The RNA-binding Motif Protein 15B (RBM15B/OTT3) Acts as Cofactor of the Nuclear Export Receptor NXF1

Hiroaki Uranishi,Andrei S Zolotukhin,Susan Lindtner,Soren Warming,Gen-Mu Zhang,Jenifer Bear,Neal G Copeland,Nancy A Jenkins,George N Pavlakis,Barbara K Felber

doi:10.1074/jbc.m109.040113

Abstract

The human SPEN family proteins SHARP, RBM15/OTT1, and RBM15B/OTT3 share the structural domain architecture but show distinct functional properties. Here, we examined the function of OTT3 and compared it with its paralogues RBM15 and SHARP. We found that OTT3, like RBM15, has post-transcriptional regulatory activity, whereas SHARP does not, supporting a divergent role of RBM15 and OTT3. OTT3 shares with RBM15 the association with the splicing factor compartment and the nuclear envelope as well as the binding to mRNA export factors NXF1 and Aly/REF. Mutational analysis revealed direct interaction of OTT3 and RBM15 with NXF1 via their C-terminal regions. Biochemical and subcellular localization studies showed that OTT3 and RBM15 also interact with each other in vivo, further supporting a shared function. Genetic knockdown of RBM15 in mouse is embryonically lethal, indicating that OTT3 cannot compensate for the RBM15 loss, which supports the notion that these proteins, in addition to sharing similar activities, likely have distinct biological roles.

Highlights

The SPEN family proteins share a domain architecture comprising of three N-terminal RNA-binding domains (RRMs)4 and a Spen paralogue and orthologue C-terminal (SPOC) domain [1,2,3]
NXF1 (Fig. 4C, mutants containing the C-terminal regions are Vivo—By screening known NXF1 cofactors using coimmunoindicated in bold type). These experiments revealed that OTT3 precipitation assays, we found that OTT3 and RBM15 interacted with NXF1 via its C-terminal region, associated with Aly/REF, a RNA-binding factor that was implisimilarly to RBM15
The SPEN family had diversified in the course of evolution, giving rise to three clearly orthologous groups in vertebrates that are represented by SHARP, RBM15, and OTT3 in humans, and recent work has linked the “small” RBM15 and OTT3 proteins to post-transcriptional gene control

Summary

Introduction

The SPEN (split end) family proteins share a domain architecture comprising of three N-terminal RNA-binding domains (RRMs)4 and a Spen paralogue and orthologue C-terminal (SPOC) domain [1,2,3]. We have reported that RBM15, a primarily nuclear protein, binds to the general mRNA export factor NXF1 [20], suggesting that RBM15 can tether the RTE-containing RNA to the NXF1 export pathway. We found that OTT3, like RBM15 but unlike SHARP, acts at the post-transcriptional level to activate reporter gene expression.

Results

Conclusion