An RRM–ZnF RNA recognition module targets RBM10 to exonic sequences to promote exon exclusion

Abstract

RBM10 is an RNA-binding protein that plays an essential role in development and is frequently mutated in the context of human disease. RBM10 recognizes a diverse set of RNA motifs in introns and exons and regulates alternative splicing. However, the molecular mechanisms underlying this seemingly relaxed sequence specificity are not understood and functional studies have focused on 3΄ intronic sites only. Here, we dissect the RNA code recognized by RBM10 and relate it to the splicing regulatory function of this protein. We show that a two-domain RRM1–ZnF unit recognizes a GGA-centered motif enriched in RBM10 exonic sites with high affinity and specificity and test that the interaction with these exonic sequences promotes exon skipping. Importantly, a second RRM domain (RRM2) of RBM10 recognizes a C-rich sequence, which explains its known interaction with the intronic 3΄ site of NUMB exon 9 contributing to regulation of the Notch pathway in cancer. Together, these findings explain RBM10's broad RNA specificity and suggest that RBM10 functions as a splicing regulator using two RNA-binding units with different specificities to promote exon skipping.

A Heterologous Cell Model for Studying the Role of T-Cell Intracellular Antigen 1 in Welander Distal Myopathy

“…These findings may be relevant for understanding why zinc improves TIA1 selfmultimerization. Zinc ions interact with proteins through electrostatic interactions in a coordination-dependent manner, known as ZFs (51)(52)(53). This structural element has been described in several transcription factors.…”

“…This structural element has been described in several transcription factors. The C 2 H 2 (Cys-Cys-His-His motif and variants thereof) zinc finger proteins are members of a superfamily of nucleic acidbinding proteins in eukaryotes (51)(52)(53). There are data revealing large diversities in both the structures of ZF domains and the mechanisms through which they can interact with molecular partners.…”

“…There are data revealing large diversities in both the structures of ZF domains and the mechanisms through which they can interact with molecular partners. Thus, there are both classical and nonclassical ZF domains that retain RNA-binding activity (51)(52)(53). Accordingly, it is possible that the positive charge of the lysine residue introduced by the WDM mutation in TIA1 somewhat reinforces the pool of positive charges of TIA1, improving and stabilizing the interaction between zinc, TIA1, and/or RNA-binding proteins and RNAs.…”

RBM10: Harmful or helpful‐many factors to consider

“…Furthermore, RBM10 NLSs can work cooperatively to influence nuclear localization of RBM10 . In terms of RNA binding, the RBM10 RanBP2‐type zinc finger domain, and both RRM domains can bind RNA; however, while the RBM10 RanBP2‐type zinc finger and RRM1 domains can act together to recognize and bind specific RNA consensus sequences, the RBM10 RRM2 domain is structurally independent from both the RanBP2‐type zinc finger and RRM1 domains, and can bind independently . Taken together, determining how each RBM10 consensus functional motif contributes to the functionality of RBM10 will enable stronger predictions regarding the impact of RBM10 mutations on RBM10 function.…”

The devil is in the domain: understanding protein recognition of multiple RNA targets

“…In addition, a second RRM domain selects for C-rich sequences found in a second set of targets, including the cancerrelated Numb mRNA. Recognition of these targets does not mediate, in this case, a different function: protein binding to both GGA-containing and C-rich targets results in exon inclusion 46,47,48 . Instead, the two recognition modes of the protein have been proposed to target regions on opposite sides of the intron-exon boundary with different nucleobase enrichment.…”