All-Atom Simulations Elucidate the Impact of U2AF2 Cancer-Associated Mutations on Pre-mRNA Recognition

Abstract

The U2AF2 splicing factor, made of two tandem RNA recognition motifs (RRMs) joined by a flexible linker, selects the intronic polypyrimidine sequence of premature mRNA, thus ensuring splicing fidelity. Increasing evidence links mutations of key splicing factors, including U2AF2, to a variety of cancers. Nevertheless, the impact of U2AF2 cancer-associated mutations on polypyrimidine recognition remains unclear. Here, we combined extensive (18 μs-long) all-atom molecular dynamics simulations and dynamical network theory analysis (NWA) of U2AF2, in its wild-type form and in the presence of the six most frequent cancer-associated mutations, bound to a poly-U strand. Our results reveal that the selected mutations affect the pre-mRNA binding at two hot spot regions, irrespectively of where these mutants are placed on the distinct U2AF2 domains. Complementarily, NWA traced the existence of cross-communication pathways, connecting each mutation site to these recognition hot spots, whose strength is altered by the mutations. Our outcomes suggest the existence of a structural/dynamical interplay of the two U2AF2’s RRMs underlying the recognition of the polypyrimidine tract and reveal that the cancer-associated mutations affect the polypyrimidine selection by altering the RRMs’ cooperativity. This mechanism may be shared by other RNA binding proteins hallmarked, like U2AF2, by multidomain architecture and high plasticity.

Conformational Heterogeneity of RNA Stem-Loop Hairpins Bound to FUS-RNA Recognition Motif with Disordered RGG Tail Revealed by Unbiased Molecular Dynamics Simulations

“…At the same time though, we wish to stress that the FUS-RNA system is challenging in many aspects and that the poor performance of the force field reported here cannot be simply generalized for a broad range of RNA–protein systems. To be more optimistic, for many less challenging RNA–protein complexes, the OL3+ff14SB­(ff12SB) combination with SPC/E or similar TIP3P water model is sufficiently accurate without any such modifications. ,,, In the Supporting Information, we provide a recommended best-practice protocol for simulating RNA–protein complexes based on our up-to-date experience.…”

“…To be more optimistic, for many less challenging RNA−protein complexes, the OL3+ff14SB-(ff12SB) combination with SPC/E or similar TIP3P water model is sufficiently accurate without any such modifications. 35,55,68,69 In the Supporting Information, we provide a recommended best-practice protocol for simulating RNA− protein complexes based on our up-to-date experience.…”

Monovalent Ionic Atmosphere Modulates the Selection of Suboptimal RNA Sequences by Splicing Factors’ RNA Recognition Motifs

“…Indeed, a high cation density exists between two loops at the RRM1−RRM2 interface close to the binding region of U/C6, a second cation density is placed at the RRM1 α-helix nearby the U8 backbone, and another is placed between the U8 aromatic ring and linker loop. Remarkably, two cancer-associated mutations (i.e., N196K and D231N) 14 affecting the overall charge of U2AF2 surface lie in the vicinity of these cation density hotspots. Interestingly, in the polyC system, these density peaks are complemented by two additional Na + density hotspots placed nearby C3 and C4 (Figure 2B).…”

“…In our calculations, we have not considered the entropic contribution of the free energy as it was reported that this term does not substantially improve the quality of the results . Moreover, a per-residue decomposition analysis was done following a procedure applied in previous publications . In the Results section, we only qualitatively discuss the results of the MM-PBSA calculation …”

Celebrating Diversity, Equity, Inclusion, and Respect in Computational and Theoretical Chemistry

“…Rozza et al. identity cross-communication pathways in mutational hotspots and a dynamical interplay between U2AF2 splicing factor recognition domains and premature mRNA . Finally, Bartocci et al simulate the dynamical “walk” of sulfonated calix-[4]-arenes on cytochrome c and find an allosteric reinforcement of residue interactions with the quasi-harmonic multibasin method …”