Messenger RNAs are exported from the nucleus as large ribonucleoprotein complexes (mRNPs). To date, proteins implicated in this process

Vertebrate TAP and its yeast ortholog Mex67p are involved in the export of messenger RNAs from the nucleus. TAP has also been implicated in the export of simian type D viral RNAs bearing the constitutive transport element (CTE). Although TAP directly interacts with CTE-bearing RNAs, the mode of interaction of TAP/Mex67p with cellular mRNAs is different from that with the CTE RNA and is likely to be mediated by protein-protein interactions. Here we show that Mex67p directly interacts with Yra1p, an essential yeast hnRNP-like protein. This interaction is evolutionarily conserved as Yra1p also interacts with TAP. Conditional expression in yeast cells implicates Yra1 p in the export of cellular mRNAs. Database searches revealed that Yra1p belongs to an evolutionarily conserved family of hnRNP-like proteins having more than one member in Mus musculus, Xenopus laevis, Caenorhabditis elegans, and Schizosaccharomyces pombe and at least one member in several species including plants. The murine members of the family directly interact with TAP. Because members of this protein family are characterized by the presence of one RNP-motif RNA-binding domain and exhibit RNA-binding activity, we called these proteins REF-bps for RNA and export factor binding proteins. Thus, Yra1p and members of the REF family of hnRNP-like proteins may facilitate the interaction of TAP/Mex67p with cellular mRNAs.

Vertebrate TAP (also called NXF1) and its yeast orthologue, Mex67p, have been implicated in the export of mRNAs from the nucleus. The TAP protein includes a noncanonical RNP-type RNA binding domain, four leucine-rich repeats, an NTF2-like domain that allows heterodimerization with p15 (also called NXT1), and a ubiquitin-associated domain that mediates the interaction with nucleoporins. Here we show that TAP belongs to an evolutionarily conserved family of proteins that has more than one member in higher eukaryotes. Not only the overall domain organization but also residues important for p15 and nucleoporin interaction are conserved in most family members. We characterize two of four human TAP homologues and show that one of them, NXF2, binds RNA, localizes to the nuclear envelope, and exhibits RNA export activity. NXF3, which does not bind RNA or localize to the nuclear rim, has no RNA export activity. Database searches revealed that although only one p15 (nxt) gene is present in the Drosophila melanogaster and Caenorhabditis elegans genomes, there is at least one additional p15 homologue (p15-2 [also called NXT2]) encoded by the human genome. Both human p15 homologues bind TAP, NXF2, and NXF3. Together, our results indicate that the TAP-p15 mRNA export pathway has diversified in higher eukaryotes compared to yeast, perhaps reflecting a greater substrate complexity. mRNAs are exported from the nucleus as large ribonucleoprotein complexes (mRNPs). To date, proteins directly implicated in this process include several nucleoporins and RNA binding proteins (hnRNPs), an RNA helicase of the DEADbox family (Dbp5), and the nuclear pore complex (NPC)-associated proteins Gle1p, TAP and Mex67p, and RAE1 (also called Gle2p) (reviewed in references 22, 28, and 32). Mex67p is essential for mRNA export in Saccharomyces cerevisiae, while RAE1 is essential for mRNA export in Schizosaccharomyces pombe (9,27,36). Their vertebrate homologues, TAP and RAE1, have also been implicated in the export of cellular mRNAs (6,8,12,15,20,31).We identified TAP as the cellular factor which is recruited by the constitutive transport element (CTE) of simian type D retroviruses to promote nuclear export of their genomic RNAs (12). In Xenopus oocytes, titration of TAP with an excess of CTE RNA prevents cellular mRNAs from exiting the nucleus (12,30,33). This suggests a role for this protein in the export of cellular mRNA.Considerable progress has been made in defining TAP structural and functional domains (see Fig. 1) and in identifying its binding partners. TAP partners include various nucleoporins (4, 17); p15 (also called NXT1), a protein related to nuclear transport factor 2 (NTF2) (7, 17); transportin, which mediates TAP nuclear import (4); and several mRNP-associated proteins, such as E1B-AP5, RAE1 (4), and members of the Yra1p/ REF family of proteins (37, 39). Binding of TAP to these mRNP-associated proteins is mediated by its N-terminal domain (residues 1 to 372) (4, 39). This domain includes a noncanonical RNP-type RNA binding domain (RBD)...

TAP-p15 heterodimers have been implicated in the export of mRNAs through nuclear pore complexes (NPCs). We report a structural analysis of the interaction domains of TAP and p15 in a ternary complex with a Phe-Gly (FG) repeat of an NPC component. The TAP-p15 heterodimer is structurally similar to the homodimeric transport factor NTF2, but unlike NTF2, it is incompatible with either homodimerization or Ran binding. The NTF2-like heterodimer functions as a single structural unit in recognizing an FG repeat at a hydrophobic pocket present only on TAP and not on p15. This FG binding site interacts synergistically with a second site at the C terminus of TAP to mediate mRNA transport through the pore. In general, our findings suggest that FG repeats bind with a similar conformation to different classes of transport factors.