Identification and characterization of the mouse nuclear export factor (Nxf) family members

Abstract

TAP/hNXF1 is a key factor that mediates general cellular mRNA export from the nucleus, and its orthologs are structurally and functionally conserved from yeast to humans. Metazoans encode additional proteins that share homology and domain organization with TAP/hNXF1, suggesting their participation in mRNA metabolism; however, the precise role(s) of these proteins is not well understood. Here, we found that the human mRNA export factor hNXF2 is specifically expressed in the brain, suggesting a brain-specific role in mRNA metabolism. To address the roles of additional NXF factors, we have identified and characterized the two Nxf genes, Nxf2 and Nxf7, which together with the TAP/hNXF1's ortholog Nxf1 comprise the murine Nxf family. Both mNXF2 and mNXF7 have a domain structure typical of the NXF family. We found that mNXF2 protein is expressed during mouse brain development. Similar to TAP/hNXF1, the mNXF2 protein is found in the nucleus, the nuclear envelope and cytoplasm, and is an active mRNA export receptor. In contrast, mNXF7 localizes exclusively to cytoplasmic granules and, despite its overall conserved sequence, lacks mRNA export activity. We concluded that mNXF2 is an active mRNA export receptor similar to the prototype TAP/hNXF1, whereas mNXF7 may have a more specialized role in the cytoplasm.

The fragile X mental retardation protein interacts with a distinct mRNA nuclear export factor NXF2

“…2, panel d). This pattern resembles that seen in transient transfection experiments using epitope-tagged NXF2 fusion proteins (Herold et al 2000;Tan et al 2005). Further, FMRP and NXF2 colocalize at the perinuclear region in spermatogonia in our double staining immunofluorescence experiments (Fig.…”

“…1A). Metazoans encode additional NXF1 orthologs, including NXF2 (Herold et al 2000;Sasaki et al 2005;Tan et al 2005). The overall domain organization of NXF2 follows a highly conserved modular architecture (Fig.…”

“…Like NXF1, NXF2 interacts with p15 and with components of the nuclear pore complex (Herold et al 2000), and is able to stimulate the nuclear export of reporter mRNAs in transient transfection assays (Sasaki et al 2005;Tan et al 2005). Intriguingly, NXF2 is present in both nucleus and cytoplasm of transfected cells at steady state (Tan et al 2005), suggesting a possible role in the cytoplasm in addition to the nucleus. In the process of studying RNA-binding proteins in the mouse testis, we unexpectedly found that NXF2 and FMRP specifically interact with each other.…”

RNA-binding Motif Protein 15 Binds to the RNA Transport Element RTE and Provides a Direct Link to the NXF1 Export Pathway

“…We found that the human NXF1 bound to RBM15 in vitro, whereas no interactions with luciferase, UAP56 or "empty" GST beads were observed. Similarly, we found that the mouse NXF2, a highly related mRNA export factor (27), interacted with RBM15 (data not shown). The binding assays were performed in the presence of RNase A, demonstrating that the identified interactions of RBM15 with the NXF proteins are RNA-independent.…”

“…In Vitro Protein Binding Assays-Metabolically labeled reticulocyte-produced proteins were synthesized in a coupled transcription/translation system (TNT T7 Coupled Reticulocyte Lysate System, Promega) and used in binding reactions con- taining ϳ0.5 g of E. coli-produced GST-tagged proteins (27). The binding was performed in 200 l of RBB-400 buffer in the presence of 100 g of RNase A.…”