The role of specific protein-RNA and protein-protein interactions in positive and negative control of pre-mRNA splicing by Transformer 2.
We have investigated the function of different structural domains of the Drosophila splicing regulator Transformer 2 (Tra2). We find that the ribonucleoprotein consensus sequence (RNP-CS) of Tra2 is required for male fertility and positive and negative control of alternative splicing in transgenic flies, as well as for in vitro binding of recombinant Tra2 to doublesex and tra2 pre-mRNAs. Thus, all of the known functions of Tra2 require specific protein-RNA interactions. We also show that one of the two arginine-serine (RS)-rich domains of Tra2 is dispensable, while the other is essential for all of the in vivo functions. Part of this domain is also required for RNA binding in vitro. Significantly, the essential RS domain is also required for specific protein-protein interactions. We find that Tra2 interacts with itself, with the splicing regulator Transformer, and with the general splicing factor SF2 in vitro and in the yeast two-hybrid system. These results demonstrate that both protein-RNA and protein-protein interactions are involved in tra2-dependent activation and repression of alternative splicing.
Amrein
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Howard Hughes Medical Institute, Columbia University, New York, New York 10032.
Hedley
M L
ML
Maniatis
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T
D000818:Animals; D001483:Base Sequence; D017931:DNA Primers / Q000737:chemistry; D004331:Drosophila melanogaster / Q000235:genetics*; D005260:Female; D005298:Fertility; D008297:Male; D008969:Molecular Sequence Data; D009694:Nucleic Acid Precursors / Q000378:metabolism; D011485:Protein Binding; D012326:RNA Splicing; D012333:RNA, Messenger / Q000235:genetics*; D016601:RNA-Binding Proteins / Q000502:physiology; D012261:Ribonucleoproteins / Q000502:physiology*; D017384:Sequence Deletion; D012732:Sex Determination Analysis; D013329:Structure-Activity Relationship
D017931:DNA Primers; D009694:Nucleic Acid Precursors; D012333:RNA, Messenger; D016601:RNA-Binding Proteins; D012261:Ribonucleoproteins