Functional domains of the human splicing factor ASF/SF2.
The human splicing factor ASF/SF2 displays two predominant activities in in vitro splicing assays: (i) it is an essential factor apparently required for all splices and (ii) it is able to switch utilization of alternative 5' splice sites in a concentration-dependent manner. ASF/SF2 is the prototype of a family of proteins typified by the presence of one or two RNP-type RNA binding domains (RBDs) and a region highly enriched in repeating arginine-serine dipeptides (RS regions). Here we describe a functional analysis of ASF/SF2, which defines several regions essential for one, or both, of its two principal activities, and provides insights into how this type of protein functions in splicing. Two isoforms of the protein, which arise from alternative splicing, are by themselves inactive, but each can block the activity of ASF/SF2, thereby functioning as splicing repressors. Some, but not all, mutations in the RS region prevent ASF/SF2 from functioning as an essential splicing factor. However, the entire RS region can be deleted without reducing splice site switching activity, indicating that it is not absolutely required for interaction with other splicing factors. Experiments with deletion and substitution mutants reveal that the protein contains two related, but highly diverged, RBDs, and that both are essential for activity. Each RBD by itself retains the ability to bind RNA, although optimal binding requires both domains.
Zuo
P
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Department of Biological Sciences, Columbia University, New York, NY 10027.
Manley
J L
JL
D000595:Amino Acid Sequence; D001483:Base Sequence; D001665:Binding Sites; D003001:Cloning, Molecular; D004247:DNA; D004926:Escherichia coli; D006801:Humans; D008969:Molecular Sequence Data; D009687:Nuclear Proteins / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D017354:Point Mutation; D012326:RNA Splicing; D016601:RNA-Binding Proteins / Q000737:chemistry / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D000068103:Serine-Arginine Splicing Factors
D009687:Nuclear Proteins; D016601:RNA-Binding Proteins; D000068103:Serine-Arginine Splicing Factors; D004247:DNA