A serine/arginine-rich domain in the human U1 70k protein is necessary and sufficient for ASF/SF2 binding

Wenhong Cao, Mariano A. Garcia-Blanco

Research output: Contribution to journalArticlepeer-review

49 Scopus citations

Abstract

Critical protein-protein interactions among pre-mRNA splicing factors determine splicing efficiency and specificity. The serine/arginine proteins, a family of factors characterized by the presence of an RNA recognition motif and an arginine/serine domain, are essential for constitutive splicing and required for some alternative splicing decisions. ASF/SF2, SC35, and other members of the serine/arginine family, interact with the 70k protein of the U1 small nuclear ribonucleoprotein. The binding of this protein with ASF/SF2 is thought to enhance recognition of the 5' splice site of pre-mRNAs by the U1 small nuclear ribonucleoprotein. It has been clearly documented that the arginine/serine domain of ASF/SF2 is responsible for binding to the U1 70k protein. In this manuscript we characterize the segment in the human U1 70k protein that is both necessary and sufficient for ASF/SF2 binding. A domain within this segment, which begins with Arg240 and ends with Asp270, was shown to bind specifically to the arginine/serine domain of ASF/SF2 using a yeast two-hybrid system and a far Western assay. Mutational analysis of this segment suggested that several arginines are critical for the interaction with ASF/SF2 and for phosphorylation by SRPK1. Inspection of the sequence of the Arg248 to Asp270 region suggested this as an arginine/serine-like domain in U1 70k protein, and the data presented in this manuscript strongly support this view. Inspection of the human U1 70k protein sequence, comparison with homologues in other animal species, and mutational analysis indicated the importance of the sequence Arg-Arg-Arg-Ser-Arg-Ser-Arg-Asp, which is found repeated twice in the region from Arg248 to Asp270 in the human protein.

Original languageEnglish (US)
Pages (from-to)20629-20635
Number of pages7
JournalJournal of Biological Chemistry
Volume273
Issue number32
DOIs
StatePublished - Aug 7 1998
Externally publishedYes

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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