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A splicing enhancer exhibits both constitutive and regulated activities
1Department of Biochemistry and Molecular Biology, Harvard University, Cambridge, Massachusetts 02138.
Genes & Development
|July 15, 1994
Summary
Transformer (Tra) and Transformer2 (Tra2) proteins regulate Drosophila doublesex (dsx) splicing. The dsx repeat element (dsxRE) acts as a splicing enhancer, with Tra and Tra2 stabilizing its function at a distance.
Area of Science:
- Molecular Biology
- Developmental Biology
- Genetics
Background:
- Transformer (Tra) and Transformer2 (Tra2) are key regulators of sex-specific alternative splicing in Drosophila.
- These proteins bind to the dsx repeat element (dsxRE), a splicing enhancer crucial for regulating doublesex (dsx) pre-mRNA splicing.
- The dsxRE is located downstream of a female-specific 3' splice site, and its precise function in relation to Tra/Tra2 and distance has been investigated.
Purpose of the Study:
- To investigate the mechanism by which the dsxRE functions as a splicing enhancer.
- To determine the role of Transformer (Tra) and Transformer2 (Tra2) in mediating the enhancer activity of the dsxRE.
- To explore the influence of the distance between the dsxRE and the splice site on its regulatory function.
Main Methods:
- In vitro splicing assays were performed to assess the enhancer activity of the dsxRE.
- Experiments were conducted to evaluate the dsxRE's function at varying distances from the 3' splice site.
- Protein binding studies were used to analyze the interactions of SR proteins and Tra/Tra2 with the dsxRE.
Main Results:
- The dsxRE can function as a Tra and Tra2-independent splicing enhancer when positioned within 150 nucleotides of the 3' splice site.
- The constitutive activity of the dsxRE is attributed to weak interactions with SR proteins and general splicing factors.
- Transformer (Tra) and Transformer2 (Tra2) enhance the dsxRE's function at a distance by stabilizing protein-dsxRE interactions.
Conclusions:
- The dsxRE possesses intrinsic enhancer activity mediated by general splicing factors.
- Transformer (Tra) and Transformer2 (Tra2) play a critical role in enabling the dsxRE to function over longer distances, thereby fine-tuning sex-specific alternative splicing.
- This study elucidates the intricate mechanism of splicing regulation by enhancer elements and specific protein factors in Drosophila development.