The solution structure of Dead End bound to AU-rich RNA reveals an unprecedented mode of tandem RRM-RNA recognition required for mRNA regulation

Abstract

A previous version of this manuscript contained an error in the normalization of the luciferase activity in the cell-based assays in Figures 5 and S8. This has been corrected and the text has been updated accordingly. AbstractDead End (DND1) is an RNA-binding protein essential for germline development through its role in posttranscriptional gene regulation. The molecular mechanisms behind selection and regulation of its targets are unknown. Here, we present the solution structure of DND1's tandem RNA Recognition Motifs (RRMs) bound to AU-rich RNA. The structure reveals how an NYAYUNN element is specifically recognized, reconciling seemingly contradictory sequence motifs discovered in recent genome-wide studies. RRM1 acts as a main binding platform, including unusual extensions to the canonical RRM fold.RRM2 acts cooperatively with RRM1, capping the RNA using an unusual binding pocket, leading to an unprecedented mode of tandem RRM-RNA recognition. We show that the consensus motif is sufficient to mediate upregulation of a reporter gene in human cells and that this process depends not only on RNA binding by the RRMs, but also on DND1's double-stranded RNA binding domain (dsRBD), that we show to be dispensable for target binding in cellulo. Our results point to a model where DND1 target selection is mediated by an uncanonical mode of AU-rich RNA recognition by the tandem RRMs and a role for the dsRBD in the recruitment of effector complexes responsible for target regulation.

Dead-End (dnd) Gene Cloning and Gonad-Specific Expression Pattern in Starry Flounder (Platichthys stellatus)

“…The second RRM motif, RRM2, plays a role in promoting nanos1 translation and germline survival in xenopus [ 26 ]. DSRM is not essential for RNA binding, although it increases the target specificity of the RRM regions in DND1 in human [ 27 ]. Zebrafish dnd has a binding site to bind miR-430 for preventing degradation from miRNA [ 8 ].…”

The structure of yeast Npl3 bound to RNA reveals a cooperative sequence-specific recognition and an RNA chaperone role in splicing

“…To keep this unusual orientation, it was hypothesized that a long inter-domain linker (15 aa) is required to allow the two RRMs to lie side-by-side (β2-β4 type) and form an extended β-sheet RNA binding surface (Afroz et al, 2015). Recently, two structures (Dnd1 (Duszczyk, Wischnewski et al, 2021) and Npl3 in this work) revealed new ways for tandem RRMs to bind RNA cooperatively with RRM1 binding to the 5'end despite having a short interdomain linker (5 and 7 aa, respectively). In such cases, the canonical β-sheet surface of the RRM is not used but rather both sides of the domain.…”