A splicing silencer that regulates smooth muscle specific alternative splicing is active in multiple cell types

Area of Science:

• Molecular Biology
• Gene Regulation
• Alternative Splicing

Background:

• Alternative splicing of alpha-tropomyosin (alpha-TM) involves mutually exclusive selection of exons 2 and 3.
• Exon 2 selection in smooth muscle (SM) cells depends on the inhibition of exon 3.

Purpose of the Study:

• To investigate the role of upstream regulatory elements (UREs) in alpha-TM alternative splicing.
• To determine if UREs are smooth muscle-specific and identify potential binding proteins.

Main Methods:

• Transfection of cells with tropomyosin reporters containing UREs.
• Point mutations and multimerization of UREs.
• Co-transfection with CUG-binding protein family members.

Main Results:

• Mutations or substitutions of the UGC-containing URE disrupted alpha-TM splicing patterns.
• Multimerization of UREs enhanced exon skipping in both SM and non-SM cells.
• The strength of UREs is modulated by cell-specific regulatory factors, not intrinsically SM-specific.

Conclusions:

• The URE's functional strength is fine-tuned by differences in regulatory factor activities between cell types.
• CUG-binding proteins do not appear to mediate the repression of tropomyosin exon 3.