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Science 20 September 1996:
Vol. 273. no. 5282, pp. 1706 - 1709
DOI: 10.1126/science.273.5282.1706
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Reports

Interaction of U2AF65 RS Region with Pre-mRNA of Branch Point and Promotion Base Pairing with U2 snRNA

Juan Valcárcel, * Rajesh K. Gaur, Ravinder Singh, Michael R. Green dagger

The mammalian splicing factor U2AF65 binds to the polypyrimidine tract adjacent to the 3' splice site and promotes assembly of U2 small nuclear ribonucleoprotein on the upstream branch point, an interaction that involves base pairing with U2 small nuclear RNA (snRNA). U2AF65 contains an RNA binding domain, required for interaction with the polypyrimidine tract, and an arginine-serine-rich (RS) region, required for U2 snRNP recruitment and splicing. Here it is reported that binding of U2AF65 to the polypyrimidine tract directed the RS domain to contact the branch point and promoted U2 snRNA-branch point base pairing even in the absence of other splicing factors. Analysis of RS domain mutants indicated that the ability of U2AF65 to contact the branch point, to promote the U2 snRNA-branch point interaction, and to support splicing are related activities, requiring only a few basic amino acids. Thus, the U2AF65 RS domain plays a direct role in modulating spliceosomal RNA-RNA interactions.

Howard Hughes Medical Institute, Program in Molecular Medicine, University of Massachusetts Medical Center, 373 Plantation Street, Worcester, MA 01605, USA.
*   Present address: Gene Expression Programme, European Molecular Biology Laboratory, Meyerhofstrasse 1, D-69117 Heidelberg, Germany.

dagger    To whom correspondence should be addressed.

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Interaction between Subunits of Heterodimeric Splicing Factor U2AF Is Essential In Vivo.
D. Z. Rudner, R. Kanaar, K. S. Breger, and D. C. Rio (1998)
Mol. Cell. Biol. 18, 1765-1773
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Molecular genetic analysis of the heterodimeric splicing factor U2AF: the RS domain on either the large or small Drosophila subunit is dispensable in vivo.
D. Z. Rudner, K. S. Breger, and D. C. Rio (1998)
Genes & Dev. 12, 1010-1021
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A cooperative interaction between U2AF65 and mBBP/SF1 facilitates branchpoint region recognition.
J. A. Berglund, N. Abovich, and M. Rosbash (1998)
Genes & Dev. 12, 858-867
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SR proteins are sufficient for exon bridging across an intron.
J. M. Stark, D. P. Bazett-Jones, M. Herfort, and M. B. Roth (1998)
PNAS 95, 2163-2168
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A Spliceosomal Recycling Factor That Reanneals U4 and U6 Small Nuclear Ribonucleoprotein Particles.
P. L. Raghunathan and C. Guthrie (1998)
Science 279, 857-860
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Sip1, a Novel RS Domain-Containing Protein Essential for Pre-mRNA Splicing.
W.-J. Zhang and J. Y. Wu (1998)
Mol. Cell. Biol. 18, 676-684
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U2AF65 recruits a novel human DEAD box protein required for the U2 snRNP-branchpoint interaction..
J Fleckner, M Zhang, J Valcarcel, and M R Green (1997)
Genes & Dev. 11, 1864-1872
   Abstract »    PDF »
Targeting of U2AF65 to Sites of Active Splicing in the Nucleus.
M. Gama-Carvalho, R. D. Krauss, L. Chiang, J. Valcarcel, M. R. Green, and M. Carmo-Fonseca (1997)
J. Cell Biol. 137, 975-987
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Phosphorylation of the ASF/SF2 RS domain affects both protein-protein and protein-RNA interactions and is necessary for splicing..
S H Xiao and J L Manley (1997)
Genes & Dev. 11, 334-344
   Abstract »    PDF »
SC35-mediated reconstitution of splicing in U2AF-depleted nuclear extract.
A. M. MacMillan, P. S. McCaw, J. D. Crispino, and P. A. Sharp (1997)
PNAS 94, 133-136
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Kinetic Role for Mammalian SF1/BBP in Spliceosome Assembly and Function after Polypyrimidine Tract Recognition by U2AF.
S. Guth and a. J. Valcarcel (2000)
J. Biol. Chem. 275, 38059-38066
   Abstract »    Full Text »    PDF »
Mutational Analysis of 3' Splice Site Selection during trans-Splicing.
H. S. Hummel, R. D. Gillespie, and J. Swindle (2000)
J. Biol. Chem. 275, 35522-35531
   Abstract »    Full Text »    PDF »
Nucleocytoplasmic Shuttling of Heterodimeric Splicing Factor U2AF.
M. Gama-Carvalho, M. P. Carvalho, A. Kehlenbach, J. Valcarcel, and M. Carmo-Fonseca (2001)
J. Biol. Chem. 276, 13104-13112
   Abstract »    Full Text »    PDF »
Three-dimensional Interaction of Phi29 pRNA Dimer Probed by Chemical Modification Interference, Cryo-AFM, and Cross-linking.
Y. Mat-Arip, K. Garver, C. Chen, S. Sheng, Z. Shao, and P. Guo (2001)
J. Biol. Chem. 276, 32575-32584
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N. A. Patel, C. E. Chalfant, J. E. Watson, J. R. Wyatt, N. M. Dean, D. C. Eichler, and D. R. Cooper (2001)
J. Biol. Chem. 276, 22648-22654
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J. Rappsilber, P. Ajuh, A. I. Lamond, and M. Mann (2001)
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M. A. Skabkin, V. Evdokimova, A. A. M. Thomas, and L. P. Ovchinnikov (2001)
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C. Chen, S. Sheng, Z. Shao, and P. Guo (2000)
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M. Li and P. H. Pritchard (2000)
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