond to an important stacking pattern in mRNA motifs, i.e., absence of stack formation in mRNA, with UPF1 isoform_2 results in the ‘open conformation’. Particularly, for UPF1 isoform_1, the increased distance between 1B and RecA2 domains has resulted in reducing the mRNA–UPF1 interactions. Lower fluctuating GC-rich mRNA motifs have better binding with UPF1, compared with AU-rich sequences. Except CCUGGGG, all other GC-rich motifs formed a 4-stack pattern with UPF1. High occupancy R363, D364, T627, and G862 residues were common binding GC-rich motifs, as were R363, N535, and T627 for the AU-rich motifs. The GC-rich motifs behave distinctly when bound to either of the isoforms; lower stability was observed with UPF1 isoform_2. The cancer-associated UPF1 variants (P533L/T and A839T) resulted in decreased protein–mRNA binding efficiency. Lack of mRNA stacking poses in the UPF1P533T system significantly decreased UPF1-mRNA binding efficiency and increased distance between 1B-RecA2. These novel findings can serve to further inform NMD-associated mechanistic and kinetic studies. View Full-Text
Keywords: UPF1; GC-rich; AU-rich; isoform; regulatory loop; mRNA; PTC; molecular dynamics; degradation; stability; NMD; alternatively spliced; motifs UPF1; GC-rich; AU-rich; isoform; regulatory loop; mRNA; PTC; molecular dynamics; degradation; stability; NMD; alternatively spliced; motifs
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MDPI and ACS Style

Padariya, M.; Fahraeus, R.; Hupp, T.; Kalathiya, U. Molecular Determinants and Specificity of mRNA with Alternatively-Spliced UPF1 Isoforms, Influenced by an Insertion in the ‘Regulatory Loop’. Int. J. Mol. Sci. 2021, 22, 12744. https://doi.org/10.3390/ijms222312744

AMA Style

Padariya M, Fahraeus R, Hupp T, Kalathiya U. Molecular Determinants and Specificity of mRNA with Alternatively-Spliced UPF1 Isoforms, Influenced by an Insertion in the ‘Regulatory Loop’. International Journal of Molecular Sciences. 2021; 22(23):12744. https://doi.org/10.3390/ijms222312744

Chicago/Turabian Style

Padariya, Monikaben, Robin Fahraeus, Ted Hupp, and Umesh Kalathiya. 2021. "Molecular Determinants and Specificity of mRNA with Alternatively-Spliced UPF1 Isoforms, Influenced by an Insertion in the ‘Regulatory Loop’" International Journal of Molecular Sciences 22, no. 23: 12744. https://doi.org/10.3390/ijms222312744

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