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Functional analysis of a C. elegans trans-splice acceptor

R Conrad1, R F Liou, T Blumenthal

  • 1Department of Biology, Indiana University, Bloomington 47405.

Nucleic Acids Research
|February 25, 1993
PubMed
Summary

Researchers investigated trans-splicing in the C. elegans rol-6 gene. They found that altering splice sites and RNA sequence can shift splicing to a cryptic upstream site, revealing new insights into gene regulation.

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Area of Science:

  • Molecular Biology
  • Genetics
  • RNA Biology

Background:

  • Trans-splicing is a crucial RNA processing event in C. elegans.
  • The SL1 trans-splice leader typically joins downstream coding sequences.
  • The rol-6 gene serves as a model for studying trans-splicing mechanisms.

Purpose of the Study:

  • To investigate the factors influencing trans-splice site selection in the rol-6 gene.
  • To identify the role of the conserved AG dinucleotide and upstream pyrimidines in splice site recognition.
  • To understand how RNA sequence composition affects trans-splicing site choice.

Main Methods:

  • Analysis of splicing in C. elegans transformants with mutated rol-6 trans-splice acceptor sites.
  • Site-directed mutagenesis to alter the conserved AG and upstream pyrimidine sequences.
  • Manipulation of the RNA sequence composition between potential splice sites.

Main Results:

  • Inactivating the canonical trans-splice acceptor site (UUUCAG) of rol-6 led to splicing at a cryptic site 20 nt upstream.
  • Mutating the conserved AG dinucleotide at the normal splice site resulted in upstream cryptic site usage.
  • Changing upstream pyrimidines to adenines also promoted splicing at the cryptic site, suggesting a role similar to polypyrimidine tracts.
  • Altering the RNA sequence to be less A+U rich favored splicing at the upstream site over the downstream site.

Conclusions:

  • The conserved AG dinucleotide is essential for canonical trans-splice site recognition in rol-6.
  • Upstream pyrimidine sequences can influence splice site selection, potentially acting as regulatory elements.
  • The local RNA sequence environment, specifically A+U richness, plays a significant role in favoring the downstream splice site.

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