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A protein related to splicing factor U2AF35 that interacts with U2AF65 and SR proteins in splicing of pre-mRNA

Nature volume 388, pages 397–400 (1997)Cite this article

Abstract

Recognition of a functional 3′ splice site in pre-mRNA splicing requires a heterodimer of the proteins U2AF65/U2AF35. U2AF65 binds to RNA at the polypyrimidine tract1,2, whereas U2AF35 is thought to interact through its arginine/serine-rich (RS) domain with other RS-domain-containing factors bound at the 5′ splice site, assembled in splicing enhancer complexes, or associated with the U4/U6.U5 small nuclear ribonucleoprotein complex3,4,5,6,7. It is unclear, however, how such network interactions can all be established through the small RS domain in U2AF35. Here we describe the function of a U2AF35-related protein (Urp), which is the human homologue of a mouse imprinted gene. Nuclear extracts depleted of Urp are defective in splicing, but activity can be restored by addition of recombinant Urp. U2AF35 could not replace Urp in complementation, indicating that their functions do not overlap. Co-immunodepletion showed that Urp is associated with the U2AF65/U2AF35 heterodimer. Binding studies revealed that Urp specifically interacts with U2AF65 through a U2AF35-homologous region and with SR proteins (a large family of RS-domain-containing proteins) through its RS domain. Therefore, Urp and U2AF35 may independently position RS-domain-containing factors within spliceosomes.

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Figure 1: Urp structure and sequence comparison with U2AF35.
Figure 2: Urp is an essential splicing factor.
Figure 3: Urp is associated with the U2AF heterodimer.
Figure 4: Urp interacts with U2AF65 and SR proteins in vitro and in yeast.

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Acknowledgements

We thank G. Hannon for yeast two-hybrid libraries; J.-L. Guan for raising anti-Urp antibodies; J. Fleckner and M. Green for U2AF cDNAs; P. Zuo and T. Maniatis for anti-U2AF antibodies and purified U2AF35 protein; J. Wu for two-hybrid plasmids and for advice; and members of our laboratory, especially H.-Y. Wang and J. Dyck, for suggestions and contributions. X.-D.F. is a Searle Scholar. H.T. was supported by postdoctoral fellowships from the Human Frontier Science Program and La Societé de Secours des Amis de la Science. J.W. was supported by a postdoctoral fellowship from the NIH. This work was supported by grants to X.-D.F. from the NIH.

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Author notes

  1. Hlne Tronchre and Jiwu Wang: These authors contributed equally to this work.

  2. Correspondence and requests for materials should be addressed to X.-D.F.

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  1. Division of Cellular and Molecular Medicine, Department and School of Medicine, University of California, San Diego, 9500 Gilman Drive, La Jolla, 92093-0651, California, USA

    Xiang-Dong Fu

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Tronchre, H., Wang, J. & Fu, XD. A protein related to splicing factor U2AF35 that interacts with U2AF65 and SR proteins in splicing of pre-mRNA. Nature 388, 397–400 (1997). https://doi.org/10.1038/41137

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